Outbreak of Salmonella Typhimurium associated with feeder rodents

In December 2012, an increase in human Salmonella Typhimurium cases was identified in the province of Ontario, Canada launching an outbreak investigation. The outbreak spanned 3 years (2012–2014), with 134 cases reported from five Canadian provinces. There was a substantial burden of illness among children: 45% of cases were children 12 years old or under, and 23% of cases were under 5 years old. Epidemiologic, traceback and laboratory findings linked this outbreak to feeder rodents (used to feed snakes) supplied by a network of rodent breeders in Ontario. Cases likely acquired their illness through either direct or indirect contact with feeder rodents. This investigation not only contributes to the weight of evidence on the risk that feeder rodents pose, but also underscores the importance of investigating indirect animal contact and associated risks, especially for high‐risk individuals.     

Characterisation of streptomycin resistance determinants in Danish isolates of Salmonella Typhimurium

Fifty six Danish streptomycin (Sm) resistant isolates of Salmonella enterica serotype Typhimurium from pigs (n=34), calves (n=3) and humans (n=19) were characterised with respect to co-resistances (14 drugs), transferability of Sm-resistance by conjugation, genetic determinants encoding Sm-resistance and diversity with respect to localisation of genes in the genome and DNA-sequences. Forty-six strains carried resistance(s) other than Sm-resistance. Nineteen different co-resistance patterns were observed and tetracycline was the most commonly observed resistance in these patterns. In 22 of the strains, Sm-resistance was transferred by conjugation. Eleven strains contained the gene aadA only, six strains contained aadA+strA+strB, and 35 strains contained strA+strB. Partial sequences of aadA were obtained from four strains. Three strains showed identical sequences to a published aadA sequence from the transposon Tn7, and in one strain the sequence showed one synonymous substitution compared to this sequence. Partial sequences were obtained of strA and strB in seven strains. The sequence of strB was identical to the published sequence of the plasmid RSF1010 in all strains. All seven sequences of strA were identical and differed from the sequence of strA in RSF1010 by two non-synonymous substitutions.     

Prevalence of the virulence plasmid in Salmonella Typhimurium isolates from pigs

To determine the prevalence of the virulence plasmid in Salmonella Typhimurium isolates from pigs in Japan, a total of 106 porcine isolates were subjected to PCR amplification for the detection of the virulence plasmid. Out of the isolates of S. Typhimurium, 38 (35.8%) harbored the virulence plasmid. The presence of the virulence plasmid was widely observed in the isolates from systemically infected pigs (92.0%, 23/25), compared with diarrheic (18.8%, 12/64) and apparently healthy pigs (17.6%, 3/17) (P<0.01).     

Characterization of antimicrobial resistant Salmonella enterica serovars Enteritidis and Typhimurium isolates from animal and food in Southern Italy

In the last two decades, the emergence and spread of antimicrobial-resistant pathogens, among them Salmonella, has become a serious health hazard worldwide, and specifically the high incidence of multidrug resistance has been encountered widely in many European countries. This study examines the antimicrobial supsceptibility of Salmonella enterica strains Typhimurium and Enteritidis isolated in Campania and Calabria region (Southern Italy) from animal and food of animal origin. The relationship of antibiotic resistance phenotype and the presence of some resistance genes has been also investigated. As espected, our results showes that resistance to ampicillin, chloramphenicol, streptomycin, sulphonamides and tetracycline is common, although resistance to other antibiotics (i.e.:nalidixic acid) and other resistance patterns occur. The genetic resistant patterns have been partially described for this food-borne pathogen but efforts are needed to realize the complete characterization of antimicrobial resistance genes.     

Protection of neonatal broiler chicks against Salmonella Typhimurium septicemia by DNA containing CpG motifs

