Molecular Epidemiology of Nontyphoidal Salmonella in Poultry and Poultry Products in India: Implications for Human Health

Human infections with non-typhoidal Salmonella (NTS) serovars are increasingly becoming a threat to human health globally. While all motile Salmonellae have zoonotic potential, Salmonella Enteritidis and Salmonella Typhimurium are most commonly associated with human disease, for which poultry are a major source. Despite the increasing number of human NTS infections, the epidemiology of NTS in poultry in India has not been fully understood. Hence, as a first step, we carried out epidemiological analysis to establish the incidence of NTS in poultry to evaluate the risk to human health. A total of 1215 samples (including poultry meat, tissues, egg and environmental samples) were collected from 154 commercial layer farms from southern India and screened for NTS. Following identification by cultural and biochemical methods, Salmonella isolates were further characterized by multiplex PCR, allele-specific PCR, enterobacterial repetitive intergenic consensus (ERIC) PCR and pulse field gel electrophoresis (PFGE). In the present study, 21/1215 (1.73 %) samples tested positive for NTS. We found 12/392 (3.06 %) of tissue samples, 7/460 (1.52 %) of poultry products, and 2/363 (0.55 %) of environmental samples tested positive for NTS. All the Salmonella isolates were resistant to oxytetracycline, which is routinely used as poultry feed additive. The multiplex PCR results allowed 16/21 isolates to be classified as S. Typhimurium, and five isolates as S. Enteritidis. Of the five S. Enteritidis isolates, four were identified as group D Salmonella by allele-specific PCR. All of the isolates produced different banding patterns in ERIC PCR. Of the thirteen macro restriction profiles (MRPs) obtained by PFGE, MRP 6 was predominant which included 6 (21 %) isolates. In conclusion, the findings of the study revealed higher incidence of contamination of NTS Salmonella in poultry tissue and animal protein sources used for poultry. The results of the study warrants further investigation on different type of animal feed sources, food market chains, processing plants, live bird markets etc., to evaluate the risk factors, transmission and effective control measures of human Salmonella infection from poultry products.     

Molecular Characterization of Motile Serovars of Salmonella enterica from Breeder and Commercial Broiler Poultry Farms in Bangladesh

Contaminated poultry and poultry products are a major source of motile Salmonellae for human salmonellosis worldwide. Local circulation of any motile Salmonella serovar in poultry has a wider public health impact beyond its source of origin for being dispersed elsewhere through poultry trades or human travels. To investigate the status of motile Salmonella serovars in breeder farms in Bangladesh, multiple flocks of two breeder farms were observed for a period of six months. In addition, a cross-sectional survey was carried out to determine the prevalence and serovar distribution of motile Salmonella by randomly selecting 100 commercial broiler poultry farms. Five pooled faecal samples representing an entire housed flock of breeders or broilers were screened for presence of motile Salmonella following conventional bacteriological procedures. The Salmonella isolates obtained were subsequently serotyped, and characterized by plasmid profiling and pulsed-field gel electrophoresis (PFGE). The results revealed that both the breeder farms were positive with three Salmonella serovars: S. Virchow, S. Paratyphi B var Java (S. Java) and S. Enteritidis. Eleven of the 100 broiler farms investigated were positive for motile Salmonella, giving a farm-level prevalence of 11% (95% confidence interval 5–17%). S. Virchow and S. Kentucky were the two predominant serovars isolated from the broiler farms. The PFGE genotyping demonstrated that the isolates belonging to the same serovars were closely related due to variation in only 1–4 bands. All the S. Virchow and S. Java isolates, irrespective of breeder or broiler farm origin, were plasmid-free, except for one S. Virchow isolate from a broiler farm that harboured a 9.7 kb-sized plasmid. The S. Kentucky isolates belonged to three plasmid profiles having plasmids of four different sizes, ranging from 2.7 to 109 kb. This is the first report of any motile Salmonella serovars from breeder and commercial broiler poultry farms in Bangladesh.     

Diversity and antimicrobial susceptibility of Salmonella enterica serovars isolated from pig farms in Ibadan, Nigeria

