Le metabolisme de l'acide citrique est a l'origine des proprietes technologiques des Leuc... more Le metabolisme de l'acide citrique est a l'origine des proprietes technologiques des Leuconostoc dans l'industrie laitiere (production de Co 2 et des composes d'arome tel que le diacetyle). Nous avons clone et sequence les genes impliques dans l'activite citrate lyase chez Leuconostoc mesenteroides subsp. Cremoris 195. Ces genes sont regroupes sur le chromosome et appartiennent au locus : clyR-mae-citCDEFGOP. En amont de ce locus, une sequence d'insertion appartenant a la famille des IS3 a ete identifiee. Les trois sous-unites de la citrate lyase (γ : acyl carrier protein ; β : citryl-s-ACP lyase ; α : citrate : acetate-ACP transferase) sont codees par les genes citD, citE et citF respectivement. L'acetylation du groupement prosthetique du citrate lyase est catalysee par l'acetate : SH-CL ligase (EC 6. 2. 2. 22) codee par le gene citC. Le gene mae codant pour une enzyme malique NAD +-dependante presumee (EC 1. 1. 1. 38) est transcrit avec le locus citCDEF. L’ARNm de 5. 2 kb n'est detecte que lorsque les cellules sont cultivees en presence de citrate. Deux promoteurs divergents localises sur la region intergenique clyR-mae, sont susceptibles de controler la transcription du locus mae-citCDEF et de clyR. Ce dernier code pour une proteine similaire aux regulateurs de la famille SorC. Une experience de retard sur gel a suggere des interactions entre la proteine purifiee clyR et la region inter genique riche en A + T. En aval du cluster citrate lyase, trois autres genes sont identifies, citG, citO et citP. Ce dernier code pour citrate permease chromosomique. CitG code pour une proteine a fonction inconnue, mais similaire aux proteines impliquees dans la synthese du groupement prosthetique de la malonate decarboxylase et citO code pour une proteine similaire aux lipases/esterases. Un role dans la modulation de l'activite citrate lyase est discute pour ces deux proteines. Parmi les bacteries dont le locus citrate lyase est desormais connu, Leuconostoc mesenteroides subsp. Cremoris 195 se distingue par une organisation genetique et une regulation originales.
Listeria monocytogenes is the etiological agent of listeriosis, a major foodborne disease and an ... more Listeria monocytogenes is the etiological agent of listeriosis, a major foodborne disease and an important public health concern. Contamination of meat with L. monocytogenes occurs frequently at the slaughterhouse. Our aims were; 1) to investigate the distribution of L. monocytogenes in the processing areas of four swine slaughterhouses; 2) to describe the diversity of L. monocytogenes strains by pulsed-field gel electrophoresis; 3) to identify persistent L. monocytogenes strains and describe their distribution; 4) to investigate the associations between persistence of strains and their following characteristics: detection in food isolates, detection in human clinical isolates, and the presence of benzalkonium chloride (BAC) resistance genes. Various operation areas within the four swine slaughterhouses were sampled on four occasions. A total of 2496 samples were analyzed, and L. monocytogenes was successfully isolated from 243 samples. The proportion of positive samples ranged from 32 to 58% in each slaughterhouse and from 24 to 68% in each operation area. Fifty-eight different pulsotypes were identified and eight pulsotypes, present in samples collected during 4 visits, were considered persistent. The persistent pulsotypes were significantly more likely to be detected in food (P < 0.01, exact χ 2) and human clinical cases (P < 0.01, exact χ 2), respectively. Among pulsotypes harboring the BAC bcrABC resistance cassette or the emrE multidrug transporter gene, 42.8% were persistent compared to 4.5% for pulsotypes without these resistance genes (P < 0.01, exact χ 2). Our study highlights the importance of persistent L. monocytogenes strains in the environmental contamination of slaughterhouses, which may lead to repeated contamination of meat products. It also
International Conference on the Epidemiology and Control of Biological, Chemical and Physical Hazards in Pigs and Pork, 2013
Listeria monocytogenes is a major public health concern. It has been associated with highly media... more Listeria monocytogenes is a major public health concern. It has been associated with highly mediatised outbreaks. It was the case in 2008, when an important episode (23 deaths) in Canada showed the need to reinforce the policies to better control Listeria in ready-to-eat products. Improved surveillance in the production plants environment increased the detection levels and represents a challenge for both industries and control agencies. In Canada, there is currently no surveillance policy of this microorganism in the steps preceeding ready-to-eat production. The distribution, diversity and characterisation of this microorganism in the slaughter and cutting environments are not well documented either. The aim of the present study is to provide such data, starting with a plant in Quebec. Moreover, recent studies suggested that the ability to produce biofilm and some virulence attributes (presence of SNP in InlA) are linked with "environmental" strains compared to virulent ones. Such characterization will be presented. The plant was sampled at three occasions, after the cleaning and disinfecting procedures, during a two-year period. A total of 874 samples were collected. Listeria detection followed the Health Canada standard method, geno-serogrouping was obtained by PCR and isolates were compared by PFGE after Apa1 and Asc1 restriction. Four serogroups (mainly I/IIb) were found, and 6 out of 21 pulsotypes were regularly or sporadically associated to human clinical strains profiles. We observed an increase in occurence of Listeria monocytogenes following the processing steps (χ 2 p<0.05). On the contrary, the diversity strongly and repeatedly decrease from step to step : 96.1% of the strains recovered from the cutting room presented the same profile. The "resident strains" have not been represented in clinical cases surveillance. Biofilm formation ability of strains alone could not explain strain transitions. Our results indicate that better knowledge of Listeria monocytogenes before food processing could be helpfull to optimise Listeria monocytogenes control under a risk analyse approach.
