Genome Multilocus Sequence Typing Analysis Research Articles

Rapid detection of the source of a Listeria monocytogenes outbreak in Switzerland through routine interviewing of patients and whole-genome sequencing.

Listeriosis is a notifiable disease in Switzerland. In summer 2022, the Swiss Federal Office of Public Health noticed an increase in reports of listeriosis cases, indicating a possible ongoing outbreak. Here we present the approaches applied for rapidly confirming the outbreak, detecting the underlying source of infection and the measures put in place to eliminate it and contain the outbreak. For close surveillance and early detection of outbreak situations with their possible sources, listeriosis patients in Switzerland are systematically interviewed about risk behaviours and foods consumed prior to the infection.Listeria monocytogenesisolates derived from patients in medical laboratories are sent to the National Reference Laboratory for Enteropathogenic Bacteria and Listeria, where they routinely undergo whole-genome sequencing. Interview and whole-genome sequencing data are continuously linked for comparison and analysis. In summer 2022, 20 patient-derivedL. monocytogenesserotype 4b sequence type 388 strains were found to belong to an outbreak cluster (≤10 different alleles between neighbouring isolates) based on core genome multilocus sequence typing analysis. Geographically, 18 of 20 outbreak cases occurred in northeastern Switzerland. The median age of patients was 77.4 years (range: 58.1-89.7), with both sexes equally affected. Rolling analysis of the interview data revealed smoked trout from a local producer as a suspected infection source, triggering an on-site investigation of the production facility and sampling of the suspected products by the responsible cantonal food inspection team on 15 July 2022. Seven of ten samples tested positive forL. monocytogenesand the respective cantonal authority ordered a ban on production and distribution as well as a product recall. The Federal Food Safety and Veterinary Office released a nationwide public alert covering the smoked fish products concerned. Whole-genome sequencing analysis confirmed the interrelatedness of theL. monocytogenessmoked trout product isolates and the patient-derived isolates. Following the ban on production and distribution and the product recall, reporting of new outbreak-related cases rapidly dropped to zero. This listeriosis outbreak could be contained within a relatively short time thanks to identification of the source of contamination through the established combined approach of timely interviewing of every listeriosis patient or a representative and continuous molecular analysis of the patient- and food-derivedL. monocytogenesisolates. These findings highlight the effectiveness of this well-established, joint approach involving the federal and cantonal authorities and the research institutions mandated to contain listeriosis outbreaks in Switzerland.

Exploring multi-antibiotic resistance in Arcobacter butzleri isolates from a poultry processing plant in northern Italy: An in-depth inquiry

Arcobacter butzleri is a foodborne pathogen that has been identified in various animal-derived foods, notably in poultry meat. The prevalence of this bacterium in poultry underscores the need for comprehensive investigations into its dissemination within poultry meat processing plants. Such assessments are critical due to the potential cross-contamination that may occur, impacting the safety of the final food product. This study endeavours to evaluate the genomic similarity of 56 A. butzleri isolates obtained from chicken carcasses (specifically neck skins and caecum, post-slaughter) and equipment surfaces following cleaning and sanitizing procedures. All samples originate from a poultry slaughterhouse in Northern Italy. The genomic analysis includes single nucleotide polymorphism, average nucleotide identity, and core genome multi-locus sequence typing analysis, aiming to discern potential cross-contamination between carcasses and equipment. The study evaluates antibiotic resistance, biofilm production, and host cell colonization of A. butzleri strains through in vitro assays. Genomic de-replication reveals the presence of 31 distinct strains. Results elucidate that a portion of these strains is evident in multiple sources, indicative of cross-contamination even post-cleaning and sanitization. The in vitro tests underline multi-antibiotic resistance of A. butzleri with genes associated with antibiotic resistance, among them mexAB-oprM, showing a correlation to the observed resistance. This study establishes the colonization capability of A. butzleri strains on a human gut mucus-secreting cell model, suggesting a potential virulence factor. Detection of various putative virulence genes further supports the hypothesis of strains virulence potential. This study holds importance of A. butzleri as it unveils the antibiotic resistance and pathogenic capabilities inherent in this species, identifying associated genetic traits. Through comprehensive genomic analysis, it was conclusively validated instances of cross-contamination between various sources and equipment surfaces, underscoring the pervasive dissemination of this bacterium within a food processing plant and its persistence. This research serves as an exploration into the intricate dynamics of A. butzleri within poultry processing environments. It accentuates the imperative for stringent hygiene protocols and ongoing surveillance to curtail the risk of cross-contamination, thereby ensuring the safety of poultry products for discerning consumers.

