Multidrug-resistant Serovar Research Articles

Biocontrol characteristics and application of phage SEP4 against multidrug-resistant Salmonella biofilm on food matrix

The emergence of antibiotic resistance and biofilm formation among Salmonella strains, especially in chicken meat, poses substantial risks to economic and public health. To mitigate these challenges, we explored interventions and biofilm inhibition strategies against multidrug-resistant serovars (B, C1, and D groups), by Salmonella phage SEP4, an isolate with broad host-spectrum. The morphological and genomic analyses classified it as a member of the Siphoviridae family with an icosahedral head of 124.56 ± 8.9 nm diameter and 66.28 ± 2.01 nm tail length, respectively. Phage SEP4 displayed robust survivability over a pH range of 3–10 and temperatures from 30 °C to 60 °C. Phage adsorption peaked at 73.20% in 15 min, with a burst size of 119 PFU/CFU at optimal MOI-1. Moreover, a linear double-stranded DNA genome of 41,828 bp, characterized by 49.84% GC content and putative 57 open reading frames (ORFs), and lacked antibiotic or virulence factor genes. Anti-Salmonella phage efficacy demonstrated a significant reduction in viable cell counts at 25 °C compared to 4 °C against tested MDR strains in food models. The bactericidal activity of phage SEP4 ranged from 14% to 46% and 25%–33.8% against host at MOI-1 and 100, while 22%–37% for other MDR strains. Simultaneously, SEP4 inhibited the growth of matured biofilms on lettuce leaves and chicken surfaces, achieving complete reduction within 4–6 h. SEM images confirmed the drastic destruction of cohesive biofilm architectures by phage treatment. The findings show that phage SEP4 has promising potential as a biocontrol agent, ensuring food safety by controlling MDR Salmonella contaminations and associated biofilms.

Intrinsic insights to antimicrobial effects of Nitrofurantoin to multi drug resistant Salmonella enterica serovar Typhimurium ms202

Emerging multidrug resistant (MDR) serovar of Salmonella has raised the concern of their impactful effect on pathogenic infection and mortality in human lead by the enteric diseases. In order to combat the battle against these MDR Salmonella pathogen, new drug molecules need to be evaluated for their potent antibacterial application. This study evaluates the mechanistic antimicrobial effect of nitrofurantoin against a MDR strain of Salmonella named S. enterica Typhimurium ms202. The antimicrobial effect of nitrofurantoin was studied through experimental and computational approach using standard microbiological and molecular techniques like growth curve analysis, live-dead analysis, oxidative stress evaluation using high throughput techniques like flow cytometry and fluorescent microscopy. The result showed a potent dose dependent antibacterial effect of nitrofurantoin against S. enterica Typhimurium ms202 with a MIC value of 64 µg/ml. Moreover, the mechanistic excavation of the phenomenon described the mechanism as an effect of molecular interaction of nitrofurantoin molecule with membrane receptor proteins OmpC of S. enterica Typhimurium ms202 leading to internalization of the nitrofurantoin heading towards the occurrence of cellular physiological disturbances through oxidative stress impeded by nitrofurantoin-Sod1 C protein interaction. The results indicated towards a synergistic effect of membrane damage, oxidative stress and genotoxicity for the antibacterial effect of nitrofurantoin against S. enterica Typhimurium ms202. The study described the potent dose-dependent application of nitrofurantoin molecule against MDR strains of Salmonella and guided towards their use in further discovered MDR strains.

Evidence of international transmission of mobile colistin resistant monophasic Salmonella Typhimurium ST34

