Typhimurium Strains Research Articles

Comparative virulence of the worldwide ST19 and emergent ST213 genotypes of Salmonella enterica serotype Typhimurium strains isolated from food

Salmonella enterica Typhimurium represents one of the most frequent causal agents of food contamination associated to gastroenteritis. The sequence type ST19 is the founder and worldwide prevalent genotype within this serotype, but its replacement by emerging genotypes has been recently reported. Particularly, the ST213 genotype has replaced it as the most prevalent in clinical and contaminated food samples in Mexico and has been recently reported in several countries. In this study, the in vitro and in vivo virulence of ST213 and ST19 strains isolated from food samples in Mexico was evaluated. Three out of the five analyzed ST213 strains, showed a greater internalization capacity and increased secretion of interleukins IL-8 and IL-6 of Caco-2 cells than the ST19 strains. Microbiological counts in feces and tissues showed the ability of all strains tested to establish infection in the rat model. The ST213 strains also caused histopathological damage, characteristic of gastroenteritis in Wistar rats. In contrast to the in vitro result, one of the ST19 strains showed marked damage in the test animals. The ST213 genotype strains showed in vitro and in vivo virulence variability, but significantly higher than the observed in the ST19 genotype strains, thus such emergent genotype represents a public health concern.

Salmonella phage akira, infecting selected Salmonella enterica Enteritidis and Typhimurium strains, represents a new lineage of bacteriophages

Some serovars of Salmonella can cause life-threatening diarrhoeal diseases and bacteriemia. The emergence of multidrug-resistant strains has led to a need for alternative treatments such as phage therapy, which requires available, well-described, diverse, and suitable phages. Phage akira was found to lyse 19 out of 32 Salmonella enterica serovars and farm isolates tested, although plaque formation was observed with only two S. Enteritidis and one S. Typhimurium strain. Phage akira encodes anti-defence genes against type 1 R-M systems, is distinct (<65% nucleotide sequence identity) from related phages and has siphovirus morphology. We propose that akira represents a new genus in the class Caudoviricetes.

Phenotypic and molecular characterization of blaCTX-M-3 and blaCTX-M-55-producing monophasic Salmonella enterica serovar Typhimurium in Greece

To characterize three Salmonella enterica serovar Typhimurium strains using whole genome sequencing (WGS) and conventional methods. The isolates were recovered from three pediatric patients in Greece as part of the hospital's epidemiological surveillance system during 2016 to 2018. Bacterial identification and antimicrobial susceptibility testing was performed using the VITEK 2 automated system, disc diffusion test, and MIC gradient test while serotyping by the slide agglutination method. Detection of resistance genes, eBurst groups (eBG), assignment to sequence types, single nucleotide polymorphism (SNP) typing, location and characterization of drug resistance regions, and in silico plasmid detection were carried out using WGS. All strains were identified as S. Typhimurium-monophasic, ST34, eBG1 with antigenic formula 1,4, [5], 12:i:-. They were phenotypically resistant to most antibiotics tested except piperacillin/tazobactam, imipenem, and co-trimoxazole. WGS revealed the chromosomally located genes encoding the ASSuT (ampicillin, streptomycin, sulfonamides, and tetracycline) resistant pattern in all three strains. WGS revealed extended spectrum ß-lactamase (ESBL) production in all three strains, the presence of blaCTX-M-3 on an IncI1 plasmid in two strains isolated in 2018, and the chromosomally encoded blaCTX-M-55 plus qnrS1 (resistance to ciprofloxacin) in the strain isolated in 2016. The two strains from 2018 were isolated from the same hospital ward and were genetically related. The emergence of ESBL among S. 1,4,[5], 12:i:- is worrisome due to its increasing antimicrobial resistance phenotype, making clinical treatment difficult. WGS provides an alternative to traditional methods of identification and genomic characterisation of strains, and serves to better understand their epidemiological dynamics and bacterial pathogenesis.

Salmonella enterica serovar Typhimurium chitinases modulate the intestinal glycome and promote small intestinal invasion.