Ralstonia pickettii bloodstream infections with potential genomic link to internationally distributed contaminated saline solution, Germany, October 2023.

Ralstonia pickettii is a Gram-negative rod which may cause invasive infections when they contaminate liquid medical products. After R. pickettii was detected in blood cultures and a stem cell product from three patients in a tertiary care hospital in Germany, whole genome sequencing of these three isolates and two water isolates from the environment was performed. Core genome multilocus sequence typing analysis showed that the three patient isolates were closely related and there was a large distance to the environmental isolates. In a genomic comparison, the patients' isolates were distantly related to an R. pickettii strain from a cluster in Australia suspected to be caused by contaminated saline produced in India, while all liquid medical products with a link to all patients were produced in Europe or the United States. Our data point towards an ongoing risk by an unknown common source that could be traced back to medical products contaminated with R. pickettii and potentially distributed worldwide. Investigating invasive R. pickettii infections, identifying and testing medical products administered to the patients and timely whole genome sequencing may help identify the exact source of this potentially global outbreak.

Klebsiella quasipneumoniae subsp. similipneumoniae ST1859 O5:KL35 from Soil: First Report of qnrE1 in the Environment.

A Klebsiella quasipneumoniae subsp. similipneumoniae strain, named S915, belonging to the ST1859 O5:KL35, and harboring the plasmid-mediated quinolone resistance qnrE1 gene, was isolated from a soil sample cultivated with lettuce in Brazil. The core genome multilocus sequence typing analysis revealed that S915 strain was most related to a clinical strain of Brazil. Comparative genomic analysis showed that ST1859 O5:KL35 strains have been circulating in clinical settings and are closely related to multidrug resistance and multimetal tolerance. Strain S915 presented a plasmid contig co-harboring the qnrE1 gene and tellurite tolerance operon. The region harboring the qnrE1 gene (ISEcp1-qnrE1-araJ-ahp) shared high similarity with others from infected humans, ready-to-eat dish, and food-producing animals in Brazil. This is the first report of the plasmid-mediated qnrE1 gene in the environment. Our findings evidence the initial dissemination of the qnrE1 gene in the environment by the introduction of a clinical strain, which may be spread to different sectors, representing a One Health challenge.

Listeria monocytogenes clones circulating in the natural environment of the Czech Republic and Slovakia

<i>Listeria monocytogenes</i> is not only a pathogen causing a serious food-borne disease in humans but can also occur as a saprophyte in the natural environment. This study aimed to evaluate the occurrence of <i>L.</i> <i>monocytogenes</i> obtained from the natural environment of the Czech Republic (hereinafter Czechia) and Slovakia in 2016–2018 and to compare the clonal relationship of strains circulating in the environment with the strains originating from the food chain and humans. Altogether, 217 samples of mud, surface water, vegetation and soil were collected in 61 locations. Samples were processed according to the modified EN ISO 11290-1 standard. The obtained <i>L.</i> <i>monocytogenes</i> isolates were characterised using serotyping, macrorestriction analysis, followed by pulsed-field gel electrophoresis, whole genome sequencing (WGS), and antimicrobial susceptibility testing. <i>L. monocytogenes</i> were detected in 8.8% of the examined samples and were isolated in 15 locations, mainly from the mud from the banks of the surface water sources. Altogether, 25 <i>L. monocytogenes</i> strains were obtained from 19 positive samples. Serotypes 1/2a, 4b, and 1/2b were detected among the strains. Twenty combined <i>Asc</i>I<i>/Apa</i>I pulsotypes were obtained by macrorestriction analysis. Altogether, 12 sequence types (STs) were detected using Multi-locus sequence typing (MLST) by WGS, with ST451 being the most frequent. The core genome MLST analysis revealed a heterogeneous population of environmental strains. No phenotype resistance was detected by antimicrobial susceptibility testing. Screening of antimicrobial-resistance genes using the platform ResFinder revealed the genes <i>fosX</i> in 24 isolates and <i>bla</i>TEM-116 in one isolate. The occurrence of <i>L.</i> <i>monocytogenes</i> in various samples from natural environments within wide altitude range during different seasons of the year may highlight this bacterium's remarkable adaptability and exceptional tolerance to external factors. Serotype distribution of the strains circulating in the natural environment of Czechia and Slovakia seems to reflect distribution in the human population more than in the food chain.