S. 4,[5],12:i:-, a monophasic variant of S. enterica serovar Typhimurium, is an important multidrug resistant serovar. Strains of colistin-resistant S. 4,[5],12:i:- have been reported in several countries with patients occasionally had recent histories of travels to Southeast Asia. In the study herein, we investigated the genomes of S. 4,[5],12:i:- carrying mobile colistin resistance (mcr) gene in Thailand. Three isolates of mcr-3.1 carrying S. 4,[5],12:i:- in Thailand were sequenced by both Illumina and Oxford Nanopore platforms and we analyzed the sequences together with the whole genome sequences of other mcr-3 carrying S. 4,[5],12:i:- isolates available in the NCBI Pathogen Detection database. Three hundred sixty-nine core genome SNVs were identified from 27 isolates, compared to the S. Typhimurium LT2 reference genome. A maximum-likelihood phylogenetic tree was constructed and revealed that the samples could be divided into three clades, which correlated with the profiles of fljAB-hin deletions and plasmids. A couple of isolates from Denmark had the genetic profiles similar to Thai isolates, and were from the patients who had traveled to Thailand. Complete genome assembly of the three isolates revealed the insertion of a copy of IS26 at the same site near iroB, suggesting that the insertion was an initial step for the deletions of fljAB-hin regions, the hallmark of the 4,[5],12:i:- serovar. Six types of plasmid replicons were identified with the majority being IncA/C. The coexistence of mcr-3.1 and blaCTX-M-55 was found in both hybrid-assembled IncA/C plasmids but not in IncHI2 plasmid. This study revealed possible transmission links between colistin resistant S. 4,[5],12:i:- isolates found in Thailand and Denmark and confirmed the important role of plasmids in transferring multidrug resistance.

The use of thymol, carvacrol and sorbic acid in microencapsules to control Salmonella Heidelberg, S. Minnesota and S. Typhimurium in broilers.

The control of Salmonella spp. in poultry involves different biosecurity actions and lately has been complicated by the emergence of multidrug resistant serovars. The application of organic acids and essential oils has been used with different approaches due to the antibacterial properties as food preservatives. The use of these molecules in animal feed to control enteric pathogens is a major interest within the poultry industry. The use of a blend containing nature-identical compounds of sorbic acid (25%), thymol (9.5%) and carvacrol (2.5%) microencapsulated in a lipid matrix, was investigated in the present work, for the control of three Salmonella serovars (S. ser. Typhimurium, S. ser. Heidelberg and S. ser. Minnesota). Commercial broilers were challenged at 3 or at 33 days of age. Groups SH-1, SM-1 and ST-1, received treatment in the feed, at 2 kg/ton from 1-21 days of age and at 1 kg/ton from 35-42 days of age (last week), while groups SH-2, SM-2 and ST-2, were treated only during the last week receiving 2 kg/ton. Each treated group had an untreated control group, that was challenged at the same moment with the respective serovar (groups PCH, PCM and PCT). The challenge strains were enumerated in liver and cecal contents, weekly after challenge, at 7, 14, 21, 28, 35 and 42 days-of-age. Significant reduction was noticed at 7 and 14 days of age in all groups that received treatment during the initial phase (p < 0.05). Moreover, the body weight was significantly higher at the last experimental day (p < 0.05) in chickens that received treatment at the initial and at the final growth stages.

Effect of NaCl, high iron, iron chelator and antibiotics on growth, virulence gene expression and drug susceptibility in non-typhoidal Salmonella: an in vitro fitness study.

Salmonella is one among the most versatile and resilient enteric pathogens that is known to have developed various survival strategies within the host system. The ability of the bacteria to circumvent the physiological parameters as well as dodge the antimicrobial stress environment within the host is one of the most crucial steps in establishing an infection. With an alarming rise in multi-drug resistant serovars of non-typhoidal Salmonella and lack of vaccine for combatting the infections, behaviour of the bacteria in the presence of host physiological conditions (NaCl, high and low iron) and antibiotics will help in understanding the survival strategies as well as mechanisms of resistance. Two multi-drug resistant and two sensitive serovars of Salmonella Weltevreden and Salmonella Newport isolated from poultry and seafood were used for growth kinetics and virulence gene expression study. The results obtained revealed that despite similar resistance pattern, effect of individual class of antibiotics on the growth of serovars varied. On the contrary, no significant difference was observed in growth pattern on exposure to these in vitro experimental conditions. Nevertheless, coupling these conditions with antibiotics drastically reduced the minimum inhibitory concentration (MIC) of antibiotics in resistant strains. A first of its kind study that draws attention on the significant effect of antibiotics and physiological conditions on MIC between resistant and sensitive non-typhoidal Salmonella serovars and expression of virulence genes from Salmonella pathogenicity island (SPI) 1 and 2 (invA, hilC, fliC2, sseA and ssrB).