Salmonella enterica serovar Typhimurium (S. Typhimurium) is one of the leading causes of food-borne illnesses worldwide. To colonize the gastrointestinal tract, S. Typhimurium produces multiple virulence factors that facilitate cellular invasion. Chitinases have been recently emerging as virulence factors for various pathogenic bacterial species, and the S. Typhimurium genome contains two annotated chitinases: STM0018 (chiA) and STM0233. However, the role of these chitinases during S. Typhimurium pathogenesis is unknown. The putative chitinase STM0233 has not been studied previously, and only limited data exists on ChiA. Chitinases typically hydrolyze chitin polymers, which are absent in vertebrates. However, chiA expression was detected in infection models and purified ChiA cleaved carbohydrate subunits present on mammalian surface glycoproteins, indicating a role during pathogenesis. Here, we demonstrate that expression of chiA and STM0233 is upregulated in the mouse gut and that both chitinases facilitate epithelial cell adhesion and invasion. S. Typhimurium lacking both chitinases showed a 70% reduction in invasion of small intestinal epithelial cells in vitro. In a gastroenteritis mouse model, chitinase-deficient S. Typhimurium strains were also significantly attenuated in the invasion of small intestinal tissue. This reduced invasion resulted in significantly delayed S. Typhimurium dissemination to the spleen and the liver, but chitinases were not required for systemic survival. The invasion defect of the chitinase-deficient strain was rescued by the presence of wild-type S. Typhimurium, suggesting that chitinases are secreted. By analyzing N-linked glycans of small intestinal cells, we identified specific N-acetylglucosamine-containing glycans as potential extracellular targets of S. Typhimurium chitinases. This analysis also revealed a differential abundance of Lewis X/A-containing glycans that is likely a result of host cell modulation due to the detection of S. Typhimurium chitinases. Similar glycomic changes elicited by chitinase deficient strains indicate functional redundancy of the chitinases. Overall, our results demonstrate that S. Typhimurium chitinases contribute to intestinal adhesion and invasion through modulation of the host glycome.

Comparative Genomic Analysis of Salmonella enterica Serovar Typhimurium Isolates from Passerines Reveals Two Lineages Circulating in Europe, New Zealand, and the United States.

Salmonella enterica serovar Typhimurium strains from passerines have caused wild bird deaths and human salmonellosis outbreaks in Europe, Oceania, and North America. Here, we performed comparative genomic analysis to explore the emergence, genetic relationship, and evolution of geographically dispersed passerine isolates. We found that passerine isolates from Europe and the United States clustered to form two lineages (EU and US passerine lineages), which were distinct from major S. Typhimurium lineages circulating in other diverse hosts (e.g., humans, cattle, pigs, chickens, and other avian hosts, such as pigeons and ducks). Further, passerine isolates from New Zealand clustered to form a sublineage (NZ passerine lineage) of the US passerine lineage. We inferred that the passerine isolates mutated at a rate of 3.2 × 10-7 substitutions/site/year, and the US, EU, and NZ passerine lineages emerged in approximately 1952, 1970, and 1996, respectively. Isolates from the three lineages presented genetic similarity, such as lack of antimicrobial resistance genes and accumulation of the same virulence pseudogenes. In addition, genetic diversity due to microevolution existed in the three passerine lineages. Specifically, pseudogenization in the type 1 fimbrial gene fimC (deletion of G at position 87) was detected only in the US and NZ passerine isolates, while single-base deletions in type 3 secretion system effector genes (i.e., gogB, sseJ, and sseK2) cooccurred solely in the EU passerine isolates. These findings provide insights into the evolution, host adaptation, and epidemiology of S. Typhimurium in passerines. IMPORTANCE Passerine-associated S. Typhimurium strains have been linked to human salmonellosis outbreaks in recent years. Here, we investigated the phylogenetic relationship of globally distributed passerine isolates and profiled their genomic similarity and diversity. Our study reveals two passerine-associated S. Typhimurium lineages circulating in Europe, Oceania, and North America. Isolates from the two lineages presented phylogenetic and genetic signatures that were distinct from those of isolates from other hosts. The findings shed light on the host adaptation of S. Typhimurium in passerines and are important for source attribution of S. Typhimurium strains to avian hosts. Further, we found that S. Typhimurium definitive phage type 160 (DT160) from passerines, which caused decades-long human salmonellosis outbreaks in New Zealand and Australia, formed a sublineage of the US passerine lineage, suggesting that DT160 might have originated from passerines outside Oceania. Our study demonstrates the importance of whole-genome sequencing and genomic analysis of historical microbial collections to modern epidemiologic surveillance.

