Induction of seroconversion and persistence of Salmonella Typhimurium in pigs are strain dependent

Foodborne salmonellosis is one of the most important bacterial zoonotic diseases worldwide. Salmonella Typhimurium is the serovar most frequently isolated from persistently infected slaughter pigs in Europe. Salmonella Typhimurium pathogenesis is host species specific. In addition, differences in in vitro behaviour of Salmonella Typhimurium strains have also been described, which may be reflected by a different course of infection within a host species. We compared the course of a Salmonella Typhimurium infection in pigs, using two Salmonella Typhimurium strains that were able to interfere with MHC II expression on porcine macrophages to a different extent in vitro. After experimental inoculation, blood and faecal samples from all pigs were collected at regular time points. At 40 days post inoculation (pi), animals were euthanized and tissue samples were bacteriologically analysed. The proportion of serologically positive piglets at 33 days pi was significantly higher in pigs that were inoculated with the strain that did not downregulate MHC II expression in vitro. Furthermore, this strain was less frequently shed and isolated in lower numbers from tonsils and ileocaecal lymph nodes than the strain that was able to markedly downregulate MHC II expression in vitro. We thus found that the delayed onset of seroconversion after oral inoculation of piglets with a particular Salmonella Typhimurium strain coincided with higher faecal shedding and increased persistence. Strain specific differences in Salmonella pathogenesis might thus have repercussions on the serological detection of Salmonella Typhimurium infections in pigs.     

Expression of inflammatory cytokines (TNF-alpha, IL-1, IL-6 and IL-8) in colon of pigs naturally infected with Salmonella typhimurium and S. choleraesuis

The expression of mRNA encoding tumour necrosis factor-alpha (TNF-alpha), interleukin-1 (IL-1), IL-6 and IL-8 was studied, by in situ hybridization with a non-radioactive digoxigenin-labelled probe, in formalin-fixed, paraffin wax-embedded colonic tissue from pigs naturally infected with Salmonella typhimurium and S. choleraesuis. By in situ hybridization, a distinct positive signal for TNF-alpha, IL-1, IL-6 and IL-8 was detected in colon from all 12 infected pigs. Hybridization signals for all four inflammatory cytokines were detected primarily inflammatory cells infiltrating the lamina propria and submucosa. In comparison, expression of all four inflammatory cytokines was minimal in non-lesional colon of infected pigs and in normal colon from control pigs. The results suggest that these cytokines play an important role in the pathophysiological processes in porcine salmonellosis.     

The significance of the serotype in the clinical and pathological features of naturally occurring porcine salmonellosis.

Salmonellosis was the diagnosis in 63 of 327 consecutive porcine necropsy accessions during a one year period in Indiana, an incidence four to ten times that previously reported in other areas. A herd outbreak was usually either a septicemic disease characterized by sudden mortality. Little overlap between these two syndromes were seen. Animals dying with the septicemic form usually yielded Salmonella choleraesuis var. kunzendorf on culture. Salmonella typhimurium was the usual isolate from cases of enteric salmonellosis. Oral inoculation of SPF pigs with S. typhimurium resulted in a disease similar to naturally occurring enteric salmonellosis. The pathology of porcine salmonellosis due to S. typhimurium is described.     

Attenuation of virulence of Lawsonia intracellularis after in vitro passages and its effects on the experimental reproduction of porcine proliferative enteropathy

