Evolutionary Biology by Siegfried Scherer
Background: Gene duplication is believed to be the classical way to form novel genes, but overpri... more Background: Gene duplication is believed to be the classical way to form novel genes, but overprinting may be an important alternative. Overprinting allows entirely novel proteins to evolve de novo, i.e., formerly non-coding open reading frames within functional genes become expressed. Only three cases have been described for Escherichia coli. Here, a fourth example is presented.
Results: RNA sequencing revealed an open reading frame weakly transcribed in cow dung, coding for 101 residues and embedded completely in the −2 reading frame of citC in enterohemorrhagic E. coli. This gene is designated novel overlapping gene, nog1. The promoter region fused to gfp exhibits specific activities and 5’ rapid
amplification of cDNA ends indicated the transcriptional start 40-bp upstream of the start codon. nog1 was strandspecifically arrested in translation by a nonsense mutation silent in citC. This Nog1-mutant showed a phenotype in competitive growth against wild type in the presence of MgCl2. Small differences in metabolite concentrations were also found. Bioinformatic analyses propose Nog1 to be inner membrane-bound and to possess at least one membrane-spanning domain. A phylogenetic analysis suggests that the orphan gene nog1 arose by overprinting
after Escherichia/Shigella separated from the other γ-proteobacteria.
Conclusions: Since nog1 is of recent origin, non-essential, short, weakly expressed and only marginally involved in E. coli’s central metabolism, we propose that this gene is in an initial stage of evolution. While we present specific experimental evidence for the existence of a fourth overlapping gene in enterohemorrhagic E. coli, we believe that this may be an initial finding only and overlapping genes in bacteria may be more common than is currently assumed by microbiologists.
Keywords: Overprinting, Overlapping gene, de novo evolution, Coding reserve, Orphan, EHEC, nog1/citC
Microbial Ecology by Siegfried Scherer
Microbial taxonomy by Siegfried Scherer
Food Microbiology by Siegfried Scherer
Papers by Siegfried Scherer
Das Endosporen bildende Bakterium Bacillus cereus wird in der Milchindustrie und weiteren Zweigen... more Das Endosporen bildende Bakterium Bacillus cereus wird in der Milchindustrie und weiteren Zweigen der Lebensmittelindustrie zunehmend als ein ernstzunehmendes Problem erkannt. Einige Stämme produzieren Toxine, die gastrointestinale Erkrankungen hervorrufen. Das Toxizitätspotential reicht von Stämmen, die als Probiotika Futtermitteln zugesetzt werden, bis hin zu stark toxischen Stämmen, die bereits für Todesfälle verantwortlich waren [1].
Frontiers in Immunology, 2021
Antibiotic-resistant bacterial pathogens have become a serious threat worldwide. One of these pat... more Antibiotic-resistant bacterial pathogens have become a serious threat worldwide. One of these pathogens is methicillin-resistant Staphylococcus aureus (MRSA), a major cause of skin and soft tissue infections. In this study we identified a strain of Staphylococcus equorum producing a substance with high antimicrobial activity against many Gram-positive bacteria, including MRSA. By mass spectrometry and whole genome sequencing the antimicrobial substance was identified as the thiopeptide bacteriocin micrococcin P1 (MP1). Based on its properties we developed a one-step purification protocol resulting in high yield (15 mg/L) and high purity (98%) of MP1. For shorter incubation times (5-7 h) MP1 was very potent against MRSA but the inhibitory effect was overshadowed by resistance development during longer incubation time (24h or more). To overcome this problem a synergy study was performed with a number of commercially available antibiotics. Among the antibiotics tested, the combination ...
International Journal of Systematic and Evolutionary Microbiology, 2021
Two strains of a Gram-staining-positive species were isolated from German bulk tank milk. On the ... more Two strains of a Gram-staining-positive species were isolated from German bulk tank milk. On the basis of their 16S rRNA sequences they were affiliated to the genus Facklamia but could not be assigned to any species with a validly published name. Facklamia miroungae ATCC BAA-466T (97.3 % 16S rRNA sequence similarity), Facklamia languida CCUG 37842T (96.9 %), and Facklamia hominis CCUG 36813T (96.6 %) are the closest relatives. In the 16S rRNA phylogeny and in the core-genome phylogeny strains WS 5301T and WS 5302 form a well-supported, separate lineage. Pairwise average nucleotide identity calculated using MUMmer (ANIm) between WS 5301T and type strains of other Facklamia species is well below the species cut-off (95 %) and ranges from 83.4 to 87.7 %. The DNA G+C content of the type strain is 36.4 mol% and the assembly size of the genome is 2.2 Mb. Cells of WS 5301T are non-motile, non-endospore-forming, oxidase-negative, catalase-negative and facultatively anaerobic cocci. The fast...