Increased Drug Resistance and Biofilm Formation Ability in ST34-Type Salmonella Typhimurium Exhibiting Multicellular Behavior in China

Salmonella Typhimurium is an important food-borne pathogen. In this paper, multicellular behavior and associated characteristics of S. Typhimurium isolated from human and animal source food were studied. All the S. Typhimurium strains exhibiting multicellular behavior (100%) belonged to the ST34 type. In addition, most of the ST34-type multicellular behavior S. Typhimurium strains had a human origin (69.11%) and 98% of the ST34-type multicellular behavior strains exhibited strong biofilm formation capacity, which was much higher than that of non-multicellular behavior strains (7%, P < 0.01). Antibiotic resistance in ST34-type multicellular behavior strains was significantly higher than in strains with non-multicellular behavior for most conventional drugs (P < 0.05); notably, Polymyxin B (8%) and Imipenem (1%) resistances were also observed in the ST34-type strains. Furthermore, all the ST34-type multicellular behavior strains (100%) exhibited Multiple Drug Resistance (resistance to ≥3antibiotics), which was much higher than that of the non-multicellular behavior strains (P < 0.05). Consistent with the drug-resistant phenotype, the carrying rates of most drug-resistant genes in ST34-type multicellular behavior strains were higher than that those in non-multicellular behavior strains (P < 0.05). Therefore, this study revealed the emergence of a prevalent ST34-type multicellular behavior S. Typhimurium strains with increased biofilm formation ability and drug resistance rate, which poses a threat to public health safety, and highlights the need for comprehensive monitoring of the strains.

Genetic Characterization of mcr-1-Positive Multidrug-Resistant Salmonella enterica Serotype Typhimurium Isolated From Intestinal Infection in Children and Pork Offal in China.

With the rapid emergence of plasmid-mediated colistin resistance gene mcr-1, the increased resistance of Salmonella has attracted extensive attention. This study reports on 11 multidrug-resistant Salmonella enterica serovar Typhimurium strains harboring mcr-1 in China. They all presented resistance to colistin, and additionally, one that was isolated from a child’s stool sample was also resistant to ceftriaxone and azithromycin. We screened 1454 strains of Salmonella for mcr-1 gene through PCR, and these strains are all preserved in our laboratory. Antimicrobial sensitivity analysis was carried out for the screened mcr-1 positive strains. Genetic polymorphism analysis of S. Typhimurium was performed by using the Pulsed-Field Gel Electrophoresis (PFGE). The plasmids harboring mcr-1 were identified by S1-PFGE and southern blotting. Plasmid conjugation assays were used to analyze the transferability of colistin resistance. The plasmids harboring mcr-1 were characterized by sequencing and bioinformatic analysis. Eleven S. Typhimurium strains harboring mcr-1 with colistin resistance (MICs 4μg/ml) were detected, which were isolated from children and pig offal in China. All of them were multidrug-resistant strains. PFGE results revealed that the strains isolated from different samples or locations have identical genotypes. S1-PFGE and southern blotting experiments showed that three plasmids of different sizes (33, 60, and 250 kb) all carried the mcr-1 gene. The plasmid conjugation assays revealed that Salmonella acquired mcr-1 harboring plasmids by horizontal transfer. Sequencing and plasmid type analysis revealed that these plasmids were types IncX4, IncI2, and IncHI2. Among them, IncX4 and IncI2 plasmids had extremely similar backbones and contained one resistant gene mcr-1. IncHI2 plasmid contained multiple resistant genes including blaCTX–M, oqxB, sul, aph, aadA, and blaTEM. We identified 11 mcr-1 harboring S. Typhimurium strains in China and described their characteristics. Our findings indicate that the mcr-1 gene can effectively spread among intestinal bacteria by horizontal transfer of three types of plasmids. Moreover, the IncHI2 plasmid can also mediate the transfer of other drug resistance genes. These results reveal that constant surveillance of mcr-1 harboring S Typhimurium is imperative to prevent the spread of colistin resistance.

