Gastroenteritis cases associated with non-toxigenic strains of Vibrio parahaemolyticus have been reported in many countries, suggesting the contribution of novel virulence factors. One candidate is zonula occludens toxin (Zot), which increases the intestinal permeability by other bacteria. Recently we identified prophages belonging to the Inoviridae family encoding putative Zot-like toxins in Chilean strains. Based on this information we performed sequence-based analyses of these toxins, followed by phylogenetic and structural analyses using computational tools. Our results showed that Zots found in Chilean V. parahaemolyticus strains are grouped into three different phylogenetic clusters, sharing two conserved motifs (Walker A and B) in their N-terminal region. These motifs are also conserved in Zots from the human pathogens Vibrio cholerae, Neisseria meningitidis and Campylobacter concisus. Although Zots of V. parahaemolyticus do not possess the FCIGRL sequence responsible for the effects produced by V. cholerae, they do possess a conserved secondary structure within their C-terminal region with Zots proteins able to disrupt the intestinal barrier, which is interesting since it has been suggested that the structure and not the Zot sequence would be responsible for the biological effects. This preliminary study provides the basis to study the function of Zots found in V. parahaemolyticus on the intestinal barrier and their possible role as a virulence factor.

It has been regarded that Vibrio cholerae O1 inhabit in environmental water. As many cholera patients emerge in Kolkata, it has been thought that V. cholerae O1 is easily detected in environmental water in Kolkata. However, the detection of V. cholerae O1 is rare, though other V. cholerae (NAG Vibrio) is constantly detected. To clear the reason for the difference of the detection rate of two Vibrios, we examined the viability of V. cholera O1 and NAG Vibrios in low ionic strength aquatic medium. We observed greater declining viability of V. cholerae O1 possessing cholera toxin gene (ctx) in low ionic strength solution, but the decline of NAG Vibrios non-possessing ctx is small. To evaluate the concerning of ctx in the viability, we examined the viabilities of V. cholerae O1which do not possess ctx and NAG Vibrios possessing ctx under the same condition. The result indicated that the existence of the ctx induces the decrease the viability of the host in low ionic strength solution. The decrease observed in this experiment might relate with the low detection of V. cholerae O1 possessing ctx in environmental water, though NAG Vibrio is constantly detected.

PirAB is a binary protein complex secreted by specific strains of Vibrio parahaemolyticus (Vp) that harbor the pVA1 virulence plasmid and express PirAVp and PirBVp toxins. PirABVp causes acute hepatopancreatic necrosis disease (AHPND), a newly emergent penaeid shrimp disease that can cause 70–100% mortality and has resulted in great economic losses since its appearance. The cy-totoxic effect of PirABVp on the epithelial cell of the shrimp hepatopancreas has been extensively reported. Our studies found that the PirBVp subunit has lectin activity and recognizes mucin-like O-glycosidic structures in the shrimp hepatopancreas. The PirAVp subunit may have a stabiliza-tion function of the binary complex. However, we also found that Vp AHPND changes the water microbiota community structure and causes a significant reduction in several bacteria, especially Neptuniibacter spp. We propose that the PirABvp toxin could exhibit a dual role: damage the shrimp hepatopancreas and kill surrounding bacteria.

Saliva, as a diagnostic medium, offers a promising alternative to blood by virtue of its non-invasive collection, which enhances patient compliance, especially in paediatric and geriatric populations. In this study, we assessed the utility of saliva as a non-invasive medium for measuring V. cholerae-specific serum antibodies in naturally infected individuals. We tested paired serum and saliva samples obtained from a total of 63 cholera patients enrolled in a cohort study. Vibriocidal antibodies assay (IgM/IgG) as markers for accurate determination was used to determine cholera specific antibody levels. Using receiver operating characteristics (ROC) curve, we found that the best cut-off that maximizes (sensitivity + specificity) is 10 titers. At this saliva titer, the sensitivity is 76.9% (95%CI: 60.9%, 87.7%) and specificity is 80.0% (95%CI: 56.6%, 92.5%). Using Spearman’s correlation coefficient, we also found evidence of a positive correlation between Vibrio Cholerae saliva and serum antibodies (rho=0.66, p<0.001). In conclusion, saliva-based diagnostic cholera tests has high diagnostic accuracy, and would be advantageous, cheaper, and quicker for early diagnosis of severe cholera outcomes.

Vibrio parahaemolyticus is an important human pathogen that is currently the leading cause of shellfish-borne gastroenteritis in the world. Particularly, the pandemic strain has the capacity to induce cytotoxicity and enterotoxicity through its Type 3 Secretion System (T3SS2) that leads to massive cell death. However, the specific mechanism by which T3SS2 induces cell death remain unclear and the contribution in mitochondrial stress is not fully understood. In this work, we evaluated the contribution of T3SS2 of V. parahaemolyticus to generate mitochondrial stress and cell death during infection in human intestinal HT-29 cells. To evaluate this, infection assays were carried out to evaluate mitochondrial transition pore opening, mitochondrial fragmentation, ATP quantification, and cell viability during infection. Our results showed that the Δvscn1 (T3SS2+) mutant strain contributes to generating the sustained opening of the mitochondrial transition pore. Furthermore, it generates perturbations in the ATP production in infected cells, leading to a significant decrease in cell viability and loss of membrane integrity. Our results suggest that T3SS2 from V. parahaemolyticus plays a role in generating mitochondrial stress and cell death in human intestinal HT-29 cells. It is important to highlight that this study represents the first report indicating the involvement of T3SS2 from V. parahaemolyticus in generating mitochondrial stress, its impact on the mitochondrial pore, and its effect on ATP production in human cells.

Temperature is a critical factor that influences the proliferation of pathogens in hosts. The impact of temperature on pathogens is commonly explored in controlled and constant temperatures. Experiments under varying environmental temperature are becoming more frequent, however, testing every temperature scenario à la carte is unachievable. One example of this is the human pathogen Vibrio parahaemolyticus (Vp) in oysters. Here, a predictive model was developed for predicting the growth of Vp in oysters under varying ambient temperature. The model was fitted and evaluated against data from experiments studying growth and inactivation of Vp in oysters at eight constant temperatures. Once evaluated, Vp dynamics in oysters were estimated at different post-harvest varying temperature scenarios affected by water and air temperature, and different ice treatment timing. The model performed adequately under varying temperature, reflecting that (i) increasing temperature, particularly in hot summers, favors a rapid Vp growth in oysters, resulting in a very high risk of gastroenteritis in humans after consumption of a serving of raw oysters, (ii) pathogen inactivation due to day/night oscillations, and more evidently, due to ice treatments, (iii) ice treatment is much more effective limiting risk of illness when applied immediately onboard compared to dockside. The model results to be a promising tool for improving the understanding of the Vp-oyster system and support studies on public health impact of pathogenic Vp associated with raw oyster consumption. Although robust validation of the model predictions is needed, initial results and evaluation show the potential of the model to be easily modified to match similar systems where the temperature is a critical factor shaping the proliferation of pathogens in hosts.

Acellular bacterial ghosts (BGs) are empty non-living bacterial cell envelopes, commonly generated by controlled expression of the cloned lysis gene E of bacteriophage PhiX174. In this study, Vibrio parahaemolyticus ghosts (VPGs) were generated by chemically induced lysis and the method is based on minimum inhibitory concentration (MIC) of sodium hydroxide (NaOH), acetic acid, boric acid, citric acid, maleic acid, hydrochloric acid and sulfuric acid. The MIC values of the respective chemicals were 3.125, 6.25, < 50.0, 25.0, 6.25, 1.56 and 0.781 mg/ml. Except boric acid, the lysis efficiency was reached more than 99.99% at 5 min after treatment of all chemicals. Among those chemicals, NaOH-induced VPGs showed completely DNA-free that was confirmed by quantitative real-time PCR. Besides, lipopolysaccharides (LPS) extracted from the NaOH-induced VPGs showed no distinctive band on SDS-PAGE gel after silver staining. On the other hand, LPS extracted from wild-type bacterial cells as well as the organic acids-induced VPGs showed triple major bands and LPS extracted from the inorganic acids-induced VPGs showed double bands. It suggests that some surface structures in LPS of the NaOH-induced VPGs may be lost, weakened or modified by the MIC of NaOH. Nevertheless, Limulus amoebocyte lysate assay revealed that there is no significant difference in endotoxic activity between the NaOH-induced VPGs and wild-type bacterial cells. Macrophages exposed to the NaOH-induced VPGs at 0.5 × 106 CFU/mL showed cell viability of 97.9%, however the MIC of NaOH did not reduce the cytotoxic effect of wild-type bacterial cells. Like Escherichia coli LPS, the NaOH-induced VPGs are an excellent activator of pro-inflammatory cytokines (IL-1β and iNOS), anti-inflammatory cytokine (IL-10) and dual activities (IL-6) in the stimulated macrophage cells. On the other hand, the induction of TNF-α mRNA was remarkable in the macrophages exposed with wild-type cells. Scanning electron microscopy showed the formation of trans-membrane lysis tunnel structures in the NaOH-induced VPGs. SDS-PAGE and agarose gel electrophoresis also confirmed that cytoplasmic proteins and genomic DNA released from the VPGs to culture medium through the lysis tunnel structures. Taken together, all these results indicated that the NaOH-induced VPGs show the potency of safe, economical and effective inactivated bacterial vaccine candidate.

Porins are crucial proteins located in the outer membrane that directly influence antimicrobial resistance mechanisms and virulence in bacteria. In this study, a porin gene (Vp-porin) was cloned in V. parahaemolyticus, and the function of Vp-porin on biological characteristics and virulence was investigated. The results of sequence analysis showed that Vp-porin is highly conserved in Vibrio spp. and the predicted 3D structure showed it could form a transmembrane channel with 20 β barrel. Membrane permeabilization provides evidence that membrane integrity of ∆Vp-porin was damaged and the sensitivity to tetracycline, polymyxin B, rifampicin and cephalothin of ∆Vp-porin obvious increased. In addition, loss of Vp-porin damaged motility due to down-regulated flagellar synthesis. Besides, ∆Vp-porin exhibited attenuated cytotoxicity to Tetra-hymena. The relative survival rate of Tetrahymena infection with ∆Vp-porin was 86%, which is much high than that with WT (49%). Take together, Vp-porin in V. parahaemolyticus plays various roles in biological characteristics in membrane integrity, antimicrobial resistance and motility, and contributes to virulence.

Cholera is a severe diarrheal disease caused by Vibrio cholerae, a natural inhabitant of brackish water. Effective control of cholera outbreaks depends on prompt detection of the pathogen from clinical specimens and tracking its source in the environment. Although the epidemiology of chol-era is well studied, rapid detection of V. cholerae remains a challenge, and data on its abundance in environmental sources are limited. Here, we describe a sensitive molecular quantification as-say by qPCR, which can be used on-site in low resource settings directly on water without the need for DNA extraction. This newly optimized method exhibited 100% specificity for total V. cholerae as well as V. cholerae O1 and allowed detection of as few as three target genome copies per reaction. The limit of detection is as low as 5 × 10E3 genome copies/L of water after concentrat-ing biomass from the sample. The ability to perform qPCR directly on water samples, portable features of the equipment, stability of the reagents at 4°C and user-friendly online software facili-tate fast quantitative analysis of V. cholerae. These characteristics make this assay extremely use-ful for field research in resource-poor settings and could support continuous monitoring in cholera endemic areas.

The genome of Vibrio cholerae contains three structural genes for the NhaP-type cation-proton antiporter paralogues, Vc-NhaP1, 2 and 3 mediating exchange of K+ and or Na+ for protons across the membrane. Based on phenotype analysis of chromosomal Vc-NhaP1, 2 and 3 triple deletion mutants we suggested that Vc-NhaP paralogues might play a role in the Acid Tolerance Response (ATR) of V. cholerae as it passes through the gastric acid barrier of the stomach. Comparison of the biochemical properties of Vc-NhaP isoforms revealed that Vc-NhaP2 is the most active among all three paralogues. Therefore, Vc-NhaP2 antiporter is a plausible therapeutic target for developing novel inhibitors targeting these ion exchangers. Our structural and mutational analysis of Vc-NhaP2 identified a putative cation binding pocket formed by antiparallel extended regions of two transmembrane segments (TMSs V/XII) along with TMS VI. Molecular Dynamics (MD) simulations suggested that the flexibility of TMS-V/XII is crucial for the intra-molecular conformational events in Vc-NhaP2. In this study, we developed some putative Vc-NhaP2 inhibitors from Amiloride analogs (AAs). Amiloride is a potent inhibitor of human Na+/H+ exchanger-1 (NHE1). Based on the pharmacokinetic properties and potential binding affinity scores we chose six AAs showing high binding affinity scores to Vc-NhaP2. In silico, the six AAs interacted with the functionally important amino acid residues located in TMSs III, IV, V, VI, VIIII and IX either from the cytoplasmic side (three AAs) or the periplasmic side (three AAs) of Vc-NhaP2. Four AAs were modified to reduce their toxicity profile compared to the original AAs. Molecular docking of the modified AAs revealed promising binding. The four selected drugs interacted with functionally important amino acid residues located on the cytoplasmic side of TMS VI, the extended chain region of TMS V and TMS XII and the loop region between TMSs VIIII and IX. Molecular dynamics simulations revealed that binding of the selected drugs destabilized the Vc-NhaP2 and altered the flexibility of functionally important TMS VI.

Vibrio parahaemolyticus is an important foodborne bacterium that causes severe gastroenteritis following the consumption of contaminated seafood. To identify V. parahaemolyticus and determine its pathogenicity, the U.S. Food and Drug Administration (FDA)’s Bacteriological Analytical Manual (BAM) recommends a multiplex polymerase chain reaction (PCR) protocol to simultaneously detect the species-specific thermolabile hemolysin (tlh) gene and the pathogenic thermostable-related hemolysin (trh) and thermostable-direct hemolysin (tdh) genes. However, this assay has shown two limitations: difficulty in separating the amplicons of the trh (486 bp) and tlh (450 bp) genes due to their similar sizes, and the weaker band exhibited by the tdh gene amplicon (270 bp). The present study aimed to improve the BAM’s multiplex PCR assay by separating the three amplicons with similar intensity. A new primer set was applied for the tlh gene (369 bp) alongside the existing primers for the trh and tdh genes. The amplicons for the three genes were effectively separated by electrophoresis on a 2% tris-borate-EDTA (TBE) agarose gel within 45 minutes. Primer concentrations of 0.2 µM for trh, 0.2 µM for tlh, and 0.4 µM for tdh produced a significant amount of amplicons among various combinations of primer concentrations with 35 PCR cycles (P &lt; 0.05). This assay exhibited a detection limit of 10 pg of bacterial DNA, demonstrating its high sensitivity. It did not display amplicons from nine Vibrio strains known to be human pathogens or from 18 well-documented foodborne pathogens. Therefore, the present multiplex PCR protocol could help overcome the limitations of existing assays and provide a more reliable method for detecting the three genes of V. parahaemolyticus.