Non-pathogenic Lawsonia intracellularis variants have been obtained through multiple passages in cell culture but there is no information regarding the number of passages necessary to attenuate a pathogenic isolate. The present study evaluated the susceptibility of pigs to L. intracellularis after 10, 20 and 40 passages in vitro. Three groups (six animals/group) were inoculated with pure culture of L. intracellularis on passage 10, 20 or 40 and one group with placebo. The animals were monitored for clinical signs, fecal shedding and serological IgG response during 28 days post-inoculation. Gross and histologic lesions and the level of infection based on the amount of L. intracellularis-specific antigen in the intestinal mucosa identified by immunohistochemistry were evaluated in two animals from each group on days 14, 21 and 28. Animals inoculated with passages 10 and 20 demonstrated proliferative lesions typical of porcine proliferative enteropathy associated with the presence of Lawsonia-specific antigen in the intestinal mucosa. Passage 40-inoculated pigs did not show proliferative lesions or presence of Lawsonia antigen at any time point throughout the study. Similar patterns of the fecal shedding were observed in passage 10 and 20-infected pigs but those infected with passage 40 shed for a short period. Serological IgG responses in passage 10 and 20-inoculated pigs were detected from day 14 post-infection but not at all in passage 40-inoculated animals. These results demonstrate attenuation of the virulence properties of L. intracellularis between 20 and 40 cell passages in vitro. This information will be valuable for design of future experimental models and for studying the mechanisms involved in the attenuation of L. intracellularis virulence.     

6-hydroxydopamine-mediated release of norepinephrine increases faecal excretion of Salmonella enterica serovar Typhimurium in pigs

Salmonella enterica serovar Typhimurium is an animal and zoonotic pathogen of worldwide importance. In pigs, transport and social stress are associated with reactivation and spread of Salmonella Typhimurium infection. The stress-related catecholamine norepinephrine (NE) has been reported to activate growth and virulence factor expression in Salmonella; however the extent to which NE contributes to stress-associated salmonellosis is unclear. We studied the impact of releasing NE from endogenous stores during Salmonella Typhimurium infection of pigs by administration of 6-hydroxydopamine (6-OHDA), which selectively destroys noradrenergic nerve terminals. Treatment of pigs with 6-OHDA 7 or 16 days post-oral inoculation with Salmonella Typhimurium produced elevated plasma NE levels and transiently, but significantly, increased faecal excretion of the challenge strain. Oral administration of NE to Salmonella Typhimurium-infected pigs also transiently and significantly increased shedding; however pre-culture of the bacteria with NE did not alter the outcome of infection. Salmonella has been proposed to sense and respond to NE via a homologue of the adrenergic sensor kinase QseC. A ΔqseC mutant of Salmonella Typhimurium was consistently excreted in lower numbers than the parent strain post-oral inoculation of pigs, though not significantly so. 6-OHDA treatment of pigs infected with the ΔqseC mutant also increased faecal excretion of the mutant strain, albeit to a lesser extent than observed upon 6-OHDA treatment of pigs infected with the parent strain. Our data support the notion that stress-related catecholamines modulate the interaction of enteric bacterial pathogens with their hosts.     

Evaluation of Salmonella enterica serovar Typhimurium and Choleraesuis slyA mutant strains for use in live attenuated oral vaccines

SlyA protein plays a key role in virulence in Salmonella enterica. In this study, we evaluated the ability of the slyA mutant strain of S. enterica serovar Choleraesuis (S. choleraesuis) to protect against swine salmonellosis. Using a murine model infected with S. enterica serovar Typhimurium (S. typhimurium), we showed that the Salmonella strain with a deletion of slyA could be used as a highly immunogenic, effective and safe vaccine in mice. Based on these data, a slyA mutant of S. enterica serovar Choleraesuis strain RF-1 was constructed, and the ability of this mutant to protect immunized pigs from S. choleraesuis infection was examined. As with the S. typhimurium slyA mutant, immunization of pigs with the S. choleraesuis slyA mutant strain provided significant protection against subsequent challenge by the wild-type RF-1. These results demonstrate that SlyA is a potential target in the development of a novel live attenuated vaccine against S. enterica.     