The introduction of Listeria monocytogenes into the food production chain is a concern, with nume... more The introduction of Listeria monocytogenes into the food production chain is a concern, with numerous grouped cases of listeriosis associated with milk‐derived or pork‐derived products have been documented. Management of this zoonotic pathogen considers all strains as an equal risk. Recently, a new perspective for characterisation of strain virulence was introduced with the discovery of the unaltered sequence of InlA as a determinant of strain virulence; this has also been reported as an infrequent finding among so‐called environmental strains, that is, strains isolated from food or from surfaces in food industries. The aim of this study was to differentiate L monocytogenes strains isolated from animal cases versus those from human cases and to differentiate clinical strains from environmental ones using a Caenorhabditis elegans virulence testing model. In Quebec in 2013/2014, the surveillance of L monocytogenes clinical isolates registered a total of 20 strains of animal origin and...
Background: A sudden increase in Salmonella Thompson (S. Thompson) cases distributed throughout t... more Background: A sudden increase in Salmonella Thompson (S. Thompson) cases distributed throughout three border regions in the province of Quebec in November 2016 triggered a provincial investigation to identify a common source of contamination and to put the appropriate control measures into place. Objective: To report on the outbreak and to describe the use of genomic sequencing to identify the salmonella serotype responsible. Methods: A descriptive survey of all reported cases of Salmonella serogroup C1 that had occurred between October 1, 2016 and February 15, 2017 was conducted. A case definition was developed. Pulsed field gel electrophoresis supplemented by analyses of genome sequences using the single nucleotide variant phylogenomics method were used to demarcate and manage the outbreak. Results: Eighteen cases of S. Thompson were identified through whole genome sequencing. The onset dates of symptoms for the 16 cases that presented enteric symptoms were November 21-December 2, 2016. Two cases that presented with atypical symptoms were not reported until February 2017. Among the 18 cases, 16 had eaten or probably eaten chicken shawarma at the same restaurant chain and nine of these cases ate it at the same restaurant. In total, five restaurants from this chain, spread throughout three border regions of Quebec, were identified. Conclusion: Outbreaks associated with chicken shawarma have been identified in the past. Efforts must be made to ensure that the owners of this type of restaurant know the contamination risk associated with this type of cooking and take the necessary steps to reduce this risk. The use of the genome sequencing method was very useful in defining the outbreak.
In this paper, we describe two open reading frames coding for a NAD-dependent malic enzyme ( mae ... more In this paper, we describe two open reading frames coding for a NAD-dependent malic enzyme ( mae ) and a putative regulatory protein ( clyR ) found in the upstream region of citCDEFG of Leuconostoc mesenteroides subsp. cremoris 195. The transcriptional analysis of the citrate lyase locus revealed one polycistronic mRNA covering the mae and citCDEF genes. This transcript was detected only on RNA prepared from cells grown in the presence of citrate. Primer extension experiments suggest that clyR and the citrate lyase operon are expressed from a bidirectional A-T-rich promoter region located between mae and clyR.
Whole genome sequencing (WGS) of Salmonella supports both molecular typing and detection of antim... more Whole genome sequencing (WGS) of Salmonella supports both molecular typing and detection of antimicrobial resistance (AMR). Here, we evaluated the correlation between phenotypic antimicrobial susceptibility testing (AST) and in silico prediction of AMR from WGS in Salmonella enterica (n = 1321) isolated from human infections in Canada. Phenotypic AMR results from broth microdilution testing were used as the gold standard. To facilitate high-throughput prediction of AMR from genome assemblies, we created a tool called Staramr, which incorporates the ResFinder and PointFinder databases and a custom gene-drug key for antibiogram prediction. Overall, there was 99% concordance between phenotypic and genotypic detection of categorical resistance for 14 antimicrobials in 1321 isolates (18,305 of 18,494 results in agreement). We observed an average sensitivity of 91.2% (range 80.5–100%), a specificity of 99.7% (98.6–100%), a positive predictive value of 95.4% (68.2–100%), and a negative pre...
Analyzing the genetic environment of clinically relevant MDR genes can provide information on the... more Analyzing the genetic environment of clinically relevant MDR genes can provide information on the way in which such genes are maintained and disseminated. Understanding this phenomenon is of interest for clinicians as it can also provide insight on where these genes might have been sourced, possibly supporting outbreak investigations.
Objectives: We investigated whether the increased prevalence of gentamicin resistance in Salmonel... more Objectives: We investigated whether the increased prevalence of gentamicin resistance in Salmonella from human infections was related to a similar increased prevalence in isolates from broiler chickens and whether this increase may have been due to co-selection from use of lincomycin-spectinomycin in chickens on farms. Methods: Whole genome sequencing was performed on gentamicin-resistant (gen-R) Salmonella isolates from human and chicken sources collected from 2014-2017 by the Canadian Integrated Program for Antimicrobial Resistance Surveillance (CIPARS). We determined the genomic relatedness of strains and characterized resistance genes and plasmids. Results: From 2014-2017, 247 isolates of gen-R Salmonella were identified by CIPARS: 188 were from humans and 59 from chicken sources (26 from live animals on farm and 33 from retail meat). The five most common gen-R serovars were Heidelberg (n=93, 31.5%), 4,[5],12:i:- (n=42, 14.2%), Kentucky (n=37, 12.5%), Infantis (n=33, 11.2%), and...