Background: Acute hepatopancreatic necrosis disease (AHPND), is a bacterial disease of whiteleg shrimp, which has a high mortality rate (100%) and incurs economic losses. Our objective was to identify the genes which lead to cell and organ damage and investigate bioproducts to prevent and treat. Methods: Litopenaeus vannamei shrimp in Thua Thien Hue province, Vietnam were collected from an infected pond and analysed at the Institute of Biotechnology, Hue University. The PirA gene of Vibrio parahaemolyticus strain K5 was isolated and analyzed for nucleotide sequence and paired with the expression vector pQE30. The expression vector was transformed into E. coli strain M15, the PirA recombinant protein was expressed in the form of 6xHis-PirA fusion protein of about 15 kDa. PirA recombinant protein was purified and determined the PirAvp binding ratio, cloning and sequencing of PirA gene from Vibrio parahaemolyticus strain K5 causing AHPND by PCR method with specific primers and molecular weights of PirAvp and the PirAvp complex. Results: PirA gene from Vibrio parahaemolyticus strain K5 was cloned into pGEM-T easy vector (Promega, USA) and screened E. coli TOP10 colonies containing pGEM T easy/PirA recombinant plasmid on LB agar/ampicillin/IPTG/X-Gal medium. PCR showing a band of about 347 bp, matching the size of PirA gene and two nucleotide sequences (BamHI and HindIII). The results showed that PirA gene has a length of 336 bp and similar to PirA gene on GenBank (Code: KU556825.1). The results of protein extracted from E. coli M15 recombinant cells and 6xHis-PirA target protein was collected in elution fractions from EF2 to EF6, showed that the concentration of 6xHis-PirA protein and EF3 elution fraction collected a highest protein concentration (1,586.54 µg/ml). Conclusions: The purified PirA recombinant protein will provide materials for development research to create biological products to prevent and treat AHPND.

Campylobacter is one of the most commonly reported foodborne bacteria worldwide. Although C. jejuni and C. coli have been reported to be responsible for the great majority of campylobacterio-sis, the burden of infections by species other than C. jejuni and C. coli have been increasing as a result of a transition to diagnostic test methods that enable the isolation of emerging species. The aim of the present study was to recover C. jejuni, C. coli, and emerging species from the stool samples of 500 patients with gastroenteritis and 100 healthy subjects by use of a filtration meth-od and culture techniques using Butzler agar and mCCDA under microaerophilic or hydrogen enriched atmosphere, identify the species by multiplex PCR methods and assess the significance of emerging species in enteric diseases. Thirty-one (6.2%) Campylobacter spp. were isolated from the stool samples of diarrheic patients but none from healthy individuals. Of 31 isolates, 21 (67.8%), 9 (29%) and 1 (3.2%) were identified as C. jejuni, C. coli and C. concisus by multiplex PCR, respectively. Filtration method was superior to the culture technique using mCCDA under mi-croaerophilic atmosphere. C. concisus was evaluated as the etiology of gastroenteritis as a result of laboratory and clinical evaluations. The present study was the first to indicate that emerging Campylobacter species are rarely detected and C. concisus is linked to acute gastroenteritis in Tur-key where additional studies are warranted to clarify the significance of emerging species in gastroenteritis.

The human innate immune response to the pore-forming toxin of Vibrio cholerae VCC, is currently under study. Here, in vitro studies on a human macrophage cell line (THP-1), helped explore the activated pathways involved on the onset the innate immune response towards the cytotoxin. The secreted monomeric 65 KDa form interacts with mature macrophages in pg/ml concentrations, determined by dose response experiments after treatments under 1 h. Non vacuolating concentrations (pg/ml) were applied to the cells; immunoblots revealed activation of MAPKs: early overexpression of p38 and ERK. Cell lysis by release of lactate dehydrogenase (LDH) was not apparent in the first hour, nonetheless it was positive after 24 h. Finally, to discern whether the VCC stimulates transcriptional activators via MAPKs pathway, NF-κB and AP-1 were studied by real time quantitation. Increased expression of p50 (NF-κB), cJun and cFos (AP-1) was observed. Given that NF-κB is the transcription factor initiating inflammation of innate immune response and in turn, AP-1 is responsible for cell surviving response, results from this study lead us to conclude that VCC in vitro treatments, induce a pro-inflammatory and a surviving response, in less than one hour on activated macrophages.

Vibrio splendidus is the main pathogen infect a wide range of hosts, especially sea cucumber Apostichopus japonicus. Our previous studies had shown that the level of L-glutamic acid (L-Glu) was significantly increased under heat stress and it was one of the best carbon sources that could be used by V. splendidus AJ01. In this study, the effects of exogenous L-Glu on the coelomocyte viability, tissue status and individual mortality of sea cucumbers were analyzed. The results showed that 10 mM L-Glu decreased coelomocyte viability and increased mortality, with tissue rupture and pyknosis, while 0.1 mM L-Glu slightly affected the survival of sea cucumbers without obvious damages at both cellular and tissue levels. Transcriptomic analysis showed that there were 343 upregulated and 206 downregulated genes regulated by exogenous L-Glu. GO analysis showed that differentially expressed genes (DEGs) were mainly enriched in signaling and membrane formation, while KEGG analysis showed that DEGs were significantly enriched in the upregulated endocytosis and downregulated lysosomal pathways. The coelomocyte viability further decreased 20% in the simultaneous presence of exogenous L-Glu and V. splendidus AJ01, compared to that of V. splendidus AJ01 infection alone. Consequently, a higher mortality of sea cucumber was also observed in the presence of exogenous L-Glu challenged by V. splendidus AJ01. Real time reverse transcriptase PCR showed that L-Glu specifically upregulated the expression of the fliC gene, conding the subunit protein of flagellar filament, with promoted swimming motility activity of V. splendidus. Our result indicated that L-Glu should be kept in a state of equilibrium, and the excess L-Glu in the host-pathogen interface would prompt the virulence of V. splendidus via increasing bacterial motility.

The global shrimp farming industry is impacted by various infectious diseases, leading to marked production losses. Virulent Vibrio parahaemolyticus strains (VPAHPND) and other Vibrio species cause acute hepatopancreatic necrosis disease (AHPND) in shrimp. We isolated a binary toxin PirA/B-bearing V. campbellii strain (HJ-2023) from a Korean shrimp farm. The biochemical characteristics and antibiotic susceptibility of HJ-2023 were analyzed and compared to those of PirA/B-bearing VPAHPND. Non-AHPND-causing V. campbellii was the control. The pathogenicity of each strain was analyzed in white-leg shrimp. Phylogenetic analysis classified the isolated HJ-2023 strain as V. campbellii. Biochemical tests confirmed distinct properties of HJ-2023, compared to the genetically similar V. harveyi group. The AHPND-causing toxin gene of HJ-2023 showed 100% similarity to the rpoD gene of VCAHPND. An antibiotic disc susceptibility test indicated that HJ-2023 showed resistance to ampicillin, clindamycin, lincomycin, and erythromycin, which is effective against gram-positive bacteria, and showed weak sensitivity to gentamycin. Furthermore, HJ-2023 was 25% less toxic than VPAHPND but could be lethal during aquaculture-related outbreaks. These results highlight the complexity and virulence of AHPND-causing strains through multiple pathways and confirm the pathogenicity of V. campbellii, which increases the risk posed by AHPND to shrimp aquaculture. Our findings could help prevent AHPND.

Vibrio is a bacterial genus widely distributed in natural aquatic systems. Some Vibrio species are pathogenic and can cause severe diseases in both marine organisms and humans. Previous studies revealed a link between the current climate change and increased incidence of the Vibrio-associated diseases recently causing sanitary, economic and/or ecological problems worldwide. The conventional culture-based methods (e.g. cultivation of TCBS agar) used to monitor the presence of Vibrio spp. in environmental samples are not always straightforward and can underestimate the number of cells, especially if the microbial population contains a fraction of ‘dormant’ cells (e.g. cells in Viable but not Culturable (VBNC) state). However, this problem can be overcome by using alternative culture-free approaches such as Catalyzed Reporter Deposition-Fluorescence In situ Hybridization (CARD-FISH). To optimize CARD-FISH for efficient and reliable detection of Vibrio spp. in environmental samples, we have used both computer-assisted and experimental approaches. Our results demonstrate that the use of the optimized protocol along with a very specific probe, ViB572a, can offer the high sensitivity and selectivity of CARD-FISH detection of marine vibrios in natural seawater.

Vibrio is a bacterial genus widely distributed in natural aquatic systems. Some Vibrio species are pathogenic and can cause severe diseases in both marine organisms and humans. Previous studies revealed a link between the current climate change and increased incidence of the Vibrio-associated diseases recently causing sanitary, economic and/or ecological problems worldwide. The conventional culture-based methods (e.g. cultivation of TCBS agar) used to monitor the presence of Vibrio spp. in environmental samples are not always straightforward and can underestimate the number of cells, especially if the microbial population contains a fraction of ‘dormant’ cells (e.g. cells in Viable but not Culturable (VBNC) state). However, this problem can be overcome by using alternative culture-free approaches such as Catalyzed Reporter Deposition-Fluorescence In situ Hybridization (CARD-FISH). To optimize CARD-FISH for efficient and reliable detection of Vibrio spp. in environmental samples, we have used both computer-assisted and experimental approaches. Our results demonstrate that the use of the optimized protocol along with a very specific probe, ViB572a, can offer the high sensitivity and selectivity of CARD-FISH detection of marine vibrios in natural seawater.

The string of bacteria, Vibrio. sp. QD-5 utilizing alginate, was separated from rotten kelp. The results of genome sequencing showed that the strain QD-5 contained four alginate lyase genes.One of the alginate genes Aly-IV was cloned and linked to the plasmid pET-22b (+). The heterologous expressed alginate lyase aly-IVwas characterized，which possessed the theoretical molecular mass of 62 kDa, and theoretical isoelectric point (pI) of 5.12. - The enzyme aly-IV was purified and the activity reached 1256.78 U/mg, with optimal temperature of 35 ^oC and pH value of 8.9. Nurtured in the temperature below 25 ^oC for 30 minutes, the activity was almost stable. The result also suggested that the activity of enzyme was strongly affected by - NaCl whose optimal concentration was 15 mM in a lab environment The TLC and ESI-TOF-MS analysis suggested that the enzyme aly-IV could degrade sodium alginate and polyG in endo-lytic type, producing monomer, dimer and trimmer. So, aly-IV can also be widely applied to make large scale preparation of alginate oligosaccharides with low degree of polymerization (DP).

Oral immunization with the choleric toxin (CT) elicits a high level of protection against its enterotoxin activities and can control cholera in endemic settings. However, the complete B-cell epitope map of the CT responsible for protection remains to be clarified. Here, we have mapped the B-cell linear epitopes of the three chains of the CT protein (CTP) prepared by Spot synthesis. The immunoreactivity of sera from mice immunized with an oral, inactivated vaccine (Schankol†™) was measured against membrane-bound peptides for mapping. Results: Eighteen IgG epitopes were identified; eight in the CTA, three in the CTB, and seven in the protein P. Three epitopes, TQTGFVRHDDGYVST (aa 66-77, Vc/TxA-3), KNGAIFQVE VPGSQN (aa 64-78, Vc/TxB-11), and LNDEHK (aa 90-95, Vc/TxP-16), were chosen to synthesize a multiple antigen peptide that was used to coat ELISA plates to screen immunized mouse sera as a test for an in vitro diagnostics for cholera. Conclusion: Vaccination with inactive CT-generated antibodies against multiple linear epitopes and the selected epitopes can be used to construct immunological reagents for rapid serological diagnosis of cholera with high sensitivity and specificity.

A challenging problem in the aquaculture industry is bacterial disease outbreaks which results in the global reduction of fish supply and foodborne outbreaks. Biofilms in marine pathogens protect against antimicrobial treatment and host immune defence. Zeolites are minerals of volcanic origin made from crystalline aluminosilicates which are useful in agriculture and in environmental management. In this study, silver ion-exchanged zeolite A of four concentrations; 0.25M (AgZ1), 0.50M (AgZ2), 1.00M (AgZ3)and 1.50M (AgZ4) were investigated for biofilm inhibition and antimicrobial properties against two predominant marine pathogens V. campbelli and V. parahemolyticus by employing the Minimum Inhibitory Concentration(MIC), Crystal Violet Biofilm Quantification assays as well as Scanning Electron Microscopy. In the first instance, all zeolite samples AgZ1-AgZ4 showed antimicrobial activity for both pathogens. For V. campbellii AgZ4 exhibited the highest MIC at 125.00 µg/ml while for V. parahaemolyticus the highest MIC was observed for AgZ3 at 62.50 µg/ml. At sublethal concentration, biofilm inhibition of V. campbelli and V. parahemolyticus by AgZ4 were observed at 60.2% and 77.3% inhibition respectively. Scanning electron microscopy exhibited profound structural alteration of the biofilm matrix by AgZ4. This is the first known study that highlights the potential application of ion-exchanged Zeolite A against marine pathogens and their biofilms.

Heavy metal contamination is listed among the most alarming threats to the environment and human health. The detrimental effects of heavy metals in the natural environment span from a reduction of biodiversity to toxic effects on marine life - through microplastic born heavy metals -, to impairment of microbial activity in the soil, and to detrimental effects on animal reproduction. A host of different chemical and biological technologies have been proposed to alleviate environmental contamination by heavy metals. Relatively less attention has been paid to the microbial precipitation of heavy metals, as a side mechanism of the most general process of microbially induced calcite precipitation (MICP). This process is currently receiving a great deal of interest from both a theoretical and practical standpoint, because of its possible practical applications in concrete healing and soil consolidation, and its importance in the more general framework of microbial induced mineral precipitation. In this study, we analyse the ability of the marine bacteria Vibrio harveyi in co-precipitating CaCO3 minerals, together with Cd, Cr, Pb, and Zn added in form of nitrates, from solutions containing CaCl2. The precipitated carbonatic minerals were a function of the different heavy metals present in the solution. The process of co-precipitation appears to be rather effective and fast, as the concentrations of the 4 heavy metals were reduced in 2 days by 97.2%, on average, in the solutions.

Microbial production is a promising method that can overcome major limitations in conventional methods of lycopene production, such as low yields and variations in product quality. Significant efforts have been made to improve lycopene production by engineering either the 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway or mevalonate (MVA) pathway in microorganisms. To further improve lycopene production, it is critical to utilize metabolic enzymes with high specific activities. Two enzymes, 1-deoxy-D-xylulose-5-phosphate synthase (Dxs) and farnesyl diphosphate synthase (IspA), are required in lycopene production using MEP pathway. Here, we evaluated the activities of Dxs and IspA of Vibrio sp. dhg, a newly isolated and fast-growing microorganism. Considering that the MEP pathway is closely related to the cell membrane and electron transport chain, the activities of the two enzymes of Vibrio sp. dhg were expected to be higher than the enzymes of E. coli. We found that Dxs and IspA in Vibrio sp. dhg exhibited 1.08-fold and 1.38-fold higher catalytic efficiencies, respectively. Consequently, the heterologous overexpression improved the specific lycopene production by 1.88-fold. Our findings could be widely utilized to enhance production of lycopene and other carotenoids.