Phage Types of Salmonella enterica ssp. enterica serovar Typhimurium Isolated from Production Animals and Humans i Denmark

S. Typhimurium is one of the 2 most common salmonella serotypes causing human salmonellosis in Denmark. In order to illustrate the significance of different production animals as a source of infection, 1461 isolates were characterized by phage typing. The isolates originated from human patients and from cattle, pigs and poultry. By phage typing the isolates could be separated in 35 different phage types. Five types (10, 12, 66, 110 and 135) predominated and comprised 78.8% of the isolates. In humans, 57.3% of the isolates were phage type 12. This phage type was also predominant in pig herds and, to a lesser degree, in cattle. Phage types 110, 120, 135 and 193 constituted 86.5% of the poultry isolates while these phage types only made up 12.9% of the human isolates. The investigation showed that pigs are probably a major source of S. Typhimurium infection in humans in Denmark today.     

Isolation and identification of Salmonella spp. from red foxes (Vulpes vulpes) and badgers (Meles meles) in northern Italy

Background

Salmonella spp. have been isolated from a wide range of wild animals. Opportunistic wild carnivores such as red foxes (Vulpes vulpes) and badgers (Meles meles) may act as environmental indicators or as potential sources of salmonellosis in humans. The present study characterizes Salmonella spp. isolated from the intestinal contents of hunted or dead red foxes (n = 509) and badgers (n = 17) in northern Italy.

Findings

Thirty-one strains of Salmonella belonging to 3 Salmonella enterica subspecies were isolated. Fourteen different serovars of S. enterica subsp. enterica were identified, among which were serovars often associated with human illness.

Conclusions

Wild opportunistic predators can influence the probability of infection of both domestic animals and humans through active shedding of the pathogen to the environment. The epidemiological role of wild carnivores in the spread of salmonellosis needs to be further studied.     

Seroprevalence of antibodies to Salmonella spp in semidomesticated reindeer in Norway,determined by enzyme-linked immunosorbent assay

An indirect ELISA was developed as a possible tool to detect the seroprevalence of antibodies to Salmonella spp in semidomesticated reindeer. To cover a broad spectrum of serogroups a lipopolysaccharide mix of S. typhimurium and S. choleraesuis was used as antigen in this pilot study. Sera from 31 culture-negative reindeer with no clinical or historical evidence of salmonellosis were used as negative serum control. After immunisation with an inactivated S. typhimurium vaccine, pooled sera from 6 reindeer were used as positive serum control as no serum from naturally infected animals was available. A seroprevalence of 0.6% in 2000 clinically healthy, slaughter-reindeer from Norway was determined by using this ELISA. No more information on Salmonella in reindeer in Norway is known to the authors. This is the first ELISA established for indirect detection of Salmonella in reindeer.     

Occurrence of Giardia and Cryptosporidium in pigs on Prince Edward Island, Canada

In a cross-sectional study of 633 pigs from 21 herds on Prince Edward Island, Canada (PEI), the prevalence of infection with Cryptosporidium and Giardia, and the genotypes and species of isolates were determined in order to establish the zoonotic potential of pigs in this region. As determined by direct immunofluorescence microscopy (DFA), 18 herds (86%) and 163 animals (26%; 95% CI: 22-29%) tested positive for Cryptosporidium, while just 3 herds (14%) and 6 animals (1%; 95% CI: 0.4-2%) tested positive for Giardia. Cryptosporidium spp. isolates were detected in 39% (95% CI: 34-44%) of weanlings (1-3 months of age) and 9% (95% CI: 6-13) of sows (>8 months of age). Molecular characterization using the 18S rDNA and HSP70 gene fragments revealed the presence of Cryptosporidium sp. pig genotype II, C. suis, C. parvum, and Cryptosporidium sp. mouse genotype. Among the 113 isolates of Cryptosporidium spp. successfully genotyped, pig genotype II (61%) predominated, with C. suis (36%) being the next most prominant isolate. C. parvum (2%; two isolates) and Cryptosporidium sp. mouse genotype (0.9%; one isolate) were only occasionally isolated. The only two Cryptosporidium-positive genotyped isolates from sows included one each of C. suis and Cryptosporidium sp. pig genotype II.All but one of the six Giardia positive isolates were detected in weanling pigs. None of the Giardia-positive isolates was amenable to PCR. This study demonstrates that Cryptosporidium spp. are highly prevalent in pigs on PEI, Canada, are found mostly in weanlings (1-3 months of age). Furthermore, the pigs are primarily infected by the host-specific genotypes and species, Cryptosporidium sp. pig genotype II and C. suis, whereas the zoonotic C. parvum is rare. Giardia duodenalis is only occasionally found in pigs. These findings suggest that domestic pigs on PEI, Canada, likely do not pose a significant health risk to humans from these parasites.     