The persuasiveness of genomic evidence has pressured scientific agencies to supplement or replace... more The persuasiveness of genomic evidence has pressured scientific agencies to supplement or replace well-established methodologies to inform public health and food safety decision-making. This study of 52 epidemiologically defined Listeria monocytogenes isolates, collected between 1981 and 2011, including nine outbreaks, was undertaken (1) to characterize their phylogenetic relationship at finished genome-level resolution, (2) to elucidate the underlying genetic diversity within an endemic subtype, CC8, and (3) to re-evaluate the genetic relationship and epidemiology of a CC8delimited outbreak in Canada in 2008. Genomes representing Canadian Listeria outbreaks between 1981 and 2010 were closed and manually annotated. Single nucleotide variants (SNVs) and horizontally acquired traits were used to generate phylogenomic models. Phylogenomic relationships were congruent with classical subtyping and epidemiology, except for CC8 outbreaks, wherein the distribution of SNV and prophages revealed multiple co-evolving lineages. Chronophyletic reconstruction of CC8 evolution indicates that prophage-related genetic changes among CC8 strains manifest as PFGE subtype reversions, obscuring the relationship between CC8 isolates, and complicating the public health interpretation of subtyping data, even at maximum genome resolution. The size of the shared genome interrogated did not change the genetic relationship measured between highly related isolates near the tips of the phylogenetic tree, illustrating the robustness of these approaches for routine public health applications where the focus is recent ancestry. The possibility exists for temporally and epidemiologically distinct events to appear related even at maximum genome resolution, highlighting the continued importance of epidemiological evidence.
We report a novel RNase H2-dependent PCR (rhPCR) genotyping assay for a small number of discrimin... more We report a novel RNase H2-dependent PCR (rhPCR) genotyping assay for a small number of discriminatory single-nucleotide polymorphisms (SNPs) that identify lineages and sub-lineages of the highly clonal pathogen Salmonella Heidelberg (SH). Standard PCR primers targeting numerous SNP locations were initially designed in silico, modified to be RNase H2-compatible, and then optimized by laboratory testing. Optimization often required repeated cycling through variations in primer design, assay conditions, reagent concentrations and selection of alternative SNP targets. The final rhPCR assay uses 28 independent rhPCR reactions to target 14 DNA bases that can distinguish 15 possible lineages and sub-lineages of SH. On evaluation, the assay correctly identified the 12 lineages and sub-lineages represented in a panel of 75 diverse SH strains. Non-specific amplicons were observed in 160 (15.2%) of the 1050 reactions, but due to their low intensity did not compromise assay performance. Furthermore, in silico analysis of 500 closed genomes from 103 Salmonella serovars and laboratory rhPCR testing of five prevalent Salmonella serovars including SH indicated the assay can identify Salmonella isolates as SH, since only SH isolates generated amplicons from all 14 target SNPs. The genotyping results can be fully correlated with whole genome sequencing (WGS) data in silico. This fast and economical assay, which can identify SH isolates and classify them into related or unrelated lineages and sub-lineages, has potential applications in outbreak identification, source attribution and microbial source tracking.
The Canadian journal of infectious diseases & medical microbiology = Journal canadien des maladies infectieuses et de la microbiologie médicale, 2016
Necrotizing fasciitis is a serious disease characterized by the necrosis of the subcutaneous tiss... more Necrotizing fasciitis is a serious disease characterized by the necrosis of the subcutaneous tissues and fascia. E. coli as the etiologic agent of necrotizing fasciitis is a rare occurrence. A 66-year-old woman underwent total abdominal hysterectomy with bilateral salpingo-oophorectomy. She rapidly developed necrotizing fasciitis which led to her death 68 hours following surgery. An E. coli strain was isolated from blood and fascia cultures. DNA microarray revealed the presence of 20 virulence genes.
Salmonella enterica serovar Heidelberg is the second most frequently occurring serovar in Quebec ... more Salmonella enterica serovar Heidelberg is the second most frequently occurring serovar in Quebec and the third-most prevalent in Canada. Given that conventional pulsed-field gel electrophoresis (PFGE) subtyping for common Salmonella serovars, such as S . Heidelberg, yields identical subtypes for the majority of isolates recovered, public health laboratories are desperate for new subtyping tools to resolve highly clonal S . Heidelberg strains involved in outbreak events. As PFGE was unable to discriminate isolates from three epidemiologically distinct outbreaks in Quebec, this study was conducted to evaluate whole-genome sequencing (WGS) and phylogenetic analysis as an alternative to conventional subtyping tools. Genomes of 46 isolates from 3 Quebec outbreaks (2012, 2013, and 2014) supported by strong epidemiological evidence were sequenced and analyzed using a high-quality core genome single-nucleotide variant (hqSNV) bioinformatics approach (SNV phylogenomics [SNVphyl] pipeline). O...