Animals including food animals play a significant role in the epidemiology of Salmonella enterica. The control requires identification of sources and institution of targeted interventions. This study investigates the diversity of S. enterica serovars, antimicrobial susceptibility, and occurrence of plasmid-mediated quinolone resistance (PMQR) genes in pigs in Ibadan, Nigeria. Pooled fresh pen floor fecal samples of pigs collected from 31 pig farms were cultured; the Salmonella isolates were serotyped and their antimicrobial susceptibility was determined. PMQR genes were screened by polymerase chain reaction. The 229 Salmonella isolates were made of 50 serovars predominated by rare serovars Salmonella Give (n?=?36; 15.7 %), Salmonella Brancaster (n?=?17; 7.4 %), Salmonella Colindale (n?=?15; 6.6 %), Salmonella Elisaberthville (n?=?13; 5.7 %), Salmonella Hillingdon (n?=?13; 5.7 %), and Salmonella Kingston (n?=?13; 5.7 %). The most widely distributed serovars among the farms were Salmonella Give (six farms) and Salmonella Elisaberthville (six farms). Resistance to chloramphenicol, sulfonamides, nalidixic acid, streptomycin, and tetracycline ranged from 11.6 % (n?=?26) to 22.8 % (n?=?51). Resistance ciprofloxacin and gentamicin was low (n?=?2; 0.9 %). Multiply resistant isolates included Salmonella Kentucky, the most resistant serovar. qnrB19 was found in two isolates of Salmonella Corvallis and one isolate of Salmonella Larochelle, respectively, while qnrS1 was found in two isolates of Salmonella Derby. Other PMQR genes were not detected. Pigs constitute an important source of diverse Salmonella serovars in Ibadan. The isolates were more resistant to old antimicrobials with some multiple resistant. Control measures and regulation of antimicrobials are warranted.     

Emergence of new variants of antibiotic resistance genomic islands among multidrug-resistant Salmonella enterica in poultry

Non-typhoidal Salmonella enterica (NTS) are diverse and important bacterial pathogens consisting of more than 2600 different serovars, with varying host-specificity. Here, we characterized the poultry-associated serovars in Israel, analysed their resistome and illuminated the molecular mechanisms underlying common multidrug resistance (MDR) patterns. We show that at least four serovars including Infantis, Muenchen, Newport and Virchow present a strong epidemiological association between their temporal trends in poultry and humans. Worrisomely, 60% from all of the poultry isolates tested (n = 188) were multidrug resistant, mediated by chromosomal SNPs and different mobile genetics elements. A novel streptomycin-azithromycin resistance island and previously uncharacterized versions of the mobilized Salmonella genomic island 1 (SGI1) were identified and characterized in S. Blockley and S. Kentucky isolates respectively. Moreover, we demonstrate that the acquisition of SGI1 does not impose fitness cost during growth under nutrient-limited conditions or in the context of Salmonella infection in the mouse model. Overall, our data emphasize the role of the poultry production as a pool of specific epidemic MDR strains and autonomous genetic elements, which confer resistance to heavy metals and medically relevant antibiotics. These are likely to disseminate to humans via the food chain and fuel the increasing global antibiotic resistance crisis.     

An Association of Genotypes and Antimicrobial Resistance Patterns among Salmonella Isolates from Pigs and Humans in Taiwan

We collected 110 Salmonella enterica isolates from sick pigs and determined their serotypes, genotypes using pulsed-field gel electrophoresis (PFGE), and antimicrobial susceptibility to 12 antimicrobials and compared the data with a collection of 18,280 isolates obtained from humans. The pig isolates fell into 12 common serovars for human salmonellosis in Taiwan; S. Typhimurium, S. Choleraesuis, S. Derby, S. Livingstone, and S. Schwarzengrund were the 5 most common serovars and accounted for a total of 84% of the collection. Of the 110 isolates, 106 (96%) were multidrug resistant (MDR) and 48 (44%) had PFGE patterns found in human isolates. S. Typhimurium, S. Choleraesuis, and S. Schwarzengrund were among the most highly resistant serovars. The majority of the 3 serovars were resistant to 8–11 of the tested antimicrobials. The isolates from pigs and humans sharing a common PFGE pattern displayed identical or very similar resistance patterns and Salmonella strains that caused severe infection in pigs were also capable of causing infections in humans. The results indicate that pigs are one of the major reservoirs to human salmonellosis in Taiwan. Almost all of the pig isolates were MDR, which highlights the necessity of strictly regulating the use of antimicrobials in the agriculture sector in Taiwan.     

High Levels of Antimicrobial Resistance among Escherichia coli Isolates from Livestock Farms and Synanthropic Rats and Shrews in the Mekong Delta of Vietnam