Salmonella enterica Serovar Dublin from Cattle in California from 1993-2019: Antimicrobial Resistance Trends of Clinical Relevance.

Salmonella enterica subsp. enterica serovar Dublin (S. Dublin) is a cattle-adapted pathogen that has emerged as one of the most commonly isolated and multidrug resistant (MDR) serovars in cattle. S. Dublin may be shed in feces, milk, and colostrum and persist in asymptomatic cattle, leading to spread and outbreaks in herds. Though infections with S. Dublin in humans are rare, they are frequently severe, with extraintestinal spread that requires hospitalization and antimicrobial therapy. To determine minimum inhibitory concentration (MIC) and antimicrobial resistance (AMR) patterns and trends in cattle in California, broth microdilution testing was performed on 247 clinical S. Dublin isolates recovered from cattle at the California Animal Health and Food Safety Laboratory System (CAHFS) over the last three decades (1993–2019). Mean MICs and classification of resistance to antimicrobial drugs using a clinical livestock panel and the National Antimicrobial Resistance Monitoring System (NARMS) Gram-negative drug panels were utilized to assess prevalence and trends in AMR. Findings indicate an increase in AMR for the years 1993 to 2015. Notably, compared to the baseline year interval (1993–1999), there was an increase in resistance among quinolone and cephalosporin drugs, as well as an increased number of isolates with an MDR profile.

Higher tolerance of predominant Salmonella serovars circulating in the antibiotic-free feed farms to environmental stresses

To counteract the dramatic increase in antibiotic-resistant bacterial pathogens, many countries, including China, have banned the use of antibiotic-supplemented feed for farming animals. However, the exact consequences of this policy have not been systematically evaluated. Therefore, Salmonella isolates from farms that ceased using antibiotics 1–5 years ago were compared with isolates from farms that continue to use antimicrobials as growth promotors. Here, we used whole-genome sequencing combined with in-depth phenotypic assays to investigate the ecology, epidemiology, and persistence of multi-drug resistant (MDR) Salmonella from animal farms during the withdrawal of antibiotic growth promotors. Our results showed that the prevalence of Salmonella was significantly lower in antibiotic-free feed (AFF) farms compared to conventional-feed (CF) farms, even though all isolates obtained from AFF farms were MDR (>5 classes) and belonged to well-recognized predominant serovars. The additional phylogenomic analysis combined with principal component analysis showed high similarity between the predominant serovars in AFF and CF farms. This result raised questions regarding the environmental persistence capabilities of MDR strain despite AFF policy. To address this question, a representative panel of 20 isolates was subjected to disadvantageous environmental stress assays. These results showed that the predominant serovars in AFF and CF farms were more tolerant to stress conditions than other serovars. Collectively, our findings suggest that AFF helps eliminate only specific MDR serovars, and future guiding policies would benefit by identifying predominant Salmonella clones in problematic farms to determine the use of AFF and additional targeted interventions.

Genomic diversity of antimicrobial resistance in non-typhoidal Salmonella in Victoria, Australia.

Non-typhoidal Salmonella (NTS) is the second most common cause of foodborne bacterial gastroenteritis in Australia with antimicrobial resistance (AMR) increasing in recent years. Whole-genome sequencing (WGS) provides opportunities for in silico detection of AMR determinants. The objectives of this study were two-fold: (1) establish the utility of WGS analyses for inferring phenotypic resistance in NTS, and (2) explore clinically relevant genotypic AMR profiles to third generation cephalosporins (3GC) in NTS lineages. The concordance of 2490 NTS isolates with matched WGS and phenotypic susceptibility data against 13 clinically relevant antimicrobials was explored. In silico serovar prediction and typing was performed on assembled reads and interrogated for known AMR determinants. The surrounding genomic context, plasmid determinants and co-occurring AMR patterns were further investigated for multidrug resistant serovars harbouring bla CMY-2, bla CTX-M-55 or bla CTX-M-65. Our data demonstrated a high correlation between WGS and phenotypic susceptibility testing. Phenotypic-genotypic concordance was observed between 2440/2490 (98.0 %) isolates, with overall sensitivity and specificity rates >98 % and positive and negative predictive values >97 %. The most common AMR determinants were bla TEM-1, sul2, tet(A), strA-strB and floR. Phenotypic resistance to cefotaxime and azithromycin was low and observed in 6.2 % (151/2486) and 0.9 % (16/1834) of the isolates, respectively. Several multi-drug resistant NTS lineages were resistant to 3GC due to different genetic mechanisms including bla CMY-2, bla CTX-M-55 or bla CTX-M-65. This study shows WGS can enhance existing AMR surveillance in NTS datasets routinely produced in public health laboratories to identify emerging AMR in NTS. These approaches will be critical for developing capacity to detect emerging public health threats such as resistance to 3GC.

