A multiplex real-time PCR assay targeting virulence and resistance genes in Salmonella enterica serotype Typhimurium

<p>Abstract</p> <p>Background</p> <p>Typhimurium is the main serotype of <it>Salmonella enterica </it>subsp. <it>enterica </it>implicated in food-borne diseases worldwide. This study aimed to detect the prevalence of ten markers combined in a macro-array based on multiplex real-time PCR. We targeted characteristic determinants located on pathogenicity islands (SPI-2 to -5, virulence plasmid <it>pSLT </it>and <it>Salmonella </it>genomic island 1 (SGI1)) as well as a specific 16S-23S rRNA intergenic spacer sequence of definitive type 104 (DT104). To investigate antimicrobial resistance, the study also targeted the presence of genes involved in sulfonamide (<it>sul1</it>) and beta-lactam (<it>bla</it>_TEM) resistance. Finally, the <it>intI1 </it>determinant encoding integrase from class 1 integron was also investigated.</p> <p>Results</p> <p>A total of 538 unrelated <it>S</it>. Typhimurium strains isolated between 1999 and 2009 from various sources, including food animals, food products, human and environmental samples were studied. Based on the combined presence or absence of these markers, we distinguished 34 different genotypes, including three major genotypes encountered in 75% of the studied strains, Although SPI determinants were almost always detected, SGI1, <it>intI1</it>, <it>sul1 </it>and <it>bla</it>_TEMdeterminants were found 47%, 52%, 54% and 12% of the time respectively, varying according to isolation source. Low-marker patterns were most often detected in poultry sources whereas full-marker patterns were observed in pig, cattle and human sources.</p> <p>Conclusion</p> <p>The GeneDisc^®assay developed in this study madeit easier to explore variability within serotype Typhimurium by analyzing ten relevant gene determinants in a large collection of strains. This real-time multiplex method constitutes a valuable tool for strains characterization on epidemiological purposes.</p

Antimicrobial susceptibility of S. aureus strains isolated from a ham plant in France

As MRSA (methicillin resistant Staphylococcus aureus) were recently isolated from pigs in the Netherlands (1), we were incited to evaluate the resistance gene circulation inside S. aureus strains isolated from swine food products. A study, build in 2002, established the biodiversity of S. aureus isolates originated from a single raw ham industrial unit located in western France. Biotypes and pulsotypes from 106 isolates from raw material ham, end product and plant environment were determined. Isolates split up into 7 biotypes and about forty pulsotypes, which delineated a total of 23 sub-groups with 80% of homology and indicated the presence of numerous S. aureus strains in the plant

2005 French Salmonella Network data on antimicrobial resistance in the swine channels

The Salmonella Network is gathering, on a voluntary participation scheme, from approximately 150 public and private laboratories dissemmated throughout France, Salmonella strains and/or epidemiological information. Those non-human Salmonella strains are isolated either from animal health and production or food, feed and the environment sectors. Thus, in 2005, a total of 527 isolations from the swine channels were reported. The top 5 prevalent serotypes were : Typhimurium, Derby, Manhattan, lnfantis and Kedougou. Two-hundred and ninety five strains were received at the laboratory and, after double clearance, 185 strains were tested for their antimicrobial resistance against 16 antibiotics by the disk diffusion method. Twenty-one strains were associated to the animal health and production sector and 164 to the food sector

A multi-country One Health foodborne outbreak simulation exercise: cross-sectoral cooperation, data sharing and communication

IntroductionThe awareness of scientists and policy makers regarding the requirement for an integrated One Health (OH) approach in responding to zoonoses has increased in recent years. However, there remains an overall inertia in relation to the implementation of practical cross-sector collaborations. Foodborne outbreaks of zoonotic diseases continue to affect the European population despite stringent regulations, evidencing the requirement for better ‘prevent, detect and response’ strategies. Response exercises play an essential role in the improvement of crisis management plans, providing the opportunity to test practical intervention methodologies in a controlled environment.MethodsThe One Health European Joint Programme simulation exercise (OHEJP SimEx) aimed at practicing the OH capacity and interoperability across public health, animal health and food safety sectors in a challenging outbreak scenario. The OHEJP SimEx was delivered through a sequence of scripts covering the different stages of a Salmonella outbreak investigation at a national level, involving both the human food chain and the raw pet feed industry.ResultsA total of 255 participants from 11 European countries (Belgium, Denmark, Estonia, Finland, France, Italy, Norway, Poland, Portugal, Sweden, the Netherlands) took part in national level two-day exercises during 2022. National evaluations identified common recommendations to countries aiming to improve their OH structure to establish formal communication channels between sectors, implement a common data sharing platform, harmonize laboratory procedures, and reinforce inter-laboratory networks within countries. The large proportion of participants (94%) indicated significant interest in pursuing a OH approach and desire to work more closely with other sectors.DiscussionThe OHEJP SimEx outcomes will assist policy makers in implementing a harmonized approach to cross-sector health-related topics, by highlighting the benefits of cooperation, identifying gaps in the current strategies and suggesting actions required to better address foodborne outbreaks. Furthermore, we summarize recommendations for future OH simulation exercises, which are essential to continually test, challenge and improve national OH strategies