Genetic Characterization of Listeria from Food of Non-Animal Origin Products and from Producing and Processing Companies in Bavaria, Germany

Reported cases of listeriosis from food of non-animal origin (FNAO) are increasing. In order to assess the risk of exposure to Listeria monocytogenes from FNAO, the genetic characterization of the pathogen in FNAO products and in primary production and processing plants needs to be investigated. For this, 123 samples of fresh and frozen soft fruit and 407 samples of 39 plants in Bavaria, Germany that produce and process FNAO were investigated for Listeria contamination. As a result, 64 Listeria spp. isolates were detected using ISO 11290-1:2017. Environmental swabs and water and food samples were investigated. L. seeligeri (36/64, 56.25%) was the most frequently identified species, followed by L. monocytogenes (8/64, 12.50%), L. innocua (8/64, 12.50%), L. ivanovii (6/64, 9.38%), L. newyorkensis (5/64, 7.81%), and L. grayi (1/64, 1.56%). Those isolates were subsequently sequenced by whole-genome sequencing and subjected to pangenome analysis to retrieve data on the genotype, serotype, antimicrobial resistance (AMR), and virulence markers. Eight out of sixty-four Listeria spp. isolates were identified as L. monocytogenes. The serogroup analysis detected that 62.5% of the L. monocytogenes isolates belonged to serogroup IIa (1/2a and 3a) and 37.5% to serogroup IVb (4b, 4d, and 4e). Furthermore, the MLST (multilocus sequence typing) analysis of the eight detected L. monocytogenes isolates identified seven different sequence types (STs) and clonal complexes (CCs), i.e., ST1/CC1, ST2/CC2, ST6/CC6, ST7/CC7, ST21/CC21, ST504/CC475, and ST1413/CC739. The core genome MLST analysis also showed high allelic differences and suggests plant-specific isolates. Regarding the AMR, we detected phenotypic resistance against benzylpenicillin, fosfomycin, and moxifloxacin in all eight L. monocytogenes isolates. Moreover, virulence factors, such as prfA, hly, plcA, plcB, hpt, actA, inlA, inlB, and mpl, were identified in pathogenic and nonpathogenic Listeria species. The significance of L. monocytogenes in FNAO is growing and should receive increasing levels of attention.

Genetic Diversity and Relationships of Listeria monocytogenes Serogroup IIa Isolated in Poland.

In the present study, 100 L. monocytogenes isolates of serogroup IIa from food and food production environments in Poland were characterized towards the presence of virulence, resistance, and stress response genes using whole-genome sequencing (WGS). The strains were also molecularly typed and compared with multi-locus sequence typing (MLST) and core genome MLST analyses. The present isolates were grouped into 6 sublineages (SLs), with the most prevalent SL155 (33 isolates), SL121 (32 isolates), and SL8 (28 isolates) and classified into six clonal complexes, with the most prevalent CC155 (33 strains), CC121 (32 isolates), and CC8 (28 strains). Furthermore, the strains were grouped to eight sequence types, with the most prevalent ST155 (33 strains), ST121 (30 isolates), and ST8 (28; strains) followed by 60 cgMLST types (CTs). WGS data showed the presence of several virulence genes or putative molecular markers playing a role in pathogenesis of listeriosis and involved in survival of L. monocytogenes in adverse environmental conditions. Some of the present strains were molecularly closely related to L. monocytogenes previously isolated in Poland. The results of the study showed that food and food production environments may be a source of L. monocytogenes of serogroup IIa with pathogenic potential.