In Mekong Delta farms (Vietnam), antimicrobials are extensively used, but limited data are available on levels of antimicrobial resistance (AMR) among Escherichia coli isolates. We performed a structured survey of AMR in E. coli isolates (n = 434) from 90 pig, chicken, and duck farms. The results were compared with AMR among E. coli isolates (n = 234) from 66 small wild animals (rats and shrews) trapped on farms and in forests and rice fields. The isolates were susceptibility tested against eight antimicrobials. E. coli isolates from farmed animals were resistant to a median of 4 (interquartile range [IQR], 3 to 6) antimicrobials versus 1 (IQR, 1 to 2) among wild mammal isolates (P < 0.001). The prevalences of AMR among farmed species isolates (versus wild animals) were as follows: tetracycline, 84.7% (versus 25.6%); ampicillin, 78.9% (versus 85.9%); trimethoprim-sulfamethoxazole, 52.1% (versus 18.8%); chloramphenicol, 39.9% (versus 22.5%); amoxicillin-clavulanic acid, 36.6% (versus 34.5%); and ciprofloxacin, 24.9% (versus 7.3%). The prevalence of multidrug resistance (MDR) (resistance against three or more antimicrobial classes) among pig isolates was 86.7% compared to 66.9 to 72.7% among poultry isolates. After adjusting for host species, MDR was ∼8 times greater among isolates from wild mammals trapped on farms than among those trapped in forests/rice fields (P < 0.001). Isolates were assigned to unique profiles representing their combinations of susceptibility results. Multivariable analysis of variance indicated that AMR profiles from wild mammals trapped on farms and those from domestic animals were more alike (R² range, 0.14 to 0.30) than E. coli isolates from domestic animals and mammals trapped in the wild (R² range, 0.25 to 0.45). The results strongly suggest that AMR on farms is a key driver of environmental AMR in the Mekong Delta.     

Distributions of Salmonella Subtypes Differ between Two U.S. Produce-Growing Regions

Salmonella accounts for approximately 50% of produce-associated outbreaks in the United States, several of which have been traced back to contamination in the produce production environment. To quantify Salmonella diversity and aid in identification of Salmonella contamination sources, we characterized Salmonella isolates from two geographically diverse produce-growing regions in the United States. Initially, we characterized the Salmonella serotype and subtype diversity associated with 1,677 samples collected from 33 produce farms in New York State (NYS). Among these 1,677 samples, 74 were Salmonella positive, yielding 80 unique isolates (from 147 total isolates), which represented 14 serovars and 23 different pulsed-field gel electrophoresis (PFGE) types. To explore regional Salmonella diversity associated with production environments, we collected a smaller set of samples (n = 65) from South Florida (SFL) production environments and compared the Salmonella diversity associated with these samples with the diversity found among NYS production environments. Among these 65 samples, 23 were Salmonella positive, yielding 32 unique isolates (from 81 total isolates), which represented 11 serovars and 17 different PFGE types. The most common serovars isolated in NYS were Salmonella enterica serovars Newport, Cerro, and Thompson, while common serovars isolated in SFL were Salmonella serovars Saphra and Newport and S. enterica subsp. diarizonae serovar 50:r:z. High PFGE type diversity (Simpson''s diversity index, 0.90 ± 0.02) was observed among Salmonella isolates across both regions; only three PFGE types were shared between the two regions. The probability of three or fewer shared PFGE types was <0.000001; therefore, Salmonella isolates were considerably different between the two sampled regions. These findings suggest the potential for PFGE-based source tracking of Salmonella in production environments.     

Prevalence of multidrug resistance in Pseudomonas spp. isolated from wild bird feces in an urban aquatic environment

Antimicrobial resistance (AMR) has been detected in the microbiota of wildlife, yet little is known about the origin and impact within the ecosystem. Due to the shortage of nonepizootic surveillance, there is limited understanding of the natural prevalence and circulation of AMR bacteria in the wild animal population, including avian species. In this surveillance study, feces from wild birds in proximity to the River Cam, Cambridge, England, were collected and Pseudomonas spp. were isolated. Of the 115 samples collected, 24 (20.9%; 95% CI, 12.6%‒29.2%) harbored Pseudomonas spp. of which 18 (75%; 95% CI, 58%‒92%) had a multiple antibiotic resistance (MAR) index greater than 0.2. No Pseudomonas spp. isolate in this study was pansusceptible. Resistance was found among the 24 isolates against ciprofloxacin (87.5%; 95% CI, 74.3%‒100%) and cefepime (83.3%; 95% CI, 68.4%‒98.2%), both of which are extensively used to treat opportunistic Pseudomonas spp. infections. The prevalence of Pseudomonas spp. in the wild bird feces sampled during this study is greater than previous, similar studies. Additionally, their multidrug resistance profile provides insight into the potential risk for ecosystem contamination. It further highlights the importance of a One Health approach, including ongoing surveillance efforts that help to develop the understanding of how wildlife, including avifauna, may contribute and disperse AMR across the ecosystem.     

Genetic Diversity of Food-Isolated Salmonella Strains through Pulsed Field Gel Electrophoresis (PFGE) and Enterobacterial Repetitive Intergenic Consensus (ERIC-PCR)

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.     