A key component of efforts to identify the biological and drug-specific aspects that contribute to therapeutic failure or unexpected exposure-associated toxicity is the study of drug-intestinal barrier interactions. While methods supporting such assessments are widely described for human therapeutics, there is relatively little information available for similar evaluation in support of veterinary pharmaceuticals. There is, therefore, a critical need to develop novel approaches for evaluating drug-gut interactions in veterinary medicine. Three-dimensional (3D) organoids can address these difficulties in a reasonably affordable system that circumvents the need for more invasive in vivo assays in live animals. However, a first step in the development of such systems is understanding organoid interactions in a 2D monolayer. Given the importance of orally administered medications for meeting the therapeutic need of companion animals, we demonstrate growth conditions under which canine colonoid-derived intestinal epithelial cells survive, mature, and differentiate into confluent cell systems with high monolayer integrity. We further examine the applicability of this canine colonoid-derived 2D model to assess the permeability of three structurally diverse, passively absorbed β-blockers (e.g., propranolol, metoprolol, and atenolol). Both absorptive and secretive apparent permeability (Papp) of these drugs at two different pH conditions were evaluated in canine colonoid-derived monolayers and were compared with that of Caco-2 cells. This proof-of-concept study provides promising preliminary results with regard to the utility of canine-derived organoid monolayers for species-specific assessments of therapeutic drug passive permeability.

This study is the first report of the successful cultivation of Dinophysis norvegica isolated from Japanese coastal waters (7 of 48 isolated cells were established as the clonal cultures, 14.5%) and their toxin content and production. The strains were maintained at relatively high abundance (>2,000 cells mL-1) for more than one year when fed on Mesodinium rubrum with the addition of Teleaulax amphioxeia. At the end of the one-month incubation, the total amounts of pectenotoxin-2 (PTX2), dinophysistoxin-1 (DTX1) and okadaic acid (OA) were 132.0-375.0 ng mL-1 (n = 7), 0.7-3.6 ng mL-1 (n = 3) and trace level OA (n = 1), respectively. Similarly, the cell quota of PTX2 and DTX1 were 60.6-152.4 pg cell-1 (n = 7) and 0.5-1.2 pg cell-1 (n = 3), respectively. These data indicated that toxin production varies depending on the strains in this species. In the growth experiment, D. norvegica grew, but it was very slow first 12 days. However, after that they grew exponentially with the maximum growth rate of 0.56 divisions day-1 (during Day 24-27), reaching a maximum concentration of 3,000 cells mL-1 at the end of incubation (Day 36), suggesting that they have a long lag phase. In the toxin production, OA was not at a detectable level (≤ 0.010 ng mL-1) during the 36 days of incubation except for Day 6. The concentration of DTX1 and PTX2 showed similar patterns as described for vegetative growth, but the toxin production still increased on Day 36 (1.3 ng mL-1 and 154.7 ng mL-1 in DTX1 and PTX2, respectively). The findings of this study provide novel information on the toxin content and production in D. norvegica as well as details on the culturing and maintenance of the species.

Europium is one of the most reactive lanthanides and humans use it in many different applica-tions, but we still know little about its potential toxicity and cellular response to its exposure. Two strains of the eukaryotic microorganism model Tetrahymena thermophila adapted to high concentrations of two Eu(III) compounds (EuCl3 or Eu2O3) were obtained and compared to a control strain and cultures treated with both compounds. In this ciliate, EuCl3 is more toxic than Eu2O3. LC50 values show that this microorganism is more resistant to these Eu(III) compounds than other microorganisms. Oxidative stress originated mainly by Eu2O3 is minimized by overexpression of genes encoding important antioxidant enzymes. The overexpression of metallothionein genes under treatment with Eu(III) compounds supports the possibility that this lanthanide may interact with the -SH groups of the cysteine residues from metallothi-oneins and/or displace essential cations of these proteins during their homeostasis function. Both lipid metabolism (lipid droplets that fuse with europium-containing vacuoles) and au-tophagy are involved in the cellular response to europium stress. Bioaccumulation, together with a possible biomineralization to europium phosphate, seems to be the main mechanism of Eu(III) detoxification in these cells.

Vibrio parahaemolyticus carrying a plasmid encoding toxins PirA and PirB is one of the causative agents leading to the severe disease of acute hepatopancreatic necrosis disease (AHPND) in shrimp aquaculture. However, there is a lack of deep understanding of the host resistant characteristics against V. parahaemolyticus infection. Here, we performed comparative transcriptome analysis on the expression profiles at the background level of hemocytes from shrimp with different resistant abilities against V. parahaemolyticus infection in two independent populations. Principal component analysis and sample clustering results showed that samples from the same population had a closer relationship than that from shrimp with similar disease-resistant abilities. Differentially expressed genes (DEGs) analysis revealed that the number of DEGs between two populations was much more than that between V. parahaemolyticus¬-resistant and susceptible shrimp. A total of 31 DEGs and 5 DEGs were identified from the comparison between V. parahaemolyticus¬-resistant and susceptible shrimp from population 1 and 2, respectively. There was no DEGs overlap between two comparisons. DEGs from population 1 were mainly cytoskeleton related genes, metabolic related genes, and immune related genes. DEGs from population 2 included tubulin alpha-3, dihydropyrimidinase-like gene, diacylglycerol kinase 1, prohibitin and iroquois-class homeodomain protein IRX-2-like. The data suggest that shrimp from different populations might adopt distinct disease-resistant mechanisms, which provided new insights into understanding disease-resistant mechanisms at population level against pathogen infection. A new strategy for screening disease resistant genes and disease resistant breeding was proposed based on the present study.

The European Green Deal poses a two-pronged challenge for the automotive industry: migrating to solutions based in light structures, requiring light weight concepts and light materials, while at the same time avoiding dependence towards the importation of these advanced materials. Aluminium alloys are lightweight and cost-effective materials that can successfully cover the requirements of many structural applications, however their production requires from bauxite and other Critical Raw Materials (CRM), such as Si and Mg. Aluminium alloys are fully recyclable, but scrap is usually contaminated and its use is related to an increment of impurities, tramp element and undesired inclusions. Traditionally, the use of secondary alloys has been restricted to low performance applications. The present work analyses the effect that the use of scrap has in the quantity of inclusions present in the alloy and in other properties relevant for the material processing. The study was carried out for common alloys belonging to 3 of the most common aluminium processes used in the car manufacturing industry: High Pressure Die Casting (HPDC) (EN AC-43500), extrusion (6063) and sheet metal forming (5754 and 6181). Reference alloy were mixed with different level of scrap (0, 20, 40, 60, 80 and 100%) trying to keep the chemical composition as unaffected as possible. The inclusions level of the alloy was characterized with the Prefil Foot-printer® test. In addition, the obtained materials, after being cast in an open mould, were subjected to metallographic characterization. Relevant properties to assess the formability quality of the alloys for the corresponding transforming process, were obtained: flowability test for HPDC alloy and high temperature compression for extrusion alloys. The results obtained suggest that the number of inclusions present in the melt highly increase with the amount of scrap used to produce the alloy. These inclusions are also related with an important lost of flowability, but do not have a noticeable impact on microstructure.

Exogenous proteases are well established for hydrolysing proteins and improving the digestibility of amino acids, which, in animal production, enhances growth performance and diet formulation flexibility. Attention has now turned to the wider, or ‘extra-proteinaceous’, benefits of proteases. Both host and (gut) microbiome utilise amino acids for diverse functions beyond just cellular growth and proliferation. For example, the host produces a diverse array of proteinaceous secretions that seek to modulate the activity of the intestinal microbiome and opportunities to interact with host (e.g., epithelial) receptors. Reciprocally, the microbiome utilises various protein-based components to perform individual lifestyle traits, such as colonising gut niches, production of virulence traits, etc., which can be influenced by protease activity and amino acid availability. In addition, microbial fermentation of proteins and amino acids produces metabolites that may be considered undesirable (e.g., toxic) for the host’s cells or, for example, can act as signalling molecules, shaping the secretion of gut hormones and function. Parallels may also exist between various antibiotic growth promoters and proteases, in that both can target/impact microbial proteins (synthesis and degradation, respectively). There are many aspects to consider, such as determining optimal intestinal levels and spectra of proteolytic activity under various scenarios, but great potential exists.

Injuries from exposure to blasts are of significant concern in military operational settings, including tactical training, and is associated with self-reported concussion-like symptomology and physiological changes as increased intestinal permeability (IP) investigated herein. Timeseries gene expression and IP biomarker data were generated from “breachers” exposed to controlled, low-level explosive blast during training. Samples from 30 male participants at pre, post, follow-up blast exposure the next day were assayed via RNA-seq and ELISA. A battery of symptom data was also collected at each of these time points that acutely showed elevated symptom reporting related to headache, concentration, dizziness, and slowed thinking, dissipating ~16hrs following blast exposure. Evidence for bacterial translocation into circulation following blast exposure was detected by significant stepwise increase in microbial diversity (measured via alpha-diversity p=0.049). Alterations in levels of IP protein biomarkers (i.e., Zonulin, LBP, Claudin-3, I-FABP) assessed in a subset of these participants (n=23) further evidenced blast exposure associates with IP. The observed symptom profile was consistent with mTBI and was further associated with changes in bacterial translocation and intestinal permeability, suggesting that IP may be linked to a decrease in cognitive performance. These preliminary findings show for the first

Objective: To examine associations between IgA responses to Gram-negative gut-commensal bacteria and peri-menstrual symptoms and sex hormone levels during the menstrual cycle in women with and without premenstrual symptoms. Methods: Forty women aged 18-45 years completed the Daily Record of Severity of Problems (DRSP) during all 28 consecutive days of the menstrual cycle. We assayed, in plasma,, IgA responses to 6 Gram-negative bacteria, i.e. Hafnei alvei, Pseudomonas aeruginosa, Morganella morganii, Klebsiella pneumoniae, Pseudomonas putida and Citrobacter pylori, progesterone and oestradiol at days 7, 14, 21 and 28 of the menstrual cycle. Results: Significant changes in Δ (actual minus one week earlier) IgA to LPS of the 6 Gram-negative bacteria during the menstrual cycle were observed with peak IgA levels at T4 (day 28) and lows at T1 or T2 (day 7 or 14). The Δ IgA changes in H. alvei, M. Morganii, P. putida during the menstrual cycle were significantly and positively associated with changes in the total DRSP score, and severity of physio-somatic, anxiety and breast-craving, but not depressive, symptoms. The changes in IgA responses to LPS were largely predicted by changes in progesterone and steady-state levels of progesterone averaged over the luteal phase. Discussion: Menstrual cycle-associated changes in IgA directed against LPS and by inference bacterial translocation may be driven by effects of progesterone on transcellular, paracellular and vascular pathways thereby contributing to the severity of physio-somatic and anxiety symptoms as well as fatigue, breast swelling and food cravings.

A novel binary toxin-positive non-027, non-078 Clostridium difficile strain designated LC693 whose sequence type was ST201 was isolated from the fecal sample of a patient with severe diarrhea in China. To understand the pathogenesis basis of C. difficile ST201, this recently recovered isolate LC693 was then chosen for whole genome sequencing. The project finally generated an estimated genome size of approximately 4.07 Mbp. The genome sequence was then analyzed together with the other two ST201 strains VL-0104 and VL-0391 and compared to the epidemic 027/ST1 and 078/ST11 strains. Phylogenetic analysis demonstrated that the ST201 strains belonged to clade 3. Genome size of the three ST201 strains ranged from 4.07 Mb~4.16 Mb, with an average GC content between 28.5%~28.9%. The ST201 genomes contained more than 40 antibiotic resistance genes and 15 of them were predicted to be associated with vancomycin-resistance, suggesting that they may have a strong antibiotic resistance. The ST201 strains contained a typical clade 3 specific PaLoc with a Tn6218 element inserted, and those genes harbored on their PaLoc that participated in the toxin expression and regulation were highly homologues to the epidemic 027 and 078 strains, with the exception of tcdC. A truncated TcdC was found in the ST201 strains, which is suggestive to have a contribution to the toxin production of the ST201 strains. In addition, the ST201 strains contained intact binary toxin coding and regulation genes, which is also proposed to contribute to the virulence. Genome comparison of the ST201 strains with the epidemic 027 and 078 strain identified 641 genes specific for the C. difficile ST201, and a number of them were predicted as fitness and virulence associated genes. The identification of those genes also contributes to the pathogenesis of the ST201 strain. To our knowledge, this is the first study that the genome sequence of C. difficile ST201 was discussed in detail, and the present study would have a contribution to understanding the pathogenesis basis of C. difficile ST201.

It is well known that the strong-evidence foodborne outbreaks of human campylobacteriosis are associated with the consumption of raw or incompletely thermally processed poultry meat, whereas broilers act as the main reservoir for Campylobacter species. Campylobacter jejuni and Campylobacter coli are the two main species of campylobacters detected in chicken meat, while they account for almost 90% of the reported cases of campylobacteriosis in humans. Over 80% of these cases are attributed to C. jejuni and about 10% of them are due to C. coli. Therefore, until recently the dominance of C. jejuni against all other Campylobacter spp. isolated from chicken meat samples was well established and unquestionable. Lately, however, C. coli has been increasingly recovered from chicken meat to such an extent that it is now evident that it often comprises the dominant species among the identified campylobacters in the meat samples. This work attempts for the first time a detailed review in the literature to deepen into this noteworthy epidemiological swift in the prevalence of C. jejuni and C. coli, along with the distribution of Campylobacter spp. in chicken meat. Factors such as the sampling method followed for screening campylobacters in broiler carcasses (e.g., swabs or carcass rinsates, skinned or skinless meat excised samples) and part of the animal carcass from which the sample is obtained (e.g., neck, breast, leg), seasonality of sampling (summer vs. winter) and environmental conditions (e.g., rainfall, relative humidity) at the farm level, the isolation procedure (enumeration or detection) and pathogen identification (biochemical or molecular), the enrichment and plating isolation media (e.g., Bolton vs. Preston broth, charcoal-based vs. chromogenic agars), as well as the biofilm-forming ability of different campylobacters, highlight the multivariate dimension of the phenomenon and are thoroughly discussed in the present review.

In the field of railroad safety, effective detection of surface cracks is critical, necessitating reliable, high-speed non-destructive testing (NDT) methods. This study introduces a hybrid Eddy Current Testing (ECT) probe, specifically engineered for railroad inspection, to address the common issue of ’lift-off noise’ due to varying distances between the probe and test material. Unlike traditional ECT methods, this probe integrates transmit and differential receiver coils, aiming to enhance detection sensitivity and minimize lift-off impact. The study optimizes ECT probes employing different driver coils, emphasizing three main objectives: a) quantitatively evaluating each probe using signal- to-noise ratio (SNR) and outlining a real-time data processing algorithm based on SNR methodology; b) exploring the frequency range proximal to the electrical resonance of the receiver coil; c) examining sensitivity variations across varying lift-off distances. The experimental outcomes indicate that the newly designed probe with figure 8-shape driver coil significantly improves sensitivity in detecting surface cracks on railroads. It achieves an impressive SNR exceeding 100 for defects with minimal dimensions of 1 mm in width and depth. Simulation results closely align with experimental findings, validating the investigation of optimal operational frequency and lift-off distance for selected probe performance, determined to be 0.3 MHz and 0.5 mm, respectively. The realization of this project would lead to notable advancements in enhancing railroad safety by improving crack detection efficiency.