Oligodeoxynucleotides (ODN) containing cytosine-phosphodiester-guanine (CpG-ODN) motifs have been shown to stimulate the innate immune system against a variety of bacterial and protozoan infections in a variety of vertebrate species. The objective of this study was to investigate the immunostimulatory effect of CpG-ODN in neonatal broilers against Salmonella Typhimurium septicemia. Day-old broiler chicks, or embryonated eggs that had been incubated for 18 days, received 50 microg of CpG-ODN, 50 microg of non-CpG-ODN, or saline. Four days after exposure to CpG-ODN or day 2 posthatch, 1 x 10(6) or 1 x 10(7) colony-forming units (cfu) of a virulent isolate of Salmonella Typhimurium was inoculated by the subcutaneous route in the neck. Clinical signs, pathology, bacterial isolations from the air sacs, and mortality were observed for 10 days following challenge with Salmonella Typhimurium. The survival rate of birds in groups receiving either non-CpG-ODN or saline following Salmonella Typhimurium infection was 40%-45%. In contrast, birds receiving CpG-ODN had significantly higher survival rate of 80%-85% (P < 0.0001). Bacterial loads and pathology were low in groups treated with CpG-ODN compared to the groups receiving saline or non-CpG-ODN. Colony-forming units of Salmonella Typhimurium in the peripheral blood were significantly lower in birds treated with CpG-ODN compared to the group that received saline. This is the first time that CpG-ODN has been demonstrated to have an immunoprotective effect against an intracellular bacterial infection in neonatal broiler chickens following in ovo delivery.     

Characterization of Salmonella Dublin and Salmonella Typhimurium (Copenhagen) Isolates from Cattle

Brackelsberg, C.A., Nolan, L.K. and Brown, J., 1997. Characterization of Salmonella dublin and Salmonella typhimurium (Copenhagen) isolates from cattle. Veterinary Research Communications, 21 (6), 409-420Eight Salmonella typhimurium (Copenhagen) and eight Salmonella dublin isolates from cattle were compared by their antibiotic resistance patterns, by their production of colicin, aerobactin, haemolysin and capsule, by their possession of transmissible R plasmids and the spvC gene, and by their ability to invade and replicate within cultured epithelial cells. The two groups differed in their antibiotic resistance profiles, with more of the host-adapted S. dublin isolates resistant to tetracycline than were the non-host-adapted S. typhimurium (Copenhagen) group, but more of the S. typhimurium (Copenhagen) isolates resistant to the other antibiotics tested. None of the isolates produced colicin, but all produced aerobactin. One isolate in each group was encapsulated. All of the S. typhimurium (Copenhagen) and S. dublin isolates contained plasmids, and all of them contained the spvC-homologous sequences. Four of the S. typhimurium (Copenhagen) isolates were able to transfer an R plasmid to a recipient organism by conjugation. One of the five S. dublin isolates, which showed resistance to some of the antibiotics tested, was able to transfer an R plasmid by conjugation. Both groups of isolates invaded cultured epithelial cells to a similar degree after 1 h, but the S. dublin isolates reached significantly higher levels within the cells than did S. typhimurium (Copenhagen) after 9 h. This ability may, in part, explain the association of S. dublin with more severe forms of salmonellosis and prolonged carrier states. Further study of the intracellular growth of these isolates seems warranted.     

Characterization of Salmonella isolates from captive lizards

Reptile-associated salmonellosis in humans is an increasing public health issue. This study aimed at characterizing Salmonella isolates from captive lizards and to compare them to human isolates. Salmonella was isolated from 25 of 33 cloacal and 47 of 79 faecal samples from captive lizards (75.8 and 59.5%, respectively). The strains belonged to 44 serotypes of subspecies I (27 serotypes), II (9), IIIb (3) and IV (5). Two strains, one of serotype Enteritidis and one of serotype Amsterdam, were resistant to nitrofurantoin. Invasion assays in Caco-2 cells were performed with 40 saurian isolates of subspecies I, 15 isolates of subspecies II, 4 strains of subspecies IIIb, 6 subspecies IV isolates and 17 human isolates of corresponding serotypes of subspecies I. Saurian isolates belonging to subspecies I invaded the Caco-2 cells to a higher extent than those from the other subspecies. The human isolates invaded the Caco-2 cells to a lesser degree compared to their saurian counterparts. In the same strains, the presence of virulence genes agfA, shdA, spvR, pefA and sopE was determined using PCR. Whereas agfA was detected in all strains, pefA was only detected in one saurian and in the human serotype Enteritidis strains. The spvR gene was detected in the same serotype Enteritidis strains and in 33% of the subspecies IV strains. The shdA gene was present in all the human isolates and in 86% of subspecies I saurian isolates. SopE was found in 17% of the human isolates, in 24% of the saurian subspecies I strains and in all of the subspecies IV strains.     