Prevalence of low-virulence Listeria monocytogenes strains from different foods and environments

International audienceVarious studies have demonstrated variations in the levels of virulence of different L monocytogenes strains. In our laboratory, a plaque-forming assay followed by subcutaneous footpad inoculation of mice enabled us to estimate the prevalence of the low-virulence strains. This value fell from 16.3% to 1.7% with bacteria collected before 1994 and after 1997 respectively. This could be related to the modification in 1997 of the reference method EN ISO 11 290-1 of Listeria detection which recommended the use of polymyxin-acriflavine-LiCl-ceftazidime-aesculin-mannitol (PALCAM) medium. The aim of this study was to determine whether the percentage of low-virulence strains detected has changed due to the modification of the detection method recommending the use of the ALOA medium. After analyzing 380 L monocytogenes strains, no increase in the percentage of low-virulence strains could be detected. The prevalence reached only 2.6% (ten of the 380 strains tested). The low virulence of L monocytogenes strains was not related to rare serotypes and was also observed in serotypes usually involved in human disease. Low-virulence strains were found in dairy, meat, ready-to-eat products and also in the environment. highlighting the absence of one specific source. These results are discussed in terms of detection methods and the definition of low virulence. (C) 2009 Elsevier B.V. All rights reserved

Antimicrobial susceptibility of S. aureus strains isolated from a ham plant in France

As MRSA (methicillin resistant Staphylococcus aureus) were recently isolated from pigs in the Netherlands (1), we were incited to evaluate the resistance gene circulation inside S. aureus strains isolated from swine food products. A study, build in 2002, established the biodiversity of S. aureus isolates originated from a single raw ham industrial unit located in western France. Biotypes and pulsotypes from 106 isolates from raw material ham, end product and plant environment were determined. Isolates split up into 7 biotypes and about forty pulsotypes, which delineated a total of 23 sub-groups with 80% of homology and indicated the presence of numerous S. aureus strains in the plant.</p

Characterization of Staphylococcus aureus strains associated with food poisoning outbreaks in France

International audienceEnterotoxins produced by Staphylococcus aureus are responsible for staphylococcal food-poisoning outbreaks (SFPO). In France, SFPO are the second cause of food-borne diseases after Salmonella. However, very little is known about the strains involved. The objective of this study was to characterize the staphylococcal strains related to these SFPO through phenotypic and genotypic analyses. A total of 178 coagulase-positive staphylococcal isolates recovered from 31 SFPO (1981-2002) were screened through biotyping. Thirty-three strains representative of the different biotypes in each SFPO were further examined for SmaI macrorestriction-type, phage-type, resistance to various antimicrobial drugs, presence of staphylococcal enterotoxin (se) genes sea to sei, and production of enterotoxins SEA to SED. All these 33 strains were identified as S. aureus species: 27 were of human biotypes and six ovine or non-host-specific biotypes. Most (74.1%) strains reacted with group III phages. Eleven strains were resistant to at least two classes of antibiotics and among them, two were resistant to methicillin. Twenty-nine strains carried one or several of the eight se genes tested; the gene sea was most common (n=23), and often linked to sed (n=12) or seh (n=5). The novel se genes seg-i were in all cases associated with se genes sea to sed except for one strain which carried only seg and sei. Pulsed-Field Gel Electrophoresis (PFGE) of SmaI macrorestriction digests of the 33 strains discriminated 32 PFGE patterns grouped into nine biotype-specific clusters. All five strains carrying sea and seh were grouped together into the same sub-cluster. Three of the four se-gene-negative strains were in one PFGE cluster: all four should be tested for se genes not included in this study and, if negative, be further investigated for the presence of unidentified SEs

NDM-1-producing Vibrio parahaemolyticus isolated from imported seafood

International audienc