Evaluation of a French ELISA for the detection of Salmonella Enteritidis and Salmonella Typhimurium in flocks of laying and breeding hens.

In France, the regular and compulsory detection of Salmonella Enteritidis (SE) and Salmonella Typhimurium (ST) in flocks of breeding and laying hens is based on bacteriological examination of environmental swabs and faeces samples. The aim of this study was to compare this bacteriological examination with a serological method (ELISA) developed in our laboratory. This ELISA was first evaluated by use of artificially infected hens. During these experimental infection studies, several groups of hens were inoculated with SE, ST, different vaccines and different Salmonella serovars to calculate the experimental parameters of our ELISA. Then, in a field study, 43 flocks were followed monthly using two bacteriological samples (environmental swab and pool of faeces) and 20 serological samples (sera or yolks). Twenty-seven flocks without SE or ST gave a negative serological response throughout their surveillance. Among the 10 various serovars different from SE and ST isolated in this study, S. Heidelberg, S. Agona and S. Hadar gave seropositive results in seven flocks. Consequently, this ELISA was not specific of SE and ST as it detected serovars sharing or not common antigens with SE and ST. Seropositive results were also obtained each month for two flocks where no Salmonella could be isolated. Finally, in seven flocks found infected with SE or ST, the positive ELISA results appeared later than the bacteriological detection. Therefore, for the detection of chicken flocks recently infected with SE or ST, bacteriological examination currently used in France seems to be more appropriate than this ELISA.

Longitudinal serological responses to Salmonella enterica of growing pigs in a subclinically infected herd.

A longitudinal survey was conducted in France in a subclinically Salmonella-infected farrow-to-finish pig farm to describe the time-course of the serological response to Salmonella enterica in growing pigs. We used three batches of sows and their corresponding litters (n = 31 litters). Among these, 256 pigs randomly selected and individually identified were followed from the first week of age until slaughter. Serial individual blood samples were submitted to indirect Salmonella-ELISA testing. Salmonella shedding was investigated by bacteriological testing of faecal material regularly collected from the sows and pigs and by environment swabs taken from the pens. Caecal contamination was checked at the slaughterhouse. Information about litter composition (filiation), location of the pigs in post-weaning and fattening pens, sanitary events, sex and body weights was recorded. 11.6% of the pigs shed S. enterica; 52% of pigs seroconverted before slaughter. The age-related variation of the natural logarithm of calibrated optical densities (COD) of pigs was described with two linear mixed models. From 8 to 61 days of age, the decrease in COD with age was fitted with a model including random effects of the animal and the dam on the intercept and slope, a batch random effect on the intercept and an individual birth-weight fixed effect on the intercept. The dam random effect was explained by the parity of the sow, Salmonella shedding by the sow during the farrowing phase and the value of the optical density of colostrum collected at parturition. A second model fitting the increase in COD from 61 days of age until slaughter included the random effect on intercept of the batch and the random effects on slope and intercept of the animal, the dam and the pen in which the followed animals were located during the fattening phase and the environmental contamination as fixed effect. In this second model, no relation was found between individual slaughter-bacteriological results and increasing COD values. Considering seroconversion time between 61 days of age and slaughter, survival function were constructed using the Cox proportional-hazards model. Both methods suggested that seroconversions generally occurred during the last third of the fattening phase (from 140 days of age to slaughter), while shedding was observed during the first half of the fattening period. The fitted models suggest the existence of clusters (such as pen and litter of origin).

Contamination of pigs by nose-to-nose contact or airborne transmission of Salmonella Typhimurium.

The aim of this study is to assess the risk of contamination by Salmonella Typhimurium of pigs by nose-to-nose contact or the airborne route. Thirty twelve-week-old SPF pigs were divided into 4 groups housed in 4 different rooms: the first room contained Salmonella-free control pigs (n = 4), the second room had 10(3) CFU S. Typhimurium inoculated pigs (n = 5) and non-inoculated "contact" pigs (n = 4), the third room had pigs (n = 8) receiving potentially contaminated air from the following room through a hole (4 pigs housed in the pen situated near the hole and 4 pigs in the pen at the opposite side of the room), and the fourth room had pigs (n = 5) inoculated with 10(6) CFU Salmonella Typhimurium and also non inoculated "contact" pigs (n = 4). The "contact" and the inoculated pigs were housed in adjacent pens allowing nose-to-nose contact. The 5 pigs orally inoculated with 10(6) CFU S. Typhimurium were bacteriologically and serologically positive 1 week later and their environment was contaminated as early as 1 day pi. The faecal samples of 4 nose-to-nose contact pigs were bacteriologically positive and one of them was seropositive 5 weeks pi before the pigs were commingled. The 8 pigs housed in the third room received S. Typhimurium by an active airflow coming from the contaminated room (1000 m3/hour). Their faecal samples remained negative until 8 weeks pi but the environmental swabs taken in the room close to the airinlet were contaminated 2 days pi and positive swabs were found elsewhere in the room 5 weeks pi. Two seropositive pigs were encountered 8 weeks pi in the pen situated near the hole. Only one among the 5 pigs inoculated with 10(3) CFU had bacteriologically positive faeces 1-week pi and the 4 pigs kept in nose-to-nose contact with them remained negative. A dose of 10(3) CFU was too small to induce persistent excretion and to stimulate a humoral immune response. However, the dose of 10(6) CFU induced contamination of nose-to-nose contact pigs and contamination of the environment by airflow.

