Response of layer and broiler strain chickens to parenteral administration of a live Salmonella Typhimurium vaccine.

Responses to the parenteral administration of a live aroA deletion Salmonella serovar Typhimurium vaccine given to three brown egg layer strains and two broiler strains were studied. Twenty-five birds of each strain were reared together in floor pens to 6 weeks of age and then moved as individual strains to new floor pens and injected with 10(8) colony forming units (CFU) per bird of the vaccine bacteria intramuscularly or subcutaneously, 10(6) CFU per bird subcutaneously, or phosphate buffered saline (PBS) subcutaneously as a vaccination control. Three birds of one layer strain were injected intramuscularly with 0.5mg/ bird S. Typhimurium lipopolysaccharide (LPS) to evaluate whether response was similar for vaccine and endotoxin. Birds were weighed, and rectal temperatures recorded at the time of injection, then observed over 24 hours. Rectal temperatures were measured and blood samples collected for serum IL-6 assay at 3 hours post injection (PI). At 12 hours PI blood samples were drawn for analyses for plasma phosphorus (P), glucose (Glu), cholesterol (Cho), aspartate transaminase (AST), total protein (Ptn) and creatinine kinase (CK). Blood was sampled 14 days PI and tested for serum antibody to S. Typhimurium. Vaccination resulted in significant seroconversion by 14 days PI in all strains compared to the controls. The three layer strains exhibited a clinical malaise, evident within 90 minutes of injection, lasting for 12 hours, with complete recovery by 24 hours PI. Only the 10(8) CFU dose given subcutaneously produced an increase in rectal temperature 3 hours PI. Vaccination had no effect on IL-6 or Ptn. All vaccine doses increased P and the higher dose by either route decreased Cho in all bird strains. The 10(8) vaccine dose increased Glu and intramuscular injection markedly elevated CK only in the layer strains. The response was not completely congruous with that to LPS alone. The results highlight the need for consideration of differences in response of bird strain when consideration is given to the parenteral administration of live Salmonella vaccines.

Development and validation of a drag swab method using tampons and different diluents for the detection of members of Salmonella in broiler houses.

Members of the genus Salmonella represent a significant public health concern and also a colonizer of commercial poultry. Therefore, the early detection and management of colonized broiler breeders and their progeny is essential. There have been numerous methods for farm-based detection, with gauze-based drag swabs being the most commonly used. In the present study, the wet (boiled water, buffered peptone water and double-strength skin milk) tampon was compared with the gauze to determine the recovery rate (10(2) colony-forming units/swab) of five common poultry serovars of Salmonella and after cold (4°C/48 h) storage. The recovery was found to be equivalent when tested using the ISO6572:2002 method, for all diluents (Cohen's κ =1.0; sensitivity = 1.0; specificity = 1.0). The subsequent field trial (n = 15 farms) compared the tampon drag swab (TDS) with a statistically appropriate (90% confidence, detect 10% prevalence) number of faecal swabs (n = 22), which also showed high agreement between the TDS and faecal sampling (κ = 0.86; McNemar's χ(2) = 1.0; sensitivity = 0.9; specificity = 1.0). However, direct faecal sampling showed a wider diversity of serovars of Salmonella than the corresponding TDS. The TDS is a very sensitive, readily available and cost-effective screening method for salmonellas in broiler breeder houses. This TDS technique may be used for routinely screening of broiler houses, and faecal sampling would only be used to confirm colonization or contamination, and to measure flock serovar variance.

Utilization of a novel autologous killed tri-vaccine (serogroups B [Typhimurium], C [Mbandaka] and E [Orion]) for Salmonella control in commercial poultry breeders.