Pulsed field gel electrophoresis for animal Salmonella enterica serovar Typhimurium isolates in Taiwan

Salmonella enterica subspecies enterica serovar Typhimurium is a common pathogen for humans and animals. In order to trace the clonal relationship and to find the circulating strains between human and animal isolates, chromosomal DNAs from 87 serovar Typhimurium strains isolated from animals (pigs were the majority) were subjected to XbaI and SpeI digestion and pulsed field gel electrophoresis (PFGE). For the 87 animal isolates, 38 PFGE pattern combinations were obtained. As the subtyping results from animal isolates were compared with those from the 45 human isolates, it was found that 14 of the animal isolates and 13 of the human isolates shared a common PFGE pattern combination, i.e., pattern XgSf (or called X5S4). When these human and animal isolates were subjected to antibiotic susceptibility test using 11 antibiotics, it was found that strains of pattern XgSf (X5S4) belong to a common antibiogram pattern which is tetracycline, gentamicin, ampicillin, streptomycin and chloramphenicol resistant. Since most of the animal and human strains in pattern XgSf were originally isolated from various areas over different years, strains of this PFGE pattern may be the most epidemic strains which circulating between human and animal sources.     

Early epithelial invasion by Salmonella enterica serovar Typhimurium DT104 in the swine ileum

Salmonella enterica serovar Typhimurium is an important intestinal pathogen in swine. This study was performed to document the early cellular invasion of Salmonella serovar Typhimurium in swine ileum. Ileal gut-loops were surgically prepared in ten 4- to 5-week-old mixed-breed pigs and inoculated for 0-60 minutes. Loops were harvested and prepared for both scanning and transmission electron microscopy (SEM and TEM, respectively). Preferential bacterial adherence to microfold cells (M cells) was seen within 5 minutes, and by 10 minutes bacterial invasion of the apical membrane was seen in M cells, goblet cells, and enterocytes. This multicellular invasion was observed throughout the course of infection. In addition, SEM revealed a specific affinity of Salmonella serovar Typhimurium to sites of cell extrusion. Using TEM, bacteria in these areas were focused in the crevices formed by the extruding cell and the adjacent cells and in the cytoplasm immediately beneath the extruding cell. Our results suggest that early cellular invasion by Salmonella serovar Typhimurium is nonspecific and rapid in swine. Furthermore, the combination of SEM and TEM data suggests that Salmonella serovar Typhimurium may use sites of cell extrusion as an additional mechanism for early invasion.     

Assessment of antibiotic resistance phenotype and integrons in Salmonella enterica serovar Typhimurium isolated from swine

Salmonella enterica serovar Typhimurium (S. Typhimurium) isolated and identified from swine were subjected for the analysis of antibiotic resistance pattern and clinically important class 1 and 2 integrons. In addition, S. Typhimurium isolates exhibiting ampicillin, chloramphenicol, streptomycin, sulfamethoxazole, tetracycline and florfenicol (ACSSuTF) resistance pattern as described in most Salmonella enterica serotype Typhimurium definitive type 104 (DT104) were characterized by polymerase chain reaction. All the isolates were resistant to more than four antibiotics and showed the highest resistance to streptomycin (94.1%), followed by tetracycline (90.1%), ampicillin (64.7%), chloramphenicol (56.8%) and gentamicin (54.9%). MIC value for the ten isolates ranged between 0.125-2 mug/ml for ciprofloxacin. Among the beta-lactams used, only one of the isolate exhibited resistance to ceftiofur (MIC 8 microg/ml). Sixty eight percent of these multi drug resistance (MDR) S. Typhimurium isolates carried clinically important class 1 integron with 1kb (aadA) and/or 2kb (dhfrXII-orfF-aadA2) resistance gene cassettes. This study reports the increasing trend of multi drug resistance (MDR) S. Typhimurium with clinically important class 1 integron in pigs. In addition, emergence of the ACSSuTF-type resistance in S. Typhimurium PT other than DT104 may limit the use of resistance gene markers in its detection methods by PCR.     

Abomasitis associated with multiple antibiotic resistant Salmonella enterica serotype Typhimurium phagetype DT104

Salmonella enterica serotype Typhimurium phagetype DT104 is a multiple antibiotic resistant pathogen that has been purported to be more pathogenic than other Salmonella. In this study, we evaluated the possibility that DT104 is the causative agent of veal calf abomasitis observed in four independent outbreaks of salmonellosis. This study was undertaken to determine if the outbreaks might be due to hypervirulent S. enterica serotype Typhimurium phagetype DT104 (DT104) since Salmonella does not usually cause abomasitis. Tissues and fluids from these calves were subjected to bacteriologic culture. Pure Salmonella cultures were then used in bovine challenge experiments. DT104 was identified as the causative agent of abomasitis in calves. Thus, abomasitis is a potential indicator of infection with multiple antibiotic resistant DT104 and adds credence to the apparent hypervirulence of this pathogen.     