We analysed 254 Shigella spp., isolates collected in Québec, Canada, during 2013-2014. Overall, 2... more We analysed 254 Shigella spp., isolates collected in Québec, Canada, during 2013-2014. Overall, 23.6% of isolates showed reduced susceptibility to azithromycin (RSA) encoded by mphA (11.6%), ermB (1.7%) or by both genes (86.7%). Shigella strains with RSA were mostly isolated from men who have sex with men (68.8% or higher) from the Montreal region. Complete sequence analysis of six selected plasmids from Shigella sonnei and different serotypes of Shigella flexneri emphasized the role of IS26 in RSA dissemination.
Contexte : Les changements climatiques jouent un rôle important dans l’établissement et l’expansi... more Contexte : Les changements climatiques jouent un rôle important dans l’établissement et l’expansion géographiques des zoonoses. Il est donc essentiel de connaître les populations à risque de contracter ces maladies pour ensuite orienter les politiques et les pratiques en santé publique. Au Québec, 14 zoonoses ont été identifiées importantes pour la santé publique afin de guider les efforts d’adaptation aux changements climatiques des décideurs et des chercheurs. Cette étude vise donc à dresser un portrait des professions et des secteurs d’activité économique à risque pour ces zoonoses. Méthode : Un examen rapide de la littérature scientifique a été réalisé. Des recherches ont été effectuées dans les bases de données des plateformes de recherche Ovid et EBSCO afin de repérer des articles publiés entre 1995 et 2018, en anglais et en français, sur 14 zoonoses (campylobactériose, cryptosporidiose, Escherichia coli vérocytotoxinogène, giardiase, listériose, salmonellose, encéphalite équi...
Résultats : Dix-huit cas de S. Thompson ont été identifiés par séquençage complet du génome. Les ... more Résultats : Dix-huit cas de S. Thompson ont été identifiés par séquençage complet du génome. Les dates de début des symptômes pour seize cas qui ont présenté des symptômes entériques s’étendaient du 21 novembre au 2 décembre 2016. Deux cas ont présenté des symptômes atypiques et n’ont pas été déclarés avant février 2017. Parmi les dix-huit cas, seize avaient consommé ou probablement consommé du poulet Shawarma dans une même chaîne de restauration, dont neuf dans le même restaurant. Au total, cinq restaurants de cette chaîne ont été identifiés et étaient répartis dans trois régions limitrophes du Québec.
Many public health laboratories across the world have implemented whole-genome sequencing (WGS) f... more Many public health laboratories across the world have implemented whole-genome sequencing (WGS) for the surveillance and outbreak detection of foodborne pathogens. PulseNet-affiliated laboratories have determined that most single-strain foodborne outbreaks are contained within 0–10 multi-locus sequence typing (MLST)-based allele differences and/or core genome single-nucleotide variants (SNVs). In addition to being a food- and travel-associated outbreak pathogen, most Shigella spp. cases occur through continuous person-to-person transmission, predominantly involving men who have sex with men (MSM), leading to long-term and recurrent outbreaks. Continuous transmission patterns coupled to genetic evolution under antibiotic treatment pressure require an assessment of existing WGS-based subtyping methods and interpretation criteria for cluster inclusion/exclusion. An evaluation of 4 WGS-based subtyping methods [SNVPhyl, coreMLST, core genome MLST (cgMLST) and whole-genome MLST (wgMLST)] ...
The emergence of multidrug-resistant bacterial strains worldwide has become a serious problem for... more The emergence of multidrug-resistant bacterial strains worldwide has become a serious problem for public health over recent decades. The increase in antimicrobial resistance has been expanding via plasmids as mobile genetic elements encoding antimicrobial resistance (AMR) genes that are transferred vertically and horizontally. This study focuses on Salmonella enterica, one of the leading foodborne pathogens in industrialized countries. S. enterica is known to carry several plasmids involved not only in virulence but also in AMR. In the current paper, we present an integrated strategy to detect plasmid scaffolds in whole genome sequencing (WGS) assemblies. We developed a two-step procedure to predict plasmids based on i) the presence of essential elements for plasmid replication and mobility, as well as ii) sequence similarity to a reference plasmid. Next, to confirm the accuracy of the prediction in 1750 S. enterica short-read sequencing data, we combined Oxford Nanopore MinION long...
Salmonella spp. - A Global Challenge [Working Title]
Salmonella enterica serovar Enteritidis (or Salmonella Enteritidis, SE) is one of the oldest memb... more Salmonella enterica serovar Enteritidis (or Salmonella Enteritidis, SE) is one of the oldest members of the genus Salmonella, based on the date of first description and has only gained prominence as a significant bacterial contaminant of food over the last three or four decades. Currently, SE is the most common Salmonella serovar causing foodborne illnesses. Control measures to alleviate human infections require that food isolates be characterized and this was until recently carried out using Pulsed-Field Gel Electrophoresis (PFGE) and phage typing as the main laboratory subtyping tools for use in demonstrating relatedness of isolates recovered from infected humans and the food source. The results provided by these analytical tools were presented with easy-to-understand and comprehensible nomenclature, however, the techniques were inherently poorly discriminatory, which is attributable to the clonality of SE. The tools have now given way to whole genome sequencing which provides a f...