Comparative plasmid profile analysis of Salmonella typhimurium var. Copenhagen strains from a Salmonella outbreak in hospitalized horses]

From April 1990 through June 1991 clinical salmonellosis and asymptomatic faecal excretion of Salmonella spp. were seen in hospitalized horses at two veterinary hospitals. 76 Salmonella strains from hospitalized horses and 18 strains from horses without any clinical contact were characterized by serotyping and plasmid profile analysis. From April 1990 through January 1991 97.8% of the hospitalized horses were infected with strains of S. typhimurium var. Copenhagen, which were closely related according to their similar plasmid patterns. Other strains of S. typhimurium var. Copenhagen and serotype S. enteritidis were isolated not before February 1991. In the same period various plasmid profile types of S. typhimurium var. Copenhagen and strains of S. typhimurium, S. enteritidis and S. lexington were isolated from horses of the control group. Our results suggest that the high incidence of salmonellosis and latent salmonella infection in hospitalized horses was mainly due to the spread of one particular strain of S. typhimurium var. Copenhagen and that this strain was obviously acquired during hospitalisation.     

Prevalence and antimicrobial susceptibility of Salmonella spp. in small Indian mongooses (Herpestes auropunctatus) in Grenada,West Indies

Intestinal samples from 156 small Indian mongooses (Herpestes auropunctatus) collected island-wide in Grenada from April 2011 to March 2013 were examined for the presence of Salmonella enterica spp. Nineteen (12%) mongooses were culture-positive for S. enterica spp. of which five serotypes were identified. Salmonella javiana and S. Montevideo were the most commonly isolated serotypes. The other serotypes isolated were S. Rubislaw, S. Panama and S. Arechavaleta. All isolates were susceptible to amoxicillin–clavulanic acid, ampicillin, cefotaxime, ceftazidime, ciprofloxacin, enrofloxacin, gentamicin, nalidixic acid, imipenem and trimethoprim–sulfamethoxazole. One isolate (S. Montevideo) showed resistance to tetracycline and intermediate resistance to streptomycin. The five isolated Salmonella serotypes are potential human pathogens suggesting that the mongoose may play a role in the epidemiology of human salmonellosis in Grenada.     

A report of an outbreak of Salmonella oranienburg in a hybrid mouse colony

Mice in a hybrid breeding colony developed symptoms compatible with a diagnosis of salmonellosis. Salmonella oranienburg was recovered from feces by conventional bacteriological techniques. Active salmonellosis appeared to be precipitated by the stress of pregnancy among breeder female BALB/c mice. Necropsy examination of the BALB/c breeder females revealed diarrhea, roughened hair coats, enlarged spleens, white foci in the liver, and skin abscesses. S. oranienburg was recovered from the spleen, cecum, oropharynx, uterus, and skin abscesses of affected mice. DBA/2N breeder males were asymptomatic and S. oranienburg was recovered from the cecum only. Breeding productivity indices dropped to unacceptable levels in rooms containing infected animals. Relatively normal production was observed among uninfected animals of the same strains in adjacent rooms, thus demonstrating the detrimental effect S. oranienburg had on this breeder colony.     