Prevalence, Serotypes, and Antimicrobial Resistance of Salmonella from Mink Feces and Feed in the United States.

Nontyphoidal Salmonella is a leading cause of foodborne illnesses, and concentrated animal production such as commercial mink farming can be a reservoir. The objective of this study was to determine the prevalence, serotypes, antimicrobial resistance, virulence genes and mobile genetic elements, and phylogenetic characterization of Salmonella from mink fecal (n = 42) and feed (n = 8) samples at a commercial mink farm in the United States. Salmonella was detected from 11 (26.2%) fecal and 1 (12.5%) feed samples. Twelve fecal isolates obtained from the positive samples were identified into four serotypes: Salmonella Uganda (eight isolates; two isolates were obtained from one sample), Salmonella Reading (two isolates), Salmonella Heidelberg (one isolate), and Salmonella Orion (one isolate). Two isolates from the positive feed sample were identified as Salmonella Cerro. Twelve isolates, one isolate per positive sample, were whole genome sequenced for further characterization. The two Salmonella Reading isolates were multidrug resistant (MDR) with the classical ASSuT (ampicillin, streptomycin, sulfisoxazole, and tetracycline) phenotype. Genotypically, the isolates were correspondingly positive, both by polymerase chain reaction and by whole-genome sequencing (WGS), for blaTEM-1, aph(3″)-Ib and aph(6)-Id, sul2, and tet(A). WGS additionally identified trimethoprim resistance gene dfrA8 in Salmonella Reading, and fosfomycin resistance gene fosA7 in Salmonella Heidelberg. All isolates carried Salmonella-specific multidrug resistance (mdsA/B), gold resistance, and intimin-like virulence genes. In addition, Salmonella Orion carried copper and silver resistance gene clusters. The two Salmonella Reading isolates also carried a cytolethal distension toxin (cdtB) gene. Salmonella Reading isolates belonged to ST412, a predominant sequence type among turkey and human isolates obtained in relation to recent salmonellosis outbreaks in North America. Phylogenetically, Salmonella Uganda, Salmonella Heidelberg, and Salmonella Reading were mostly associated with historic human/clinical, and poultry and swine source isolates. This study indicates that mink production can be a reservoir for foodborne pathogens such as Salmonella with MDR serovars commonly associated with human illnesses.

Isolation and Characterization of Potential Salmonella Phages Targeting Multidrug-Resistant and Major Serovars of Salmonella Derived From Broiler Production Chain in Thailand.

