Publications by authors named "Helmut Hotzel"

110 Publications

Characterisation of S. aureus/MRSA CC1153 and review of mobile genetic elements carrying the fusidic acid resistance gene fusC.

Sci Rep 2021 Apr 14;11(1):8128. Epub 2021 Apr 14.

Leibniz Institute of Photonic Technology (IPHT), Jena, Germany.

While many data on molecular epidemiology of MRSA are available for North America, Western Europe and Australia, much less is known on the distribution of MRSA clones elsewhere. Here, we describe a poorly known lineage from the Middle East, CC1153, to which several strains from humans and livestock belong. Isolates were characterised using DNA microarrays and one isolate from the United Arab Emirates was sequenced using Nanopore technology. CC1153 carries agr II and capsule type 5 genes. Enterotoxin genes are rarely present, but PVL is common. Associated spa types include t504, t903 and t13507. PVL-positive CC1153-MSSA were found in Egyptian cattle suffering from mastitis. It was also identified among humans with skin and soft tissue infections in Saudi Arabia, France and Germany. CC1153-MRSA were mainly observed in Arabian Gulf countries. Some isolates presented with a previously unknown SCCmec/SCCfus chimeric element in which a mec B complex was found together with the fusidic acid resistance gene fusC and accompanying genes including ccrA/B-1 recombinase genes. Other isolates carried SCCmec V elements that usually also included fusC. Distribution and emergence of CC1153-MRSA show the necessity of molecular characterization of MRSA that are resistant to fusidic acid. These strains pose a public health threat as they combine resistance to beta-lactams used in hospitals as well as to fusidic acid used in the community. Because of the high prevalence of fusC-positive MRSA in the Middle East, sequences and descriptions of SCC elements harbouring fusC and/or mecA are reviewed. When comparing fusC and its surrounding regions from the CC1153 strain to available published sequences, it became obvious that there are four fusC alleles and five distinct types of fusC gene complexes reminiscent to the mec complexes in SCCmec elements. Likewise, they are associated with different sets of ccrA/B recombinase genes and additional payload that might include entire mec complexes or SCCmec elements.
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http://dx.doi.org/10.1038/s41598-021-86273-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8046974PMC
April 2021

Antimicrobial Susceptibility and Genomic Analysis of and , Two Rarely Detected Species.

Front Cell Infect Microbiol 2021 17;11:532989. Epub 2021 Mar 17.

IBIZ, Friedrich-Loeffler-Institut Jena, Jena, Germany.

and are two rarely detected species. In the study, we analyzed the antimicrobial susceptibility and provide detailed insights into the genotype and phylogeny of both species using whole-genome sequencing. Thermophilic species are the most common bacterial foodborne pathogens causing gastroenteritis in humans worldwide. The genus is part of the family and includes the species , and the rarely described , and are emergent enteropathogens and potential zoonotic agents. Here, we generated, analyzed, and characterized whole-genome sequences of and They were isolated from water poultry farms in Germany, cultured and identified by MALDI-TOF MS. With PCR the identity was verified. Antibiotic susceptibility testing was carried out with erythromycin, ciprofloxacin, doxycycline, tetracycline, gentamicin, streptomycin, ampicillin, and cefotaxime using the gradient strip method (E-test). Whole-genome sequences were generated including those of reference strains. Complete genomes for six selected strains are reported. These provide detailed insights into the genotype. With these, we predicted known AMR genes, virulence-associated genes, and plasmid replicons. Phenotypic analysis of resistance showed differences between the presence of resistance genes and the prediction of phenotypic resistance profiles. In , the nucleotide sequence of the A gene (DQ464331) can show a signature mutation resulting in an amino acid change T85>I. and showed the same gene as assessed by similarity annotation of the mutations 254C>G. Most of the isolates were found to be sensitive to ciprofloxacin. The ciprofloxacin-resistant isolate was associated with the amino acid change T85>I. But this was not predicted with antibiotic resistance databases, before. Ultimately, a phylogenetic analysis was done to facilitate in future outbreak analysis.
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http://dx.doi.org/10.3389/fcimb.2021.532989DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8010192PMC
March 2021

Newly Isolated Animal Pathogen Is Cytotoxic to Human Epithelial Cells.

Int J Mol Sci 2021 Mar 29;22(7). Epub 2021 Mar 29.

Microbiology Division, Department of Biology, Friedrich-Alexander-Universität Erlangen-Nürnberg, 91054 Erlangen, Germany.

is a newly identified animal pathogen of forest animals such as roe deer and wild boars. The species is closely related to the emerging human pathogen and the widely distributed animal pathogen . In this study, strain W25 was characterized with respect to its interaction with human cell lines. Microscopy, measurement of transepithelial electric resistance and cytotoxicity assays revealed detrimental effects of to different human epithelial cell lines and to an invertebrate animal model, larvae, comparable to diphtheria toxin-secreting Furthermore, the results obtained may indicate a considerable zoonotic potential of this newly identified species.
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http://dx.doi.org/10.3390/ijms22073549DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8037504PMC
March 2021

Characterization of Enterococci- and ESBL-Producing Isolated from Milk of Bovides with Mastitis in Egypt.

Pathogens 2021 Jan 21;10(2). Epub 2021 Jan 21.

Friedrich-Loeffler-Institut, Institute of Bacterial Infections and Zoonoses, Naumburger Str. 96a, 07743 Jena, Germany.