Detection of Actinobacillus pleuropneumoniae Infection in Pigs

It is difficult to control the spread of porcine haemophilus pleuropneumonia caused by Actinobacillus pleuropneumoniae because there is no sensitive and specific way to accurately determine whether or not a pig herd is infected. This paper reports bacteriological and serological techniques used to detect A. pleuropneumoniae infection in pigs from a herd with endemic disease.

The bacteria were isolated from the anterior nasal mucosa of grower pigs, but not from younger or older pigs. Bacteriological culture of several tissues from the respiratory tract showed that nine of ten young finishing pigs were infected, but culture of lung tissue from slaughtered hogs detected infection in only 39 of 288 (13.5%). Both cooler storage temperature and use of selective medium prolonged the time that lung tissue could be stored and the organism still recovered. An enzyme-linked immunosorbent assay detected serotype-specific antibodies in serum of infected pigs.

     

Influence of long-time transportation stress on re-activation of Salmonella typhimurium DT104 in experimentally infected pigs

In this study a Salmonella Typhimurium infection model in swine was used in order to investigate the influence of pre-mortal stress induced by long time period transportation on the re-activation of Salmonella in experimentally infected pigs. Salmonella free pigs were exposed to a highly virulent strain of Salmonella Typhimurium DT104 by direct intragastrical administration. Clinical parameters were monitored and the shedding rate in faeces was qualitatively and quantitatively determined by standard bacteriological procedures for 21 days. The distribution of the challenge organism in 14 different internal organs of transported and nontransported animals was determined. All infected animals developed clinical signs of salmonellosis 12 to 24 hours post infection. About 88 to 100% of the fecal samples were culture-positive up to post exposure day 6, and then varied from 71 to 92% until slaughter, respectively. At necropsy S. Typhimurium was recovered most frequently from caecum and ileocolic lymph nodes (83%), colon (79%), palatine tonsils (71%) and mandibular lymph nodes (62.5%). A negative impact of transportation stress on the shedding rate and the general condition of the animals was observed.     

The rate of Salmonella spp. infection in zoo animals at Seoul Grand Park, Korea

Salmonellosis is an important zoonotic disease that affects both people and animals. The incidence of reptile-associated salmonellosis has increased in Western countries due to the increasing popularity of reptiles as pets. In Korea, where reptiles are not popular as pets, many zoos offer programs in which people have contact with animals, including reptiles. So, we determined the rate of Salmonella spp. infection in animals by taking anal swabs from 294 animals at Seoul Grand Park. Salmonella spp. were isolated from 14 of 46 reptiles (30.4%), 1 of 15 birds (6.7%) and 2 of 233 mammals (0.9%). These findings indicate that vigilance is required for determining the presence of zoonotic pathogen infections in zoo animals and contamination of animal facilities to prevent human infection with zoonotic diseases from zoo facilities and animal exhibitions. In addition, prevention of human infection requires proper education about personal hygiene.     

Novel insights into transmission routes of Mycobacterium avium in pigs and possible implications for human health

Mycobacterium avium infection is a severe condition in humans, whereas pigs are often subclinically infected. Pig carcasses represent a possible source of human infection. Faecal excretion of M. avium was recently demonstrated in experimentally infected pigs, along with detection of M. avium in apparently normal lymph nodes. The present study investigates faecal excretion in naturally infected herds and the presence of live mycobacteria in lymph nodes. Two pig herds (A and B), with a history of sporadically suspected M. avium infection were sampled. Herd B used peat, as opposed to Herd A. Samples from peat, sawdust, drinking water, faeces and lymph nodes were collected. Identification of mycobacteria was performed by 16S rDNA sequencing and PCR. Mycobacterium avium isolates were analysed by Multi-Locus Variable Number of Tandem repeat Analysis (MLVA). Mycobacterium avium subsp. hominissuis was detected in samples of faeces, peat and lymph nodes from Herd B, often with identical MLVA profiles. Additionally, other non-tuberculous mycobacteria (NTM) were found in the same material. The absence of macroscopic lymph node lesions in the presence of M. avium subsp. hominissuis was frequently demonstrated. In Herd A, only one NTM isolate, which proved not to be M. avium, was found. Faeces might facilitate transmission of M. avium subsp. hominissuis between pigs and maintain the infection pressure in herds. The low incidence of macroscopic lesions together with the massive presence of M. avium subsp. hominissuis in lymph nodes from pigs kept on peat raises questions related to animal husbandry, food safety and human health.     