Whole-Genome Analysis of Multidrug-Resistant Salmonella Enteritidis Strains Isolated from Poultry Sources in Korea.

The Salmonella Enterica subsp. Enterica serovar Enteritidis is one of main serovars isolated from human patients with food poisoning and poultry without clinical signs. Consumption of poultry products contaminated with Salmonella Enteritidis is a common source of human salmonellosis; 82 Salmonella spp. were isolated from 291 samples of retail chicken meat, 201 one-day-old chicks, 30 internal organs of chickens, 156 chicken eggs, 100 duck eggs, 38 straw bedding samples, 18 samples of retail duck meat, and 19 swab samples from slaughterhouses in 2019 and 2020. An antibiotic susceptibility test was performed for all isolates, revealing 33 multidrug-resistant (MDR) strains. The whole genome of 33 MDR strains isolated in 2019 and 2020 and 10 strains isolated in 2011, 2012, and 2017 was sequenced using the MinION sequencing protocol. Within these 43 samples, 5 serovars were identified: S. Enteritidis, S. Agona, S. Virchow, S. Albany, and S. Bareilly. The most common serovar was S. Enteritidis (26/43), which showed the highest resistance to ampicillin (100%), followed by nalidixic acid (90%) and colistin (83%). Core genome multilocus sequence typing analysis showed that the S. Enteritidis strains isolated from different sources and in different years were clustered together. In addition, the S. Enteritidis strains isolated since 2011 consistently harbored the same antibiotic resistance patterns.

Molecular characterisation and antibiotic susceptibility of meningococcal isolates from healthy men who have sex with men

ObjectivesTo evaluate and characterise meningococcal carriage among healthy men who have sex with men (MSM) within a screening programme for Neisseria gonorrhoeae infection at the San Gallicano Dermatological Institute, Italy.MethodsA...

Use of an Ecosystem-Based Approach to Shed Light on the Heterogeneity of the Contamination Pattern of Listeria monocytogenes on Conveyor Belt Surfaces in a Swine Slaughterhouse in the Province of Quebec, Canada.

The role of the accompanying microbiota in the presence of Listeria monocytogenes on meat processing surfaces is not yet understood, especially in industrial production conditions. In this study, 300 conveyor belt samples from the cutting room of a swine slaughterhouse were collected during production. The samples were subjected to the detection of L. monocytogenes. Recovered strains were characterized by serogrouping-PCR, InlA Sanger sequencing and for their ability to form biofilm. A selection of isolates was compared with core genome multi-locus sequence typing analysis (cgMLST). The sequencing of the V4 region of the 16S RNA gene of the microorganisms harvested from each sample was carried out in parallel using the Illumina MiSeq platform. Diversity analyses were performed and MaAsLin analysis was used to assess the link between L. monocytogenes detection and the surrounding bacteria. The 72 isolates collected showed a low genetic diversity and important persistence characteristics. L. monocytogenes isolates were not stochastically distributed on the surfaces: the isolates were detected on three out of six production lines, each associated with a specific meat cut: the half carcasses, the bostons and the picnics. MaAsLin biomarker analysis identified the taxa Veillonella (p ≤ 0.0397) as a bacterial determinant of the presence of L. monocytogenes on processing surfaces. The results of this study revealed a heterogenous contamination pattern of the processing surfaces by L. monocytogenes and targeted a bacterial indicator of the presence of the pathogen. These results could lead to a better risk assessment of the contamination of meat products.

Serotyping, MLST, and Core Genome MLST Analysis of Salmonella enterica From Different Sources in China During 2004-2019.