Salmonella is a major foodborne pathogen that causes foodborne disease in humans through consumption of contaminated foods, especially those of animal origin. Multiple Salmonella strains are antibiotic-resistant due to the common use of antibiotics in farm animals, including broiler farms. In this study, an alternative strategy using phage-based treatment was evaluated against Salmonella isolated from the broiler production. The prevalence of Salmonella spp. showed up to 46.2 and 44.4% in bedding samples from the broiler farms located in eastern and southern Thailand, respectively. Overall, 21 samples (36.2%) were positive for Salmonella and eight serovars were recovered from cloacal swabs, bedding materials (rice husk), and boot swabs collected from five farms. Up to 20 Salmonella phages were isolated from seven water samples from wastewater treatment ponds, a river, and a natural reservoir in Songkhla province. Isolated phages were investigated, as well as their lysis ability on eight target Salmonella serovars derived from broiler farms, five foodborne outbreak-related serovars, and 10 multidrug-resistant (MDR) serovars. All phages showed a strong lytic ability against five serovars of Salmonella derived from broiler farms including Kentucky, Saintpaul, Schwarzengrund, Corvalis, and Typhimurium; three foodborne outbreak serovars including Enteritidis, Typhimurium, and Virchow; and eight MDR serovars including Agona, Albany, Give, Kentucky, Typhimurium, Schwarzengrund, Singapore, and Weltevreden. Three phages with the highest lysis potential including vB_SenS_WP109, vB_SenS_WP110, and vB_SenP_WP128 were selected for a phage cocktail preparation. Overall, a phage cocktail could reduce Salmonella counts by 2.2–2.8 log units at 6 h of treatment. Moreover, Salmonella did not develop a resistant pattern after being treated with a phage cocktail. Findings here suggest that a phage cocktail is an effective biocontrol to combat Salmonella derived from broiler production chain, other serovars linked to foodborne outbreaks, and MDR serovars.

Porcine Response to a Multidrug-Resistant Salmonella enterica serovar I 4,[5],12:i:- Outbreak Isolate.

Salmonella enterica serovar I 4,[5],12:i:- has emerged as a common nontyphoidal Salmonella serovar to cause human foodborne illness. An interesting trait of serovar I 4,[5],12:i:- is that it only expresses the fliC gene for bacterial motility (i.e., monophasic), while most Salmonella strains alternately express two flagellin genes (fliC and fljB). The goal of this study was to characterize the porcine response following inoculation with a multidrug-resistant (MDR) serovar I 4,[5],12:i:- isolate associated with a multistate pork outbreak to determine if the increased prevalence of serovar I 4,[5],12:i:- in swine is due to enhanced pathogenicity. Pigs were inoculated and subsequently evaluated for the ability of the isolate to colonize intestinal tissues, cause clinical symptoms, induce an immune response, and alter the fecal microbiota over a 7-day period. Pigs exhibited a significant increase in rectal temperature (fever) (p < 0.01) and fecal moisture content (diarrhea) (p < 0.05) at 2 days postinoculation (d.p.i.) compared with preinoculation (day 0). Serum analyses revealed significantly increased interferon-gamma (IFN-γ) levels at 2 (p ≤ 0.0001) and 3 (p < 0.01) d.p.i. compared with day 0, and antibodies against Salmonella lipopolysaccharide (LPS) were present in all pigs by 7 d.p.i. Serovar I 4,[5],12:i:- colonized porcine intestinal tissues and was shed in the feces throughout the 7-day study. Analysis of the 16S rRNA gene sequences demonstrated that the fecal microbiota was significantly altered following MDR serovar I 4,[5],12:i:- inoculation, with the largest shift observed between 0 and 7 d.p.i. Our data indicate that the pork outbreak-associated MDR serovar I 4,[5],12:i:- isolate induced transient clinical disease in swine and perturbed the gastrointestinal microbial community. The porcine response to MDR serovar I 4,[5],12:i:- is similar to previous studies with virulent biphasic Salmonella enterica serovar Typhimurium, suggesting that the absence of fljB does not substantially alter acute colonization or pathogenesis in pigs.

Isolation and characterization of polyvalent bacteriophages infecting multi drug resistant Salmonella serovars isolated from broilers in Egypt