Toxin/antitoxin (TA) systems are used primarily to inhibit phage, reduce metabolic activity during stress, and maintain genetic elements. Given the extreme toxicity of some of the toxins of these TA systems, we were curious how the cell silences toxins, if the antitoxin is inactivated or when toxins are obtained without antitoxins via horizontal gene transfer. Here we find that the RalR (type I), MqsR (type II), GhoT (type V), and Hha (type VII) toxins are inactivated primarily by a mutation that inactivates the toxin promoter or via the chromosomal mutations iraM and mhpR.

The intestinal microbiota, comprised of bacteria, archaea, and phages, inhabits the gastrointestinal tract of the organism. Male reproductive sterility is currently a prominent topic in medical research. Increasing research suggests that gut microbiota dysbiosis can result in various reproductive health problems. This article specifically investigates the impact of gut microbiota dysbiosis on male reproductive infertility development. Gut microbiota imbalances can disrupt the immune system and immune cell metabolism, affecting testicular growth and sperm production. This dysfunction can compromise the levels of hormones produced and secreted by the endocrine glands, affecting male reproductive health. Furthermore, imbalance of the gut microbiota can disrupt the gut-brain-reproductive axis, resulting in male reproductive infertility. This article will explore how the imbalance of the gut microbiota impacts male reproductive infertility through immune regulation, endocrine regulation, and interactions of the gut-brain-reproductive axis, concluding with recommendations for prevention and treatment.

The global usage of veterinary antibiotics is significant. These antibiotics can be released into the aquatic environment and exert toxic effects on non-target organisms. To explore the physiological effects of tetracycline antibiotics on aquatic life, the growth characteristics of and toxin release from the cyanobacterium Microcystis aeruginosa (M. aeruginosa) were studied. Results showed that the toxicity order of the three target antibiotics was TC (tetracycline) > CTC (chlortetracycline hydrochloride) > OTC (oxytetracycline) in terms of inhibition occurrence time and the EC₁₀ and EC₂₅ values. Further, the target antibiotics regulated the production of MC-LR (microcystin-LR) to different degrees. CTC destroyed the M. aeruginosa cells and resulted in a decreased MC-LR release but stimulated the ability to synthesise MC-LR. OTC had a relatively weaker toxicity compared with CTC, while TC was the most toxic among the three antibiotics. Therefore, TC is friendly to the aquatic environment because it simultaneously reduced the intracellular and extracellular MC-LR content. These results aid our understanding of the effects of tetracycline antibiotics on Microcystis aeruginosa, which is important for environmental evaluation and protection. These results are also helpful for guiding the application of veterinary antibiotics in agricultural settings.

The sharing of genome sequences in online data repositories, allows for large scale analyses of specific genes or gene families. This can result in the detection of novel gene subtypes as well as development of improved detection methods. Here we used publicly available WGS data to detect a novel Stx subtype, Stx2n in two clinical E. coli strains isolated in the USA. During this process, additional Stx2 subtypes were detected; six Stx2j one Stx2m strain and one Stx2o, all were analyzed for variability from the originally described subtypes [1,2]. Complete genome sequences were assembled from short or long read sequencing and analyzed for serotype, and ST types. The stx2n and Stx2o WGS were further analyzed for virulence genes pro-phage analysis and phage insertion sites. Nucleotide and amino acid maximum parsimony trees showed expected clustering of the previously described subtypes and a clear separation of the novel Stx2n subtype. WGS data was used to design OMNI PCR primers for the detection of all known Stx1 (283 bp amplicon), Stx2 (400 bp amplicon), eae (222bp amplicon) and Stx2f (438 bp amplicon) subtypes. These primers were tested in three different laboratories, using standard reference strains. Analysis of complete genome sequence showed variability in serogroup, virulence genes, ST type, and Stx2 pro-pages showed variability in size, gene composition, and phage insertion sites. The isolates with Stx2n and Stx2o in this study demonstrated toxicity for Vero cells. The Stx2n strain were induced when grown with sub-inhibitory concentrations of Cip, and toxicity was detected. Taken together, these data highlight the need to reinforce genomic surveillance to identify the emergence of potential new Stx2 or Stx1 variants. The importance of this surveillance has a paramount impact in public health. Per our description in this study, we suggest that 2017C-4317 be designated as the Stx2n type-strain.

Background: Chronic lateral epicondylitis challenges the therapeutical approach; underlying mechanisms are incompletely understood; neuropathic pain and central and peripheral sensitization may explain the fact that botulinum toxin was found to play a role in pain and function management. Methods: We searched the literature for MeSH terms: lateral epicondylitis or synonyms and botulinum toxin. Results: We found 14 papers containing trials on botulinum toxin injection into the tendon or into the extensor muscles (particularly extensor carpi radialis brevis and extensor communis digitorum). We followed the administration pathways, doses, timing and side effects. Conclusion: With chronic course, the focus of the therapy shifts from the afflicted tendon to the inserting muscles, as muscle contracture may create a vicious loop to perpetuate and aggravate the disease. Doses, timing and side effects are discussed.

Campylobacter is one of the major foodborne pathogens of concern in its growing trend of antimicrobial resistance. C. jejuni and C. coli are the major causative agents, with C. jejuni contributing to most of the cases in approximately 90% in the world. Infection is transmitted to humans due to consumption of contaminated food and water. Campylobacteriosis caused by C. jejuni is commonly presented with severe diarrhoea, abdominal pain, fever, headache, nausea, and vomiting with some extreme cases resulting in Guillain–Barré syndrome (GBS) and acute flaccid paralysis. Symptoms are severe in cases of children below 5 years, elderly and individuals who are immunocompromised. The infection is usually sporadic, and self-limiting and thus does not require antibiotics for treatment. Still, the antimicrobial resistance in Campylobacter is a major concern because of the transmission of resistance from animal sources to humans. This review highlights the recent epidemiology, geographical impact, resistance mechanisms, spread of Campylobacter spp. and the strategies to control the transmission of Campylobacter from veterinary sources and its antimicrobial resistance.

Campylobacter infection affects more than 200,000 people every year in Europe and in the last four years trend shows an increase of campylobacteriosis. Main vehicle for transmission of the bacterium is contaminated food like meat, milk, fruit and vegetables. In this study, the aim was to find characteristic VOCs of C. jejuni in order to detect its presence with an array of MOX gas sensors. Using a starting concentration of 10³ CFU/mL, VOCs were analyzed using GC-MS with SPME technique at the initial time (T0) and after 20 hours (T20). It has been found that Campylobacter sample at T20 is characterized by a higher number of alcohol compounds that the one at T0 and this is due to sugar fermentation. Sensors results showed the ability of the system to follow bacteria curve growth from T0 to T20 using PCA. In particular, this results in a decrease of ΔR/R₀ value over time. For this reason, MOX sensors are a promising technology for the development of a rapid and sensitive system for C. jejuni.

Mechanically homogenized composite of expanded graphite and cobalt(II) rhenate(VII) has been described. Cobalt(II) rhenate(VII) was obtained in a reaction of perrhenic acid with cobalt(II) nitrate. A simple mortar homogenization method was used to enhance intercalation of cobalt species within carbon matrix. The specific capacity of composite was enhanced by 50% in comparison to bare expanded graphite. The electrochemical characteristic was significantly improved including better cyclability, lower resistance of electrode material, lower iR drop, with respect to expanded graphite without cobalt(II) rhenate(VII) active species. Expanded graphite with its unique specific surface area and pore size diameter was proved as potential and cheap carbon support.

AB toxins have historically been associated with significant morbidity, mortality through infections such as botulinum, anthrax, cholera, and diphtheria. These AB toxin-mediated diseases remain prevalent in low and middle income countries, with intermittent outbreaks of Shiga toxin-producing Escherichia coli (STEC) or whooping cough by Bordetella pertussis in high-income countries. These reports warrant an investigation to better understand the distinct characteristics of AB toxins derived from different pathogens. As toxigenic pathogens broaden their scope and diversity, it amplifies the complexity of the problems posed by their AB toxins. Here, we discuss the history, structure and characteristics of key AB toxins, and report on historical and ongoing research on these toxins. We also explore research avenues that hold great promise in potentially improving clinical management of toxin-mediated diseases in the future.

Aquaporins (AQPs) are water channels widely distributed in living organisms and involved in many pathophysiologies as well as in cell volume regulations (CVR). In the present study, based on the structural homology existing between mineralocorticoids receptors (MR), glucocorticoids receptors (GR), cholesterol consensus motif (CCM) and the extra-cellular vestibules of AQPs, we investigated the binding of corticosteroids on AQP family through in silico molecular dynamics simulations of AQP2 interactions with cortisol. We propose for the first time a putative AQPs corticosteroids binding site (ACBS) and discussed its conservation through structural alignment. Corticosteroids can mediate non-genomic effects, nonetheless, the transduction pathways involved are still misunderstood. Moreover, a growing body of evidence is pointing out toward the existence of a novel membrane receptor mediating part of these rapid corticosteroids effects. Our results suggest that the naturally produced glucocorticoid cortisol inhibits channel water permeability. Based on these results, we propose a detailed description of a putative underlying molecular mechanism. In this process, we also bring new insights on the regulatory function of AQPs extra-cellular loops and on the role of ions in tuning the water permeability. Altogether, this work brings new insights into corticosteroids non-genomic effects through the proposition of AQPs as membrane receptor of this family of regulatory molecules. This original result is the starting point for future investigations to define more in depth and in vivo the validity of this functional model.

Sialylated milk oligosaccharides (SMOs) is the most important oligosaccharides in human milk oligosaccharides (HMOs), and have a multifunctional health benefit, and the most abundant SMOs are sialyllactose (SLs). The previous studies indicate that the growth of tumor cells, and angiogenesis on tumor tissues with Lewis lung carcinoma, melanoma, and colon carcinoma cells could be prevented by SL. Thus, it has been proposed that SL might be an ideal candidate for the development of anti-angiogenic drugs without any side effects. It has been proposed that the Sialic acid (Sia) can be produced through SL’s hydrolysis, and the dietary Sia is further transported to the brain and other tissues though blood for formation of CMP-Sia, the activated precursor of Sia, and then is catalyzed by polysialyltransferases (polySTs) for synthesis of polysialic acid (polySia) or polysialylation of neural cell adhesion molecule (NCAM). Thus, it’s necessary to study and develop new inhibitors, which not only modulate a proper balance between the gut health and neuros system development, but also inhibit the overexpression of polySia, and thus block the tumor cell migration. In this study, we proposed that 3’-SL and 6’-SL may be more ideal inhibitors comparing with two previous inhibitors, the low molecular weight heparin (LMWH) and cytidine monophosphate (CMP) in the previous studies.

Most of the mid-high permeability sandstone oilfields in China have entered the ultra-high water cut stage, accompanied with the highly-dispersed residual oil, and the control effect of mi-cro-heterogeneity on the residual oil is increasingly prominent. Understanding the microscopic characteristics of pore structural and its evolution during the long-term waterflooding is critical for enhancing the accuracy and efficiency of oil field exploitation. While the previous insights paid more attention to the geometrical and morphological features of pore structure and ignores the relation of pore structure with the fluid flow capacity. In this study, we propose a microscopic pore structure classification method considering pore size and local permeable capacity, which realizes the physical classification and quantitative evaluation of pore structure characteristics for different permeability rocks. The micro-pore structure is classified into 6 types based on the key evaluation parameters of pore diameter and flow flux area. By taking six mid-high permea-bility sandstone core samples from Daqing oilfield in China, a series of long-term waterflooding experiments are carried out and the interior pore structure was captured using high precision CT scanning technique. The results reveals that high-permeability reservoirs exhibit concentrated pores diameters and finer throats, whereas mid-permeability reservoirs exhibit smaller pores and throats in greater quantity. With the increase of water injection volumes, the permeable ca-pacity and connectivity of the high-connectivity pore space in ultra-high- and high-permeability reservoirs are enhanced, and in mid-permeability reservoirs, the micro-heterogeneity intensifies the increase in low-connectivity small pores. These findings provide new insights for the micro-scopic heterogeneity and a solid theoretical basis for investigating the occurrence of microscopic residual oil and optimising enhanced oil recovery strategies at the late stage of ultra-high water cut.

Infrastructure development is rapidly increasing, driving up demand for concrete, a key construction material. Cement and aggregate are essential constituents, with global cement production reaching 4.1 billion metric tons in 2011 and CO2 emissions hitting 1.7 billion metric tons by 2024, a significant contributor to carbon emissions. To address this, alternative constituents like Rice Husk Ash (RHA) and Animal Bone Powder (ABP) are proposed. With rice husk comprising 20% of rice weight and a substantial cattle population in Bangladesh, RHA and ABP offer promising alternatives. This study examines the benefits and limitations of incorporating RHA and ABP into cement, focusing on concrete strength and permeability. By replacing varying percentages of cement with RHA and ABP simultaneously, experiments evaluate their impact on concrete properties after 28 days of curing. The research aims to identify the optimal combination for sustainable concrete production, reducing environmental impact by curbing waste generation and pollution. Results show that compressive strength increases and permeability decreases with up to 5% replacement of RHA and ABP, highlighting their potential for enhancing concrete performance while mitigating environmental concerns.

Salmonella infections can lead to intestinal inflammation and metabolic disorders in birds. However, whether ARA metabolism is involved in Salmonella-induced intestinal inflammation remains unclear. This experiment investigated the changes in cecal flora and arachidonic acid (ARA) metabolism in Hainan Wenchang chickens infected with S. Typhimurium using 16s rDNA sequencing and targeted metabolomics. The results showed that the levels of ARA metabolites were increased in the cecum tissue of Wenchang chickens after infection with S. Typhimurium, including prostaglandin E2, prostaglandin F2α, lipoxin A4, ±8(9)-EET, ±11(12)-EET, and ±8,9-DiHETrE. The content of key enzymes for ARA production and metabolism (PLA2 and COX-2) in chicken cecum tissues was increased after S. Typhimurium infection. The relative mRNA levels of inflammatory factors were also increased after infection, including IFN-γ, TGF-β1, IL-4, and IL-6. In HD11 cells, the use of a cyclooxygenase (COX) inhibitor reduced the increased levels of COX-2 and PGF2α induced by S. Typhimurium infection and effectively reduced the inflammatory response. In addition, the number of beneficial genera (e.g., Bifidobacterium, Lactobacillus, and Odorobacterium) in the cecum of Wenchang chickens was significantly reduced after infection with S. Typhimurium. In this study, we revealed the structure of the cecal flora of Wenchang chickens infected with S. Typhimurium and demonstrated that S. Typhimurium can mediate the onset and development of intestinal inflammation through the activation of the metabolic pathway of ARA cyclooxygenase in Wenchang chickens. The results can provide data support and theoretical support for the prevention and control of avian salmonellosis.