Are Salmonella-Induced Gastroenteritis Neglected in Developing Countries? Feedback from Microbiological Investigations in N’Djamena Hospitals,Chad

Salmonella is considered to be one of the main pathogens causing human gastroenteritis worldwide. Looking for Salmonella in Africa in patients suffering from gastroenteritis is rather unusual, and the use of antibiotics is not subject to any regulation. This study intends for stressing the possible prominent importance of Salmonella in digestive diseases in Africa as well as identifying antimicrobial resistance of Salmonella isolates from faeces samples of human origin. All samples were collected from five N’Djamena hospitals, from patients suffering from diarrhoea. The collecting was undertaken over two periods of six months each: from August 2010 to January 2011 and from September 2011 to February 2012. Salmonella isolates were obtained by standard cultivation and serotyping methods. A total of 43 Salmonella isolates were identified, belonging to 21 different serovars. The most prevalent serovar was Salmonella Stanleyville (n = 7), followed by S. Anatum (n = 4) and S. Kottbus (n = 3). The other serovars were under-represented. The majority of these isolates were susceptible to all antibiotics tested (CLSI Standards), except two S. Enteritidis isolates that exhibited resistance to fluoroquinolones. The different serovars and antibiotic resistance profiles that were observed highlight the substantial diversity of Salmonella in N’Djamena, Chad. Roughly, one out of ten patients who consulted for gastroenteritis was shedding Salmonella spp. and none of them would have been diagnosed outside the context of this research program. This study may encourage local clinicians to explore more often salmonellosis suspicion in their daily practice.     

Salmonella enterica isolates from pasture‐raised poultry exhibit antimicrobial resistance and class I integrons

Aims: While considerable foodborne pathogen research has been conducted on conventionally produced broilers and turkeys, few studies have focused on free‐range (organic) or pastured poultry. The current surveillance study was designed to isolate, identify and genetically characterize Salmonella from pastured poultry farm environment and from retail samples. Methods and Results: In this study, 59 isolates were collected from two pastured poultry farms (n = 164; pens, feed, water and insect traps) and retail carcasses (n = 36) from a local natural foods store and a local processing plant. All isolates were serotyped and analysed phenotypically (antimicrobial resistance profiles) and genotypically (DNA fingerprints, plasmid profiles and integron analysis). Salmonella enterica was detected using standard microbiological methods. Salmonella Kentucky was the most prevalent serotype detected from the sampled sources (53%), followed by Salmonella Enteritidis (24%), Bareilly (10%), Mbandaka (7%), Montevideo (5%) or Newport (2%). All isolates were resistant to sulfisoxazole and novobiocin, and the majority (40/59) possessed class I integrons shown by PCR detection. Each Salmonella serotype elicited a distinct pulsed‐field gel electrophoresis fingerprint profile, and unique differences were observed among the serotypes. Conclusions: The findings of this study show that Salmonella serotypes isolated from pasture‐raised poultry exhibit antimicrobial resistance and class I integrons. Significance and Impact of the Study: This study demonstrates that despite the cessation of antibiotic usage in poultry production, antibiotic resistant Salmonella may still be recovered from the environment and poultry products.     

Antimicrobial Resistance-Conferring Plasmids with Similarity to Virulence Plasmids from Avian Pathogenic Escherichia coli Strains in Salmonella enterica Serovar Kentucky Isolates from Poultry