In this study, we isolated and characterized three phages named as Salmacey1, Salmacey2 and Salmacey3, infecting multi drug resistant Salmonella serovars isolated from broilers in Egypt. The most prevalent Salmonella serovars were S. typhimurium, S. enteritidis, and S. kentucky. All these Salmonella serovars were found to be resistant to more than two of the ten antimicrobial agents tested. Only S. kentucky was found to be resistant to seven antimicrobial agents. Examination of these phage particles by transmission electron microscopy (TEM), demonstrated that two phages (Salmacey1, Salmacey2) were found to belong to family Siphoviridae, and Salmacey3 was assigned to the family Myoviridae. The results of host range assay revealed that these bacteriophages were polyvalent and thus capable of infecting four strains of Salmonella serovars and Citrobacter freundii. Moreover, the two phages (Salmacey1, Salmacey2) had a lytic effect on Enterobacter cloacae and Salmacey3 was able to infect E. coli. All phages could not infect S. para Typhi, Staphylococus aureus and Bacillus cereus. One-step growth curves of bacteriophages revealed that siphovirus phages (Salmacey1, Salmacey2) have burst size (80 and 90pfu per infected cell with latent period 35min and 40min respectively), and for the myovirus Salmacey3 had a burst size 110pfu per infected cell with latent period 60min. Molecular analyses indicated that these phages contained double-stranded DNA genomes. The lytic activity of the phages against the most multidrug resistant serovars S. kentucky as host strain was evaluated. The result showed that these bacteriophages were able to completely stop the growth of S. kentucky in vitro. These results suggest that phages have a high potential for phage application to control Salmonella serovars isolated from broilers in Egypt.

Draft Genome Sequence of Multidrug Resistant Salmonella enterica serovar Weltevreden Isolated from Seafood.

Salmonella enterica subsp. enterica serovar Weltevereden is the most frequent serovar isolated from Asia. Here, we report a draft genome sequence of multidrug resistant Salmonella Weltevreden 9 isolated from seafood. Whole-genome of this isolate and annotation will help enhance the understanding of this pathogenic multidrug-resistant serovar.

Characterization of integrons and resistance genes in multidrug-resistant Salmonella enterica isolated from meat and dairy products in Egypt

Foodborne pathogens are a leading cause of illness and death, especially in developing countries. The problem is exacerbated if bacteria attain multidrug resistance. Little is currently known about the extent of antibiotic resistance in foodborne pathogens and the molecular mechanisms underlying this resistance in Africa. Therefore, the current study was carried out to characterize, at the molecular level, the mechanism of multidrug resistance in Salmonella enterica isolated from 1600 food samples (800 meat products and 800 dairy products) collected from different street venders, butchers, retail markets and slaughterhouses in Egypt. Forty-seven out of 69 isolates (68.1%) showed multidrug resistance phenotypes to at least three classes of antimicrobials. The incidence of multidrug-resistant isolates was higher in meat products (37, 69.8%) than in dairy products (10, 62.5%). The multidrug-resistant serovars included, S. enterica serovar Typhimurium (24 isolates, 34.8%), S. enterica serovar Enteritidis, (15 isolates, 21.8%), S. enterica serovar Infantis (7 isolates, 10.1%) and S. enterica non-typable serovar (1 isolate, 1.4%). The highest resistance was to ampicillin (95.7%), then to kanamycin (93.6%), spectinomycin (93.6%), streptomycin (91.5%) and sulfamethoxazole/trimethoprim (91.5%). PCR and DNA sequencing were used to screen and characterize integrons and antibiotic resistance genes and 39.1% and 8.7% of isolates were positive for class 1 and class 2 integrons, respectively. β-lactamase-encoding genes were identified in 75.4% of isolates and plasmid-mediated quinolone resistance genes were identified in 27.5% of isolates. Finally, the florphenicol resistance gene, floR, was identified in 18.8% of isolates. PCR screening identified S. enterica serovar Typhimurium DT104 in both meat and dairy products. This is the first study to report many of these resistance genes in dairy products. This study highlights the high incidence of multidrug-resistant S. enterica in meat and dairy products in Egypt, with the possibility of their transfer to humans leading to therapeutic failure. Therefore, the overuse of antibiotics in animals should be drastically reduced in developing countries.

Extended-spectrum -lactamase- and AmpC -lactamase-producing D-tartrate-positive Salmonella enterica serovar Paratyphi B from broilers and human patients in Belgium, 2008-10