Ascaris suum in Saskatchewan Pigs: An Abattoir Survey of Prevalence and Intensity of Infection

In a survey of 2500 market weight pigs in a Saskatchewan abattoir, 37% were infected with adult Ascaris suum and in 46% there were milkspot hepatic lesions. A total of 60% of the pigs examined had some evidence of A. suum infection. Most infected pigs contained less than 50 adult ascarids.

In a second abattoir survey of a further 2500 market weight pigs, 44% of the animals had milk-spot hepatic lesions. In approximately 12% of the affected pigs these lesions were very severe and a very similar proportion of pigs' livers were condemned or sent for use as animal feed.

The results of these surveys are discussed with particular reference to the significance of A. suum infection in growing pigs in Saskatchewan.

     

Individual risk factors for Mycoplasma hyopneumoniae infections in suckling pigs at the age of weaning

Background

In recent years, the occurrence and the relevance of Mycoplasma hyopneumoniae infections in suckling pigs has been examined in several studies. Whereas most of these studies were focused on sole prevalence estimation within different age groups, follow-up of infected piglets or assessment of pathological findings, none of the studies included a detailed analysis of individual and environmental risk factors. Therefore, the aim of the present study was to investigate the frequency of M. hyopneumoniae infections in suckling pigs of endemically infected herds and to identify individual risk factors potentially influencing the infection status of suckling pigs at the age of weaning.

Results

The animal level prevalence of M. hyopneumoniae infections in suckling pigs examined in three conventional pig breeding herds was 3.6% (41/1127) at the time of weaning. A prevalence of 1.2% was found in the same pigs at the end of their nursery period. In a multivariable Poisson regression model it was found that incidence rate ratios (IRR) for suckling pigs are significantly lower than 1 when teeth grinding was conducted (IRR: 0.10). Moreover, high temperatures in the piglet nest during the first two weeks of life (occasionally >40°C) were associated with a decrease of the probability of an infection (IRR: 0.23-0.40). Contrary, the application of PCV2 vaccines to piglets was associated with an increased infection risk (IRR: 9.72).

Conclusions

Since single infected piglets are supposed to act as initiators for the transmission of this pathogen in nursery and fattening pigs, the elimination of the risk factors described in this study should help to reduce the incidence rate of M. hyopneumoniae infections and thereby might contribute to a reduced probability of high prevalences in older pigs.     

Salmonella typhimurium DT104: a virulent and drug-resistant pathogen.

Salmonella typhimurium phage type (PT) or definitive type (DT) 104 is a virulent pathogen for humans and animals, particularly cattle. It has been isolated increasingly from humans and animals in the United Kingdom and several other European countries and, more recently, in the United States and Canada. Humans may acquire the infection from foods of animal origin contaminated with the infective organism. Farm families are particularly at risk of acquiring the infection by contact with infected animals or by drinking unpasteurized milk. The symptoms in cattle are watery to bloody diarrhea, a drop in milk production, pyrexia, anorexia, dehydration and depression. Infection may result in septicemic salmonellosis and, upon necropsy, a fibrinonecrotic enterocolitis may be observed. The infection occurs more commonly in the calving season than at other times. Feedlot cattle and pigs may also be affected. Prolonged carriage and shedding of the pathogen may occur. Symptoms in humans consist of diarrhea, fever, headache, nausea, abdominal pain, vomiting, and, less frequently, blood in the stool. Salmonella typhimurium DT104 strains are commonly resistant to ampicillin, chloramphenicol, streptomycin, sulfonamides, and tetracycline.