Salmonella enterica, a leading cause of food-borne gastroenteritis worldwide, may be found in any raw food of animal, vegetable, or fruit origin. Salmonella serovars differ in distribution, virulence, and host specificity. Salmonella enterica serovar Kentucky, though often found in the food supply, is less commonly isolated from ill humans. The multidrug-resistant isolate S. Kentucky {"type":"entrez-protein","attrs":{"text":"CVM29188","term_id":"985925135","term_text":"CVM29188"}}CVM29188, isolated from a chicken breast sample in 2003, contains three plasmids (146,811 bp, 101,461 bp, and 46,121 bp), two of which carry resistance determinants (pCVM29188_146 [strAB and tetRA] and pCVM29188_101 [bla_CMY-2 and sugE]). Both resistance plasmids were transferable by conjugation, alone or in combination, to S. Kentucky, Salmonella enterica serovar Newport, and Escherichia coli recipients. pCVM29188_146 shares a highly conserved plasmid backbone of 106 kb (>90% nucleotide identity) with two virulence plasmids from avian pathogenic Escherichia coli strains (pAPEC-O1-ColBM and pAPEC-O2-ColV). Shared avian pathogenic E. coli (APEC) virulence factors include iutA iucABCD, sitABCD, etsABC, iss, and iroBCDEN. PCR analyses of recent (1997 to 2005) S. Kentucky isolates from food animal, retail meat, and human sources revealed that 172 (60%) contained similar APEC-like plasmid backbones. Notably, though rare in human- and cattle-derived isolates, this plasmid backbone was found at a high frequency (50 to 100%) among S. Kentucky isolates from chickens within the same time span. Ninety-four percent of the APEC-positive isolates showed resistance to tetracycline and streptomycin. Together, our findings of a resistance-conferring APEC virulence plasmid in a poultry-derived S. Kentucky isolate and of similar resistance/virulence plasmids in most recent S. Kentucky isolates from chickens and, to lesser degree, from humans and cattle highlight the need for additional research in order to examine the prevalence and spread of combined virulence and resistance plasmids in bacteria in agricultural, environmental, and clinical settings.Nontyphoidal Salmonella enterica infections are one of the leading causes of bacterial food-borne gastroenteritis worldwide and an important public health problem in the United States, causing an estimated 1.4 million cases of infection, 15,000 hospitalizations, and more than 400 deaths annually in the United States alone (41). Although Salmonella infection usually presents with self-limiting diarrhea, in some patient populations, such as the immunocompromised, it can lead to life-threatening systemic infections that require effective and immediate antimicrobial therapy (21). The global emergence of multidrug resistance in S. enterica isolates from agricultural and clinical settings has therefore raised concerns and resulted in the establishment of several national resistance surveillance programs, such as the European Antimicrobial Resistance Surveillance System and the National Antimicrobial Monitoring System (NARMS) in the United States.In the United States, antimicrobial compounds are widely used in agriculture, not only to treat and prevent disease in plants, fruits, vegetables, and animals but also to promote growth in poultry and other livestock (25, 38). As a consequence, multidrug resistance is commonly detected in enterobacteria isolated from veterinary sources, including nontyphoidal Salmonella and other food-borne pathogens (40). Several studies have indicated the possibility that resistance reservoirs in animals can promote the transmission of resistance determinants from agricultural to clinical settings via food contaminants (1, 2, 45). Whether antimicrobial use in agriculture enhances the distribution and spectrum of antimicrobial resistance phenotypes in clinical settings has been the focus of vigorous debate within the public health and research communities (15, 19, 24, 27, 43). The extent of multidrug resistance in food-borne pathogens (10, 40), however, remains a concern. In 2006, only 17.7%, 25.0%, 38.8%, and 73.7% of the nontyphoidal Salmonella isolates from ground turkey, pork chop, chicken breast, and ground beef samples, respectively, showed susceptibility to all 16 antimicrobial compounds tested as part of the NARMS program (10). On the other hand, the overall prevalence of antimicrobial resistance phenotypes in nontyphoidal Salmonella isolates from human sources has slightly decreased, from 33.8% of all 876 isolates tested in 1996 to 19.4% of all 1,654 isolates tested in 2005 displaying a detectable resistance phenotype to at least one out of five antimicrobial subclasses as defined by the Clinical and Laboratory Standards Institute. However, in the same interval, increases from 0.4% to 2.4% and 0.2% to 2.9% in resistance to the clinically important subclasses of quinolones (nalidixic acid [Nal]) and cephalosporins (ceftiofur), respectively, were observed for the same set of human Salmonella isolates (6). Altogether, these reports demonstrate the need for further investigations on the influence of antimicrobial selection on the evolution, distribution, and transmission of resistance and virulence phenotypes among enteric bacteria derived from agricultural and clinical settings in order to prevent or at least limit the future spread of resistant zoonotic pathogens between these environments.Salmonella enterica subsp. enterica serovar Kentucky is widespread in poultry meats but is relatively uncommon in human cases of salmonellosis (7). S. Kentucky did not rank among the 20 most frequent Salmonella serotypes isolated from human sources in 2006 (7). In food-related sources, however, it is often found in animal samples and has been the most common serotype isolated from chickens (48.8%) (40) and chicken meat (38.8%) (10). At lower proportions, it is also present in turkey and cattle (2.6% and 3.6% of all nontyphoidal Salmonella isolates, respectively) (40). In the past decade, the fraction of S. Kentucky isolates from chickens, compared with other serotypes, has been increasing steadily, from 25% in 1997 to almost 50% in 2006 (40). Interestingly, while on average only 63 (0.174%) S. Kentucky isolates were reported between 1996 and 2004 among all nontyphoidal salmonellae from human samples, this number increased in 2005 (81 isolates [0.224%]) and 2006 (123 isolates [0.302%]) (7). Antimicrobial resistance phenotypes in S. Kentucky isolates from chicken meat are overrepresented compared to those in other serovars, with resistance to tetracycline (72.9%) and streptomycin (69.5%) being most commonly found (10). Although a causal connection between the increase of S. Kentucky in chickens and the number of human infections caused by the same serovar has not been demonstrated, further investigation is warranted, particularly in light of the high prevalence of antimicrobial resistance phenotypes in S. Kentucky isolates from chickens and the increasing resistance to beta-lactam compounds.Here, we describe the plasmid component of the genome of S. Kentucky {"type":"entrez-protein","attrs":{"text":"CVM29188","term_id":"985925135","term_text":"CVM29188"}}CVM29188, a multidrug-resistant strain that was isolated in 2003 from a chicken meat sample with resistance to streptomycin, tetracycline, ampicillin, and ceftiofur. Using a combination of in silico and in vivo approaches, including comparative plasmid sequence analysis, conjugative plasmid transfer, and PCR-based plasmid screenings, we present new insights into the genetic basis for multidrug resistance phenotypes of this isolate that provide new clues about virulence evolution and host adaptation in this Salmonella serovar.     