Salmonella enterica (S. enterica) is an important foodborne pathogen, causing food poisoning and human infection, and critically threatening food safety and public health. Salmonella typing is essential for bacterial identification, tracing, epidemiological investigation, and monitoring. Serotyping and multilocus sequence typing (MLST) analysis are standard bacterial typing methods despite the low resolution. Core genome MLST (cgMLST) is a high-resolution molecular typing method based on whole genomic sequencing for accurate bacterial tracing. We investigated 250 S. enterica isolates from poultry, livestock, food, and human sources in nine provinces of China from 2004 to 2019 using serotyping, MLST, and cgMLST analysis. All S. enterica isolates were divided into 36 serovars using slide agglutination. The major serovars in order were Enteritidis (31 isolates), Typhimurium (29 isolates), Mbandaka (23 isolates), and Indiana (22 isolates). All strains were assigned into 43 sequence types (STs) by MLST. Among them, ST11 (31 isolates) was the primary ST. Besides this, a novel ST, ST8016, was identified, and it was different from ST40 by position 317 C → T in dnaN. Furthermore, these 250 isolates were grouped into 185 cgMLST sequence types (cgSTs) by cgMLST. The major cgST was cgST235530 (11 isolates), and only three cgSTs contained isolates from human and other sources, indicating a possibility of cross-species infection. Phylogenetic analysis indicated that most of the same serovar strains were putatively homologous except Saintpaul and Derby due to their multilineage characteristics. In addition, serovar I 4,[5],12:i:- and Typhimurium isolates have similar genomic relatedness on the phylogenetic tree. In conclusion, we sorted out the phenotyping and genotyping diversity of S. enterica isolates in China during 2004–2019 and clarified the temporal and spatial distribution characteristics of Salmonella from different hosts in China in the recent 16 years. These results greatly supplement Salmonella strain resources, genetic information, and traceability typing data; facilitate the typing, traceability, identification, and genetic evolution analysis of Salmonella; and therefore, improve the level of analysis, monitoring, and controlling of foodborne microorganisms in China.

Prevalence and Genotyping of Campylobacter jejuni and Campylobacter coli from Ovine Carcasses in New Zealand

A pilot survey was performed to determine the prevalence of Campylobacter jejuni and Campylobacter coli on three age classes (lamb, hogget, and mutton) of ovine carcass trim postdressing and prechill. Sampling of hogget carcasses was undertaken 6 months before sampling of lamb and mutton carcasses. A total of 120 trim samples were collected from 11 processing plants across New Zealand. All samples were enriched and screened using PCR for the presence of C. jejuni and C. coli, and isolation was attempted for all screen-positive samples. Enumeration of Campylobacter from lamb trim samples showed that Campylobacter bacteria were present in very low numbers (<10 CFU/g). The overall prevalence of Campylobacter for ovine trim based on PCR detection was 33% (39 of 120 samples), with prevalences for hogget, lamb, and mutton carcass trim of 56% (28 of 50), 11% (4 of 35), and 20% (7 of 35), respectively. Whole genome sequencing was performed on a selection of C. jejuni and C. coli isolates, and the data were used to subtype using multilocus sequence typing (MLST) and whole genome MLST. Twenty-five MLST sequence types (STs) were identified among 44 isolates, including ST42, ST50, ST3222, and ST3072, which have been previously reported to be associated with ruminant sources. Four novel STs were also identified. Whole genome MLST analysis further discriminated isolates within a single ST type and demonstrated a genetic diversity among the ovine isolates collected. Genes associated with the oxacillinase class of β-lactamase enzymes were identified in 41 of 44 Campylobacter isolates. This study provides preliminary data that can be incorporated into existing source attribution models to assist in determining the potential contribution of ovine sources to the burden of campylobacteriosis in New Zealand.