Salmonella Typhimurium fepB negatively regulates C. elegans behavioral plasticity

ObjectivesDauer is an alternative developmental stage of Caenorhabditis elegans (C. elegans) that gives survival benefits under unfavorable environmental conditions. Our study aims to decipher C. elegans dauer larvae development upon Salmonella Typhimurium infection and how the bacterial gene regulating the worm's behavioural plasticity for better survival. MethodsAge-synchronized L4 C. elegans worms were infected with Salmonella Typhimurium 14028s (WT-STM) strain and mutant strains to check the dauer larvae development using 1% SDS. Besides, bacterial load in animals’ gut, pharyngeal pumping rate and viability were checked. Worm's immune genes (e.g., ilys-3, lys-7, pmk-1, abf-2, clec-60) and dauer regulatory genes (e.g., daf-7, daf-11, daf-12, daf-16, daf-3) were checked by performing qRT-PCR under infection conditions. ResultsWe found that deletion of the fepB gene in S. Typhimurium strain became less pathogenic with reduced flagellar motility and biofilm-forming ability. Besides, there was decreased bacterial burden in the worm's gut with no damage to their pharynx. The fepB mutant strain was also able to enhance the immune responses for better survival of worms. Infection with mutant strain could activate dauer signaling via the TGF-β pathway leading to a significant increase in dauer formation than WT-STM infection. ConclusionOur study indicated that the bacteria act as a food source for the growth of C. elegans and development and can act as a signal that might be playing an essential role in regulating the host physiology for their survival. Such a study can help us in understanding the complex host-pathogen interaction benefiting pathogen in host dissemination.

Characterization of Salmonella Typhimurium and monophasic Salmonella Typhimurium isolated in Abruzzo and Molise regions, Italy, from 2012 to 2017.

Salmonellosis is currently the second most common zoonosis in European Union but in the 6-years periods, between 2012 and 2017, there has been a significant decrease trend in the yearly number of infections caused by Salmonella. In Italy, S. Typhimurium and monophasic S. Typhimurium represent the most prevalent serotypes. In this paper, we investigated these two serovars isolated from 2012 to 2017 in Abruzzo and Molise regions. A set of 345 strains isolated from human sporadic cases, surface water, food and animals were collected and analyzed. Monophasic S. Typhimurium strains were found to be resistant to streptomycin, sulfisoxazole, ampicillin, tetracycline and nalidixic acid, while S. Typhimurium isolates showed high levels of resistance to tetracycline, sulfisoxazole and ampicillin. The 5-loci Multilocus Variable-Number Tandem Repeat Analysis (MLVA) identified 88 genotypes (GT), six of which were common for the two serovars. Several MLVA profiles were shared by human and non-human isolates. MLVA had sufficient typing resolution for epidemiological studies of S. Typhimurium but demonstrated poor discriminatory in trace-back study of monophasic S. Typhimurium.

Lactiplantibacillus plantarum 1201 Inhibits Intestinal Infection of Salmonella enterica subsp. enterica Serovar Typhimurium Strain ATCC 13311 in Mice with High-Fat Diet.

Salmonella Typhimurium is widely distributed in food. It can colonise the gastrointestinal tract after ingestion, causing lamina propria edema, inflammatory cell infiltration, and mucosal epithelial decomposition. A high-fat diet (HFD) can induce an inflammatory response, but whether HFD can increase the infection level of S. Typhimurium is unknown. We established a model of Salmonella enterica subsp. enterica serovar Typhimurium strain ATCC 13311 ATCC 13311 infection in healthy adult mice with a maintenance diet (MD) or HFD to explore the effect of Lactiplantibacillus plantarum 1201 intervention on S. Typhimurium ATCC 13311 colonization and its protective effects on mice. HFD exacerbated the infection of S. Typhimurium ATCC 13311, while the intervention of L. plantarum 1201 effectively mitigated this process. L. plantarum 1201 can reduce the colonies of S. ATCC 13311 in the intestines and tissues; and reduce intestinal inflammation by down-regulating the level of TLR4/NF-κB pathway related proteins in serum and the expression of related inflammatory factors in the colon and jejunum. Since L. plantarum 1201 can inhibit the colonization of S. Typhimurium ATCC 13311 and relieve inflammation in HFD, current research may support the use of L. plantarum 1201 to prevent S. Typhimurium infection.