Pozol is a Mexican beverage prepared from fermented nixtamalized maize dough. To contribute to understanding its complex microbial ecology, the effect of inoculating on MRS-starch pure and mixed cultures of amylolytic Sii-25124 and non-amylolytic W. confusa 17, isolated from pozol, were studied on their interactions and fermentation parameters. These were compared with L. plantarum A6, an amylolytic strain isolated from cassava. Microbial growth, kinetic parameters, amylolytic activity, lactic acid production, and hydrolysis products from starch fermentation were measured. The population dynamics were followed by qPCR. L. plantarum A6 showed higher enzymatic activity, lactic acid, biomass production, and kinetic parameters than pozol LAB in pure cultures. Mixed culture of each pozol LAB with L. plantarum A6 showed a significant decrease in amylolytic activity, lactic acid yield, specific growth rate, and specific rate of amylase production. The interaction between Sii-25124 and W. confusa 17 increased the global maximum specific growth rate (µ), the lactic acid yield from starch (Ylac/s), lactic acid yield from biomass (Ylac/x), and specific rate of lactic acid production (qlac) by 15, 30, 30, and 40%, respectively compared with the pure culture of Sii-25124. Interactions between the two strains are essential for this fermentation.

Recent interest in intestinal hormones has risen with the idea that they modulate glucose tolerance and food intake through a variety of mechanisms, and such hormones like peptide YY (PYY), ghrelin, glucagon-like peptide (GLP)-1 and 2, and cholecystokinin (CKK) are therefore excellent therapeutic candidates for the treatment of diabetes and obesity. Furthermore, in the recent years, multiple studies suggest that the microbiota is critically important for normal host functions, while impaired host microbiota interactions contribute to the pathogenesis of numerous common metabolic disorders. In this study, we considered the nutraceutical effects of β-glucans added to pasta at the concentration of 6g\100g. Ten participants have been recruited and hematochemical analyses and intestinal hormones tests have been performed before and after 30 days of pasta intake. Stool specimens have been studied for Lactobacillus Fermentum, Lactobacillus acidophilus, Lactobacillus salivarius, Bifidobacterium longum, and Enterococcus faecium presence before and after 30 days of nutritional intervention. After 30 days of regular intake of pasta enriched by β-glucans results have been evaluated. In conclusion, pasta prepared from barley flour enriched with β-glucans at 6% exhibit promising responses on glucose metabolism, on intestinal hormones responses and on microbiota modification.

The emergence of antibiotic-resistant Campylobacter jejuni, a leading cause of gastroenteritis worldwide, presents a significant public health challenge requiring vigilant surveillance and disease control. This study aimed to characterize C. jejuni strains isolated in Romania from 2017-2020, focusing on genetic diversity, virulence, and antibiotic resistance determinants. The isolates underwent phenotypical testing, PCR, and antibiotic resistance assessment using Disc Diffusion Kirby–Bauer for ciprofloxacin, nalidixic acid, erythromycin, and tetracycline. Genetic analysis identified resistance and virulence genes, point mutations, and performed sequence typing (7-gene MLST) to determine genetic relatedness. Results indicated substitutions at position 86 in the amino acid sequence or position 257 in the nucleotide sequence of the gyrA gene in 47 fluoroquinolone-resistant isolates. Additionally, mutations in the rRNA23S gene at positions 2074 and 2075, associated with macrolide resistance, were found in 12 of the 66 isolates. Allelic profiles generated 38 sequence types (STs), including three new STs not present in the reference database. The sequence data analysis revealed a genetically diverse C. jejuni population with a weak clonal structure. This study provides crucial insights into the genetic diversity and antibiotic resistance of C. jejuni strains in Romania, highlighting the need for ongoing surveillance and control measures.

Gas-water relative permeability curve is the foundation of gas reservoir dynamic analysis and numerical simulation. It is generally obtained through gas displacing water experiment. Referring to the existing national standards, gas-water relative permeability test is carried out for low-permeability and tight sandstone. The flow is discontinuous and the curve smoothness is poor, which affects the application of gas-water relative permeability curve. Based on the comparative analysis of a large number of mercury injection and NMR experimental results of low-permeability and tight sandstone, a method to characterize the throat radius distribution of low-permeability tight sandstone based on NMR T2 spectrum is explored. Combined with high-speed centrifugal experiment, the calculation formulas of core water saturation, phase relative permeability of gas-water under different centrifugal force are established. A new method of "nuclear magnetic resonance + high-speed centrifugation" for gas-water relative permeability curve test of low-permeability and tight sandstone is formed, and the test is carried out on Sulige low-permeability tight sandstone core. The results show that: (1) The shape of cumulative distribution curve of throat radius by mercury injection method is basically consistent with that of T2 relaxation time by NMR, and there is a good corresponding relationship between them. (2) The centrifugation process is similar to the gas displacing water process. With the decrease of water saturation, water centrifugation in rock core becomes more and more difficult until it become irreducible. (3) The test process of the new method is unified and standard and up to 6 cores can be tested at the same time. The calculation results of water saturation and relative permeability curve are reliable and the integral curve is smooth, which greatly improves the test efficiency of gas-water relative permeability curve in low-permeability and tight sandstone. The new method can be widely used to study the gas-water two-phase seepage law in low-permeability and tight sandstone gas reservoir.

This study aimed to explore the fermentation behavior of selected potential probiotic strains during sorghum fermentation, considering their microbial counts, pH levels, and antibacterial activity, their ability to break down starch. The fermentation processes were conducted in the presence and absence of glucose, and the lactic acid bacterial strains underwent controlled fermentation to assess microbial counts, amylase activity, pH, and lactic acid levels in the presence and absence of glucose. In addition, the inhibitory activity of these potential probiotic strains against pathogenic bacteria (Salmonella typhimurium, Escherichia coli, and Staph-ylococcus aureus) was examined through co-culturing. The results demonstrated that all nine Lactobacillus strains exhibited robust growth in both sugar and sugar-free fermentation experiments. Glucose supple-mentation significantly enhanced lactic acid yield which was in a range of 0.19-0.52 % compared to fer-mentation without glucose with a range of 0.03-0.29 %. The selected Lactobacillus strains effectively lowered the pH to less than 4 after 24 hours, producing substantial lactic acid. Notably, in the absence of glucose, only Lb. helveticus D7 and Lb. amylolyticus D12 achieved pH levels below 4 after 8 hours, producing the highest lactic acid amounts of 0.27 and 0.29 % after 24 hours, respectively. Amylase activity was detected on two strains D7 and D12. Furthermore, most of the tested Lactobacillus strains demonstrated complete inhibition of pathogen growth after 24 hours of co-culturing, suggesting their potential for enhancing the safety quality of sorghum-based fermented products.

The expression of polysialic acid (polySia) on the neuronal cell adhesion molecule (NCAM), is called NCAM-polysialylation, which is strongly related to the migration and invasion of tumor cells and with aggressive in the clinic. Thus, it’s important to select a proper drug to block tumor cell migration for clinic treatment. In this study, we proposed that lactoferrin (LFcinB11) may be a more ideal candidate for inhibition of NCAM polysialylation comparing with CMP and low molecular weight heparin (LMWH), which were determined based on our NMR studies. On the other hand, because neutrophil extracellular traps (NET) is the most dramatic stage in cell death process, and NETs’ release are related to pathogenesis of autoimmune and inflammatory disorders, with proposed involvement in glomerulonephritis, chronic lung disease, sepsis and vascular disorders. In this study, the molecular mechanism of inhibition of NETs’ release using LFcinB11 as inhibitor was also determined. Based on these results, LFcinB11 is proposed to be bifunctional inhibitor for inhibiting both NCAM polysialylation and NETs releases.

We propose novel spectrometer designs that aim to enhance the measured spectral range 1 of ions on a finite-sized detector. In contrast to the traditional devices that use uniform magnetic 2 field, in which the deflection of particles increases inversely proportional to their momentum, in a 3 gradient magnetic field, the deflection of particles will be decreased due to the reduction of the 4 magnetic field along their propagation. Low energy ions are particularly impacted, as they are 5 deflected less and are more likely to reach the detector compared to being driven out of it while 6 using a uniform magnetic field. By the means of a gradient magnetic field, the nonlinear dispersion 7 of ions in a homogeneous magnetic field can approach a nearly linear dispersion. Nonetheless, 8 the dispersion of low energy ions in a homogeneous magnetic field remains unnecessarily high. 9 In this article, we explore linear, quadratic, and higher-order field profiles in comparison to the 10 homogeneous field case. We discuss the employed methodology and present simulation results of 11 the spectrometer with an extended ion spectral range, focusing on the minimum detectable energy 12 (energy dynamic range) and energy resolution

(1) Background: Formation of biofilms on food-contact surfaces by Shiga-toxigenic Escherichia coli (STEC) can pose a significant challenge to the food industry, making conventional control-methods insufficient. Targeted use of bacteriophages to disrupt these biofilms could reduce this problem. Previously isolated and characterized bacteriophages (n=52) were evaluated against STEC biofilms in-vitro and on food-contact surfaces. (2) Methods: Phage-treatments (9 logs PFU/ml), in phosphate-buffered-saline, were used individually or as cocktails. Biofilms of STEC (O157, O26, O45, O103, O111, O121, and O145) were formed in 96-well micro-titer plates (7 logs CFU/ml; 24 h) or on stainless-steel (SS) and high-density-polyethylene (HDPE) coupons (9 logs CFU/cm2; 7 h), followed by phage-treatment. Biofilm-disruption was measured in-vitro at 0, 3, and 6 h as a change in optical-density (A595). Coupons were treated with STEC-serotype-specific phage-cocktails or a 21-phage cocktail (3 phages/serotype) for 0, 3, 6 and 16 h and surviving STEC populations enumerated. (3) Results: Of the 52 pages, 77% showed STEC-biofilm disruption in-vitro. Serotype-specific phage-treatments reduced pathogen population within the biofilms by 1.9-4.1 and 2.3-5.6 logs CFU/cm2, while the 21-pahge cocktail reduced it by 4.0 and 4.8 logs CFU/cm2 on SS and HDPE, respectively. (4) Conclusions: Bacteriophages can be used to reduce STEC and their biofilms.

Several pathotypes of enteric E. coli have been identified. The group represented by Shiga toxin-producing E. coli (STEC) is of particular interest. Raw milk and raw milk products are significant sources of STEC infection in humans; therefore, identifying pathogens at the herd level is crucial for public health. Most national surveillance programs focus solely on raw milk and raw milk cheeses that are ready for retail sale, neglecting the possibility of evaluating the source of contamination directly at the beginning of the dairy chain. In order to assess the viability of new molecular methodologies applied to raw milk filters enabling the identification of the presence of STEC serotypes in milk production and apply the same methods to identify the presence of these pathogens in calf feces we analyzed, through a commercial real-time PCR assay, a total of 290 samples coming from 18 different dairy herds; 88 bulk tank milk (BTM), 104 raw milk filters (RMF) and 98 calves’ feces samples. In total 3.4% of BTM, 41.4% of RMF and 73.4% of calves’ feces tested positive for stx presence, supporting our hypothesis that BTM is not a suitable matrix to assess the STEC presence at herd level, underestimating it. Our conclusion is that the surveillance program needs critical and extensive improvements such as RMF and calves’ feces analysis implementation in order to be more efficient in detecting and preventing STEC infections. The epidemiology of these infections and the characteristics of the pathogen clearly show how a One Health approach will be pivotal to improve our capabilities to control the spread of these infections.

As one of the typical deposits in Sichuan-Yunnan-Guizhou Pb-Zn metallogenic province, the Daliangzi Pb-Zn deposit has a close genetic relation with the structural system of “black/fracture zone” formed under the action of NWW-approx. EW strike-slip structures in the metallogenic province, but the mechanism of rock and ore-controlling action of the structural system remains unclear. Based on detailed analysis of tectonite-mineralized alteration lithofacies of R1 “black/fracture zone”, the tectonite-mineralized alteration lithofacies zones can be divided into 4 types in succession outward from the Pb-Zn mineralization center, i.e., ① brecciated and stockwork-like Pb-Zn mineralization-complex breccia facies zone, ② stockwork-like Pb-Zn mineralization-simple breccia and cataclasite facies zone, ③ veined pyrite-sulfide-dolomitic cataclasite facies zone, and ④ fine-veined calcite-black carbonized dolomite facies zone; Cl- and SO42--rich high-temperature oxidizing ore-bearing fluid mixes with reducing fluid in Zone ① while migration and precipitation occur in Zone ④. With the evolution of ore-forming fluid, the homogenization temperature decreases from Zone ① to Zone ④, the salinity increases from Zone ① to Zone ② and then decreases to Zones ③ and ④, and the fluid density has little change on the whole; the contents of Zn, Pb, Cu, Ga, Ge, Cd, Ag, and other metallogenic elements, the Zn/Pb ratio, and the CaO/MgO mole ratio decrease gradually from Zone ① to Zone ④; and REE fractionation, calcilization, silicification, and pyritization enhance gradually from Zone ① to Zone ④. This series of changes are the product of diapirism (cryptoexplosion) of strike-slip structures and “black/fracture zone”, among which the second-order structures (F5, F100, and other faults) derived from NWW-approx. EW striking dextral shear-tension faults F1 and F15 control the brecciated and stockwork-like Pb-Zn mineralized complex breccia facies zones and the stockwork-like Pb-Zn mineralized simple breccia and cataclasite facies zones. Therefore, this paper establishes the zoning mode of tectonite-mineralized alteration lithofacies of “black/fracture zone” and proposes that Zones ① and ② are important prospecting criteria. conclusions.

This study conducted an extensive analysis of campylobacteriosis surveillance in Portugal from 2009 to 2021, aiming to investigate demographic shifts, seasonal variations, and antimicrobial resistance (AMR) within Campylobacter isolates. Surveillance network and sentinel laboratory-based system data revealed a substantial under-notification of campylobacteriosis cases, suggesting an underestimated disease burden. A paradigm shift was observed in notification rates, with a notable prevalence among the pediatric population, particularly in children aged 1-4 years, diverging from European reports. However, the study also highlighted an emerging trend of Campylobacter infections in younger adults (15-44 years). The study unveiled a unique seasonal distribution of cases, defying typical summer peaks seen elsewhere. AMR analysis revealed extremely to very high-level resistance to ciprofloxacin and tetracycline, both in C. jejuni (93.7% and 79.2%, respectively) and in C. coli (96.5% and 93.2%, respectively), stable throughout the studied period (2013-2021). C. coli also exhibited significantly higher resistance to erythromycin, gentamicin, ampicillin, amoxicillin-clavulanic acid (decreased resistance), and ertapenem compared to C. jejuni (P&lt;0.001). In conclusion, the study highlights the need for enhanced surveillance efforts and emphasizes worrying AMR levels. The implementation of WGS-based surveillance is recommended to attain a more profound comprehension of disease patterns and the evolving landscape of AMR.