International Journal of Systematic and Evolutionary Microbiology, 2020
Two strains, WS 5063T and WS 5067, isolated from raw cow’s milk and skimmed milk concentrate, cou... more Two strains, WS 5063T and WS 5067, isolated from raw cow’s milk and skimmed milk concentrate, could be affiliated as members of the same, hitherto unknown, Pseudomonas species by 16S rRNA and rpoD gene sequences. Multilocus sequence and average nucleotide identity (ANIm) analyses based on draft genome sequences confirmed the discovery of a novel Pseudomonas species. It was most closely related to Pseudomonas synxantha DSM 18928T with an ANIm of 91.4 %. The DNA G+C content of WS 5063T was 60.0 mol %. Phenotypic characterizations showed that the isolates are rod-shaped, motile, catalase- and oxidase-positive, and aerobic. Growth occurred at 4–34 °C and at pH values of pH 5.5–8.0. Both strains showed strong β-haemolysis on blood agar. The major cellular polar lipids were phosphatidylethanolamine, phosphatidylglycerol and diphosphatidylglycerol. The dominant quinone was Q-9 (90 %), but noticeable amounts of Q-8 (9 %) and traces of Q-7 were also detected. Fatty acid profiles were typical...
Angewandte Chemie International Edition, 2018
Zeitschrift für Naturforschung C, 2016
During a 1-year longitudinal study, water, sediment and water plants from two creeks and one pond... more During a 1-year longitudinal study, water, sediment and water plants from two creeks and one pond were sampled monthly and analyzed for the presence ofListeriaspecies. A total of 90 % of 30 sediment samples, 84 % of 31 water plant samples and 67 % of 36 water samples were tested positive. Generally, most probable number counts ranged between 1 and 40 g−1, only occasionally >110 cfu g−1were detected. Species differentiation based on FT-IR spectroscopy and multiplex PCR of a total of 1220 isolates revealedL. innocua(46 %), L. seeligeri(27 %),L. monocytogenes(25 %) andL. ivanovii(2 %). Titers and species compositions were similar during all seasons. While the species distributions in sediments and associatedRanunculus fluitansplants appeared to be similar in both creeks, RAPD typing did not provide conclusive evidence that the populations of these environments were connected. It is concluded that (i) the fresh-water sediments and water plants are year-round populated byListeria, (ii...
Genome Announcements, 2016
Escherichia coli O157:H7 EDL933, isolated in 1982 in the United States, was the first enterohemor... more Escherichia coli O157:H7 EDL933, isolated in 1982 in the United States, was the first enterohemorrhagic E. coli (EHEC) strain sequenced. Unfortunately, European labs can no longer receive the original strain. We checked three European EDL933 derivatives and found major genetic deviations (deletions, inversions) in two strains. All EDL933 strains contain the cryptic EHEC-plasmid, not reported before.
Toxins, Jan 9, 2015
The ability of Bacillus cereus to cause foodborne toxicoinfections leads to increasing concerns r... more The ability of Bacillus cereus to cause foodborne toxicoinfections leads to increasing concerns regarding consumer protection. For the diarrhea-associated enterotoxins, the assessment of the non-hemolytic enterotoxin B (NheB) titer determined by a sandwich enzyme immunoassay (EIA) correlates best with in vitro cytotoxicity. In general, the regulation of enterotoxin expression of B. cereus is a coordinately-regulated process influenced by environmental, and probably also by host factors. As long as these factors are not completely understood, the currently-applied diagnostic procedures are based on indirect approaches to assess the potential virulence of an isolate. To date, sandwich EIA results serve as a surrogate marker to categorize isolates as either potentially low or highly toxic. Here, we report on a single amino acid exchange in the NheB sequence leading to an underestimation of the cytotoxic potential in a limited number of strains. During the screening of a large panel of ...