Antimicrobial Resistance in Salmonella enterica Serovar Heidelberg Isolates from Retail Meats, Including Poultry, from 2002 to 2006

Salmonella enterica serovar Heidelberg frequently causes food-borne illness in humans. There are few data on the prevalence, antimicrobial susceptibility, and genetic diversity of Salmonella serovar Heidelberg isolates in retail meats. We compared the prevalences of Salmonella serovar Heidelberg in a sampling of 20,295 meats, including chicken breast (n = 5,075), ground turkey (n = 5,044), ground beef (n = 5,100), and pork chops (n = 5,076), collected during 2002 to 2006. Isolates were analyzed for antimicrobial susceptibility and compared genetically using pulsed-field gel electrophoresis (PFGE) and PCR for the bla_CMY gene. A total of 298 Salmonella serovar Heidelberg isolates were recovered, representing 21.6% of all Salmonella serovars from retail meats. One hundred seventy-eight (59.7%) were from ground turkey, 110 (36.9%) were from chicken breast, and 10 (3.4%) were from pork chops; none was found in ground beef. One hundred ninety-eight isolates (66.4%) were resistant to at least one compound, and 49 (16.4%) were resistant to at least five compounds. Six isolates (2.0%), all from ground turkey, were resistant to at least nine antimicrobials. The highest resistance in poultry isolates was to tetracycline (39.9%), followed by streptomycin (37.8%), sulfamethoxazole (27.7%), gentamicin (25.7%), kanamycin (21.5%), ampicillin (19.8%), amoxicillin-clavulanic acid (10.4%), and ceftiofur (9.0%). All isolates were susceptible to ceftriaxone and ciprofloxacin. All ceftiofur-resistant strains carried bla_CMY. PFGE using XbaI and BlnI showed that certain clones were widely dispersed in different types of meats and meat brands from different store chains in all five sampling years. These data indicate that Salmonella serovar Heidelberg is a common serovar in retail poultry meats and includes widespread clones of multidrug-resistant strains.     

Characterization of multiple-antimicrobial-resistant Salmonella enterica Subsp. enterica isolated from indigenous vegetables and poultry in Malaysia

Aims: The aims of this communication were to study characterization of serogroups among Salmonella isolates and the relationship of antimicrobial resistance to serogroups. Multiple antimicrobial resistance (MAR) was performed on 189 Salmonella enterica isolates associated with 38 different serovars that were recovered from poultry and four types of indigenous vegetables. Methods and Results: Disc diffusion analysis was performed with a selection of 10 different antimicrobial agents. Isolates recovered from indigenous vegetables showed 100% (134/134) resistant to erythromycin and followed by 42%, 34%, 19% for tetracycline, streptomycin and trimethroprim‐sulfamethoxazole respectively. In general, 90·1% (50/55) and 56·7% (76/134) of Salmonella isolated from poultry and indigenous vegetables, respectively, exhibited MAR index more than 0·2. Conclusions: Characterization of Salmonella isolates based on the MAR results indicated that poultry still remains as the main reservoir for multi‐drug‐resistant Salmonella. Four isolates from the indigenous vegetables showed the highest MAR index in this study. Further investigations need to be conducted to determine if Salmonella isolates recovered from indigenous vegetables were gaining more antimicrobial resistance. Significance and Impact of the Study: The study enabled us to determine antimicrobial patterns and trends in Salmonella from poultry and indigenous vegetables in Malaysia.     