This study aimed to investigate the prevalence and antimicrobial resistance of enterococci- and ESBL-producing isolated from milk of bovine mastitis cases in Egypt. Fifty milk samples of dairy animals were collected from localities in the Nile Delta region of Egypt. Isolates were identified using MALDI-TOF MS, and antibiotic susceptibility testing was performed by the broth microdilution method. PCR amplifications were carried out, targeting resistance-associated genes. Seventeen isolates and eight coliform isolates could be cultivated. Vancomycin resistance rate was high in The VITEK 2 system confirmed all isolates as ESBL-producing. All . isolates harbored (B), L and D genes. The A gene was detected in isolate, B was found in other while one isolate of exhibited the A gene. isolates exhibited high prevalence of erm(B) and tetL. isolates were analyzed by DNA microarray analysis. Four isolates were determined by O-serotyping as O8 (n = 1), O86 (n = 2) and O157 (n = 1). H-serotyping resulted in H11, H12, H21 (two isolates each) and one was of H16 type. Different virulence-associated genes were detected in isolates including A, A, B, L, I1 and I2, and the N gene was identified by DNA microarray analysis.
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http://dx.doi.org/10.3390/pathogens10020097DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7909756PMC
January 2021

Antimicrobial Resistance and Virulence Profiling of Strains From German Water Poultry.

Front Microbiol 2020 14;11:617685. Epub 2020 Dec 14.

Institute of Bacterial Infections and Zoonoses (IBIZ), Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Jena, Germany.

is an emerging foodborne and zoonotic pathogen that is usually transmitted via contaminated food or water. is not only the most prevalent species, it is also closely related to thermophilic , which have shown increasing resistance in recent years. Therefore, it is important to assess its resistance and virulence profiles. In this study, 45 strains from water poultry farms in Thuringia, Germany, were subjected to an antimicrobial susceptibility test using the gradient strip diffusion method and whole-genome sequencing. In the phylogenetic analysis, the genomes of the German strains showed high genetic diversity. Thirty-three isolates formed 11 subgroups containing two to six strains. The antimicrobial susceptibility testing showed that 32 strains were resistant to erythromycin, 26 to doxycycline, and 20 to tetracycline, respectively. Only two strains were resistant to ciprofloxacin, while 39 strains were resistant to streptomycin. The prediction of the antimicrobial resistance profiles identified a large repertoire of potential resistance mechanisms. A strong correlation between a A point mutation (Thr-85-Ile) and ciprofloxacin resistance was found in 11 strains. A partial correlation was observed between the presence of the 3 gene and ampicillin resistance. virulence profiling revealed a broad spectrum of putative virulence factors, including a complete lipid A cluster in all studied genomes.
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http://dx.doi.org/10.3389/fmicb.2020.617685DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7767855PMC
December 2020

Phylogenomic Analysis of Reveals a Clonal Structure of Insertion Element IS Positive Genomes.

Front Microbiol 2020 12;11:585374. Epub 2020 Nov 12.

Institute of Bacterial Infections and Zoonoses, Friedrich-Loeffler-Institut, Jena, Germany.

Subspecies of the species are associated with specific host niches including mammals and reptiles. subsp. is a zoonotic pathogen infecting humans. Infections can vary from an acute intestinal illness to severe systemic infections, with sheep and cattle as major reservoirs. In contrast, subsp. causes bovine genital campylobacteriosis, which leads to abortion in cattle and a high economic burden for the farmers. Therefore, high-quality molecular subtyping is indispensable for interventional epidemiology. We used whole-genome sequencing (WGS) data of 283 strains from 18 countries and compared several methods for subtyping, including WGS, multilocus sequence typing, PCR assays, and the presence of the insertion element IS. We identified a highly clonal clade (designated as clade 1) that harbors the insertion sequence IS. The presence of this insertion sequence is an essential diagnostic tool for the identification of the subspecies subsp. , serving as a target for several PCR assays. However, we have found a high sequence variability for the IS besides the presence of IS-paralogues in certain other genomes ( = 7) which may cause incorrect diagnostic results. Clade 1 seems to be the cattle-specific clade of this species. We propose that only this clade might be designated as subsp. as it harbors the IS marker sequence, which is a major target for molecular methods currently used for subspecies identification. Fostering this proposal, we defined eleven stable nucleotide markers specific for this clade. Additionally, we developed a bioinformatics toolbox for the fast identification of this clade based on WGS data. In conclusion, our results demonstrate that WGS can be used for subtyping overcoming limitations of current PCR and MLST protocols.
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http://dx.doi.org/10.3389/fmicb.2020.585374DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7688749PMC
November 2020

Molecular investigations on a chimeric strain of Staphylococcus aureus sequence type 80.

PLoS One 2020 14;15(10):e0232071. Epub 2020 Oct 14.

Institute for Medical Microbiology and Hygiene, Technical University of Dresden, Dresden, Germany.

A PVL-positive, methicillin-susceptible Staphylococcus aureus was cultured from pus from cervical lymphadenitis of a patient of East-African origin. Microarray hybridisation assigned the isolate to clonal complex (CC) 80 but revealed unusual features, including the presence of the ORF-CM14 enterotoxin homologue and of an ACME-III element as well as the absence of etD and edinB. The isolate was subjected to both, Illumina and Nanopore sequencing allowing characterisation of deviating regions within the strain´s genome. Atypical features of this strain were attributable to the presence of two genomic regions that originated from other S. aureus lineages and that comprised, respectively, 3% and 1.4% of the genome. One deviating region extended from walJ to sirB. It comprised ORF-CM14 and the ACME-III element. A homologous but larger fragment was also found in an atypical S. aureus CC1/ST567 strain whose lineage might have served as donor of this genomic region. This region itself is a chimera comprising fragments from CC1 as well as fragments of unknown origin. The other deviating region comprised the region from htsB to ecfA2, i.e., another 3% of the genome. It was very similar to CC1 sequences. Either this suggests an incorporation of CC1 DNA into the study strain, or alternatively a recombination event affecting "canonical" CC80. Thus, the study strain bears witness of several recombination events affecting supposedly core genomic genes. Although the exact mechanism is not yet clear, such chimerism seems to be an additional pathway in the evolution of S. aureus. This could facilitate also a transmission of virulence and resistance factors and therefore offer an additional evolutionary advantage.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0232071PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7556507PMC
November 2020

Using affinity propagation clustering for identifying bacterial clades and subclades with whole-genome sequences of Francisella tularensis.