Optimized Attenuated Salmonella Typhimurium Suppressed Tumor Growth and Improved Survival in Mice.

The gram-negative facultative anaerobic bacteria Salmonella enterica serovar Typhimurium (hereafter S. Typhimurium) has always been considered as one candidate of anti-tumor agents or vectors for delivering drug molecules. In this study, we compared several widely studied S. Typhimurium strains in their anti-tumor properties aiming to screen out the best one for further optimization and use in cancer therapy. In terms of the motility, virulence and anti-tumor efficacy, the three strains 14028, SL1344, and UK-1 were similar and obviously better than LT-2, and UK-1 showed the best phenotypes among them. Therefore, the strain UK-1 (D) was selected for the following studies. Its auxotrophic mutant strain (D1) harboring ∆aroA and ∆purM mutations was further optimized through the modification of lipid A structure, generating a new strain named D2 with stronger immunostimulatory activity. Finally, the ∆asd derivative of D2 was utilized as one live vector to deliver anti-tumor molecules including the angiogenesis inhibitor endostatin and apoptosis inducer TRAIL and the therapeutic and toxic-side effects were evaluated in mouse models of colon carcinoma and melanoma. After intraperitoneal infection, engineered Salmonella bacteria equipped with endostatin and/or TRAIL significantly suppressed the tumor growth and prolonged survival of tumor-bearing mice compared to PBS or bacteria carrying the empty plasmid. Consistently, immunohistochemical studies confirmed the colonization of Salmonella bacteria and the expression of anti-tumor molecules inside tumor tissue, which were accompanied by the increase of cell apoptosis and suppression of tumor angiogenesis. These results demonstrated that the beneficial anti-tumor efficacy of attenuated S. Typhimurium bacteria could be improved through delivery of drug molecules with powerful anti-tumor activities.

A Novel Dibenzoxazepine Attenuates Intracellular Salmonella Typhimurium Oxidative Stress Resistance.

ABSTRACTSalmonella enterica serovar Typhimurium is the leading cause of invasive nontyphoidal salmonellosis. Additionally, the emergence of multidrug-resistant S. Typhimurium has further increased the difficulty of controlling its infection. Previously, we showed that an antipsychotic drug, loxapine, suppressed intracellular Salmonella in macrophages. To exploit loxapine’s antibacterial activity, we simultaneously evaluated the anti-intracellular Salmonella activity and cytotoxicity of newly synthesized loxapine derivatives using an image-based high-content assay. We identified that SW14 exhibits potent suppressive effects on intramacrophagic S. Typhimurium with an 50% effective concentration (EC50) of 0.5 μM. SW14 also sensitized intracellular Salmonella to ciprofloxacin and cefixime and effectively controlled intracellular multidrug- and fluoroquinolone-resistant S. Typhimurium strains. However, SW14 did not affect bacterial growth in standard microbiological broth or minimal medium that mimics the phagosomal environment. Cellular autophagy blockade by 3-methyladenine (3-MA) or shATG7 elevated the susceptibility of intracellular Salmonella to SW14. Finally, reactive oxygen species (ROS) scavengers reduced the antibacterial efficacy of SW14, but the ROS levels in SW14-treated macrophages were not elevated. SW14 decreased the resistance of outer membrane-compromised S. Typhimurium to H2O2. Collectively, our data indicated that the structure of loxapine can be further optimized to develop new antibacterial agents by targeting bacterial resistance to host oxidative-stress defense.IMPORTANCE The incidence of diseases caused by pathogenic bacteria with resistance to common antibiotics is consistently increasing. In addition, Gram-negative bacteria are particularly difficult to treat with antibiotics, especially those that can invade and proliferate intracellularly. In order to find a new antibacterial compound against intracellular Salmonella, we established a cell-based high-content assay and identified SW14 from the derivatives of the antipsychotic drug loxapine. Our data indicate that SW14 has no effect on free bacteria in the medium but can suppress the intracellular proliferation of multidrug-resistant (MDR) S. Typhimurium in macrophages. We also found that SW14 can suppress the resistance of outer membrane compromised Salmonella to H2O2, and its anti-intracellular Salmonella activity can be reversed by reactive oxygen species (ROS) scavengers. Together, the findings suggest that SW14 might act via a virulence-targeted mechanism and that its structure has the potential to be further developed as a new therapeutic against MDR Salmonella.