Prevalence and Fimbrial Genotype Distribution of Poultry Salmonella Isolates in China (2006 to 2012)

In this study, a total of 323 Salmonella enterica strains were isolated from 3,566 rectal swab samples of 51 poultry farms in seven regions of 12 provinces of China between 2006 and 2012. The prevalences of Salmonella sp. carriage were 12.4% in geese (66 positive/533 samples), 10.4% in turkeys (32/309), 9.8% in chickens (167/1,706), 6.8% in ducks (41/601), and 4.1% in pigeons (17/417), respectively. These isolates belonged to 20 serovars, in which the most frequent serovars were S. enterica serovar Gallinarum biovar Pullorum (herein, S. Pullorum) (55 isolates, 17.0%), S. enterica serovar Typhimurium (50 isolates, 15.5%), and S. enterica serovar Enteritidis (39 isolates, 12.1%). Overall, S. Typhimurium was the most commonly detected serovar; among the individual species, S. Pullorum was most commonly isolated from chickens, S. Enteritidis was most common in ducks, S. Typhimurium was most common in geese and pigeons, and S. enterica serovar Saintpaul was most common in turkeys. PCR determination of 20 fimbrial genes demonstrated the presence of bcfD, csgA, fimA, stdB, and sthE genes and the absence of staA and stgA genes in these isolates, and other loci were variably distributed, with frequency values ranging from 11.8 to 99.1%. These 323 Salmonella isolates were subdivided into 41 different fimbrial genotypes, and of these isolate, 285 strains (88.2%) had 12 to 14 fimbrial genes. Our findings indicated that the Salmonella isolates from different poultry species were phenotypically and genetically diverse and that some fimbrial genes are more frequently associated with serovars or serogroups.     

Clonal differences between Non-Typhoidal Salmonella (NTS) recovered from children and animals living in close contact in the Gambia

Background

Non-Typhoidal Salmonella (NTS) is an important cause of invasive bacterial disease and associated with mortality in Africa. However, little is known about the environmental reservoirs and predominant modes of transmission. Our study aimed to study the role of domestic animals in the transmission of NTS to humans in rural area of The Gambia.

Methodology

Human NTS isolates were obtained through an active population-based case-control surveillance study designated to determine the aetiology and epidemiology of enteric infections covering 27,567 Gambian children less than five years of age in the surveillance area. Fourteen children infected with NTS were traced back to their family compounds and anal swabs collected from 210 domestic animals present in their households. Identified NTSs were serotyped and genotyped by multi-locus sequencing typing.

Principal Findings

NTS was identified from 21/210 animal sources in the households of the 14 infected children. Chickens carried NTS more frequently than sheep and goats; 66.6%, 28.6% and 4.8% respectively. The most common NTS serovars were S. Colindale in humans (21.42%) and S. Poona in animals (14.28%). MLST on the 35 NTS revealed four new alleles and 24 sequence types (ST) of which 18 (75%) STs were novel. There was no overlap in serovars or genotypes of NTS recovered from humans or animal sources in the same household.

Conclusion

Our results do not support the hypothesis that humans and animals in close contact in the same household carry genotypically similar Salmonella serovars. These findings form an important baseline for future studies of transmission of NTS in humans and animals in Africa.     

Antimicrobial Resistance in Invasive Non-typhoid Salmonella from the Democratic Republic of the Congo: Emergence of Decreased Fluoroquinolone Susceptibility and Extended-spectrum Beta Lactamases

Background

Co-resistance against the first-line antibiotics ampicillin, chloramphenicol and trimethoprim/sulphamethoxazole or multidrug resistance (MDR) is common in non typhoid Salmonella (NTS). Use of alternative antibiotics, such as fluoroquinolones or third generation cephalosporins is threatened by increasing resistance, but remains poorly documented in Central-Africa.

Methodology/Principal findings

As part of a microbiological surveillance study in DR Congo, blood cultures were collected between 2007 and 2011. Isolated NTS were assessed for serotype and antimicrobial resistance including decreased ciprofloxacin susceptibility and extended-spectrum beta-lactamase (ESBL) production. In total, 233 NTS isolates (representing 23.6% of clinically significant organisms) were collected, mainly consisting of Salmonella Typhimurium (79%) and Salmonella Enteritidis (18%). The majority of NTS were isolated in the rainy season, and recovered from children ≤2 years old. MDR, decreased ciprofloxacin susceptibility, azithromycin and cefotaxime resistance were 80.7%, 4.3%, 3.0% and 2.1% respectively. ESBL production was noted in three (1.3%) isolates. Decreased ciprofloxacin susceptibility was associated with mutations in codon 87 of the gyrA gene, while ESBLs all belonged to the SHV-2a type.

Conclusions/Significance

Presence of almost full MDR among NTS isolates from blood cultures in Central Africa was confirmed. Resistance to fluoroquinolones, azithromycin and third generation cephalosporins is still low, but emerging. Increased microbiological surveillance in DR Congo is crucial for adapted antibiotic therapy and the development of treatment guidelines.     