To characterize the genetic determinants responsible for extended-spectrum cephalosporin (ESC) resistance of d-tartrate-positive Salmonella enterica subsp. enterica serovar Paratyphi B (serovar Paratyphi B dT+) strains that have emerged in poultry and humans in Belgium during 2008-10. The ESC resistance genes among non-redundant serovar Paratyphi B dT+ strains were determined using PCR and sequencing. ESC phenotypes were horizontally transferred by conjugation. Extended-spectrum β-lactamase (ESBL)- or AmpC-carrying plasmids were typed by PCR-based replicon typing, plasmid multilocus sequence typing and restriction fragment length polymorphism. The genetic relationship of ESC-resistant strains was assessed by XbaI PFGE and multilocus sequence typing. Since 2008, the proportion of serovar Paratyphi B dT+ strains from broiler origin has increased significantly to reach 36.5% in 2010. Among 95 non-duplicate serovar Paratyphi B dT+ strains, 35% were resistant to ESCs. At the same time, a few ESC-resistant serovar Paratyphi B dT+ strains from humans were also detected in Belgium. The most prevalent ESBL gene, blaCTX-M-1, and the AmpC cephalosporinase gene blaCMY-2 were identified on various conjugative IncI1 plasmids of different sequence types and with different additional non-β-lactam phenotypes. Interestingly, the blaCTX-M-2 gene was located on large multireplicon IncHI2/P plasmids. In addition, highly ESC-resistant strains contained both the ESBL CTX-M-2 and the AmpC CMY-2 encoded by the IncHI2/P and IncI1 plasmids, respectively. All ESC-resistant serovar Paratyphi B dT+ strains belonged to sequence type 28 and showed the common PFGE pattern X8, as well as the chromosomal class 2 integron cassette array dfrA1-sat2-aadA1 previously described in the European poultry-associated serovar Paratyphi B dT+ clonal population. This study showed that the clonal population of multidrug-resistant serovar Paratyphi B dT+, persisting in broilers in Belgium for the last decade, recently acquired various plasmid-borne ESC resistance determinants, constituting a major concern for public health. Further surveillance programmes and research are an absolute necessity to understand their epidemiology and to propose interventions to limit the spread of ESC- and multidrug-resistant Salmonella spp.

Complete Genome Sequence of a Multidrug-Resistant Salmonella enterica Serovar Typhimurium var. 5− Strain Isolated from Chicken Breast

Salmonella enterica subsp. enterica serovar Typhimurium is a leading cause of salmonellosis. Here, we report a closed genome sequence, including sequences of 3 plasmids, of Salmonella serovar Typhimurium var. 5− CFSAN001921 (National Antimicrobial Resistance Monitoring System [NARMS] strain ID N30688), which was isolated from chicken breast meat and shows resistance to 10 different antimicrobials. Whole-genome and plasmid sequence analyses of this isolate will help enhance our understanding of this pathogenic multidrug-resistant serovar.

Salmonella enterica isolated from infections in Australian livestock remain susceptible to critical antimicrobials

Salmonella enterica is a zoonotic pathogen causing a variety of diseases in humans and animals. Many countries are reporting an increase in the prevalence of multidrug-resistant (MDR) S. enterica in food animals. The aim of this study was to determine whether S. enterica isolated from livestock in New South Wales, Australia, have similar resistance traits to those reported internationally. Salmonella enterica (n=165) from clinical infections in food animals between 2007 and 2011 were serotyped and tested for susceptibility to 18 antimicrobials. Also, 22 antimicrobial resistance genes (ARGs), 3 integrons and 18 plasmid replicon types were screened for using PCR. Most isolates (66.1%) remained susceptible to all antimicrobials; 8.5% of the isolates were resistant to four or more antimicrobials. Antimicrobials with the highest prevalence of resistance were sulfafurazole (28.5%), ampicillin (17.0%), tetracycline (15.8%) and trimethoprim (8.5%). There was no resistance to fluoroquinolones or third-generation cephalosporins. The most common ARGs were blaTEM (15.2%), sul2 (10.3%), tetB (9.1%), tetA (5.5%), aphA1 (4.8%) and dhfrV (4.8%). Class 1 integrons (7.9%) and IncFIIA (69.7%) were the most commonly detected integron and plasmid replicon types, respectively. Class 1 integrons were positively associated with MDR phenotypes and ARG carriage (P≤0.001). Internationally prominent MDR serovars associated with severe disease in humans (e.g. AmpC-positive Salmonella Newport) were not detected. Overall, the comparatively favourable resistance status of S. enterica in Australian livestock represents minimal public health risk associated with MDR strains and supports a conservative approach to the registration of antimicrobial drug classes in food-producing animals.