875. Pseudo-outbreak of Clostridioides (Clostridium) difficile amongst post-operative Oncology Patients

Background Clostridioides difficile infection rates are subject to infection prevention surveillance as a quality measure within the hospital setting. A large spike in Clostridioides difficile infections in post-operative patients, the majority of whom were gastrointestinal surgery (GIS) patients, was noted within a six month period (June through November 2019) at our comprehensive cancer center. These patients had been housed in one of two inpatient units and there was appropriate concern that this represented a C. difficile outbreak possibly related some type of infection control breach.MethodsIn an effort to query case relatedness, whole genome sequencing was performed using Illumina MiSeq instrumentation and chemistry with Illumina Nextera XT library chemistry. Assembly and core genome multilocus sequence typing analysis were performed with Ridom SeqSphere+ software. Cases were classified as community or hospital acquired based on the National Healthcare Safety Network (NHSN) definitions.ResultsThere were 23 samples submitted for possible whole genome sequencing (WGS). 5 samples were unable to be grown therefore WGS was not completed; 16 were found to be unrelated (51 or more allelic differences); 2 of the 18 isolates were found to be possibly related (7 to 50 allelic differences). There were no isolates found to be definitively related (zero to 6 allelic differences).ConclusionGiven the overwhelming unrelatedness of the isolates via whole genome sequencing, this increase of C. difficile cases, identified by routine surveillance within two inpatient units, was determined to be representative of a pseudo-outbreak rather than an outbreak. This study has implications on public health reporting. National Healthcare Safety Network definitions are used to identify healthcare facility-onset C. difficile infections (CDI). The majority of cases in this study met the definition of healthcare facility-onset, and thus were reported as such, despite being genetically unrelated. This raises the concern that a significant percentage of C. difficile infections may be currently misclassified as hospital-associated and this may have negative, unfair consequences for hospitals, such as implications on reimbursement.DisclosuresRobin Patel, MD, Accelerate Diagnostics (Grant/Research Support)CD Diagnostics (Grant/Research Support)Contrafect (Grant/Research Support)Curetis (Consultant)GenMark Diagnostics (Consultant)Heraeus Medical (Consultant)Hutchison Biofilm Medical Solutions (Grant/Research Support)Merck (Grant/Research Support)Next Gen Diagnostics (Consultant)PathoQuest (Consultant)Qvella (Consultant)Samsung (Other Financial or Material Support, Dr. Patel has a patent on Bordetella pertussis/parapertussis PCR issued, a patent on a device/method for sonication with royalties paid by Samsung to Mayo Clinic, and a patent on an anti-biofilm substance issued.)Selux Dx (Consultant)Shionogi (Grant/Research Support)Specific Technologies (Consultant)

Genome-based reclassification of Lactobacillus casei: emended classification and description of the species Lactobacillus zeae.

Taxonomic relationships between Lactobacillus casei, Lactobacillus paracasei and Lactobacillus zeae have long been debated. Results of previous analyses have shown that overall genome relatedness indices (such as average nucleotide identity and core nucleotide identity) between the type strains L. casei ATCC 393T and L. zeae ATCC 15820T were 94.6 and 95.3 %, respectively, which are borderline for species definition. However, the digital DNA‒DNA hybridization value was 57.3 %, which was clearly lower than the species delineation threshold of 70 %, and hence raised the possibility that L. casei could be reclassified into two species. To re-evaluate the taxonomic relationship of these taxa, multilocus sequence analysis (MLSA) based on the concatenated five housekeeping gene (dnaJ, dnaK, mutL, pheS and yycH) sequences, phylogenomic and core genome multilocus sequence typing analyses, gene presence and absence profiles using pan-genome analysis, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) profiling analysis, cellular fatty acid compositions, and phenotype analysis were carried out. The results of phenotypic characterization, MLSA, whole-genome sequence-based analyses and MALDI-TOF MS profiling justified an independent species designation for the L. zeae strains, and supported an emended the description of the name of Lactobacillus zeae (ex Kuznetsov 1956) Dicks et al. 1996, with ATCC 15820T (=DSM 20178T=BCRC 17942T) as the type strain.

Serological Survey and Molecular Typing Reveal New Leptospira Serogroup Pomona Strains among Pigs of Northern Italy.