Holistic Characterization of a Salmonella Typhimurium Infection Model Using Integrated Molecular Imaging.

A more complete and holistic view on host-microbe interactions is needed to understand the physiological and cellular barriers that affect the efficacy of drug treatments and allow the discovery and development of new therapeutics. Here, we developed a multimodal imaging approach combining histopathology with mass spectrometry imaging (MSI) and same section imaging mass cytometry (IMC) to study the effects of Salmonella Typhimurium infection in the liver of a mouse model using the S. Typhimurium strains SL3261 and SL1344. This approach enables correlation of tissue morphology and specific cell phenotypes with molecular images of tissue metabolism. IMC revealed a marked increase in immune cell markers and localization in immune aggregates in infected tissues. A correlative computational method (network analysis) was deployed to find metabolic features associated with infection and revealed metabolic clusters of acetyl carnitines, as well as phosphatidylcholine and phosphatidylethanolamine plasmalogen species, which could be associated with pro-inflammatory immune cell types. By developing an IMC marker for the detection of Salmonella LPS, we were further able to identify and characterize those cell types which contained S. Typhimurium.

CD4+ T cell immunity to Salmonella is transient in the circulation.

While Salmonella enterica is seen as an archetypal facultative intracellular bacterial pathogen where protection is mediated by CD4+ T cells, identifying circulating protective cells has proved very difficult, inhibiting steps to identify key antigen specificities. Exploiting a mouse model of vaccination, we show that the spleens of C57BL/6 mice vaccinated with live-attenuated Salmonella serovar Typhimurium (S. Typhimurium) strains carried a pool of IFN-γ+ CD4+ T cells that could adoptively transfer protection, but only transiently. Circulating Salmonella-reactive CD4+ T cells expressed the liver-homing chemokine receptor CXCR6, accumulated over time in the liver and assumed phenotypic characteristics associated with tissue-associated T cells. Liver memory CD4+ T cells showed TCR selection bias and their accumulation in the liver could be inhibited by blocking CXCL16. These data showed that the circulation of CD4+ T cells mediating immunity to Salmonella is limited to a brief window after which Salmonella-specific CD4+ T cells migrate to peripheral tissues. Our observations highlight the importance of triggering tissue-specific immunity against systemic infections.

Chitosan propolis nanocomposite alone or in combination with apramycin: an alternative therapy for multidrug-resistant Salmonella Typhimurium in rabbits: in vitro and in vivo study

Introduction. The emergence of multidrug-resistant Salmonella Typhimurium strains has increased the need for safe, alternative therapies from natural sources with antibacterial properties.Hypothesis/Gap Statement. There are no published data regarding the use of chitosan propolis nanocomposite (CPNP) either alone or in combination with antibiotics as antimicrobials against S. Typhimurium, especially in Egypt.Aim. This study evaluated the antibacterial activities of five antimicrobials [apramycin, propolis, chitosan nanoparticles (CNPs), chitosan propolis nanocomposite (CPNP) and CPNP +apramycin] against ten virulent and multidrug-resistant (MDR) S. Typhimurium field strains recovered from diarrheic rabbits through in vitro and in vivo study.Methodology. The expression levels of three virulence genes of S. Typhimurium strains were determined by quantitative reverse-transcription PCR (RT-qPCR) after exposure to sub-inhibitory concentrations of apramycin, propolis, CNPs, CPNP alone, and CPNP +apramycin. Additionally, 90 New Zealand rabbits were divided into control and experimentally S. Typhimurium-infected groups. The infected rabbits were orally administered saline solution (infected-untreated); 10 mg apramycin/kg (infected-apramycin-treated); 50 mg propolis/kg (infected-propolis-treated); 15 mg CPNP/kg (infected-CPNP-treated) and 15 mg CPNP +10 mg apramycin/kg (infected-CPNP +apramycin-treated) for 5 days.Results. The RT-qPCR analysis revealed different degrees of downregulation of all screened genes. Furthermore, the treatment of infected rabbits with CPNP or CPNP +apramycin significantly improved performance parameters, and total bacterial and Salmonella species counts, while also modulating both oxidative stress and altered liver and kidney parameters.Conclusion. This work demonstrates the use of CPNP alone or in combination with apramycin in the treatment of S. Typhimurium in rabbits.