BMC Evolutionary Biology, 2015
Background Gene duplication is believed to be the classical way to form novel genes, but overprin... more Background Gene duplication is believed to be the classical way to form novel genes, but overprinting may be an important alternative. Overprinting allows entirely novel proteins to evolve de novo, i.e., formerly non-coding open reading frames within functional genes become expressed. Only three cases have been described for Escherichia coli. Here, a fourth example is presented. Results RNA sequencing revealed an open reading frame weakly transcribed in cow dung, coding for 101 residues and embedded completely in the −2 reading frame of citC in enterohemorrhagic E. coli. This gene is designated novel overlapping gene, nog1. The promoter region fused to gfp exhibits specific activities and 5’ rapid amplification of cDNA ends indicated the transcriptional start 40-bp upstream of the start codon. nog1 was strand-specifically arrested in translation by a nonsense mutation silent in citC. This Nog1-mutant showed a phenotype in competitive growth against wild type in the presence of MgCl2...
Applied and Environmental Microbiology, Oct 1, 2002
Food Analytical Methods, 2015
Scientific reports, Jan 27, 2015
Cereulide and isocereulides A-G are biosynthesized as emetic toxins by Bacillus cereus via a non-... more Cereulide and isocereulides A-G are biosynthesized as emetic toxins by Bacillus cereus via a non-ribosomal peptide synthetase (NRPS) called Ces. Although a thiotemplate mechanisms involving cyclo-trimerization of ready-made D-O-Leu-D-Ala-L-O-Val-L-Val via a thioesterase (TE) domain is proposed for cereulide biosynthesis, the exact mechanism is far from being understood. UPLC-TOF MS analysis of B. cereus strains in combination with (13)C-labeling experiments now revealed tetra-, octa-, and dodecapeptides of a different sequence, namely (L-O-Val-L-Val-D-O-Leu-D-Ala)1-3, as intermediates of cereulide biosynthesis. Surprisingly, also di-, hexa-, and decadepsipeptides were identified which, together with the structures of the previously reported isocereulides E, F, and G, do not correlate to the currently proposed mechanism for cereulide biosynthesis and violate the canonical NRPS biosynthetic logic. UPLC-TOF MS metabolite analysis and bioinformatic gene cluster analysis highlighted dipe...
Microbiology, 2009
Cereulide, a depsipeptide structurally related to the antibiotic valinomycin, is responsible for ... more Cereulide, a depsipeptide structurally related to the antibiotic valinomycin, is responsible for the emetic type of gastrointestinal disease caused by Bacillus cereus. Recently, it has been shown that cereulide is produced non-ribosomally by the plasmid-encoded peptide synthetase Ces. Using deletion mutants of the emetic reference strain B. cereus F4810/72, the influence of the well-known transcription factors PlcR, Spo0A and AbrB on cereulide production and on the transcription of the cereulide synthetase gene cluster was investigated. Our data demonstrate that cereulide synthesis is independent of the B. cereus specific virulence regulator PlcR but belongs to the Spo0A-AbrB regulon. Although cereulide production turned out to be independent of sporulation, it required the activity of the sporulation factor Spo0A. The σ A-promoted transcription of spo0A was found to be crucial for cereulide production, while the σ H-driven transcription of spo0A did not affect cereulide synthesis. ...
Journal of Biological Chemistry, 1995
Journal of Applied Microbiology, 2006
Applied and Environmental Microbiology, 2005
The microbial composition of smear-ripened cheeses is not very clear. A total of 194 bacterial is... more The microbial composition of smear-ripened cheeses is not very clear. A total of 194 bacterial isolates and 187 yeast isolates from the surfaces of four Irish farmhouse smear-ripened cheeses were identified at the midpoint of ripening using pulsed-field gel electrophoresis (PFGE), repetitive sequence-based PCR, and 16S rRNA gene sequencing for identifying and typing the bacteria and Fourier transform infrared spectroscopy and mitochondrial DNA restriction fragment length polymorphism (mtDNA RFLP) analysis for identifying and typing the yeast. The yeast microflora was very uniform, and Debaryomyces hansenii was the dominant species in the four cheeses. Yarrowia lipolytica was also isolated in low numbers from one cheese. The bacteria were highly diverse, and 14 different species, Corynebacterium casei , Corynebacterium variabile , Arthrobacter arilaitensis , Arthrobacter sp., Microbacterium gubbeenense , Agrococcus sp. nov., Brevibacterium linens , Staphylococcus epidermidis , Staphy...