Development of a complete ELISA using Salmonella lipopolysaccharides of various serogroups allowing to detect all infected pigs.

Although poultry is recognized as the major source of food-poisoning caused by Salmonella, pork also contributes to human infections. This study was therefore undertaken in order to develop a reliable serological method for the evaluation of the Salmonella status of piglets. A complete ELISA was performed using lipopolysaccharides of Salmonella Typhimurium, Anatum, Hadar and Infantis because these serovars were representative of the serogroups isolated from 30 contaminated fattening farms. S. Enteritidis was also added because of its importance in human infection and to include the O:9 antigen. This method potentially detects 100% of infected pigs. A significant correlation was found between this serological method and the bacteriological data from mesenteric lymph nodes (p = 0.01). In addition, both sensitivity and specificity were high (97% and 94% respectively). The ELISA test was therefore used in a cross-sectional study on 4 farms to evaluate when pigs became contaminated: seropositive pigs were only found for the 20 week old finishing pigs. The antibody response to Salmonella in piglets was also investigated: maternal antibodies persisted until 7 weeks of age and post-Salmonella contamination seroconversion was detected from 8 weeks of age onwards.

Genetic resistance to mortality of day-old chicks and carrier-state of hens after inoculation with Salmonella enteritidis.

The heritability of resistance of poultry to Salmonella enteritidis (SE) was investigated. Three measurements of resistance were made: survival after intramuscular inoculation of 419 day-old chicks, absence versus presence of Salmonella in spleens and caeca 4 weeks after oral inoculation of 304 hens at peak of laying, and antibody response of 228 hens following two inoculations of an aroA mutant of this serotype. In the first two models of infection, resistance appeared to be heritable. The heritability was estimated from the sire and dam components, respectively, at 0.14 ± 0.10 and 0.62 ± 0.16 for chick mortality, 0.47 ± 0.21 and 0.13 ± 0.26 for resistance to spleen contamination, and 0.24 ± 0.15 and 0.53 ± 0.26 for resistance to caecal contamination in laying hens. By contrast the estimated heritability of antibody response was very low (0.03 ± 0.08 and 0.10 ± 0.08 when estimated from the sire and dam components, respectively). These results suggest that a selection for increased resistance to SE may be efficient.

Vaccination du pigeon contre Salmonella typhimurium.

Les salmonclles constituent l'une des preoccupations majeures des eleveurs de pigeons. Aussi, afin de diminuer le portage et d'assurer la protection des animaux ainsi que celle des consommateurs, des moyens de detection et de prevention doivent etre developpes. Dans ce cadre, il a ete mis au point une technique ELISA pour detecter les anticorps anti-Salmonella typhimurium chez des pigeons recevant un autovaccin. Le vaccin a entraine une protection de 85% des pigeons vaccines a 7 et 11 semaines et de 50% des sujets vaccines a 11 semaines, alors que les temoins non vaccines sont tous morts apres l'epreuve a 13 semaines. La vaccination n'exclut pas le portage ni l'excretion.

World Health Organisation--supervised interlaboratory comparison of ELISAs for the serological detection of Salmonella enterica serotype Enteritidis in chickens.

A collaborative exercise, supervised by the World Health Organisation, was set up to compare ELISAs used for the serological detection of Salmonella enteritica serotype Enteritidis in chickens. The aim was to ascertain how far agreement could be reached on the interpretation of optical density readings for high titre, intermediate titre and low titre sera. Two sets of sera were sent to 14 participants. The first set compared high, medium and low titre sera raised in specified-pathogen-free and commercial broiler breeder chickens. The second set comprised 20 sera of different antibody titres raised in commercial birds reared under laboratory conditions and sent blind. Both indirect and double-antibody sandwich blocking ELISAs were used with a number of different detecting antigens. With a few exceptions good agreement was reached on the interpretation of results obtained from high and low titre sera from the optical density obtained with a single serum dilution. Differences were observed in the interpretation of medium titre sera. The results suggested that most ELISAs produce reasonably comparable results and that practical problems may arise from interpretation of the results mainly as a result of the choice of the criteria used for differentiating sera obtained from infected and uninfected chickens. These problems are discussed.