BACKGROUND: Salmonella spp. is a major foodborne pathogen with a wide variety of serovars associated with human cases and food sources. Nevertheless, in Europe a panel of ten serovars is responsible for up to 80% of confirmed human cases. Clustering studies by single nucleotide polymorphism (SNP) core-genome phylogenetic analysis of outbreaks due to these major serovars are simplified by the availability of many complete genomes in the free access databases. This is not the case for outbreaks due to less common serovars, such as Welikade, for which no reference genomes are available. In this study, we propose a method to solve this problem. We propose to perform a core genome MLST (cgMLST) analysis based on hierarchical clustering using the free-access EnteroBase to select the most suitable genome to use as a reference for SNP phylogenetic analysis. In this study, we applied this protocol to a retrospective analysis of a Salmonella enterica serovar Welikade (S. Welikade) foodborne outbreak that occurred in France in 2016. Finally, we compared the cgMLST and SNP analyses. SNP phylogenetic reconstruction was carried out considering the effect of recombination events identified by the ClonalFrameML tool. The accessory genome was also explored by phage content and virulome analyses.
RESULTS: Our findings revealed high clustering concordance using cgMLST and SNP analyses. Nevertheless, SNP analysis allowed for better assessment of the genetic distance among strains. The results revealed epidemic clones of S. Welikade circulating within the poultry and dairy sectors in France, responsible for sporadic and non-sporadic human cases between 2012 and 2019.
CONCLUSIONS: This study increases knowledge on this poorly described serovar and enriches public genome databases with 42 genomes from human and non-human S. Welikade strains, including the isolate collected in 1956 in Sri Lanka, which gave the name to this serovar. This is the first genomic analysis of an outbreak due to S. Welikade described to date.

The International Workshop on Genotoxicity Testing (IWGT) meets every four years to seek consensus on difficult or conflicting approaches to genotoxicity testing based upon experience, available data, and analysis techniques. At the 2017 IWGT meeting in Tokyo, one working group addressed the sensitivity and selectivity of the bacterial strains specified in the Organization for Economic Cooperation and Development (OECD) Test Guideline TG471 to recommend possible modification of the test guideline. Three questions were posed: (1) Although TA100 is derived from TA1535, does TA1535 detect any mutagens that are not detected by TA100? (2) Among the options of Salmonella TA1537, TA97 or TA97a, are these strains truly equivalent? (3) Because there is a choice to use one of either E. coli WP2 uvrA, E. coli WP2 uvrA pKM101, or Salmonella TA102, are these strains truly equivalent? To answer these questions, we analyzed published bacterial mutation data in multiple strains from large (>10,000 compound) databases from Leadscope and Lhasa Limited and anonymized data for 53 compounds tested in TA1535 and TA100 provided by a pharmaceutical company. Our analysis involved (1) defining criteria for determining selective responses when using different strains; (2) identifying compounds producing selective responses based upon author calls; (3) confirming selective responses by visually examining dose-response data and considering experimental conditions; (4) using statistical methods to quantify the responses; (5) performing limited additional direct-comparison testing; and (6) determining the chemical classes producing selective responses. We found that few mutagens would fail to be detected if the test battery did not include Salmonella strains TA1535 (8/1167), TA1537 (2/247), TA102 (4/46), and E. coli WP2 uvrA (2/21). Of the mutagens detected by the full TG471 strain battery, 93% were detected using only strains TA98 and TA100; consideration of results from in vitro genotoxicity assays that detect clastogenicity increased this to 99%.

H7N3 and H7N7 are highly pathogenic avian influenza (HPAI) viruses and have posed a great threat not only for the poultry industry but for the human health as well. H7N9, a low pathogenic avian influenza (LPAI) virus, is also highly pathogenic to humans, and there is a great concern that these H7 subtypes would acquire the ability to spread efficiently between humans, thereby becoming a pandemic threat. A vaccine candidate covering all the three subtypes must, therefore, be an integral part of any pandemic preparedness plan. To address this need, we constructed a consensus hemagglutinin (HA) sequence of H7N3, H7N7, and H7N9 based on the data available in the NCBI in early 2012-2015. This artificial sequence was then optimized for protein expression before being transformed into an attenuated auxotrophic mutant of Salmonella Typhimurium, JOL1863 strain. Immunizing chickens with JOL1863, delivered intramuscularly, nasally or orally, elicited efficient humoral and cell mediated immune responses, independently of the route of vaccination. Our results also showed that JOL1863 deliver efficient maturation signals to chicken monocyte derived dendritic cells (MoDCs) which were characterized by upregulation of costimulatory molecules and higher cytokine induction. Moreover, immunization with JOL1863 in chickens conferred a significant protection against the heterologous LPAI H7N1 virus challenge as indicated by reduced viral sheddings in the cloacal swabs. We conclude that this vaccine, based on a consensus HA, could induce broader spectrum of protection against divergent H7 influenza viruses and thus warrants further study.