Journal of Bacteriology, 1999
Inverse PCR was used to amplify major cold shock protein (MCSP) gene families from a diverse rang... more Inverse PCR was used to amplify major cold shock protein (MCSP) gene families from a diverse range of bacteria, including the psychrotolerant Yersinia enterocolitica , which was found to have two almost identical MCSP coding regions ( cspA1 and cspA2 ) located approximately 300 bp apart. This tandem gene duplication was also found in Y. pestis , Y. pseudotuberculosis , and Y. ruckeri but not in other bacteria. Analysis of the transcriptional regulation of this MCSP gene in Y. enterocolitica , performed by using both reverse transcriptase-PCR and Northern blot assays, showed there to be two cold-inducible mRNA templates arising from this locus: a monocistronic template of approximately 450 bp ( cspA1 ) and a bicistronic template of approximately 900 bp ( cspA1/A2 ). The former may be due to a secondary structure between cspA1 and cspA2 causing either 3′ degradation protection of cspA1 or, more probably, partial termination after cspA1 . Primer extension experiments identified a putat...
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Evolutionary Biology by Siegfried Scherer
Results: RNA sequencing revealed an open reading frame weakly transcribed in cow dung, coding for 101 residues and embedded completely in the −2 reading frame of citC in enterohemorrhagic E. coli. This gene is designated novel overlapping gene, nog1. The promoter region fused to gfp exhibits specific activities and 5’ rapid
amplification of cDNA ends indicated the transcriptional start 40-bp upstream of the start codon. nog1 was strandspecifically arrested in translation by a nonsense mutation silent in citC. This Nog1-mutant showed a phenotype in competitive growth against wild type in the presence of MgCl2. Small differences in metabolite concentrations were also found. Bioinformatic analyses propose Nog1 to be inner membrane-bound and to possess at least one membrane-spanning domain. A phylogenetic analysis suggests that the orphan gene nog1 arose by overprinting
after Escherichia/Shigella separated from the other γ-proteobacteria.
Conclusions: Since nog1 is of recent origin, non-essential, short, weakly expressed and only marginally involved in E. coli’s central metabolism, we propose that this gene is in an initial stage of evolution. While we present specific experimental evidence for the existence of a fourth overlapping gene in enterohemorrhagic E. coli, we believe that this may be an initial finding only and overlapping genes in bacteria may be more common than is currently assumed by microbiologists.
Keywords: Overprinting, Overlapping gene, de novo evolution, Coding reserve, Orphan, EHEC, nog1/citC
Microbial Ecology by Siegfried Scherer
Microbial taxonomy by Siegfried Scherer
Food Microbiology by Siegfried Scherer
Papers by Siegfried Scherer
Results: RNA sequencing revealed an open reading frame weakly transcribed in cow dung, coding for 101 residues and embedded completely in the −2 reading frame of citC in enterohemorrhagic E. coli. This gene is designated novel overlapping gene, nog1. The promoter region fused to gfp exhibits specific activities and 5’ rapid
amplification of cDNA ends indicated the transcriptional start 40-bp upstream of the start codon. nog1 was strandspecifically arrested in translation by a nonsense mutation silent in citC. This Nog1-mutant showed a phenotype in competitive growth against wild type in the presence of MgCl2. Small differences in metabolite concentrations were also found. Bioinformatic analyses propose Nog1 to be inner membrane-bound and to possess at least one membrane-spanning domain. A phylogenetic analysis suggests that the orphan gene nog1 arose by overprinting
after Escherichia/Shigella separated from the other γ-proteobacteria.
Conclusions: Since nog1 is of recent origin, non-essential, short, weakly expressed and only marginally involved in E. coli’s central metabolism, we propose that this gene is in an initial stage of evolution. While we present specific experimental evidence for the existence of a fourth overlapping gene in enterohemorrhagic E. coli, we believe that this may be an initial finding only and overlapping genes in bacteria may be more common than is currently assumed by microbiologists.
Keywords: Overprinting, Overlapping gene, de novo evolution, Coding reserve, Orphan, EHEC, nog1/citC