Complete Genome Sequence of Salmonella enterica Serovar Typhimurium U288

Salmonella enterica serovar Typhimurium U288 has firmly established itself within the United Kingdom pig production industry. The prevalence of this highly pathogenic multidrug-resistant serovar at such a critical point in the food chain is therefore of great concern. To enhance our understanding of this microorganism, whole-genome and plasmid sequencing was performed.

Characterization of Multidrug-ResistantSalmonella entericaSerovar Heidelberg Isolated from Humans and Animals

Salmonella enterica serovar Heidelberg has been recognized as one of the most common serovar associated with foodborne infections in the United States. It is also frequently isolated from nonhuman sources and has increasingly shown resistance to various antimicrobial agents. The present study was undertaken to identify the predominant antimicrobial resistance phenotypes and genotypes of Salmonella Heidelberg (n = 95) isolates of human, swine, and turkey origin. Antimicrobial susceptibility was done using Kirby-Bauer disk diffusion method with a panel of 12 antimicrobials. Pulsed-field gel electrophoresis genotyping was used to determine the diversity of the isolates. The antimicrobial resistance genes and carriage of Class 1 and 2 integrons were determined by polymerase chain reaction. All Salmonella Heidelberg isolates from swine were resistant to one or more of the antimicrobials tested and the majority (73.3%) showed multidrug resistance to streptomycin, tetracycline, and kanamycin (R-type: StTeKm). About 80% of the Salmonella Heidelberg isolates of human origin were pan-susceptible, however, one isolate showed multidrug resistance to 10 of 12 antimicrobials tested. Among the multidrug-resistant (MDR) Salmonella Heidelberg isolates, Class 1 integrons with variable sizes of 1.2 to 1.5 kb were detected in six isolates (three each) from humans and swine. DNA sequencing revealed that Class 1 integrons of both human and swine origin carried a gene encoding aminoglycoside adenyltransferase (aadA). Resistance genes identified in other loci include aphA1-Iab, strA, bla(TEM), and tetA (B). Both human and swine MDR strains of Salmonella Heidelberg carried the resistance phenotypes on self-transferable plasmids. Dendrogram analysis of pulsotypes indicated possible clonality of Salmonella Heidelberg between isolates of human and swine origin. The findings in this study indicate the increasing significance of swine as reservoirs of emerging MDR serovars, such as MDR Salmonella Heidelberg, is of public health significance.

Association of IS26-composite transposons and complex In4-type integrons generates novel multidrug resistance loci in Salmonella genomic island 1

Clinical isolates of Salmonella enterica serovar Haifa and Newport, which displayed extended multidrug resistance phenotypes, were investigated for the presence of Salmonella genomic island 1 (SGI1) and the genetic organization of its antibiotic resistance gene clusters. The S. enterica strains were isolated from humans in France in 2003 and 2004. Antibiotic susceptibility tests and various molecular techniques were used for detection and characterization of SGI1. We identified SGI1 integrated in the 3' end of the chromosomal thdF gene in six multidrug-resistant serovar Haifa and Newport strains. Two strains, of serovar Haifa and Newport, harboured the previously described SGI1-H variant. A new variant of the novel SGI1-Ks group, named SGI1-K6, revealed IS26-mediated rearrangements of the antibiotic resistance gene cluster in two serovar Newport strains. Two other serovar Newport strains harboured the SGI1-L complex class 1 integron containing the dfrA15 and bla(PSE-1) resistance gene cassettes. In addition, these variants of SGI1 also contained large IS26-composite transposons inserted by a transposition event in the SGI1 backbone. These IS26-composite transposons showed a similar genetic structure to the SGI1-K variants containing an In4-type integron, a mercury resistance operon and parts of Tn1721 and Tn5393. These extended resistance gene clusters containing up to 10 antibiotic resistance genes were named SGI1-L1 and -L2. The serovar Haifa represents the 16th S. enterica serovar in which SGI1 has been identified. The genomic island SGI1 appears to be a hotspot of acquisition of antibiotic resistance genes by the transposition of In4-type integrons and large IS26-composite transposons.