PLoS Negl Trop Dis 2020 09 29;14(9):e0008018. Epub 2020 Sep 29.

Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Institute of Bacterial Infections and Zoonoses, Friedrich-Loeffler-Institut, Jena, Germany.

By combining a reference-independent SNP analysis and average nucleotide identity (ANI) with affinity propagation clustering (APC), we developed a significantly improved methodology allowing resolving phylogenetic relationships, based on objective criteria. These bioinformatics tools can be used as a general ruler to determine phylogenetic relationships and clustering of bacteria, exemplary done with Francisella (F.) tularensis. Molecular epidemiology of F. tularensis is currently assessed mostly based on laboratory methods and molecular analysis. The high evolutionary stability and the clonal nature makes Francisella ideal for subtyping with single nucleotide polymorphisms (SNPs). Sequencing and real-time PCR can be used to validate the SNP analysis. We investigate whole-genome sequences of 155 F. tularensis subsp. holarctica isolates. Phylogenetic testing was based on SNPs and average nucleotide identity (ANI) as reference independent, alignment-free methods taking small-scale and large-scale differences within the genomes into account. Especially the whole genome SNP analysis with kSNP3.0 allowed deciphering quite subtle signals of systematic differences in molecular variation. Affinity propagation clustering (APC) resulted in three clusters showing the known clades B.4, B.6, and B.12. These data correlated with the results of real-time PCR assays targeting canSNPs loci. Additionally, we detected two subtle sub-clusters. SplitsTree was used with standard-setting using the aligned SNPs from Parsnps. Together APC, HierBAPS, and SplitsTree enabled us to generate hypotheses about epidemiologic relationships between bacterial clusters and describing the distribution of isolates. Our data indicate that the choice of the typing technique can increase our understanding of the pathogenesis and transmission of diseases with the eventual for prevention. This is opening perspectives to be applied to other bacterial species. The data provide evidence that Germany might be the collision zone where the clade B.12, also known as the East European clade, overlaps with the clade B.6, also known as the Iberian clade. Described methods allow generating a new, more detailed perspective for F. tularensis subsp. holarctica phylogeny. These results may encourage to determine phylogenetic relationships and clustering of other bacteria the same way.
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http://dx.doi.org/10.1371/journal.pntd.0008018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7523947PMC
September 2020

from Water Poultry: Insights into Antimicrobial Resistance, Virulence and Heavy Metal Resistance.

Genes (Basel) 2020 09 21;11(9). Epub 2020 Sep 21.

Institute of Bacterial Infections and Zoonoses (IBIZ), Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, 07743 Jena, Germany.

is the most prevalent species and has been isolated from a wide variety of sources. This species is an emerging foodborne and zoonotic pathogen because the bacteria can be transmitted by contaminated food or water and can cause acute enteritis in humans. Currently, there is no database to identify antimicrobial/heavy metal resistance and virulence-associated genes specific for . The aim of this study was to investigate the antimicrobial susceptibility and resistance profile of two isolates from Muscovy ducks () reared on a water poultry farm in Thuringia, Germany, and to create a database to fill this capability gap. The taxonomic classification revealed that the isolates belong to the gen. nov. as comb. nov. The antibiotic susceptibility was determined using the gradient strip method. While one of the isolates was resistant to five antibiotics, the other isolate was resistant to only two antibiotics. The presence of antimicrobial/heavy metal resistance genes and virulence determinants was determined using two custom-made databases. The custom-made databases identified a large repertoire of potential resistance and virulence-associated genes. This study provides the first resistance and virulence determinants database for .
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http://dx.doi.org/10.3390/genes11091104DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7564025PMC
September 2020

Draft genome sequence of multi-resistant subsp. serovar Rissen strain 19CS0416 isolated from Vietnam reveals plasmid mediated resistance to colistin already in 2013.

J Genomics 2020 3;8:76-79. Epub 2020 Jul 3.

Friedrich-Loeffler-Institut, Institute of Bacterial Infections and Zoonoses (IBIZ), Naumburger Str. 96a, 07743 Jena, Germany.

We report the first draft genome sequence of a strain with plasmid-mediated resistance to colistin encoded by gene in Vietnam. serovar Rissen was isolated from a Vietnamese pig slaughterhouse in 2013. We can confirm that gene is identical to the first reported gene of the strain SHP45, isolated in 2015 from a Chinese pig. The plasmid containing this gene in the strain 19CS0416 was highly related (96.86% identity) to the plasmid (pHNSHP45) contained in this Chinese strain. Moreover, this plasmid was determined to be 100% identical to a plasmid (p13P477T-7) belonging to an (13P477T) found in Hong Kong during the same year in pigs. Our results will aid in understanding the dissemination of gene in East Asia, dating back to as early as 2013.
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http://dx.doi.org/10.7150/jgen.42790DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7425045PMC
July 2020

Complete genome and plasmid sequences of a multidrug-resistant Salmonella enterica subsp. enterica serovar Panama isolate from a German cattle farm.

J Genomics 2020 29;8:71-75. Epub 2020 Jun 29.

Friedrich-Loeffler-Institut, Institute of Bacterial Infections and Zoonoses (IBIZ), Naumburger Str. 96a, 07743 Jena, Germany.

We describe a rare isolate of Salmonella enterica subsp. enterica serovar Panama with an extended-spectrum β-lactamase (ESBL) profile from a German cattle-fattening farm. Applying two next-generation sequencing methods we generated sequences of the genome as well as the plasmids; assembled the draft genome sequence of Salmonella enterica subsp. enterica serovar Panama isolate 18PM0209. Antimicrobial resistance genes, virulence-associated genes and plasmids were analyzed using bioinformatics. Occurrence of multidrug-resistant serovars at cattle-fattening farms indicate the need of enhanced surveillance to prevent further spread of these organisms.
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http://dx.doi.org/10.7150/jgen.48656DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7425046PMC
June 2020

Genomic Analysis and Antimicrobial Resistance of Strains From German Water Poultry.