The intestinal microbial community of aquatic animals is closely related to the environment and thus affects the health of aquatic animals. Here we investigated annual variation of intestinal bacteria community structures of Ruditapes philippinarum cultured using two different methods, bottom sown and raft hanging. Bacterial diversity of seawater and sediment in different seasons was examined as well. Correlation between R. philippinarum intestinal bacteria community structures and environmental factors was analyzed. The results showed that in the two aquculture areas where R. philippinarum was differentially cultured, bacteria community structure of seawater and sediment both altered with seasons. However, neither seasonal changes nor difference in culture methods could significantly affect the intestinal bacteria community structures of R. philippinarum. Additionally, in terms of the alpha diversity of intestinal bacteria communities, there was no significant difference in R. philippinarum cultured using the two methods. For benthic cultured clams, alpha diversity of the intestinal bacterial communities was often higher than that of the environmental bacterial communities. For raft hanging cultured calms, Alpha diversity of the intestinal bacterial communities was normally comparable with that of the environmental counterparts. Beta diversity analysis showed that the intestinal bacterial community structure of R. philippinarum was mainly determined by both deterministic and stochastic processes. Correlation analysis showed that the intestinal bacterial community structures of R. philippinarum were determined by a variety of environmental factors. For clams collected from the benthic culture area, the intestinal bacterial structure was more correlated with the physicochemical properties of sediment in comparing with those of seawater. For clams collected from the raft-culture area, the intestinal bacterial structure was mainly determined by the phsicochemical characteristics of seawater. This study helps us to better understand the bacterial community structure in complex aquaculture ecosystems, providing a reference for healthy shellfish aquaculture and ecosystem protection in aquaculture area.

(1) Background: After the discovery and application of Chlamydomonas reinhardtii channelrhodopsins, the optogenetic toolbox has been greatly expanded with engineered and newly discovered natural channelrhodopsins. However, channelrhodopsins of higher Ca2+ conductance or more specific ion permeability are still in demand. (2) Methods: In this study, we mutated the conserved aspartate of the transmembrane helix 4 (TM4) within Chronos (Stigeoclonium helveticum channelrhodopsin = ShChR) and PsChR (Platymonas subcordiformis channelrhodopsin) and compared them with published ChR2 (C. reinhardtii channelrhodopsin-2) aspartate mutants. (3) Results: We found that the ChR2 D156H mutant (XXM) showed enhanced Na+ and Ca2+ conductance, which was not noticed before, while the D156C mutation (XXL) influenced the Na+ and Ca2+ conductance only slightly. Furthermore, the D173H mutant of PsChR showed a much improved photocurrent, compared to wildtype, and even higher Na+ selectivity to H+ than XXM. PsChR D173H also showed a strongly enhanced Ca2+ conductance, more than 2-fold that of the calcium translocating L132C of ChR2 (CatCh). (4) Conclusions: We found that mutating the aspartate of the TM4 to histidine influences the ion selectivity of channelrhodopsins. With the large photocurrent and enhanced Na+ selectivity and Ca2+ conductance, XXM and PsChR D139H are promising powerful optogenetic tools, especially for Ca2+ manipulation.

This study aims to investigate the interfacial tension, emulsion stability, and the impact on oil displacement effectiveness of mixed systems containing nonionic surfactants and amphoteric surfactants at different ratios. To achieve this, three different emulsion stability formulations were prepared by mixing betaine and nonionic surfactants in ratios of 1:1, 3:2, and 3:1 respectively. By adjusting the proportions of betaine and nonionic surfactants, variations in interfacial tension and emulsion formation among the three mixed systems were studied to analyze the mechanism of interaction between betaine and nonionic surfactants and evaluate their effects on crude oil displacement and enhanced oil recovery in reservoirs. Subsequently, microscopic and macroscopic oil displacement experiments were conducted on the three mixed systems, along with experiments on emulsion flow resistance. The aim was to verify the oil displacement effectiveness of different mixed systems and further understand their potential applications in reservoirs. By comprehensively analyzing the experimental results, optimal ratios and conditions for the mixed systems were determined. The experimental results indicate that the emulsification system formed by mixing betaine and nonionic surfactants at a ratio of 3:1 achieves molecular synergies through electrostatic interactions and mixed adsorption with betaine, resulting in a compact arrangement of interface membranes and demonstrating a synergistic effect in reducing interfacial tension to an ultra-low value of 10 -4 mN/m. Additionally, as the proportion of betaine surfactant increases, the viscosity of the aqueous phase liquid film increases, the rate of thinning of the liquid film slows down, the rate of water release slows down, and the stability of the emulsion increases. Microscopic oil displacement experiments verified that the three mixed systems increase oil washing efficiency through low interfacial tension and emulsification stripping action, thereby increasing the oil displacement mechanism's coefficient. Emulsion flow resistance experiments demonstrated that as the proportion of betaine surfactant increases, the system exhibits better plugging ability. However, macroscopic oil displacement experiments revealed differences in compatibility between different emulsion stability solutions and rocks with different permeabilities. The mixed system of betaine and nonionic surfactants at a ratio of 3:1 exhibits the best emulsification plugging ability in rocks with a permeability of 109mD, while the 1:1 mixed system performs best in rocks with a permeability of 12mD. This holds significant theoretical and practical significance for reservoir development and enhanced production.

The aim of our study was a comprehensive comparative assessment of the effect of consumption of 1 and 2% cholesterol as part of a standard diet with and without regular moderate physical activity on the main biomarkers of lipid and cholesterol metabolism, as well as the intestinal microbiota of male Wistar rats. The animals were randomly divided into 5 groups according to body weight and Open Field test results: Control (n=10), 1% Chol (n=10), 2% Chol (n=10), 1% Chol + RUN (n=10) and 2% Chol + RUN. The cholesterol was additionally added in the amount of 1g or 2g/100g, respectively, to the diet of animals. The rats of experimental groups 1% Chol + RUN and 2% Chol + RUN were subjected to regular (2 days a week) physical activity on a Treadmill. In rats of four experimental groups, when compared to the Control, a response of 39 indicators was revealed. The largest number of changes in biomarkers - 29 (statistically significant, p

The vast array of omics data in microbiology presents significant opportunities for studying bacterial pathogenesis and creating computational tools for predicting pathogenic potential. However, the field lacks a comprehensive, curated resource that catalogs bacterial strains and their ability to cause human infections. Current methods for identifying pathogenicity determinants often introduce biases and miss critical aspects of bacterial pathogenesis.In response to this gap, we introduce BacSPaD (Bacterial Strains’ Pathogenicity Database), a thoroughly curated database focusing on pathogenicity annotations for a wide range of high-quality, complete bacterial genomes. Our rule-based annotation workflow combines metadata from trusted sources with automated keyword matching, extensive manual curation, and detailed literature review. Our analysis classified 5,502 genomes as pathogenic to humans (HP) and 490 as non-pathogenic to humans (NHP), encompassing 532 species, 193 genera, and 96 families. Statistical analysis demonstrated a significant but moderate correlation between virulence factors and HP classification, highlighting the complexity of bacterial pathogenicity and the need for ongoing research. This resource is poised to enhance our understanding of bacterial pathogenicity mechanisms and aid in the development of predictive models. To improve accessibility and provide key visualization statistics, we developed a user-friendly web interface, accessible at https://bacspad.altrabio.com.

Esophageal cancer is an extremely deadly tumor that accounts for 5% of all cancer deaths. The two main subtypes of the disease are esophageal squamous cell carcinoma (ESCC) and esophageal adenocarcinoma (EAC). To date, few studies analyse EAC for transcriptional signatures associated with diagnosis and/or prognosis, while most focus on the analysis of transcriptional profiles of ESCC. In this study, we used single-cell RNA sequencing (scRNA-seq) analysis of CD45+ cells enriched from Tumors and matched Non Tumor tissues from therapy naїve patients to identify all types of immune cells present in tumor immune infiltrate and its transcriptional profile. In addi-tion, we analysed the entire transcriptome in a cohort of 23 patients whose tissue biopsies were taken from Tumors and matched with Non Tumor tissue. The transcriptional signatures we obtained were then used to stratify a large cohort of TCGA EAC patients, demonstrating a strong association with their prognosis, as well as the ability to predict a patient's clinical outcome and better define the prognosis of EAC after surgery. In addition, these features may lead patients to effective therapies including immunotherapy approaches.

Our prior studies have shown that B type Natriuretic Peptide (BNP) decreases absorption from the gastrointestinal (GI) tract. Since non-muscle myosin-type II (NMM-II) is known to have a role in epithelial tight junction regulation, we aimed to test whether BNP has an effect on the localization and expression of NMM-II in the intestinal villi. We treated wild type mice with a 10 ng/g intravenous bolus of BNP followed by infusion of BNP at 1ng/g/minute vs. vehicle control. The mice were then euthanized and intestinal tissue isolated and sectioned. The tissue was immune-stained for NMM-II and examined using fluorescence microscopy. The tissue section not treated with antibody to NMM-II was used as a methodological control to evaluate the non-specific binding. Scanning electron micrographic images were taken to compare structural differences between the two groups. Western blotting was performed to compare the regional protein expression of NMM-II and associated proteins including smooth muscle actin, kinesin, and E cadherin in small intestinal tissue between BNP vs. vehicle infused mice. Fluorescence microscopy revealed markedly increased localization of NMM-II at the crypts and core of intestinal villi of the jejunum, ileum and colon of BNP treated mice compared to vehicle. Transmission electron microscopy revealed that the microvilli of BNP treated mice assumed the distinctive appearance of ‘relaxed’ microvilli compared to vehicle. Such decrease in contractility was previously shown to decrease para-cellular permeability of epithelia. Our observation indicates that BNP alters the localization of NMM-II in intestinal villi and makes the intestinal villi structures assume a state of decreased permeability of the apical junctional complex. Further characterization of this process and understanding the specific mediators involved could lead to the identification of novel therapeutic targets for various disease states such as heart failure and malabsorption.

Recent technical advances in the stem cell field have enabled the in vitro generation of complex structures resembling whole organs termed organoids. Most of these approaches employ culture systems that allow stem cell-derived or tissue progenitor cells to self-organize into three-dimensional (3D)-structures. Since organoids can be grown from various species, organs and from patient-derived induced pluripotent stem cells, they create significant prospects for modelling development and diseases, for toxicology and drug discovery studies, and in the field of regenerative medicine. Here, we report on intestinal stem cells, organoid culture, organoid disease modeling, transplantation, current and future uses of this exciting new insight model to veterinary medicine field.

Pore Forming Toxins (PFTs), formed mainly by virulence factors of bacteria, belongs to Pore Forming Protein (PFP) family. Secreted as soluble monomers, they bind specific targets in membranes where their oligomerization and insertion place. Lysenin, a member of the PFTs, forms and oligomer after sphingomyelin binding, the so-called prepore, which become inserted forming a pore after a conformational change triggered by a pH decrease. In crowded conditions, oligomers tends to stay in prepore form because the prepore-to-pore transition is sterically blocked. In this study, we investigate the effect of calcium ions in those crowded conditions, finding that calcium act as a trigger for lysenin insertion. We localize the residues responsible for calcium sensitivity in a small α-helix. Our results are not only one of the few complete structural descriptions of prepore-to-pore transitions but the very first that involves a calcium triggering mechanism. The presence of glutamic or aspartic acids in the insertion domains could be an indication that calcium may be a general trigger for PFTs and more generally PFP.

Heavy metal accumulation in soils used for agricultural purposes is a major issue as it can affect the growth of plants and contain health risks as plants containing heavy metals are taken as food. This study provided the potential of three Bacillus pumilus strains (C-2PMW-8, C-1 SSK-8, and C-1PWK-7) for mitigation of the health factors related to heavy metals on lettuce (Lactuca sativa) grown in chromium contaminated soil from activities related to mining. Lettuce seeds inoculated with B. pumilus strains were grown in pots with garden soil amended with varying amount of chromite mine soil (2.27%, 4.65%, 7.14%). Growth indicators like days to germination, percentage of germination, leaf weight, leaf area and rate of survival decreased with incremental soil chromium levels but improved highly with inoculation of B. pumilus compared to uninoculated controls. When B. pumilus strains were provided, there was also a decreased level in the buildup of chromium in soil as well as lettuce shoots. The results explains that plant growth-promoting B. pumilus strains may be able to lower down the detrimental effects of chromium on lettuce development, hence increment in the effectiveness of phytoremediation.

The toxin-antitoxin (TA) systems have been attracting attention due to their role in regulating prokaryotic cell responses to stress and their biotechnological potential. Much recognition has been given to type II TA system of mesophiles, but so far, limited attention has been given to thermophiles. Here, we are presenting the putative type II TA families encoded on the genomes of four Geobacillus strains. We employed the TA finder tool to mine for TA-coding genes and manually curated the results using various tools. We identified 28 putative TA pairs, distributed over 8 TA families. Among the identified TAs, 15 represent putative novel toxins and antitoxins that have been overlooked or annotated as hypothetical proteins in their genome records. We also identified a potentially new TA composite, AbrB-ParE. Furthermore, we are suggesting the Geobacillus acetyltransferase toxin-antitoxin (GacTA) family which potentially represents one of the unique TA families that has a reverse gene order. Moreover, we are proposing a hypothesis on the regulation of the xre-cog2856 gene expression, which seems to involve c-di-AMP. This study aims for highlighting the significance of studying TAs in Geobacillus, since they have special features. It also aims for facilitating future experimental research.

Nanostructured carriers have been widely used in pharmaceutical formulations for dermatological treatment. They offer targeted drug delivery, sustained release, improved biostability, and low toxicity, usually presenting advantages over conventional formulations. Due to its large surface area, small size and photothermal properties, graphene oxide (GO) has the potential to be used for such applications. Nanographene oxide (GOn) presented average sizes of 197.6 ± 11.8 nm, and a surface charge of -39.4 ± 1.8 mV, being stable in water for over 6 months. 55.5 % of the mass of GOn dispersion (at a concentration of 1 mg mL-1) permeated the skin after 6 h of exposure. GOn dispersions have been shown to absorb near-infrared radiation, reaching temperatures up to 45.7 °C, within mild photothermal therapy temperature range. Furthermore, GOn in amounts superior to those which could permeate the skin were shown not to affect human skin fibroblasts (HFF-1) morphology or viability, after 24 h of incubation. Due to its large size, no skin permeation was observed for graphite particles in aqueous dispersions stabilized with Pluronic P-123 (Gt-P-123). Altogether, for the first time, GOn potential as a topic administration agent and for delivery of photothermal therapy has been demonstrated.