Le metabolisme de l'acide citrique est a l'origine des proprietes technologiques des Leuc... more Le metabolisme de l'acide citrique est a l'origine des proprietes technologiques des Leuconostoc dans l'industrie laitiere (production de Co 2 et des composes d'arome tel que le diacetyle). Nous avons clone et sequence les genes impliques dans l'activite citrate lyase chez Leuconostoc mesenteroides subsp. Cremoris 195. Ces genes sont regroupes sur le chromosome et appartiennent au locus : clyR-mae-citCDEFGOP. En amont de ce locus, une sequence d'insertion appartenant a la famille des IS3 a ete identifiee. Les trois sous-unites de la citrate lyase (γ : acyl carrier protein ; β : citryl-s-ACP lyase ; α : citrate : acetate-ACP transferase) sont codees par les genes citD, citE et citF respectivement. L'acetylation du groupement prosthetique du citrate lyase est catalysee par l'acetate : SH-CL ligase (EC 6. 2. 2. 22) codee par le gene citC. Le gene mae codant pour une enzyme malique NAD +-dependante presumee (EC 1. 1. 1. 38) est transcrit avec le locus citCDEF. L’ARNm de 5. 2 kb n'est detecte que lorsque les cellules sont cultivees en presence de citrate. Deux promoteurs divergents localises sur la region intergenique clyR-mae, sont susceptibles de controler la transcription du locus mae-citCDEF et de clyR. Ce dernier code pour une proteine similaire aux regulateurs de la famille SorC. Une experience de retard sur gel a suggere des interactions entre la proteine purifiee clyR et la region inter genique riche en A + T. En aval du cluster citrate lyase, trois autres genes sont identifies, citG, citO et citP. Ce dernier code pour citrate permease chromosomique. CitG code pour une proteine a fonction inconnue, mais similaire aux proteines impliquees dans la synthese du groupement prosthetique de la malonate decarboxylase et citO code pour une proteine similaire aux lipases/esterases. Un role dans la modulation de l'activite citrate lyase est discute pour ces deux proteines. Parmi les bacteries dont le locus citrate lyase est desormais connu, Leuconostoc mesenteroides subsp. Cremoris 195 se distingue par une organisation genetique et une regulation originales.
Listeria monocytogenes is the etiological agent of listeriosis, a major foodborne disease and an ... more Listeria monocytogenes is the etiological agent of listeriosis, a major foodborne disease and an important public health concern. Contamination of meat with L. monocytogenes occurs frequently at the slaughterhouse. Our aims were; 1) to investigate the distribution of L. monocytogenes in the processing areas of four swine slaughterhouses; 2) to describe the diversity of L. monocytogenes strains by pulsed-field gel electrophoresis; 3) to identify persistent L. monocytogenes strains and describe their distribution; 4) to investigate the associations between persistence of strains and their following characteristics: detection in food isolates, detection in human clinical isolates, and the presence of benzalkonium chloride (BAC) resistance genes. Various operation areas within the four swine slaughterhouses were sampled on four occasions. A total of 2496 samples were analyzed, and L. monocytogenes was successfully isolated from 243 samples. The proportion of positive samples ranged from 32 to 58% in each slaughterhouse and from 24 to 68% in each operation area. Fifty-eight different pulsotypes were identified and eight pulsotypes, present in samples collected during 4 visits, were considered persistent. The persistent pulsotypes were significantly more likely to be detected in food (P < 0.01, exact χ 2) and human clinical cases (P < 0.01, exact χ 2), respectively. Among pulsotypes harboring the BAC bcrABC resistance cassette or the emrE multidrug transporter gene, 42.8% were persistent compared to 4.5% for pulsotypes without these resistance genes (P < 0.01, exact χ 2). Our study highlights the importance of persistent L. monocytogenes strains in the environmental contamination of slaughterhouses, which may lead to repeated contamination of meat products. It also
International Conference on the Epidemiology and Control of Biological, Chemical and Physical Hazards in Pigs and Pork, 2013
Listeria monocytogenes is a major public health concern. It has been associated with highly media... more Listeria monocytogenes is a major public health concern. It has been associated with highly mediatised outbreaks. It was the case in 2008, when an important episode (23 deaths) in Canada showed the need to reinforce the policies to better control Listeria in ready-to-eat products. Improved surveillance in the production plants environment increased the detection levels and represents a challenge for both industries and control agencies. In Canada, there is currently no surveillance policy of this microorganism in the steps preceeding ready-to-eat production. The distribution, diversity and characterisation of this microorganism in the slaughter and cutting environments are not well documented either. The aim of the present study is to provide such data, starting with a plant in Quebec. Moreover, recent studies suggested that the ability to produce biofilm and some virulence attributes (presence of SNP in InlA) are linked with "environmental" strains compared to virulent ones. Such characterization will be presented. The plant was sampled at three occasions, after the cleaning and disinfecting procedures, during a two-year period. A total of 874 samples were collected. Listeria detection followed the Health Canada standard method, geno-serogrouping was obtained by PCR and isolates were compared by PFGE after Apa1 and Asc1 restriction. Four serogroups (mainly I/IIb) were found, and 6 out of 21 pulsotypes were regularly or sporadically associated to human clinical strains profiles. We observed an increase in occurence of Listeria monocytogenes following the processing steps (χ 2 p<0.05). On the contrary, the diversity strongly and repeatedly decrease from step to step : 96.1% of the strains recovered from the cutting room presented the same profile. The "resident strains" have not been represented in clinical cases surveillance. Biofilm formation ability of strains alone could not explain strain transitions. Our results indicate that better knowledge of Listeria monocytogenes before food processing could be helpfull to optimise Listeria monocytogenes control under a risk analyse approach.