Mixed Salmonella cultures reveal competitive advantages between strains during pre‐enrichment and selective enrichment

AbstractCulture‐based Salmonella isolation consists of nonselective pre‐enrichment, followed by selective enrichment in Rappaport–Vassiliadis (RV) or tetrathionate (TT) broths, and subsequent plating on selective indicator agar. This study aimed to assess the recovery of two strains belonging to serovars Montevideo (strain ATCC‐8387) and Typhimurium (strain ATCC‐14028) when grown together in different media. The two strains were co‐inoculated 1:1, 10:1, 100:1, and the reciprocal, and CRISPR‐SeroSeq was used to assess the relative frequency of both strains after each culture step. Individually, there was no growth difference between both strains in universal pre‐enrichment (UP) or RV broths, though both strains had a higher growth in the former. In TT, both strains had reduced growth, especially ser. Montevideo‐8387. When the strains were combined, the growth of ser. Typhimurium‐14028 strain was higher in UP and TT broths, while the ser. Montevideo strain was higher in RV broth. The ser. Typhimurium strain also grew better on xylose lysine tergitol‐4 (XLT‐4) agar. Using media that are commonly used for Salmonella isolation, this work reveals strain‐to‐strain differences in growth and that in three conditions (UP, RV, XLT‐4), these differences were only manifested when the strains were in competition with each other. These findings illustrate the importance of dual selective enrichments to be able to capture all Salmonella present in a sample.

Genomic population structure associated with repeated escape of Salmonella enterica ATCC14028s from the laboratory into nature.

Salmonella enterica serovar Typhimurium strain ATCC14028s is commercially available from multiple national type culture collections, and has been widely used since 1960 for quality control of growth media and experiments on fitness ("laboratory evolution"). ATCC14028s has been implicated in multiple cross-contaminations in the laboratory, and has also caused multiple laboratory infections and one known attempt at bioterrorism. According to hierarchical clustering of 3002 core gene sequences, ATCC14028s belongs to HierCC cluster HC20_373 in which most internal branch lengths are only one to three SNPs long. Many natural Typhimurium isolates from humans, domesticated animals and the environment also belong to HC20_373, and their core genomes are almost indistinguishable from those of laboratory strains. These natural isolates have infected humans in Ireland and Taiwan for decades, and are common in the British Isles as well as the Americas. The isolation history of some of the natural isolates confirms the conclusion that they do not represent recent contamination by the laboratory strain, and 10% carry plasmids or bacteriophages which have been acquired in nature by HGT from unrelated bacteria. We propose that ATCC14028s has repeatedly escaped from the laboratory environment into nature via laboratory accidents or infections, but the escaped micro-lineages have only a limited life span. As a result, there is a genetic gap separating HC20_373 from its closest natural relatives due to a divergence between them in the late 19th century followed by repeated extinction events of escaped HC20_373.

Phylogenetic relationship and genomic characterization of Salmonella Typhimurium strains isolated from swine in Brazil.