Front Microbiol 2020 10;11:1549. Epub 2020 Jul 10.

Institute of Bacterial Infections and Zoonoses (IBIZ), Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Jena, Germany.

(formerly ) is a globally emerging foodborne and zoonotic pathogen. However, little is known about the species' genomic features and diversity, antibiotic resistance and virulence. In this study, 27 strains from water poultry in Thuringia, Germany, were investigated using whole-genome sequencing. Four of these strains were sequenced using long- and short-read sequencing methods to obtain circularized genomes. The German strains belong to the cluster I. Cluster I genomes exhibited a high degree of genetic diversity in which variable sites comprised 9.1% of the core genome. The German strains formed three subgroups that contained 2, 6, and 9 strains, respectively. The genomic analysis of cluster I revealed variable presence of mobile elements and that 65% of the strains lack CRISPR systems. The four circularized genomes carried a ∼2 Mbp chromosome and a single megaplasmid (size 98.1-154.5 Kbp). The chromosome was densely packed with coding sequences (∼92%) and showed inversions and shifts in the gene blocks between different strains. Antimicrobial resistance was assessed using a gradient strip diffusion method and showed that all 27 strains were resistant to cefotaxime and susceptible to erythromycin, gentamicin, and ampicillin. Sixteen strains were also resistant to ciprofloxacin, whereas 23 were resistant to streptomycin. The genetic prediction of antibiotic resistance identified numerous efflux pumps similar to those found in . All strains harbored two beta-lactamase genes which may explain the cefotaxime resistance. A correlation between the A point mutation (Thr-85-Ile) and ciprofloxacin resistance was partially discovered in 15 out of 16 strains. virulence profiling showed a wide range of virulence factors including a full chemotaxis system and most of the flagellar genes. In contrast to , no urease cluster was found. This study provides new insights into the genomic variability of strains of cluster I. The different genetic makeup of these strains may contribute to the virulence of strains and the severity of the infections in humans.
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http://dx.doi.org/10.3389/fmicb.2020.01549DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7365950PMC
July 2020

Characterization of Staphylococci and Streptococci Isolated from Milk of Bovides with Mastitis in Egypt.

Pathogens 2020 May 15;9(5). Epub 2020 May 15.

Friedrich-Loeffler-Institut, Institute of Bacterial Infections and Zoonoses, Naumburger Str. 96a, 07743 Jena, Germany.

The aim of this study was to characterize staphylococci and streptococci in milk from Egyptian bovides. In total, 50 milk samples were collected from localities in the Nile Delta region of Egypt. Isolates were cultivated, identified using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), and antibiotic susceptibility testing was performed by the broth microdilution method. PCR amplifications were carried out, targeting resistance-associated genes. Thirty-eight isolates and six isolates could be cultivated. isolates revealed a high resistance rate to penicillin, ampicillin, clindamycin, and erythromycin. The A gene defining methicillin-resistant , (C) and D genes was found in 87.5% of each. Coagulase-negative staphylococci showed a high prevalence of A, Z and K genes. Other resistance-associated genes were found. All isolates carried Z, (A), (B), (C) and A genes, while harbored (C), A-3L and M genes, additionally. In , most of these genes were found. The isolate harbored Z, (B), (C), A, K, L and M genes. isolate was analyzed by DNA microarray analysis. It was determined as sequence type 14, belonging to clonal complex 19 and represented capsule type VI. Pilus and cell wall protein genes, A, D and B/A genes were identified by microarray analysis.
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http://dx.doi.org/10.3390/pathogens9050381DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7281669PMC
May 2020

Phenotypic and Molecular Detection of Biofilm Formation in Isolated from Different Sources in Algeria.

Pathogens 2020 Feb 24;9(2). Epub 2020 Feb 24.

Institute of Bacterial Infections and Zoonoses, Friedrich-Loeffler-Institut, 07743 Jena, Germany.

is an opportunistic bacterium causing a wide variety of diseases. Biofilm formation of is of primary public and animal health concern. The purposes of the present study were to investigate the ability of isolated from animals, humans, and food samples to form biofilms and to screen for the presence of biofilm-associated and regulatory genes. In total, 55 isolated from sheep mastitis cases (n = 28), humans (n = 19), and from food matrices (n = 8) were identified using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The ability of aureus for slime production and biofilm formation was determined quantitatively. A DNA microarray examination was performed to detect adhesion genes (icaACD and biofilm-associated protein gene (bap)), genes encoding microbial surface components recognizing adhesive matrix molecules (MSCRAMMs), regulatory genes (accessory gene regulator (agr) and staphylococcal accessory regulator (sarA)), and the staphylococcal cassette chromosome mec elements (SCCmec). Out of 55 Staphylococcus aureus isolates, 39 (71.0%) and 23 (41.8%) were producing slime and biofilm, respectively. All strains isolated from food showed biofilm formation ability. 52.6% of the strains isolated from sheep with mastitis, and 17.9% of isolates from humans, were able to form a biofilm. Microarray analysis typed the Staphylococcus aureus into 15 clonal complexes. Among all isolates, four of the human isolates (21.1%) harbored the mecA gene (SCCmec type IV) typed into 2 clonal complexes (CC22-MRSA-IV and CC80-MRSA-IV) and were considered as methicillin-resistant, while two of them were slime-producing. None of the isolates from sheep with mastitis harbored the cna gene which is associated with biofilm production. The fnbB gene was found in 100%, 60% and 40% of biofilm-producing isolated from food, humans, and sheep with mastitis, respectively. Three agr groups were present and agr group III was predominant with 43.6%, followed by agr group I (38.2%), and agr group II (18.2%). This study revealed the capacity of Staphylococcus aureus isolates to form biofilms and highlighted the genetic background displayed by isolates from different sources in Algeria.
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http://dx.doi.org/10.3390/pathogens9020153DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7168657PMC
February 2020

Microarray-based detection of resistance and virulence factors in commensal Escherichia coli from livestock and farmers in Egypt.