The introduction of Listeria monocytogenes into the food production chain is a concern, with nume... more The introduction of Listeria monocytogenes into the food production chain is a concern, with numerous grouped cases of listeriosis associated with milk‐derived or pork‐derived products have been documented. Management of this zoonotic pathogen considers all strains as an equal risk. Recently, a new perspective for characterisation of strain virulence was introduced with the discovery of the unaltered sequence of InlA as a determinant of strain virulence; this has also been reported as an infrequent finding among so‐called environmental strains, that is, strains isolated from food or from surfaces in food industries. The aim of this study was to differentiate L monocytogenes strains isolated from animal cases versus those from human cases and to differentiate clinical strains from environmental ones using a Caenorhabditis elegans virulence testing model. In Quebec in 2013/2014, the surveillance of L monocytogenes clinical isolates registered a total of 20 strains of animal origin and...
Background: A sudden increase in Salmonella Thompson (S. Thompson) cases distributed throughout t... more Background: A sudden increase in Salmonella Thompson (S. Thompson) cases distributed throughout three border regions in the province of Quebec in November 2016 triggered a provincial investigation to identify a common source of contamination and to put the appropriate control measures into place. Objective: To report on the outbreak and to describe the use of genomic sequencing to identify the salmonella serotype responsible. Methods: A descriptive survey of all reported cases of Salmonella serogroup C1 that had occurred between October 1, 2016 and February 15, 2017 was conducted. A case definition was developed. Pulsed field gel electrophoresis supplemented by analyses of genome sequences using the single nucleotide variant phylogenomics method were used to demarcate and manage the outbreak. Results: Eighteen cases of S. Thompson were identified through whole genome sequencing. The onset dates of symptoms for the 16 cases that presented enteric symptoms were November 21-December 2, 2016. Two cases that presented with atypical symptoms were not reported until February 2017. Among the 18 cases, 16 had eaten or probably eaten chicken shawarma at the same restaurant chain and nine of these cases ate it at the same restaurant. In total, five restaurants from this chain, spread throughout three border regions of Quebec, were identified. Conclusion: Outbreaks associated with chicken shawarma have been identified in the past. Efforts must be made to ensure that the owners of this type of restaurant know the contamination risk associated with this type of cooking and take the necessary steps to reduce this risk. The use of the genome sequencing method was very useful in defining the outbreak.
In this paper, we describe two open reading frames coding for a NAD-dependent malic enzyme ( mae ... more In this paper, we describe two open reading frames coding for a NAD-dependent malic enzyme ( mae ) and a putative regulatory protein ( clyR ) found in the upstream region of citCDEFG of Leuconostoc mesenteroides subsp. cremoris 195. The transcriptional analysis of the citrate lyase locus revealed one polycistronic mRNA covering the mae and citCDEF genes. This transcript was detected only on RNA prepared from cells grown in the presence of citrate. Primer extension experiments suggest that clyR and the citrate lyase operon are expressed from a bidirectional A-T-rich promoter region located between mae and clyR.
Whole genome sequencing (WGS) of Salmonella supports both molecular typing and detection of antim... more Whole genome sequencing (WGS) of Salmonella supports both molecular typing and detection of antimicrobial resistance (AMR). Here, we evaluated the correlation between phenotypic antimicrobial susceptibility testing (AST) and in silico prediction of AMR from WGS in Salmonella enterica (n = 1321) isolated from human infections in Canada. Phenotypic AMR results from broth microdilution testing were used as the gold standard. To facilitate high-throughput prediction of AMR from genome assemblies, we created a tool called Staramr, which incorporates the ResFinder and PointFinder databases and a custom gene-drug key for antibiogram prediction. Overall, there was 99% concordance between phenotypic and genotypic detection of categorical resistance for 14 antimicrobials in 1321 isolates (18,305 of 18,494 results in agreement). We observed an average sensitivity of 91.2% (range 80.5–100%), a specificity of 99.7% (98.6–100%), a positive predictive value of 95.4% (68.2–100%), and a negative pre...
Analyzing the genetic environment of clinically relevant MDR genes can provide information on the... more Analyzing the genetic environment of clinically relevant MDR genes can provide information on the way in which such genes are maintained and disseminated. Understanding this phenomenon is of interest for clinicians as it can also provide insight on where these genes might have been sourced, possibly supporting outbreak investigations.
Objectives: We investigated whether the increased prevalence of gentamicin resistance in Salmonel... more Objectives: We investigated whether the increased prevalence of gentamicin resistance in Salmonella from human infections was related to a similar increased prevalence in isolates from broiler chickens and whether this increase may have been due to co-selection from use of lincomycin-spectinomycin in chickens on farms. Methods: Whole genome sequencing was performed on gentamicin-resistant (gen-R) Salmonella isolates from human and chicken sources collected from 2014-2017 by the Canadian Integrated Program for Antimicrobial Resistance Surveillance (CIPARS). We determined the genomic relatedness of strains and characterized resistance genes and plasmids. Results: From 2014-2017, 247 isolates of gen-R Salmonella were identified by CIPARS: 188 were from humans and 59 from chicken sources (26 from live animals on farm and 33 from retail meat). The five most common gen-R serovars were Heidelberg (n=93, 31.5%), 4,[5],12:i:- (n=42, 14.2%), Kentucky (n=37, 12.5%), Infantis (n=33, 11.2%), and...