Swine act as both maintenance and incidental hosts of pathogenic Leptospira spp. Here, a serological test was performed on 131,660 pig sera collected between 2002 and 2017 from 4715 farms in Northern Italy. A positivity rate of 13.05% was determined. Australis was the most frequently identified serogroup (77.29%), followed by Pomona (18.47%), Tarassovi (1.51%) and Icterohaemorrhagie (1.40%). Culture isolation and real-time Polymerase chain reaction (PCR) were carried out on 347 kidneys and 470 clinical samples, respectively. Overall, 133 strains were cultured successfully and 43 randomly chosen isolates were identified as serogroup Pomona. Multi-locus sequence typing (MLST) revealed that 41 isolates and 8 DNA extracted from biological samples belonged to sequence type 140. Using a multiple-locus, variable-number tandem repeat analysis, 43 samples produced identical profiles but, after 2014, three new Leptospira interrogans serogroup Pomona genotypes were observed. Interestingly, two isolates showed new MLST profiles and an unclassified identification by monoclonal antibodies. The 16S rRNA gene sequencing clustered them into L. kirschneri species and a core genome MLST analysis revealed an allelic identity of 96% compared with Mozdok strains. Genotyping allowed us to discriminate leptospires and to identify new emerging strains. The accurate identification of infective strains is required for formulating preventive methods and intervention strategies.

Whole genome sequencing confirms source of pathogens associated with bacterial foodborne illness in pets fed raw pet food.

Reports of raw meat pet food containing zoonotic foodborne bacteria, including Salmonella, Escherichia coli, and Listeria monocytogenes, are increasing. Contaminated raw pet food and biological waste from pets consuming those diets may pose a public health risk. The U.S. Food and Drug Administration Veterinary Laboratory Investigation and Response Network conducted 2 case investigations, involving 3 households with animal illnesses, which included medical record review, dietary and environmental exposure interviews, animal sample testing, and whole genome sequencing (WGS) of bacteria isolated from the pets and the raw pet food. For each case investigation, WGS with core genome multi-locus sequence typing analysis showed that the animal clinical isolates were closely related to one or more raw pet food bacterial isolates. WGS and genomic analysis of paired animal clinical and animal food isolates can confirm suspected outbreaks of animal foodborne illness.

Sequencing and phylogenetic characterization of Brucella canis isolates, Ohio, 2016

Brucella canis is one of zoonotic pathogens causing infection in human. In this study, we isolated and sequenced 38 B.canis strains from 11 cases. Core genome multilocus sequence typing analysis classified all B.canis isolates into two genogroups, GI and GII. All 38 isolates cluster together forming a 2016 Ohio cluster, in which they form five subclusters reflecting their geographical differences. Unlike GI, the isolates of the GII are from diverse geographical locations including Asia, America, Africa, and Europe and form Asia and South America clusters. Overall, our findings could be useful to investigate and track B.canis of future outbreaks.

Whole genome sequence-based serogrouping of Listeria monocytogenes isolates

Whole genome sequencing (WGS) is currently becoming the method of choice for characterization of Listeria monocytogenes isolates in national reference laboratories (NRLs). WGS is superior with regards to accuracy, resolution and analysis speed in comparison to several other methods including serotyping, PCR, pulsed field gel electrophoresis (PFGE), multilocus sequence typing (MLST), multilocus variable number tandem repeat analysis (MLVA), and multivirulence-locus sequence typing (MVLST), which have been used thus far for the characterization of bacterial isolates (and are still important tools in reference laboratories today) to control and prevent listeriosis, one of the major sources of foodborne diseases for humans. Backward compatibility of WGS to former methods can be maintained by extraction of the respective information from WGS data. Serotyping was the first subtyping method for L. monocytogenes capable of differentiating 12 serovars and national reference laboratories still perform serotyping and PCR-based serogrouping as a first level classification method for Listeria monocytogenes surveillance. Whole genome sequence based core genome MLST analysis of a L. monocytogenes collection comprising 172 isolates spanning all 12 serotypes was performed for serogroup determination. These isolates clustered according to their serotypes and it was possible to group them either into the IIa, IIc, IVb or IIb clusters, respectively, which were generated by minimum spanning tree (MST) and neighbor joining (NJ) tree data analysis, demonstrating the power of the new approach.