Vet Microbiol 2020 Jan 29;240:108539. Epub 2019 Nov 29.

Leibniz Institute of Photonic Technology (IPHT), 07745, Jena, Germany; INFECTOGNOSTICS Research Campus, 07745, Jena, Germany. Electronic address:

The objective of our study was to provide a molecular analysis using DNA-microarray based assays of commensal E. coli populations from apparently healthy livestock and their attendants to assess the virulence potential as well as multidrug resistance (MDR) genotypes. We randomly collected 132 fecal samples from seemingly healthy smallholder´s food producing animals [buffalo (n = 32) and cattle (n = 50)] as well as from contacting farmers (n = 50). Bacterial isolation and identification were performed using standard protocols, while E. coli isolates were characterized using a DNA microarray system targeting 60 different virulence and 47 antibiotic resistance genes of clinical importance and allowing assignment to most common H and O types. From the fecal samples examined, 47 E. coli isolates were obtained. The array predicted serotypes for 14 out of the 47 E. coli isolates. Six E. coli isolates were identified as STEC since Shiga toxin genes were detected. In summary, 36 different virulence genes were identified; of which, hemL, lpfA and iss were most prevalent. Thirty-four E. coli isolates were found to carry at least one antimicrobial resistance gene. Of these, 20 did exhibit genes allowing strain classification as MDR. More than half of the isolates contained antimicrobial resistance genes associated with beta lactam resistance 27/47 (57.5 %). The 13 remaining isolates did not contain any resistance gene tested with the array. Our study demonstrated the presence of antimicrobial resistance genes and virulence genotypes among commensal E. coli of human and animal sources.
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http://dx.doi.org/10.1016/j.vetmic.2019.108539DOI Listing
January 2020

Short communication: Diversity of staphylococci isolated from sheep mastitis in northern Algeria.

J Dairy Sci 2020 Jan 14;103(1):890-897. Epub 2019 Nov 14.

Leibniz Institute of Photonic Technology (IPHT), 07745 Jena, Germany; InfectoGnostics Research Campus Jena e. V., 07743 Jena, Germany; Institute for Medical Microbiology and Hygiene, Medical Faculty "Carl Gustav Carus," Technical University Dresden, 01307 Dresden, Germany.

Mastitis in ruminants is an important disease with major effects on both the economy and animal welfare. It is caused by major pathogens such as Staphylococcus aureus and minor pathogens such as coagulase-negative staphylococci. The objective of this study was to identify and characterize staphylococci as a cause of sheep mastitis in Algeria. In this study, 123 milk samples were collected directly from the udder of sheep suffering from clinical mastitis in 2 provinces in Algeria. Recovered isolates were identified using MALDI-TOF mass spectrometry. Virulence-associated and antimicrobial resistance genes as well as clonal complexes (CC) of S. aureus were determined using microarray-based analysis. A total of 45 staphylococci isolates were cultivated from sheep milk samples, and 28 S. aureus were identified as methicillin susceptible (62.2%). Seventeen other Staphylococcus isolates of different species were identified using MALDI-TOF mass spectrometry. Subsequent microarray analysis typed the methicillin-susceptible S. aureus to 6 CC: CC8-MSSA, CC97-MSSA, CC130/521-MSSA, CC479-MSSA, CC522-MSSA, and CC705-MSSA. The accessory gene regulator agrIII and the ruminant leukocidin genes lukF-P83 and lukM were found in all isolates of CC130/521, CC479, CC522, and CC705. The toxic shock syndrome toxin gene tst1 was detected exclusively in CC130/521. Additionally, virulence-associated genes (sea, sed, sak, hld, hlgA, edinB, and others) were detected. The presence of antibiotic resistance genes [blaZ, erm(B), and tet(K)] was detected in small numbers of staphylococci. Staphylococci possessing these genes are considered potential hazards for farm animals, farmers, and consumers. Data concerning the prevalence and diversity of staphylococci causing mastitis in sheep from Algeria are lacking. Presented results on different aspects about staphylococci in Algerian sheep populations should at least partially close that gap. However, further extensive studies covering more geographical regions are needed to assess the epidemiological risk.
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http://dx.doi.org/10.3168/jds.2019-16583DOI Listing
January 2020

Characterisation of a novel SCCmec VI element harbouring fusC in an emerging Staphylococcus aureus strain from the Arabian Gulf region.

PLoS One 2019 5;14(11):e0223985. Epub 2019 Nov 5.

InfectoGnostics Research Campus Jena, Jena, Germany.

Fusidic acid is a steroid antibiotic known since the 1960s. It is frequently used in topical preparations, i.e., ointments, for the treatment of skin and soft tissue infections caused by Staphylococcus aureus. There is an increasing number of methicillin-resistant S. aureus (MRSA) strains that harbour plasmid-borne fusB/far1 or fusC that is localised on SCC elements. In this study we examined a series of related CC30-MRSA isolates from the Arabian Gulf countries that presented with SCCmec elements and fusC, including a variant that-to the best of our knowledge-has not yet formally been described. It consisted of a class B mec complex and ccrA/B-4 genes. The fusidic acid resistance gene fusC was present, but contrary to the previously sequenced element of HDE288, it was not accompanied by tirS. This element was identified in CC30 MRSA from Kuwait, Saudi Arabia and the United Arab Emirates that usually also harbour the Panton-Valentin leukocidin (PVL) genes. It was also identified in CC8 and ST834 isolates. In addition, further CC30 MRSA strains with other SCCmec VI elements harbouring fusC were found to circulate in the Arabian Gulf region. It can be assumed that MRSA strains with SCCmec elements that include fusC have a selective advantage in both hospital and community settings warranting a review of the use of topical antibiotics and indicating the necessity of reducing over-the-counter sale of antibiotics, including fusidic acid, without prescription.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0223985PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6830749PMC
March 2020

Genome sequence of a pathogenic Corynebacterium ulcerans strain isolated from a wild boar with necrotizing lymphadenitis.