Salmonella Typhimurium has been transmitted between humans and animals. Although, Brazil has been one of the largest pork meat exporters worldwide, there are few studies that characterized epidemiologically S. Typhimurium strains from swine. The aims of this work were to study the phylogenetic relationship of S. Typhimurium genomes isolated from swine in Brazil among themselves and with other genomes isolated from several sources and countries using wgMLST and cgMLST and to perform the search of Salmonella pathogenicity islands (SPIs). In addition, for S. Typhimurium strains from swine to compare the virulence and antimicrobial resistance genes by VFDB and ResFinder, genetic content by BLAST Atlas and orthologous proteins clusters by OrthoVenn. The constructed phylogenetic trees by wgMLST and cgMLST grouped the majority (92.3% and 80.7%, respectively) of the strains isolated from swine in Brazil into the same group. All the isolates contained important SPIs (SPI-1, SPI-2, SPI-3, SPI-5 and SPI-9). A total of 100 and 31 virulence and resistance genes were detected in the S. Typhimurium strains isolated from swine, respectively. The BLAST Atlas and orthologous proteins analysis found regions of phages and differences in metabolic, regulatory and cellular processes among S. Typhimurium LT2 and S. Typhimurium isolates from swine. In conclusion, molecular typing based in the wgMLST and cgMLST suggested that the S. Typhimurium isolates from swine studied were genetically related. The pathogenic potential of the strains studied was corroborated by the presence of important SPIs and virulence genes. The high number of antimicrobial resistance genes detected is worrying and reinforced their potential risk in swine in Brazil. The comparison by BLAST Atlas suggested differences in mobile genetic elements among S. Typhimurium LT2 and S. Typhimurium isolates from swine in Brazil. The orthologous proteins analysis revealed unique genes related to important cellular processes in the strains from swine.

Genomic and phenotypic comparison of two Salmonella Typhimurium strains responsible for consecutive salmonellosis outbreaks in New Zealand

Salmonella enterica serovar Typhimurium DT160 was the predominant cause of notified human salmonellosis cases in New Zealand from 2000 to 2010, before it was superseded by another S. Typhimurium strain, DT56 variant (DT56v). Whole genome sequencing and phenotypic testing were used to compare 109 DT160 isolates with eight DT56v isolates from New Zealand animal and human sources. Phylogenetic analysis provided evidence that DT160 and DT56v strains were distantly related with an estimated date of common ancestor between 1769 and 1821. The strains replicated at different rates but had similar antimicrobial susceptibility profiles. Both strains were resistant to the phage expressed from the chromosome of the other strain, which may have contributed to the emergence of DT56v. DT160 contained the pSLT virulence plasmid, and the sseJ and sseK2 genes that may have contributed to the higher reported prevalence compared to DT56v. A linear pBSSB1-family plasmid was also found in one of the DT56v isolates, but there was no evidence that this plasmid affected bacterial replication or antimicrobial susceptibility. One of the DT56v isolates was also sequenced using long-read technology and found to contain an uncommon chromosome arrangement for a Typhimurium isolate. This study demonstrates how comparative genomics and phenotypic testing can help identify strain-specific elements and factors that may have influenced the emergence and supersession of bacterial strains of public health importance.

English

The emergence of antimicrobial resistance is a growing public health issue worldwide due to extensive use in agriculture, food, and veterinary medicine. The rate of antimicrobial resistance varies with different antibiotics and serotypes but Salmonella enterica serovar Enteritidis is one of the most widespread serotypes which is comparatively more susceptible to antimicrobial resistance followed by S. Typhimurium. A total of 71 Salmonella strains (S. Typhimurium, n=45; S. Enteritidis, n=26) isolated from humans, food and the environment were used in the study. Both isolates were confirmed through PCR by targeting their specific spy and sdf genes respectively. Kirby-Bauer disc diffusion method was used to determine antimicrobial susceptibility against 10 antibiotics used. An exponentially high level of resistance was found in S. Typhimurium strains. The highest level of resistance was found against amoxicillin (97.78%) followed by tetracycline (95.56%), gentamicin (93.33%), trimethoprim (86.67%), streptomycin (84.44%), nalidixic acid (77.78%), sulphafurazole (64.44%), ampicillin (62.22%), chloramphenicol (46.67%) and least resistance was found against ciprofloxacin (31.11%). Four penta MDR (ACSSuT) and two tetra MDR (ASSuT) resistance patterns were found S. Typhimurium strains. In S. Enteritidis strains, a high level of resistance was found against sulphonamides and streptomycin (92.31%) with the least resistance against ciprofloxacin (11.54%). Two penta MDR (ACSSuT) and six tetra MDR (ASSuT) resistance patterns were found. The presence of high antimicrobial resistance in zoonotic S. Typhimurium and Enteritidis in the broiler meat production chain is alarming. Immediate action and appropriate measures are required to control over the counter and irrational use of antibiotics both in poultry and humans. © 2021 Friends Science Publishers