A multi-state Salmonella Typhimurium outbreak associated with frozen vacuum-packed rodents used to feed snakes

From December 2005 through January 2006, the Minnesota Department of Health (MDH) identified four human clinical isolates of Salmonella Typhimurium that were indistinguishable by pulsed-field gel electrophoresis (PFGE). During routine interviews, three of the cases reported attending the same junior high school and two handled snakes in the science classroom. MDH collected environmental samples from the school's science classroom for Salmonella culturing; these included environmental samples and frozen vacuum-packed mice purchased over the internet to feed the classroom snakes. Through PulseNet, a national molecular subtyping surveillance network for enteric bacteria, 21 human S. Typhimurium isolates with indistinguishable PFGE patterns were identified in the United States since December 2005. Each state determined whether these human cases had recent exposure to snakes fed vacuum-packed rodents. Texas state officials conducted tracebacks of the vacuum-packed mice and collected samples at the breeding facility. Nineteen of 21 cases were interviewed, and seven reported contact with frozen vacuum-packed rodents from the same internet-based supplier in Texas. In Minnesota, the outbreak PFGE subtype of S. Typhimurium was isolated from the snakes, frozen feed rodents, and the classroom environment. Three human cases were identified in Michigan, Pennsylvania, and Wyoming. The outbreak PFGE subtype of S. Typhimurium was isolated from the Pennsylvania case's frozen rodents and the Michigan case's pet snake. The outbreak PFGE subtype of S. Typhimurium was also isolated from the supplier's rodent facility. This was a S. Typhimurium outbreak associated with frozen rodents. Human transmission likely occurred through direct contact with snakes and contaminated environmental surfaces. This report represents the second recent multi-state salmonellosis outbreak associated with commercially distributed rodents. Stronger oversight of the commercial rodent industry is warranted.     

Characterization of Pseudomonas aeruginosa isolates associated with mortality in broiler chicks

In mid-2000, a broiler chicken company in Alabama experienced high early mortality rates in chicks from two different hatcheries. Five isolates of Pseudomonas aeruginosa, obtained from these contaminated hatcheries and resulting broiler chicks with omphalitis, were selected to determine virulence of the bacteria. One-day-old specific-pathogen-free white leghorn chicks were placed into positive pressure isolation units (10 chicks per unit); feed and water were provided ad libitum. The five isolates of P. aeruginosa (1 x 10(1) or 1 x 10(1) colony-forming units/bird) were used to challenge two replicates of 10 chicks via yolk sac inoculation. Two control groups were injected with 0.1 ml of phosphate-buffered saline, and two groups received no treatment. Mortality was recorded daily, and the chicks that died were necropsied and liver and yolk sacs were cultured. After 14 days, the remaining chickens were euthanatized and necropsied. Bacterial isolates retrieved from liver and yolk sacs were identified by the API 20 NE typing system to confirm that they were the same as the challenge isolate. Virulence varied greatly among the isolates, resulting in mortality rates from 0 to 90%. The challenge isolates produced different and often distinctive postmortem lesion patterns. Antibiotic sensitivity tests showed that all five isolates were resistant to sulfisoxazole, ceftiofur, penicillin, lincomycin, bacitracin, oxytetracycline, erythromycin, naladixic acid, and tetracycline. The isolates varied in sensitivity to other antibiotics, but all isolates were sensitive to gentamicin.     

Change in antimicrobial resistance pattern in Salmonella enterica serovar Typhimurium isolates detected in a beef cattle farm

Multidrug-resistant Salmonella enterica serovar Typhimurium (S. Typhimurium) isolates with four different antimicrobial resistance patterns obtained from a beef cattle farm were characterized to determine their clonality. Macrorestriction analysis of genomic DNA revealed that these four isolates are closely related to each other and can be classified as a newly emerged pulsed-field gel electrophoresis type among cattle: cluster VII. Three of the four isolates showed resistance to extended-spectrum cephalosporins (ESCs), and this resistance was mediated by AmpC β-lactamase encoded by the bla(CMY-2) gene in a 190-kbp IncA/C plasmid. Results of restriction analysis and IncA/C backbone PCR suggest that the three 190-kbp plasmids are identical and that a 70-kbp IncA/C plasmid of the ESC-susceptible isolate is derived from the 190-kbp plasmid by a deletion event. Three isolates harboured a virulence-resistance plasmid (165 or 180 kbp), and restriction analysis revealed that these plasmids were identical or closely related to each other. These results suggest that the four S. Typhimurium cluster VII isolates originate from a common ancestor that probably invaded the farm prior to the salmonellosis outbreak. Antimicrobial resistance patterns may not necessarily reflect the relationships of the isolates.     