BMC Res Notes 2019 Oct 25;12(1):692. Epub 2019 Oct 25.

Institut für Bakterielle Infektionen und Zoonosen, Friedrich-Loeffler-Institut, Bundesforschungsinstitut für Tiergesundheit, Jena, Germany.

Objectives: Corynebacterium ulcerans can colonize a wide variety of animals and also humans are infected, typically by zoonotic transmission. Symptoms range from skin ulcers or systemic infections to diphtheria-like illness. In contrast, Corynebacterium pseudotuberculosis is widely distributed among herds of sheep, goats and other farm animals, where it causes high economic losses due to caseous lymphadenitis. Here we describe the genome sequence of an atypical C. ulcerans strain isolated from a wild boar with necrotizing lymphadenitis. This strain has similarities to C. pseudotuberculosis.

Data Description: Genome sequence data of C. ulcerans isolate W25 were generated, analyzed and taxonomical relationship to other Corynebacterium species as well as growth properties of the isolate were characterized. The genome of C. ulcerans W25 comprises 2,550,924 bp with a G+C content of 54.41% and a total of 2376 genes.
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http://dx.doi.org/10.1186/s13104-019-4704-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6815020PMC
October 2019

Evolution of Antibiotic Resistance of Coagulase-Negative Staphylococci Isolated from Healthy Turkeys in Egypt: First Report of Linezolid Resistance.

Microorganisms 2019 Oct 22;7(10). Epub 2019 Oct 22.

Friedrich-Loeffler-Institut, Institute of Bacterial Infections and Zoonoses, Naumburger Str. 96a, 07743 Jena, Germany.

Coagulase-negative staphylococci (CoNS) are gaining much attention as causative agents of serious nosocomial infections in humans. This study aimed to determine the prevalence and phenotypic antimicrobial resistance of CoNS as well as the presence of resistance-associated genes in CoNS isolated from turkey farms in Egypt. Two hundred and fifty cloacal swabs were collected from apparently healthy turkeys in Egypt. Suspected isolates were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The susceptibility testing of CoNS isolates against 20 antimicrobial agents was performed using the broth microdilution test. The presence of resistance-associated genes like A, A, Z, (A), (B), (C), -D, A, S, and was determined. Thirty-nine CoNS were identified. All isolates were phenotypically resistant to trimethoprim/sulfamethoxazole, penicillin, ampicillin, and tetracycline. The resistance rates to erythromycin, chloramphenicol, oxacillin, daptomycin, and tigecycline were 97.4%, 94.9%, 92.3%, 89.7%, and 87.2%, respectively. Thirty-one isolates were resistant to linezolid (79.5%). Low resistance rate was detected for both imipenem and vancomycin (12.8%). The (C) gene was identified in all erythromycin phenotypically resistant isolates, whereas two resistant isolates possessed three resistance-conferring genes (A), (B), and (C). The and A genes were detected in 11 (35.5%) and 12 (38.7%) of the 31 linezolid-resistant isolates. The A, -D, and Z genes were identified in 22.2%, 41.9%, and 2.6% of phenotypically resistant isolates to oxacillin, gentamicin, and penicillin, respectively. This is the first study revealing the correlation between linezolid resistance and presence of and A genes in CoNS isolates from Egypt, and it can help to improve knowledge about the linezolid resistance mechanism.
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http://dx.doi.org/10.3390/microorganisms7100476DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6843140PMC
October 2019

Reliable differentiation of a non-toxigenic tox gene-bearing Corynebacterium ulcerans variant frequently isolated from game animals using MALDI-TOF MS.

Vet Microbiol 2019 Oct 23;237:108399. Epub 2019 Aug 23.

Chemisches und Veterinäruntersuchungsamt Stuttgart (CVUAS), Schaflandstr. 3/2, 70367 Fellbach, Germany. Electronic address:

Corynebacterium (C.) ulcerans is a zoonotic member of the C. diphtheriae group and is known to cause abscesses in humans and several animal species. Toxigenic strains, expressing the tox gene encoding diphtheria toxin, are also able to cause diphtheria in humans. In recent years, a non-toxigenic but tox gene-bearing (NTTB) variant of C. ulcerans has been identified that was frequently isolated from clinically healthy as well as from diseased wildlife animals, especially wild boars (Sus scrofa scrofa) in Germany and Austria. The described clinical cases showed similar signs of disease and the isolated corynebacteria displayed common genetic features as well as similar spectroscopic characteristics, therefore being assigned to a so called wild boar cluster (WBC). This study describes the establishment and validation of a method using MALDI-TOF mass spectrometry for a reliable differentiation between various members of the C. diphtheriae group at species level as well as a reliable sub-level identification of C. ulcerans isolates of the WBC variant. For this study 93 C. ulcerans isolates from wildlife animals, 41 C. ulcerans isolates from other animals and humans, and 53 isolates from further representatives of the C. diphtheriae group, as well as 26 non-diphtheriae group Corynebacteria collected via the MALDI user platform from seven MALDI users were used. By assigning 86 C. ulcerans isolates to the WBC the extensive geographical distribution of this previously less noticed variant in two Central European countries could be shown.
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http://dx.doi.org/10.1016/j.vetmic.2019.108399DOI Listing
October 2019

Differentiation of Subspecies by Proteotyping.

Eur J Microbiol Immunol (Bp) 2019 Jun 20;9(2):62-71. Epub 2019 May 20.