Evaluation of drug permeability using cell monolayers is an important in vitro assay in the process of drug development, to identify compounds with favourable pharmacokinetics that may proceed to in vivo studies. When allowed to differentiate on permeable filters for at least 21 days, monolayers of Caco-2 cells develop morphological and functional properties similar to those of the intestinal epithelium. Permeability through Caco-2 monolayers is thus particularly relevant to assess drug absorption after oral administration. The major drawback of this assay is its very low throughput, with the cell monolayers being used in a single permeability assay. We have previously proposed a protocol to allow the re-use of the Caco-2 monolayer on additional permeability assays during the stable period and shown that cell monolayer integrity is maintained. In this work we evaluate the effect of re-use on the permeability of several reference compounds that permeate through passive pathways and show that the cell monolayers can be re-used twice in additional permeability assays. Preliminary results were also obtained regarding active transport pathways, and a new methodology is proposed to quantify the contribution from passive and active transport. This is a major step towards the full validation of the re-use methodology proposed, which at least triplicates the throughput of this important in vitro assay.

Appling the controlling relative permittivity and permeability in the equations for the gravitational-magnetic-electric field interaction, a very large variation of the gravitational acceleration of the Earth by electric/magnetic field could be arrived at. This conclusion may be supported by some of the experiments for the gravitational effect of superconductivity.

The activity of the mitochondrial Permeability Transition Pore, mPTP, a highly regulated multi-component mega-channel, is enhanced in aging and in aging-driven degenerative diseases. mPTP activity accelerates aging by releasing large amounts of cell-damaging Reactive Oxygen Species, Ca2+ and NAD+. The various pathways that control the channel activity, directly or indirectly, can therefore either, inhibit, or accelerate aging, retards, or enhance the progression of aging-driven degenerative diseases, and determine lifespan and healthspan. Autophagy, a catabolic process that removes and digests damaged proteins and organelles protects the cell against aging and disease. However, the protective effect of autophagy depends on mTORC2/SKG1 inhibition of mPTP. Autophagy is inhibited in aging cells. Mitophagy, a specialized form of autophagy, which retards aging by removing mitochondrial fragments with activated mPTP, is also inhibited in aging cells, and this inhibition leads to increased mPTP activation, which is a major contributor to neurodegenerative diseases, such as Alzheimer’s and Parkinson’s Diseases. The Increased activity of mPTP in aging turns autophagy/mitophagy into a destructive process leading to cell aging and death. Several drugs and lifestyle modifications that enhance healthspan and lifespan enhance autophagy and inhibit the activation of mPTP. Therefore, elucidating the intricate connections between pathways that activate and inhibit mPTP, in the context of aging and degenerative diseases, could enhance the discovery of new drugs and lifestyle modifications that slow aging and degenerative disease.

In this study, the relationship between air diffusivity and permeability in cementitious materials was investigated. First, we derived an equation to correlate air diffusivity and permeability in a straight circular tube. Then, we reviewed existing studies that measured both air diffusivity and permeability and compared reported data and calculated values to verify the applicability of the derived equation to cementitious materials. Although a correction factor was not used, the two sets of data showed good agreement quantitatively. This indicates that the derived equation can be applied to cementitious materials including concrete, and measured air diffusivity can be converted to permeability of concrete and vice versa using the derived equation.

Training researchers and stakeholders is crucial to mitigate the risks associated with toxins and venom. By sharing knowledge on the responsible use of toxins, venoms, chemicals, and data from biosecurity and cyberbiosecurity perspectives, we empower laboratorians to make informed choices. We need to remember that cultivating a culture of sustainable biosafety and biosecurity is vital for one health. This technical note highlights the essence of safeguarding biosecurity and cyberbiosecurity in toxin and venom research laboratories workshop designed and implemented to boost biosecurity.

We previously showed that feeding mice a diet containing 1% mantle tissue decreased food consumption, leading to death. We also isolated and identified toxic substances in the mantle tissue. In the present study, we investigated the characteristics and stability of mantle tissue toxicity. Treatment of mantle tissue with 1 mM HCl, 1 mM NaOH, 1 mM dithiothreitol, and 1 mM H2O2 and heating did not reduce the toxicity of mantle tissue in mice. These results suggest that mantle toxins are stable in tissues, particularly when exposed to acidic and digestive enzymes. We examined whether mantle tissue exhibited acute toxicity. Diets containing 1% and 20% mantle extract showed similar levels of toxicity, demonstrating that feeding of mantle tissue does not lead to acute toxicity. Finally, we examined the toxicity of the mantle tissue against small intestinal tissue. Chronic feeding of mantle tissue to mice changed the color of the small intestine. Real-time PCR analysis showed that mantle tissue feeding caused changes of inflammation and endoplasmic reticulum stress markers in the small intestine. These results suggest that feeding of mantle tissue causes toxicity after causing initial damage to the small intestinal tissue.

Bacillus anthracis, the causative agent of anthrax, has two virulence plasmids: pXO1 and pXO2. Plasmid pXO2 carries the genes of an antiphagocytic capsule synthesis. Plasmid pXO1 carries the genes pagA, lef, and cya encoding anthrax toxins, as well as atxA, which encodes the main regulator of pathogenicity factor expression. In the present paper, we evaluated the polymorphism of the pagA, lef, cya, and atxA genes for 85 B. anthracis strains belonging to different evolutionary lineages and canSNP groups and three B. cereus strains possessing pXO1-like plasmids. We found 19 genotypes (GT) strongly correlated with the main evolutionary lineages; they were mainly correlated with the canSNP group within these lines. In some cases, one GT combined the strains of several canSNP groups, and some canSNP groups were divided into several GTs. The most interesting of such cases were 1) the formation of a separate GT by strains of the A.Br.008/009 group isolated in the former USSR; 2) the presence of a unique GT in some American strains of the A.Br.Aust94 group; 3) the division of the A.Br.001/002 group into two subgroups, one of which may be a transitional link to the group A.Br.Ames; 4) the fact that European isolates of the B.Br.CNEVA group and Arctic strains of the B.Br.001/002 group have a common GT.

Codium fragile, a green algae, exhibit biological properties. In this study, we examined the effect of Codium fragile extract (CFE) on dextran sulfate sodium (DSS)-induced colitis. As the administration of CFE increased, the proliferation of Akkermansia muciniphila, which is key player in metabolic and gastrointestinal disorders, increased. After CFE administration for 10 weeks, acetic acid was revealed as the major metabolite in mouse cecum and β-glucuronidase activity in mouse feces is decreased. Further, CFE significantly alleviates the acute intestinal injury induced by DSS administration, including DAI scores and colon length. MPO activity in the colon was significantly reduced in the CFE-fed group, which indicated that CFE could inhibit neutrophils infiltration and inflammation in mice. In conclusion, a protective effect of CFE against DSS colitis suggests its clinical use for IBD patients.

Dissemination of resistant pathogens via food supply chains is a notable public health danger from farm to fork. The proposed work analysed the distribution of E. coli among various sources of animal-based food supply chain. Overall, 500 samples were heaped up from livestock, poultry sectors, environment, fisheries, and dairy. Standard microbiological procedures were used to isolate and identify E. coli. Further, isolates were confirmed by using MALDI-TOF and virulence associated genes (VAGs). The phenotypic resistance pattern of the isolates was determined by molecular identification of antibiotic resistance genes (ARGs) through PCR using specific primers. Shiga toxin producing E. coli (STEC) were subjected to PCR based O typing using specific primers against different O types. Overall results revealed that 154 (30.5%) samples were confirmed as E. coli, in which77 (50%) isolates were MDR E. coli. Among these, total 52 (67.53%) isolates showed array of VAGs in which 21 (40.38%) were confirmed as STEC based on stx1 and stx2. Furthermore, 12/52 (23.07%) isolates were detected as Non-O157 STEC co-harboring mcr-1 and blaNMD-1. Overall, O26 STEC was found to be the most prevalent among different non-O157 types. Herein, results proposed that detection of such STEC MDR E. coli causing serious public health consequences in developing countries.

The development effect of water flooding in ultra-low permeability reservoir was poor due to its poor physical property and high shale content, the experimental study of air flooding which help to complement producing energy was carried out. Based on the Accelerating Rate Calorimeter experimental results, the crude oil of N block in L oilfield can produce low-temperature oxidation reaction, which was the basic condition of air flooding. Three groups of experiment natural cylinder core were designed for oil displacement, water flooding and air flooding were used respectively, and the relationship of differential pressure, oil recovery, injection capacity with injection volume was investigated. It has been showed on the result that the recovery efficiency improved sharply than water flooding, the effect of depressurization and augmented injection was obvious, and the air displacement was validation.

The focus of this work lies in the construction of monitoring system designed in the form of a patch using a long-lasting potentiometric electrolyte concentration sensor from metal electrodes, in a galvanic cell configuration Mg(s)|NaCl(aq)|Cu(s). Initially, a mathematical analysis was conducted on the expected sensor output through the Nernst equation. The experimental phase involved conducting controlled tests in the laboratory using various molar concentrations of sodium chloride (NaCl) in aqueous solution. Results showed that the electrolyte sensor allows for monitoring electrolyte levels in NaCl in the range of 0.02 to 1 M/L, with electrode lifespan exceeding 10 days. On the other hand, when patch is using human sweat during controlled exercise routines, it showed the sensor's capability for monitoring changes in the subjects' hydration levels in real time. The sensor output potential is greater than 1.7 volts when the sensor electrodes are in contact with human sweat, which is equivalent to measuring molar concentrations at normal hydration and moderately high hydration levels. Sensor results indicate that the system can function in a self-powered manner without external batteries. Additionally, the extended lifespan of its electrodes allows for continuous monitoring of imbalances in the human body, showcasing the system capability for long-term health assessment.

Background: The incidence of metabolic dysfunction-associated steatohepatitis (MASH)-related hepatocellular carcinoma (HCC) is increasing annually with a rise in the incidence of obesity and metabolic syndrome. Based on preliminary reports regarding the potential association of HCC and periodontitis, this study aimed to analyze the involvement of periodontal bacteria as well as the oral and intestinal bacterial flora in MASH-related HCC (MASH-HCC). Methods: Forty-one patients with MASH and 19 with MASH-HCC who completed survey questionnaires, underwent periodontal examinations, and subjected to sample collection (saliva, mouth-rinsed water, feces, and peripheral blood) participated in the study. Bayesian network analysis was used to analyze the causation between various factors, including MASH-HCC, examinations, and bacteria. Results: The occupancy rate of the genus Fusobacterium in the intestinal bacterial flora was significantly higher in in the MASH-HCC group than in the MASH group (p = 0.002), whereas that of Butyricicoccus (p = 0.022) and Roseburia (p &lt; 0.05) was significantly lower. Bayesian network analysis revealed the absence of periodontal pathogenic bacteria and enteric bacteria affecting HCC. However, HCC directly affected the periodontal bacterial species Porphyromonas gingivalis, Tannerella forsythia, Fusobacterium nucleatum, and Prevotella intermedia in the saliva, as well as the genera Lactobacillus, Roseburia, Fusobacterium, Prevotella, Clostridium, Ruminococcus, Trabulsiella, and SMB53 in the intestine. Furthermore, P. gingivalis in the oral cavity directly affected the genera Lactobacillus and Streptococcus in the intestine. Conclusion: MASH-HCC directly affects periodontal pathogenic and intestinal bacteria, and P. gingivalis may affect the intestinal bacteria associated with gastrointestinal cancer.

Keywords: gut microbiota; small intestinal bacteria overgrowth; SIBO; diet, dysbiosis; microbial ecology.

Microcystis aeruginosa is a common cyanobacteria found in water blooms and often causes ecological harm. Antibiotics are also increasingly used for the prevention and treatment of bacterial infections in aquaculture. However, since most antibiotics cannot be fully metabolized, they enter the water environment and cause ecological impacts. In this paper, the effects of the two quinolone antibiotics (enrofloxacin and ciprofloxacin) on the population density and microcystins (MCs) production of Microcystis aeruginosa were studied. It is of great significance for the ecological risk assessment of antibiotics to the water environment. The results showed that the 96h EC50 values of enrofloxacin and ciprofloxacin to Microcystis aeruginosa were 56.10mg/L and 49.80mg/L respectively, and the toxicity of ciprofloxacin to Microcystis aeruginosa was slightly stronger than that of enrofloxacin. With the increase of the two quinolone antibiotic concentration, the growth inhibition rate (IR) increased, but when the concentration reached a certain height, the IR would reach its threshold. Long-term exposure to low concentrations of these two quinolone antibiotics is not only more likely to lead to the outbreak of Microcystis aeruginosa, but also increase its toxin production capacity. The highest contents of MCs in enrofloxacin and ciprofloxacin groups were 95.539 μg/g and 93.720 μg/g, respectively. The MCs value of these three enrofloxacin treatment groups was more than above 51.8 times that of control group (CK) on the 4th day; from 8th day to 14th day, the MCs value of these three enrofloxacin treatment groups was more than above 3.2 times that of CK group. For another ciprofloxacin, the MCs value of the treatment group was more than 64.98 times that of the CK group on the 4th day, and from 8th day to 14th day, the MCs value of the treatment group was more than 2.7 times that of the CK group. These findings provide crucial management rationale.

Environmental genetics-related modern methods are shown as important indicators of various cyanotoxins syntheses, and their knowledge and use are critically analyzed. Microcystins and other cyanotoxins loads and syntheses are related to different drivers, like various chemical elements and compounds (especially nutrients, such as nitrogen and phosphorus, and their ratio), then to the light, conductivity, temperature, and other climatical and hydrological factors, to which spatial and geographical features (such as the surface of the water bodies) have to be added. The biotic relationships include different specific and supraspecific, uni- and bilateral links between the cyanobacteria, and subsequently their synthesized toxins, and protozoans (or protoctists), chromists, macrophytes, different systematical and ecological groups of zooplankton, and others. The importance of, but also the gaps in, the knowledge and the scarcity of studies involving ectocrines mediated interactions between different groups of algae and plants are highlighted. The paper ends with an interesting classification of lakes' trophicity, illustrated with conceptual diagrams, based on possible scenarios of cyanobacteria behavior.

Cercospora beticola Sacc. is an ascomycete pathogen that causes Cercospora leaf spot in sugar beets (Beta vulgaris L.) and other related crops. The disease symptoms are characterized by circular lesions with an ash-gray centre and dark brown to purple margins on the leaves. It can lead to significant yield losses if not effectively managed. This study aims to characterize and assess the impact of bacterial biological control agents against C. beticola. Following a dual-culture screening, 18 bacteria exhibiting over 50% inhibition were selected, with six of them demonstrating more than 80% control. Identification of the bacteria was carried out through sequencing of the16S rDNA, revealing potentially 12 species belonging to six genera including Bacillus which was represented by four species. Additionally, the bacteria underwent in-depth characterization of biochemical and molecular properties, as well as plant-growth promotion. PCR detection of genes responsible for producing antifungal metabolites revealed that 83%, 78%, 89%, and 56% of the selected bacteria possessed the bacillomycin, iturin, fengycin, and surfactin encoding genes, respectively. Infrared spectroscopy analysis confirmed the presence of lipopeptide structure in the bacteria supernatant filtrate. Subsequently, the bacteria were assessed for their effect on sugar beet plants in controlled conditions. The bacteria exhibited notable capabilities, promoting growth in both roots and shoots, resulting in significant increases in root length, weight, and shoot length. The field experiment with four bacterial candidates demonstrated good performance against C. beticola compared to difenoconazole fungicide. These bacteria played a significant role in disease control, achieving a maximum efficacy of 77.42%, slightly below the 88.51% efficacy attained by difenoconazole. Additional field trials are necessary to verify the protective and growth-promoting effects of these candidates, whether applied individually, combined in consortia, or integrated with chemical inputs in sugar beet crop production.