All over the world, the incidence of Salmonella spp contamination on different food sources like broilers, clams and cow milk has increased rapidly in recent years. The multifaceted properties of Salomnella serovars allow the microorganism to grow and multiply in various food matrices, even under adverse conditions. Therefore, methods are needed to detect and trace this pathogen along the entire food supply network. In the present work, PFGE and ERIC-PCR were used to subtype 45 Salmonella isolates belonging to different serovars and derived from different food origins. Among these isolates, S. Enteritidis and S. Kentucky were found to be the most predominant serovars. The Discrimination Index obtained by ERIC-PCR (0.85) was slightly below the acceptable confidence value. The best discriminatory ability was observed when PFGE typing method was used alone (DI = 0.94) or combined with ERIC-PCR (DI = 0.93). A wide variety of profiles was observed between the different serovars using PFGE or/and ERIC-PCR. This diversity is particularly important when the sample origins are varied and even within the same sampling origin.

Since 2004, an increasing number of multidrug-resistant Salmonella serovar Concord infections have been isolated in Belgium among children adopted from Ethiopia. The patients or their family were interviewed and the isolates were subtyped. Between 2004 and 2009, a total of 39 Salmonella Concord infections were isolated from patients. Thirty-four isolates presented a multidrug resistance including resistance to extended-spectrum cephalosporins. Thirty-six cases involved children and 30 of these were adopted from Ethiopia. One case was due to contact with an adopted child and for the other 5 cases no direct epidemiological link with Ethiopia could be found, although four isolates displayed the same patterns observed on the adoptees\' isolates, strongly suggesting a phylogenetic relationship with the Ethiopian isolates. Our study confirmed the emergence in Europe of S. Concord isolates resistant to third-generation cephalosporin among Ethiopian adoptees. We have demonstrated that transmission (intra- and extra familial) can happen even if the frequency seems to be low. The presence and the transmission of such a multidrug-resistant Salmonella infection constitute a major concern, since such strains could jeopardize classical antibiotic therapy in patients at risk. This study provides useful information for parents adopting children and for their family practitioner.

Salmonella is one of the most important foodborne pathogens associated with severe diseases in animals and humans. Meat samples are considered as one of the main sources of Salmonella infections. Consequently, the survey of Salmonella contamination in meat samples is of outmost importance for the control and prevention of severe diseases. In this study, 250 meat samples were selected for surveys of Salmonella contaminations. Results indicated that 12% (n=30) of samples tested were positive to Salmonella. The genetic characterization of 30 Salmonella was studied by enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR), and 22 of ERIC-PCR types were found with D of 94.8%. In addition, the resistant characterization was also carried out using nine antibiotics test, and nine resistant patterns were observed with D of 88.7%. A good correlation was also observed between ERIC-PCR fingerprinting and resistant patterns in some Salmonella such as SAL 6 and SAL 7.

OBJECTIVE: To optimize the reaction conditions of enterobacteia repetitive intergenic consensus sequences-based PCR (ERIC-PCR), a molecular typing method of Salmonella, study the genomic DNA ERIC-PCR fingerprint maps of Salmonella standard strains and epidemic isolates, and supply data for a system of Salmonella epidemiology investigation and homology tracing.
METHODS: Genomic DNA of S. typhimurium was abstracted and used as the template for PCR. Enterobacteia repetitive intergenic consensus sequences were used as primers to amplify the target sequences in S. typhimurium genomic DNA: Amplification products were separated by agarose gel electrophoresis, and electrophoresis maps were analyzed by gel image analysis system. To optimize template concentration, Mg2+ concentration, primers concentration and annealing temperature of PCR, the factor to be optimized was designed in different concentration grads and other factors were fixed. Analyzed 16 Salmonella strains and one E. coli strain by PCR conditions optimized.
RESULTS: The electrophoresis bands of amplified products were entire and most clear when template concentration was 100 ng/25 microl, Mg2+ concentration was 2.0 mmol/L, primers concentrations were 0.4 micromol/L respectively in the total volume of 25 microl of the reaction system, and annealing temperature was 52 degrees C. The ERIC-PCR fingerprint maps of different Salmonella strains with different sources were different. From 250 to 5000 bp, there were 3 to 13 bands, and in those there was a specific 250 bp band.
CONCLUSIONS: Important reaction conditions of ERIC-PCR had been optimized in this study. ERIC-PCR technique could discriminate Salmonella strains isolated from different regions. It could be used in Salmonella epidemiology investigation and homology tracing, so as to make up for the flaws of the traditional classification methods for salmonella.