Institut für Medizinische Mikrobiologie, Universitätsmedizin Göttingen, Göttingen, Germany.

is a causative agent of intestinal illness and, occasionally, severe systemic infections and meningitis. currently comprises three subspecies: subspecies subspecies , and subspecies and are primarily associated with mammals whereas is associated with reptiles. To offer an alternative to laborious sequence-based techniques such as multilocus sequence typing (MLST) and polymerase chain reaction (PCR)-ribotyping for this species, the purpose of the study was to develop a typing scheme based on proteotyping. In total, 41 representative strains were analyzed by intact cell mass spectrometry and compared to MLST results. Biomarkers detected in the mass spectrum of subsp. reference strain LMG 6442 (NCTC 10842) as well as corresponding isoforms were associated with the respective amino acid sequences and added to the proteotyping scheme. In combination, the 9 identified biomarkers allow the differentiation of subspecies strains from and subspecies strains. Biomarkers to distinguish between and were not found. The results of the study show the potential of proteotyping to differentiate different subspecies, but also the limitations of the method.
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http://dx.doi.org/10.1556/1886.2019.00006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6563684PMC
June 2019

Antimicrobial Susceptibility and Genomic Structure of Isolates.

Front Microbiol 2018 14;9:3067. Epub 2018 Dec 14.

Institute of Bacterial Infections and Zoonoses (IBIZ), Friedrich Loeffler Institute, Jena, Germany.

spp. are considered the most common bacterial cause of foodborne gastroenteritis in the world. The family includes the genus with the three species , , and as emergent enteropathogens and potential zoonotic agents. Here, we characterized genome sequences of that were isolated from water poultry on farms in Germany. Isolates were cultured, identified by MALDI-TOF MS and identification was verified with PCR assays. Antibiotic susceptibility testing of isolates was carried out with erythromycin, ciprofloxacin, doxycycline, tetracycline, gentamicin, and streptomycin using the gradient strip method (-test). We also sequenced whole genomes and predicted antibiotic resistance determinants, virulence factors, performed a phylogenetic analysis to determine the genetic relatedness of these isolates and searched for plasmids.
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http://dx.doi.org/10.3389/fmicb.2018.03067DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6302008PMC
December 2018

Fast, economic and simultaneous identification of clinically relevant Gram-negative species with multiplex real-time PCR.

Future Microbiol 2019 01 12;14:23-32. Epub 2018 Dec 12.

Research & Development, Abbott (Alere Technologies GmbH), Jena, Germany.

Aim: A newly designed multiplex real-time PCR (rt-PCR) was validated to detect four clinically relevant Gram-negative bacteria (Escherichia coli, Acinetobacter baumannii, Klebsiella pneumoniae and Pseudomonas aeruginosa).

Materials & Methods: Serial dilutions of genomic DNA were used to determine the limit of detection. Colony PCR was performed with isolates of the four selected species and other species as negative controls. Isolates were characterized genotypically and phenotypically to evaluate the assay.

Results: Specific signals of all target genes were detected with diluted templates comprising ten genomic equivalents. Using colony rt-PCR, all isolates of the target species were identified correctly. All negative control isolates were negative.

Conclusion: The genes gad, basC, khe and ecfX can reliably identify these four species via multiplex colony rt-PCR.
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http://dx.doi.org/10.2217/fmb-2018-0197DOI Listing
January 2019

Complete Genome Sequence of Mycoplasma gallopavonis Type Strain WR1.

Microbiol Resour Announc 2018 Nov 29;7(21). Epub 2018 Nov 29.

Friedrich-Loeffler-Institut, Institute of Bacterial Infections and Zoonoses (IBIZ), Jena, Germany.

Here, we report the complete genome sequence of Mycoplasma gallopavonis type strain WR1 (= ATCC 33551 = NCTC 10186), which is a common microorganism in eastern wild turkeys and is considered a nonpathogenic commensal in turkeys.
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http://dx.doi.org/10.1128/MRA.01256-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6284725PMC
November 2018

Antimicrobial resistance in from healthy broilers in Egypt: emergence of colistin-resistant and extended-spectrum β-lactamase-producing .

Gut Pathog 2018 19;10:39. Epub 2018 Sep 19.

Friedrich-Loeffler-Institut (Federal Research Institute for Animal Health), Institute of Bacterial Infections and Zoonoses, Naumburger Str. 96a, 07743 Jena, Germany.

Background: Poultry remains one of the most important reservoir for zoonotic multidrug resistant pathogens. The global rise of antimicrobial resistance in Gram-negative bacteria is of reasonable concern and demands intensified surveillance.

Methods: In 2016, 576 cloacal swabs were collected from 48 broiler farms located in five governorates in northern Egypt. Isolates of could be cultivated on different media and were identified by MALDI-TOF MS and PCR. isolates were genotyped by DNA-microarray-based assays. The antimicrobial susceptibility to 14 antibiotics was determined and resistance-associated genes were detected. The VITEK-2 system was applied for phenotypical confirmation of extended-spectrum β-lactamase-producing isolates. The determination of colistin resistance was carried out phenotypically using E-test and genotypically using PCR for detection of the -1 gene.

Results: Out of 576 samples, 72 representatives of were isolated and identified as 63 (87.5%), 5 (6.9%), 2 (2.8%) and 2 spp. (2.8%). Seven out of 56 cultivated (12.5%) were confirmed as ESBL-producing and one isolate (1.8%) as ESBL/carbapenemase-producing . Five out of 63 isolates (7.9%) recovered from different poultry flocks were phenotypically resistant to colistin and harboured -1 gene.

Conclusions: This is the first study reporting colistin resistance and emergence of multidrug resistance in isolated from healthy broilers in the Nile Delta region, Egypt. Colistin-resistant in poultry is of public health significance. The global rise of ESBL- and carbapenemase-producing Gram-negative bacteria demands intensified surveillance. ESBL-producing in poultry farms in Egypt are of major concern that emphasizes the possibility of spread of such strains to humans. The results also reinforce the need to develop strategies and to implement specific control procedures to reduce the use of antibiotics.
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http://dx.doi.org/10.1186/s13099-018-0266-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6148799PMC
September 2018

Emerging of antimicrobial resistance in staphylococci isolated from clinical and food samples in Algeria.