In this study, a commercial sodium butyrate protected by a new buffer salt solution (NSB) was tested to determine whether it can be used as an antibiotic alternative in broiler production. 192 1-day-old broilers were randomly allocated to three dietary treatments: soybean meal diet (CON), antibiotic diet (ANT, basal diet + 100 mg/kg aureomycin), and NSB (basal diet + 800 mg/kg NSB). The growth performance, serum anti-inflammatory cytokines, intestinal morphology, gut barrier function, antioxidative parameters, SCFAs content, and cecal microbiota were analyzed. The result showed that NSB significantly improved ADFI, ADG (p < 0.01), and decreased FCR (p < 0.01). Serum anti-inflammatory cytokine IL-10 was up-regulated (p < 0.01), and pro-inflammatory TNF-α was down-regulated (p < 0.05) by NSB supplementation. H&E results showed that VH and VH: CD ratio significantly increased (p < 0.05) in the jejunum and ileum in the NSB group. Furthermore, ZO-1 (p < 0.01), claudin-1 (p < 0.01), and occludin (p < 0.05) in the jejunum and claudin-1 (p < 0.01) and mucin-2 (p < 0.05) in the ileum significantly up-regulated in the NSB group, respectively. Additionally, SOD (p < 0.05) and T-AOC/MDA ratio (p < 0.01) in the jejunum and SOD in the ileum were significantly increased (p < 0.05) in the NSB group. The MDA level also significantly increased (p < 0.01) in the ANT group in the jejunum. Propionic acid (p < 0.05) and butyric acid (p < 0.01) content significantly increased in the NSB group in the jejunum and ileum segments. The 16S rRNA sequencing results showed no significant difference (p > 0.05) in alpha and beta diversity among the groups. LEFSe analysis also indicated that Peptostreptococcaceae, Colidextribacter, Firmicutes, Oscillospira, and Erysipelatoclostridiaceae, which promote SCFA production (p < 0.05), were identified as dominant taxon-enriched bacterial genera in the NSB group. The Spearman correlation analysis revealed that Colidextribacter with ADFI, ADG, VH, claudin-1 (p < 0.05), and unclassified_f__Peptostreptococcaceae with ADFI, IL-10, and ZO-1 were positively correlated (p < 0.05). Furthermore, ADFI and ADG with IL-10, claudin-1, SOD, T-AOC, and butyric acid (p < 0.05), and similarly, ADG with VH (p < 0.05) showed a positive correlation. In conclusion, NSB enhanced the growth performance by improving jejunum and ileum morphology, serum anti-inflammatory cytokines, regulating intestinal barrier function and antioxidant capacity, SCFAs content, and cecum microbiota, showing its potential use as an alternative to antibiotics in poultry nutrition.

The castor plant (Ricinus communis L.) has been known since time immemorial in traditional medicine in the pharmacopeia of Mediterranean and eastern ancient cultures. Moreover, it is still used in folk medicine worldwide. Castor bean has been mainly recommended as anti-inflammatory, anthelmintic, anti-bacterial, laxative, abortifacient, for wounds, ulcers, and many other indications. Many cases of human intoxication occurred accidentally or voluntarily with the ingestion of castor seeds or derivatives. Ricinus toxicity depends on several molecules, among them the most important is ricin, a protein belonging to the family of ribosome-inactivating proteins. Ricin is the most studied of this category of proteins and it is also known to the general public, having been used for biocrimes in several cases. Here, the main steps of ricin research are reported with particular regards to its enzymatic activity, structure and cytotoxicity. Moreover, we discuss ricin toxicity for animals and humans, as well as the relation amongst bioterrorism and ricin and its impact on environmental toxicity. Ricin has also been of great utility to develop a number of immunotoxins specific for the elimination of unwanted cells, mainly cancer cells; some of these immunotoxins gave promising results also in clinical trials.

Epithelial ovarian cancer (EOC) has not significantly benefited from advances in immunotherapy, mainly because of the lack of well-defined actionable antigen targets. Using proteogenomic analyses of primary EOC tumors, we previously identified 91 aberrantly expressed tumor-specific antigens (TSAs) originating from unmutated genomic sequences. Most of these TSAs derive from non-exonic regions, and their expression results from cancer-specific epigenetic changes. The present study aimed to evaluate the immunogenicity of 48 TSAs selected according to two criteria: presentation by highly prevalent HLA allotypes and expression in a significant fraction of EOC tumors. Using targeted mass spectrometry analyses, we found that pulsing with synthetic TSA peptides leads to a high-level presentation on dendritic cells. TSA abundance correlated with the predicted binding affinity to the HLA allotype. We stimulated naïve CD8 T cells from healthy blood donors with TSA-pulsed dendritic cells and assessed their expansion with two assays: MHC-peptide tetramer staining and TCR Vβ CDR3 sequencing. We report that these TSAs can expand sizeable populations of CD8 T cells and, therefore, represent attractive targets for EOC immunotherapy.

The persistence of Salmonella and Campylobacter in chicken meat is a considerable public health risk and a future challenge. This study aimed to determine the prevalence of Salmonella and Campyl-obacter in poultry processing lines where different chlorination techniques were used. The sam-ples were collected from four types of processing plants considering the chlorine concentration used in the chill tank, which ranged from 2 ppm to 50 ppm. Salmonella and Campylobacter were isolated from carcass washings, neck skin, and, caecal samples. Subsequently, an antimicrobial susceptibility test was performed for the isolates. The results revealed the overall prevalence of Salmonella and Campylobacter was 78.25%, and 64% respectively. Positive percentages of Salmonella and Campylobacter were high in carcasses compared to neck skin and caeca. Carcass Campylobacter counts at higher chlorine concentrations ranging from 20 – 50 ppm was significantly low (p

Marine yeasts have versatile applications in industrial, medical and environmental fields, but have received little attention compared to terrestrial yeasts and filamentous fungi. In this study, a phylogenetic analysis of 11 marine-derived yeasts was conducted using internal transcribed spacers and nuclear large subunit rDNA, and their bioactivities, such as antioxidant, antibacterial, and tyrosinase inhibition activities, were investigated. The 11 marine-derived yeasts were identified to belong to seven species including Geotrichum candidum, Metschnikowia bicuspidata, Papiliotrema fonsecae, Rhodotorula mucilaginosa, Vishniacozyma carnescens, Yamadazyma olivae, and Yarrowia lipolytica, and three strains of these were candidates for new species of the genera Aureobasidium, Rhodotorula, and Vishniacozyma. Most extracts showed antioxidant activity, whereas seven strains exhibited antibacterial activities against Bacillus subtilis. Only Aureobasidium sp. US-Sd3 among the 11 isolates showed tyrosinase inhibition. Metschnikowia bicuspidata BP-Up1 and Yamadazyma olivae K2-6 showed notable radical-scavenging activity, which has not been reported previously. Among the isolates, Aureobasidium sp. US-Sd3 exhibited the highest antibacterial and tyrosinase inhibitory activities. Overall, our results demonstrate the potential of marine-derived yeasts as a source of bioactive compounds for improving industrial applications.

Hantavirus cardiopulmonary syndrome (HCPS) is characterized by capillary leak, pulmonary edema (PE) and shock that leads to death in up to 40% of patients. Treatment is supportive, including mechanical ventilation (MV) and extracorporeal membrane oxygenation (ECMO). Hemodynamic monitoring is critical to titrate therapy and to decide ECMO support. Transpulmonary thermodilution (TPTD) provides hemodynamic and PE data that has not been systematically used to understand HCPS pathophysiology. We identified 11 HCPS patients monitored with TPTD; 5 on MV, 3 on ECMO. We analyzed 133 measurements to describe the hemodynamic pattern and its association to PE. The main findings were reduced stroke volume, global ejection fraction (GEF) and preload parameters associated to increased extravascular lung water and pulmonary vascular permeability compatible with hypovolemia, myocardial dysfunction and increased permeability PE. Lung water correlated positively with heart rate (HR, r=0.20) and negatively with mean arterial pressure (r=-0.27) and GEF (r=-0.36), suggesting that PE is linked to hemodynamic impairment. Pulmonary vascular permeability correlated positively with HR (r=0.31) and negatively with cardiac index (r=-0.49), end-diastolic volume (r=-0.48) and GEF (r=-0.40), suggesting that capillary leak contributes to hypovolemia and systolic dysfunction. In conclusion, TPTD data suggests that in HCPS patients, increased permeability leads to PE, hypovolemia and circulatory impairment.

Correlations between the air permeability coefficient and various pore structure indicators in cementitious materials were examined to determine the pore structure indicator that best evaluated air permeability using data from previous studies of air permeabilities and pore structures. The determination coefficients of air permeability with total pore volume, critical pore diameter, and ordinary threshold pore diameter were low, although these have often been used as indicators. The median and threshold pore diameters obtained by percolation theory showed high determination coefficients. The equation using the threshold pore diameter better estimated the air permeability coefficient than the Katz–Thompson equation.

To understand the relationship between water permeability and the pore structure of cementitious materials, literature that studied both water permeability and the pore structure are reviewed, and their correlation is studied. Although the majority of this data is for cement paste, mortar, concrete, and cemented soil are also included. Based on this study, it is established the determination coefficient between water permeability and the total pore volume is very low; however, those between water permeability and the critical, threshold, and median pore diameters, respectively, are moderate. On the other hand, the threshold pore diameter, derived based on the percolation theory, exhibits a very high determination coefficient. The calculated water permeability, using the Katz and Thompson equation, agrees with the reported water permeability quantitatively to a certain level, but is overestimated.

Microplastics (plastic particles &lt; 5 mm) are ubiquitous pollutants that pose the ability to carry microbiota, including pathogens. Microbial adhesion is usually a sign of pathogenicity, thus we investigated the adherent strains found on 4 mm nylon strips, which were ingested and excreted by wild fish specimens. Polymer analysis of the excreted samples shown no signs of degradation, nor have their controls, represented by the nylon strips submerged in the same water tanks. Both the ingested samples and controls presented pathogens in large numbers, revealing the fact that nylon microplastics can be physical carriers for pathogens in the aquatic environment.

The nuclear factor-ĸB (NF-ĸB) transcriptional system is a major effector pathway involved in inflammatory responses. Previous studies found that a Gardenia decoction (GD) inhibited the expression of NF-κB in a lipopolysaccharide (LPS)-stimulated mouse intestinal injury model. Herein, we hypothesized that geniposide (GE), a component of Gardenia jasminoides Ellis, also exerts anti-inflammatory effects and inhibits NF-ĸB activity in LPS-induced intestinal epithelial cells (IEC-6). IEC-6 cells were stimulated with LPS, following which the effects of GE on NF-ĸB signaling in the IEC-6 cells were examined by western blotting to detect IĸB phosphorylation/degradation. The expression of NF-κB was determined by immunofluorescence assay (IFA). Enzyme-linked immunosorbent assay (ELISA) was used to detect the inhibitory effect of GE on the release of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-1β (IL-1β) activated by LPS in IEC-6 cells. In addition, the migration ability of IEC-6 cells was observed by the scratch method. These results showed that GE dose-dependently downregulated levels of the proinflammatory cytokines TNF-α, IL-6 and IL-1β that had been upregulated by LPS and suppressed the phosphorylation of IĸB and NF-ĸB induced by LPS. Our findings indicated that GE could reduce LPS-induced NF-ĸB signaling and proinflammatory expression in IEC-6 cells and significantly enhance the migration of IEC-6 cells. Moreover, GE inhibited the expression of NF-κB, nuclear transfer, and transcriptional activity in IEC-6 cells. GE could block the synthesis of inflammatory factors of IEC-6 cells by inhibiting activation of the IĸB/NF-κB signaling pathway induced by LPS.

In recent experimental studies, we reported the aggravating effects of Asian sand dust (ASD) on male reproductive function in mice. However, the effects of fetal ASD exposure on male reproductive function have not been investigated. The present study investigated the effects of fetal ASD exposure on reproductive function in male offspring. Using pregnant CD-1 mice, ASD was administered intratracheally on days 7 and 14 of gestation, and the reproductive function of male offspring was determined at 5, 10, and 15 weeks after birth. The secondary sex ratio was significantly lower in the fetal ASD-exposed mice than in the controls. Histologic examination showed partial vacuolation of seminiferous tubules in immature mice. Moreover, daily sperm production (DSP) was significantly less in the fetal ASD-exposed mice than in the controls. DSP in the fetal ASD-exposed mice was approximately 10% less than the controls at both 5 and 10 weeks. However, both the histologic changes and the DSP decrease were reversed as the mice matured. These findings suggest that ASD exposure affects both the fetal development and the reproductive function of male offspring. In the future, it will be necessary to clarify the onset mechanisms of ASD-induced male fetus death and male reproductive disorders.

Necroptosis is a caspases-independent form of programmed cell death exhibiting intermediate features between necrosis and apoptosis. Albeit some physiological roles during embryonic development, tissue homeostasis and innate immune response are documented, necroptosis is mainly considered a pro-inflammatory cell death. Key actors of necroptosis are the receptor-interacting-protein-kinases, RIPK1 and RIPK3, and their target, the mixed-lineage-kinase-domain-like protein, MLKL. The intestinal epithelium has one of the highest rates of cellular turnover in a process that is tightly regulated. Altered necroptosis at the intestinal epithelium leads to uncontrolled microbial translocation and deleterious inflammation. Indeed, necroptosis has been associated to chronic inflammatory diseases and cancer. Drugs that inhibit necroptosis could, therefore, be used therapeutically for the treatment of these diseases, and researches to develop such inhibitors are already underway. In this Review, we outline pathways for necroptosis and its role in chronic inflammation and cancer. We also discuss current and developing therapies that target necroptosis machinery.

Pattern recognition receptors such as nucleotide-binding oligomerization domain (NOD)-containing protein receptors (NLRs) and the pyrin and HIN domain (PYHIN) receptors initiate the inflammatory response following cell stress or pathogenic challenge. When activated some of these receptors oligomerize to form the structural backbone of a signalling platform known as the inflammasome. The inflammasome promotes the activation of caspase-1 and the maturation of the proinflammatory cytokines, interleukin (IL)-1β and IL-18. In the gut dysregulation of the inflammasome complex is thought to be a contributing factor in the development of inflammatory bowel diseases (IBD), such as ulcerative colitis (UC) and Crohn’s disease (CD). The importance of inflammasomes to intestinal health has been emphasized by various inflammasome deficient mice in dextran sulphate sodium (DSS) models of intestinal inflammation and by the identification of novel potential candidate genes in population based human studies. In this review we summarise the most recent finding with regard to formation, sensing and regulation of the inflammasome complex and highlight their importance in maintaining intestinal health.