Characterization of integron mediated antimicrobial resistance in Salmonella isolated from diseased swine

Forty-two Salmonella isolates obtained from diseased swine were genetically characterized for the presence of specific antimicrobial resistance mechanisms. Twenty of these isolates were characterized as S. Typhimurium DT104 strains. Pulsed-field gel electrophoresis was used to determine genetic relatedness and revealed 20 distinct genetic patterns among the 42 isolates. However, all DT104 isolates fell within 2 closely related genetic clusters. Other Salmonella isolates were genetically grouped together according to serotype. All DT104 isolates displayed the penta-resistance phenotype to ampicillin, chloramphenicol, streptomycin, sulfamethoxazole, and tetracycline. Resistance to sulfamethoxazole, tetracycline, streptomycin, kanamycin, and ampicillin was most common among the non-DT104 Salmonella isolates. All DT104 strains contained 2 chromosomal integrons of 1000 and 1200 base pairs. The DNA sequencing revealed that the 2 integrons contained genes encoding a resistance to streptomycin and ampicillin, respectively. None of the non-DT104 strains showed the same pattern, although several strains possessed integrons of 1000 base pairs or larger. However, the majority of non-DT104 Salmonella strains did not possess any integrons. Two Salmonella isolates displayed tolerance to the organic solvent cyclohexane, indicating the possibility that they are overexpressing chromosomal regulatory genes marA or soxS or the associated multidrug efflux pump, acrAB. This research suggests that integrons contribute to antimicrobial resistance among specific swine Salmonella serotypes; however, they are not as widely disseminated among non-Typhimurium swine Salmonella serotypes as previously thought.     

Clonal relatedness of Salmonella isolates associated with invasive infections in captive and wild-caught rattlesnakes

This study examines the serotype distribution and clonal relatedness among Salmonella isolates obtained from healthy and diseased snakes. Isolates from extraintestinal body sites were obtained through routine diagnostic lab submissions from snakes in two facilities that had experienced a high prevalence of osteomyelitis in Crotalus species. Gastrointestinal isolates were predominantly from fecal samples collected from healthy snakes of both crotalid and non-crotalid species in one facility. PFGE macrorestriction analysis of Salmonella isolates confirmed the clonal and species-restricted nature of Salmonella serotype IIIa 56: z4, z23: - in one facility. Fourteen of 15 isolates from suspected osteomyelitis lesions in wild-caught snakes at the second facility were also from Salmonella subgroup IIIa (serotype IIIa 18: z4, z23: -) and appeared to be closely related by PFGE. Evaluation of a PCR assay for the spvC gene in 209 isolates demonstrated that this method consistently distinguished isolates of subgroup IIIa from those of subgroup IIIb. The data presented establish that Salmonella of subgroup IIIb are abundant and regularly associated with gastrointestinal shedding in snakes but that Salmonella in subgroup IIIa disproportionately cause infections in bone or other extraintestinal sites.     

Distribution,quantitative load and characterization of Salmonella associated with swine farms in upper-northern Thailand

This study was conducted to analyze the prevalence and quantitative loads of Salmonella spp. on pig farms in Chiang Mai, Lamphun, Thailand to assess loading levels before slaughtering. The serotype diversity, antimicrobial-resistance pattern and pulse-field type of Salmonella spp. were also characterized to assess the dynamic propagation of the pathogen. The Salmonella-positive prevalence was 246/805 (30.56%), and the quantitative loads varied from 1.48~4.04 Log₁₀MPN/g, with a mean ± standard deviation of 2.11 ± 0.57. AMP/S/TE (ampicillin/streptomycin/tetracycline) was the highest frequency antimicrobial resistance pattern found in this study. In addition, Salmonella Rissen was the primary serotype in this region. PFGE results indicated the occurrence of infection by cross contamination among pig farms. Our study showed that pork is easily contaminated with this pathogen. Farm control programs must be based on strict biosecurity and hygienic measures, which could further reduce the contamination pressure at slaughterhouses or retail shops.