BMC Res Notes 2018 Sep 12;11(1):663. Epub 2018 Sep 12.

Institute of Bacterial Infections and Zoonoses, Friedrich-Loeffler-Institut, Naumburger Str. 96a, 07743, Jena, Germany.

Objective: The antimicrobial resistance of staphylococci rose worldwide. In total, 96 Staphylococcus isolates from food and clinical samples were collected from two provinces in Algeria. The antimicrobial susceptibility testing was performed and resistance-associated genes were detected.

Results: Fifty-one strains were isolated from food samples and differentiated into 33 Staphylococcus aureus and 18 coagulase-negative staphylococci. Forty-five staphylococci were collected from hospital and community-acquired infection cases. All S. aureus isolated from food were resistant to penicillin and 45.5% were resistant to tetracycline. The resistance rates of 45 clinical Staphylococcus isolates were 86.7%, 48.9%, 37.8% and 20.0% to penicillin, tetracycline, erythromycin and kanamycin, respectively. Nine isolates were confirmed as MRSA from food and clinical isolates. One S. aureus originated from food was confirmed as vancomycin-resistant. Multidrug-resistance was observed among 25.5% and 53.3% of food and clinical staphylococci, respectively. The tetM/K, blaZ, aacA-aphD, ermC and mecA genes were detected in food and clinical isolates. ermA gene was not found. This study provided insight into the status of antimicrobial resistance of staphylococci isolated from food and clinical samples in Algeria. Further investigations and surveillance programmes are mandatory.
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http://dx.doi.org/10.1186/s13104-018-3762-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6134775PMC
September 2018

Identification of Campylobacter fetus by fluorescence in situ hybridization (FISH).

J Microbiol Methods 2018 08 24;151:44-47. Epub 2018 May 24.

Institut für Medizinische Mikrobiologie, Universitätsmedizin Göttingen, Göttingen, Germany. Electronic address:

Two new DNA FISH-probes for Campylobacter fetus were designed, in silico checked for cross-reactions and successfully evaluated in a multi-centric approach with 41 Campylobacter fetus isolates including isolates of all three know subspecies: Campylobacter fetus ssp. fetus, Campylobacter fetus ssp. venerealis, and Campylobacter fetus ssp. testudinum and 40 strains of five non-target Campylobacter species.
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http://dx.doi.org/10.1016/j.mimet.2018.05.020DOI Listing
August 2018

Draft Genome Sequence of Riemerella anatipestifer Isolate 17CS0503.

Genome Announc 2018 May 17;6(20). Epub 2018 May 17.

Institute of Bacterial Infections and Zoonoses at the Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Jena, Germany.

is a Gram-negative bacterium belonging to the family It is primarily associated with acute septicemia in younger birds. The isolate 17CS0503 described here was isolated from a Peking duck () in Hannover, Germany, in 1999.
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http://dx.doi.org/10.1128/genomeA.00274-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5958266PMC
May 2018

Variability of SCCmec elements in livestock-associated CC398 MRSA.

Vet Microbiol 2018 Apr 22;217:36-46. Epub 2018 Mar 22.

Abbott (Alere Technologies GmbH), Jena, Germany; InfectoGnostics Research Campus Jena, Jena, Germany.

The most common livestock-associated lineage of methicillin-resistant Staphylococcus aureus (MRSA) in Western Europe is currently clonal complex (CC) 398. CC398-MRSA spread extensively across livestock populations in several Western European countries, and livestock-derived CC398-MRSA strains can also be detected in humans. Based on their SCCmec elements, different CC398 strains can be distinguished. SCCmec elements of 100 veterinary and human CC398-MRSA isolates from Germany and Austria were examined using DNA microarray-based assays. In addition, 589 published SCC and/or genome sequences of CC398-MRSA (including both, fully finished and partially assembled sequences) were analysed by mapping them to the probe sequences of the microarrays. Several isolates and sequences showed an insertion of a large fragment of CC9 genomic DNA into the CC398 chromosome. Fifteen subtypes of SCCmec elements were detected among the 100 CC398 isolates and 41 subtypes could be discerned among the published CC398 sequences. Eleven of these were also experimentally detected within our strain collection, while four subtypes identified in the isolates where not found among the sequences. A high prevalence of heavy metal resistance genes, especially of czrC, was observed among CC398-MRSA. A possible co-selection of resistances to antibiotics and zinc/copper supplements in animal feed as well as a spill-over of SCCmec elements that have evolved in CC398-MRSA to other, possibly more virulent and/or medically relevant S. aureus lineages might pose public health problems in future.
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http://dx.doi.org/10.1016/j.vetmic.2018.02.024DOI Listing
April 2018

Revisiting subsp. , Causative Agent of Tularemia in Germany With Bioinformatics: New Insights in Genome Structure, DNA Methylation and Comparative Phylogenetic Analysis.

Front Microbiol 2018 13;9:344. Epub 2018 Mar 13.

Institute of Bacterial Infections and Zoonoses, Friedrich-Loeffler-Institut, Jena, Germany.

() is a highly virulent, Gram-negative bacterial pathogen and the causative agent of the zoonotic disease tularemia. Here, we generated, analyzed and characterized a high quality circular genome sequence of the subsp. strain 12T0050 that caused fatal tularemia in a hare. Besides the genomic structure, we focused on the analysis of oriC, unique to the genus and regulating replication in and outside hosts and the first report on genomic DNA methylation of a strain. The high quality genome was used to establish and evaluate a diagnostic whole genome sequencing pipeline. A genotyping strategy for was developed using various bioinformatics tools for genotyping. Additionally, whole genome sequences of subsp. isolates isolated in the years 2008-2015 in Germany were generated. A phylogenetic analysis allowed to determine the genetic relatedness of these isolates and confirmed the highly conserved nature of subsp.
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http://dx.doi.org/10.3389/fmicb.2018.00344DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5859110PMC
March 2018