The persuasiveness of genomic evidence has pressured scientific agencies to supplement or replace... more The persuasiveness of genomic evidence has pressured scientific agencies to supplement or replace well-established methodologies to inform public health and food safety decision-making. This study of 52 epidemiologically defined Listeria monocytogenes isolates, collected between 1981 and 2011, including nine outbreaks, was undertaken (1) to characterize their phylogenetic relationship at finished genome-level resolution, (2) to elucidate the underlying genetic diversity within an endemic subtype, CC8, and (3) to re-evaluate the genetic relationship and epidemiology of a CC8delimited outbreak in Canada in 2008. Genomes representing Canadian Listeria outbreaks between 1981 and 2010 were closed and manually annotated. Single nucleotide variants (SNVs) and horizontally acquired traits were used to generate phylogenomic models. Phylogenomic relationships were congruent with classical subtyping and epidemiology, except for CC8 outbreaks, wherein the distribution of SNV and prophages revealed multiple co-evolving lineages. Chronophyletic reconstruction of CC8 evolution indicates that prophage-related genetic changes among CC8 strains manifest as PFGE subtype reversions, obscuring the relationship between CC8 isolates, and complicating the public health interpretation of subtyping data, even at maximum genome resolution. The size of the shared genome interrogated did not change the genetic relationship measured between highly related isolates near the tips of the phylogenetic tree, illustrating the robustness of these approaches for routine public health applications where the focus is recent ancestry. The possibility exists for temporally and epidemiologically distinct events to appear related even at maximum genome resolution, highlighting the continued importance of epidemiological evidence.
We report a novel RNase H2-dependent PCR (rhPCR) genotyping assay for a small number of discrimin... more We report a novel RNase H2-dependent PCR (rhPCR) genotyping assay for a small number of discriminatory single-nucleotide polymorphisms (SNPs) that identify lineages and sub-lineages of the highly clonal pathogen Salmonella Heidelberg (SH). Standard PCR primers targeting numerous SNP locations were initially designed in silico, modified to be RNase H2-compatible, and then optimized by laboratory testing. Optimization often required repeated cycling through variations in primer design, assay conditions, reagent concentrations and selection of alternative SNP targets. The final rhPCR assay uses 28 independent rhPCR reactions to target 14 DNA bases that can distinguish 15 possible lineages and sub-lineages of SH. On evaluation, the assay correctly identified the 12 lineages and sub-lineages represented in a panel of 75 diverse SH strains. Non-specific amplicons were observed in 160 (15.2%) of the 1050 reactions, but due to their low intensity did not compromise assay performance. Furthermore, in silico analysis of 500 closed genomes from 103 Salmonella serovars and laboratory rhPCR testing of five prevalent Salmonella serovars including SH indicated the assay can identify Salmonella isolates as SH, since only SH isolates generated amplicons from all 14 target SNPs. The genotyping results can be fully correlated with whole genome sequencing (WGS) data in silico. This fast and economical assay, which can identify SH isolates and classify them into related or unrelated lineages and sub-lineages, has potential applications in outbreak identification, source attribution and microbial source tracking.
The Canadian journal of infectious diseases & medical microbiology = Journal canadien des maladies infectieuses et de la microbiologie médicale, 2016
Necrotizing fasciitis is a serious disease characterized by the necrosis of the subcutaneous tiss... more Necrotizing fasciitis is a serious disease characterized by the necrosis of the subcutaneous tissues and fascia. E. coli as the etiologic agent of necrotizing fasciitis is a rare occurrence. A 66-year-old woman underwent total abdominal hysterectomy with bilateral salpingo-oophorectomy. She rapidly developed necrotizing fasciitis which led to her death 68 hours following surgery. An E. coli strain was isolated from blood and fascia cultures. DNA microarray revealed the presence of 20 virulence genes.
Salmonella enterica serovar Heidelberg is the second most frequently occurring serovar in Quebec ... more Salmonella enterica serovar Heidelberg is the second most frequently occurring serovar in Quebec and the third-most prevalent in Canada. Given that conventional pulsed-field gel electrophoresis (PFGE) subtyping for common Salmonella serovars, such as S . Heidelberg, yields identical subtypes for the majority of isolates recovered, public health laboratories are desperate for new subtyping tools to resolve highly clonal S . Heidelberg strains involved in outbreak events. As PFGE was unable to discriminate isolates from three epidemiologically distinct outbreaks in Quebec, this study was conducted to evaluate whole-genome sequencing (WGS) and phylogenetic analysis as an alternative to conventional subtyping tools. Genomes of 46 isolates from 3 Quebec outbreaks (2012, 2013, and 2014) supported by strong epidemiological evidence were sequenced and analyzed using a high-quality core genome single-nucleotide variant (hqSNV) bioinformatics approach (SNV phylogenomics [SNVphyl] pipeline). O...