The dissemination of enteric contaminants (generic Escherichia coli and Salmonella) associated with pork carcasses and contact surfaces within a high capacity (6,000 carcasses per day) pork slaughter line was evaluated. Sponge samples were taken periodically from the holding area floor and carcasses at different points in the line throughout an 8.75 h production period. E. coli levels within the holding area were high (ca. 6 log cfu 100 cm(-2)) during the initial phase of processing and did not significantly increase throughout the activity period. In the course of dehairing carcasses, the levels of E. coli were significantly (p<0.05) reduced by scalding but increased during the scraping process. A combination of polishing and triple singeing reduced E. coli populations and the bacterium was only recovered sporadically on eviscerated carcasses. The E. coli populations associated with the slaughter line had a low diversity considering the large number of carcasses processed. In Visit I, the 665 E. coli isolates typed using ERIC-PCR could be grouped into 41 genotypes. In Visit II, 141 genotypes were identified among the 855 E. coli isolates tested. This would suggest that contamination on incoming pigs was of only minor significance compared to that present within the slaughterhouse environment. The holding area was shown to act as a reservoir for endemic E. coli genotypes that could be systematically transferred throughout the dressing line on carcasses. Indeed, the majority of genotypes could be re-isolated throughout the 8.75-h processing period. E. coli isolated from carcasses within the evisceration area could be traced to up-stream operations. The holding area and scraper operation were found to be the most important sites of cross-contamination. Fourteen genotypes recovered (primarily within the holding area) on Visit I were re-isolated on Visit II. Despite the presence of endemic E. coli populations, Salmonella was recovered from only two sites (holding area floor and a carcass within the cooler) on a single occasion. The two Salmonella recovered were genetically distinct (similarity index=22%) suggesting that they originated from different sources and were not part of an endemic population. The study has further illustrated the utility of molecular typing of generic E. coli isolates to establish the dynamics of enteric contamination within pork slaughter lines. However, the extent to which the distribution of E. coli can be extrapolated to that of Salmonella remains uncertain.

Decay rates in coastal water and sediment for the bacterial pathogens Salmonella typhimurium and S. derby were compared in laboratory-based microcosms with results previously obtained for a number of faecal indicators. In general, the decay rates of Salmonella spp. were greater than either enterococci or coliphage in overlying water and sediment. Decay rates of E. coli were similar to Salmonella spp. in overlying water, although greater in sediment. Raised temperature resulted in an increased decay rate for all organisms in the overlying water (and to a lesser extent in the surface sediment layer). It was demonstrated that decay rates for both S. typhimurium and S. derby were greater in overlying water compared with sediment. This suggested that sediments may be acting as a reservoir for pathogenic microorganisms released into the coastal environment during recreational activity and should be considered when estimating environmental exposure. Using measured decay rates and available dose-response data, a quantitative microbial risk assessment (QMRA) utilising Monte Carlo simulation was undertaken to estimate the risk of infection to Salmonella spp. following exposure to recreational coastal water subject to a range of faecal contamination levels. In waters of extremely poor quality, subject to contamination by faecal coliforms (10(6) CFU/100 mL), the maximum probability of infection on the day of an accidental release was above 2.0 x 10(-1) and remained above 1 x 10(-3) for three days following the initial high concentration.

Discrimination of 70 Salmonella strains previously studied by ribotyping was realized by RAPD and ERIC-PCR analysis. RAPD results on the 56 S. typhimurium isolates did not closely match those of ribotyping. With ERIC-PCR, two fingerprints only were obtained. For the 14 S. enteritidis strains, a helpful discrimination was obtained with RAPD analysis, while ERIC-PCR resulted in a single fingerprint.