Fast DNA Serotyping and Antimicrobial Resistance Gene Determination of Salmonella enterica with an Oligonucleotide Microarray-Based Assay

Salmonellosis caused by Salmonella (S.) belongs to the most prevalent food-borne zoonotic diseases throughout the world. Therefore, serotype identification for all culture-confirmed cases of Salmonella infection is important for epidemiological purposes. As a standard, the traditional culture method (ISO 6579:2002) is used to identify Salmonella. Classical serotyping takes 4–5 days to be completed, it is labor-intensive, expensive and more than 250 non-standardized sera are necessary to characterize more than 2,500 Salmonella serovars currently known. These technical difficulties could be overcome with modern molecular methods. We developed a microarray based serogenotyping assay for the most prevalent Salmonella serovars in Europe and North America. The current assay version could theoretically discriminate 28 O-antigens and 86 H-antigens. Additionally, we included 77 targets analyzing antimicrobial resistance genes. The Salmonella assay was evaluated with a set of 168 reference strains representing 132 serovars previously serotyped by conventional agglutination through various reference centers. 117 of 132 (81%) tested serovars showed an unique microarray pattern. 15 of 132 serovars generated a pattern which was shared by multiple serovars (e.g., S. ser. Enteritidis and S. ser. Nitra). These shared patterns mainly resulted from the high similarity of the genotypes of serogroup A and D1. Using patterns of the known reference strains, a database was build which represents the basis of a new PatternMatch software that can serotype unknown Salmonella isolates automatically. After assay verification, the Salmonella serogenotyping assay was used to identify a field panel of 105 Salmonella isolates. All were identified as Salmonella and 93 of 105 isolates (88.6%) were typed in full concordance with conventional serotyping. This microarray based assay is a powerful tool for serogenotyping.     

Antibiotic Resistance in Food-Borne Bacterial Contaminants in Vietnam

This study was conducted to examine the rate of contamination and the molecular characteristics of enteric bacteria isolated from a selection of food sources in Vietnam. One hundred eighty raw food samples were tested; 60.8% of meat samples and 18.0% of shellfish samples were contaminated with Salmonella spp., and more than 90% of all food sources contained Escherichia coli. The isolates were screened for antibiotic resistance against 15 antibiotics, and 50.5% of Salmonella isolates and 83.8% of E. coli isolates were resistant to at least one antibiotic. Isolates were examined for the presence of mobile genetic elements conferring antibiotic resistance. Fifty-seven percent of E. coli and 13% of Salmonella isolates were found to contain integrons, and some isolates contained two integrons. Sequencing results revealed that the integrons harbored various gene cassettes, including aadA1, aadA2, and aadA5 (resistance to streptomycin and spectinomycin), aacA4 (resistance to aminoglycosides), the dihydrofolate reductase gene cassettes dhfrXII, dfrA1, and dhfrA17 (trimethoprim resistance), the beta-lactamase gene bla_PSE1 (ampicillin resistance), and catB3 (chloramphenicol resistance). Plasmids were also detected in all 23 antibiotic-resistant Salmonella isolates and in 33 E. coli isolates. Thirty-five percent of the Salmonella isolates and 76% of the E. coli isolates contained plasmids of more than 95 kb, and some of the isolates contained two large plasmids. Conjugation experiments showed the successful transfer of all or part of the antibiotic resistance phenotypes among the Salmonella and E. coli food isolates. Our results show that enteric bacteria in raw food samples from Vietnam contain a pool of mobile genetic elements and that the transfer of antibiotic resistance can readily occur between similar bacteria.     

Effect of Conventional and Organic Production Practices on the Prevalence and Antimicrobial Resistance of Campylobacter spp. in Poultry

Intestinal tracts of broilers and turkeys from 10 conventional broiler farms and 10 conventional turkey farms, where antimicrobials were routinely used, and from 5 organic broiler farms and 5 organic turkey farms, where antimicrobials had never been used, were collected and cultured for Campylobacter species. A total of 694 Campylobacter isolates from the conventional and organic poultry operations were tested for antimicrobial resistance to nine antimicrobial agents by the agar dilution method. Although Campylobacter species were highly prevalent in both the conventional and organic poultry operations, the antimicrobial resistance rates were significantly different between the organic operations and the conventional operations. Less than 2% of Campylobacter strains isolated from organically raised poultry were resistant to fluoroquinolones, while 46% and 67% of Campylobacter isolates from conventionally raised broilers and conventionally raised turkeys, respectively, were resistant to these antimicrobials. In addition, a high frequency of resistance to erythromycin (80%), clindamycin (64%), kanamycin (76%), and ampicillin (31%) was observed among Campylobacter isolates from conventionally raised turkeys. None of the Campylobacter isolates obtained in this study was resistant to gentamicin, while a large number of the isolates from both conventional and organic poultry operations were resistant to tetracycline. Multidrug resistance was observed mainly among Campylobacter strains isolated from the conventional turkey operation (81%). Findings from this study clearly indicate the influence of conventional and organic poultry production practices on antimicrobial resistance of Campylobacter on poultry farms.