We analysed 254 Shigella spp., isolates collected in Québec, Canada, during 2013-2014. Overall, 2... more We analysed 254 Shigella spp., isolates collected in Québec, Canada, during 2013-2014. Overall, 23.6% of isolates showed reduced susceptibility to azithromycin (RSA) encoded by mphA (11.6%), ermB (1.7%) or by both genes (86.7%). Shigella strains with RSA were mostly isolated from men who have sex with men (68.8% or higher) from the Montreal region. Complete sequence analysis of six selected plasmids from Shigella sonnei and different serotypes of Shigella flexneri emphasized the role of IS26 in RSA dissemination.
Contexte : Les changements climatiques jouent un rôle important dans l’établissement et l’expansi... more Contexte : Les changements climatiques jouent un rôle important dans l’établissement et l’expansion géographiques des zoonoses. Il est donc essentiel de connaître les populations à risque de contracter ces maladies pour ensuite orienter les politiques et les pratiques en santé publique. Au Québec, 14 zoonoses ont été identifiées importantes pour la santé publique afin de guider les efforts d’adaptation aux changements climatiques des décideurs et des chercheurs. Cette étude vise donc à dresser un portrait des professions et des secteurs d’activité économique à risque pour ces zoonoses. Méthode : Un examen rapide de la littérature scientifique a été réalisé. Des recherches ont été effectuées dans les bases de données des plateformes de recherche Ovid et EBSCO afin de repérer des articles publiés entre 1995 et 2018, en anglais et en français, sur 14 zoonoses (campylobactériose, cryptosporidiose, Escherichia coli vérocytotoxinogène, giardiase, listériose, salmonellose, encéphalite équi...
Résultats : Dix-huit cas de S. Thompson ont été identifiés par séquençage complet du génome. Les ... more Résultats : Dix-huit cas de S. Thompson ont été identifiés par séquençage complet du génome. Les dates de début des symptômes pour seize cas qui ont présenté des symptômes entériques s’étendaient du 21 novembre au 2 décembre 2016. Deux cas ont présenté des symptômes atypiques et n’ont pas été déclarés avant février 2017. Parmi les dix-huit cas, seize avaient consommé ou probablement consommé du poulet Shawarma dans une même chaîne de restauration, dont neuf dans le même restaurant. Au total, cinq restaurants de cette chaîne ont été identifiés et étaient répartis dans trois régions limitrophes du Québec.
Many public health laboratories across the world have implemented whole-genome sequencing (WGS) f... more Many public health laboratories across the world have implemented whole-genome sequencing (WGS) for the surveillance and outbreak detection of foodborne pathogens. PulseNet-affiliated laboratories have determined that most single-strain foodborne outbreaks are contained within 0–10 multi-locus sequence typing (MLST)-based allele differences and/or core genome single-nucleotide variants (SNVs). In addition to being a food- and travel-associated outbreak pathogen, most Shigella spp. cases occur through continuous person-to-person transmission, predominantly involving men who have sex with men (MSM), leading to long-term and recurrent outbreaks. Continuous transmission patterns coupled to genetic evolution under antibiotic treatment pressure require an assessment of existing WGS-based subtyping methods and interpretation criteria for cluster inclusion/exclusion. An evaluation of 4 WGS-based subtyping methods [SNVPhyl, coreMLST, core genome MLST (cgMLST) and whole-genome MLST (wgMLST)] ...
The emergence of multidrug-resistant bacterial strains worldwide has become a serious problem for... more The emergence of multidrug-resistant bacterial strains worldwide has become a serious problem for public health over recent decades. The increase in antimicrobial resistance has been expanding via plasmids as mobile genetic elements encoding antimicrobial resistance (AMR) genes that are transferred vertically and horizontally. This study focuses on Salmonella enterica, one of the leading foodborne pathogens in industrialized countries. S. enterica is known to carry several plasmids involved not only in virulence but also in AMR. In the current paper, we present an integrated strategy to detect plasmid scaffolds in whole genome sequencing (WGS) assemblies. We developed a two-step procedure to predict plasmids based on i) the presence of essential elements for plasmid replication and mobility, as well as ii) sequence similarity to a reference plasmid. Next, to confirm the accuracy of the prediction in 1750 S. enterica short-read sequencing data, we combined Oxford Nanopore MinION long...
Salmonella spp. - A Global Challenge [Working Title]
Salmonella enterica serovar Enteritidis (or Salmonella Enteritidis, SE) is one of the oldest memb... more Salmonella enterica serovar Enteritidis (or Salmonella Enteritidis, SE) is one of the oldest members of the genus Salmonella, based on the date of first description and has only gained prominence as a significant bacterial contaminant of food over the last three or four decades. Currently, SE is the most common Salmonella serovar causing foodborne illnesses. Control measures to alleviate human infections require that food isolates be characterized and this was until recently carried out using Pulsed-Field Gel Electrophoresis (PFGE) and phage typing as the main laboratory subtyping tools for use in demonstrating relatedness of isolates recovered from infected humans and the food source. The results provided by these analytical tools were presented with easy-to-understand and comprehensible nomenclature, however, the techniques were inherently poorly discriminatory, which is attributable to the clonality of SE. The tools have now given way to whole genome sequencing which provides a f...
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Papers by Sadjia Bekal