An autologous killed trivalent vaccine (3x10(8) colony-forming units [CFU]), based on three Salmonella serovars (Typhimurium - serogroup B, Mbandaka - serogroup C, and Orion - serogroup E) prevalent in the flocks of Australian poultry companies, was developed using Salenvac techniques. At 20 weeks, hens vaccinated at 12 and 17 weeks as well as non-vaccinated hens were challenged (250 microl of 10(7) CFU) with autologous and heterologous serovars belonging to serogroup B (Typhimurium and Agona), serogroup C (Mbandaka and Infantis) and serogroup E (Orion and Zanzibar). Overall, vaccination resulted in a significant difference in carriage of Salmonella between non-vaccinated and vaccinated commercial Cobb hens (P <0.05) for serogroups B and C. However, due to low colonization rates in the non-vaccinated birds, no significant difference (P>0.05) could be determined for serogroup E. All vaccinated flocks produced a significant antibody response (P<0.001) to the S. Typhimurium vaccine strain, measured using a S. Typhimurium enzyme-linked immunosorbent assay (Guildhay), which peaked at 20 weeks of age, with 39% of the hens positive. Maternal antibodies were detected in 16% of the yolks from eggs produced by these flocks. There was a significant difference after challenge with Salmonella (P <0.05) among 1-day-old chicks from vaccinated versus non-vaccinated parents, when challenged using 10(4) CFU but not when challenged with 10(8) CFU. The success of this trial resulted in the incorporation of this vaccine into a Salmonella control system in commercial broiler breeder production.

The interplay between Campylobacter and Helicobacter species and other gastrointestinal microbiota of commercial broiler chickens.

BACKGROUND: Poultry represent an important source of foodborne enteropathogens, in particular thermophilic Campylobacter species. Many of these organisms colonize the intestinal tract of broiler chickens as harmless commensals, and therefore, often remain undetected prior to slaughter. The exact reasons for the lack of clinical disease are unknown, but analysis of the gastrointestinal microbiota of broiler chickens may improve our understanding of the microbial interactions with the host. METHODS: In this study, the fecal microbiota of 31 market-age (56-day old) broiler chickens, from two different farms, was analyzed using high throughput sequencing. The samples were then screened for two emerging human pathogens, Campylobacter concisus and Helicobacter pullorum, using species-specific PCR. RESULTS: The gastrointestinal microbiota of chickens was classified into four potential enterotypes, similar to that of humans, where three enterotypes have been identified. The results indicated that variations between farms may have contributed to differences in the microbiota, though each of the four enterotypes were found in both farms suggesting that these groupings did not occur by chance. In addition to the identification of Campylobacter jejuni subspecies doylei and the emerging species, C. concisus, C. upsaliensis and H. pullorum, several differences in the prevalence of human pathogens within these enterotypes were observed. Further analysis revealed microbial taxa with the potential to increase the likelihood of colonization by a number of these pathogens, including C. jejuni. CONCLUSION: Depletion of these taxa and the addition of taxa that compete with these pathogens, may form the basis of competitive exclusion strategies to eliminate them from the gastrointestinal tract of chickens.

A novel miniaturized most probable number method for the enumeration of Campylobacter spp. from poultry-associated matrices.

A novel miniaturized most probable number (mMPN) method was developed for the enumeration of thermophilic Campylobacter spp. using a modification of blood-free Bolton broth (supplemented with 25mg/l of sulfamethoxazole) and CampyFood ID agar. The mMPN was evaluated by comparison with direct plating (modified ISO/TS, 10272-2:2006) for the analysis of samples (n=149) representing various poultry matrices (carcases, broiler ceca and feces, scald tank water and feed). A sensitivity of 95%, specificity of 90% and Cohen KAPPA agreement of 0.84 was determined for the mMPN method compared to direct plating. Quantitative comparison found 83% of enumerations to be less than ±1log10 different (Student's t-test P<0.001). Financial analysis showed that the mMPN required 51% less media and 60% less labor than the direct plating protocol. The mMPN provides a method that can be used for complete through-chain analysis that has a single enrichment step and multiple dilutions to extinction for a variety of samples (containing low, medium and high populations).

Diversity and toxigenicity among members of the Bacillus cereus group.

Members of the Bacillus cereus group were isolated from rice products by centrifugation-plating and conventional spread-plating methods. Random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) results showed broad diversity among the strains and revealed some associations among isolates from raw and cooked rice samples, at the genotypic level. A comparatively greater diversity among strains was observed in isolates from raw rice than those from cooked rice and, generally, the RAPD profiles of isolates from raw and cooked rice were different, with only a few of them common to both types of rice. The toxigenic potential of the isolates was also determined by molecular and immunoassay analyses. The results revealed that most isolates from the B. cereus group were potentially or actually toxigenic, and some isolates could produce both diarrhoeal and emetic toxins. Generally, isolates belonging to the B. cereus group with the same RAPD pattern were shown to have a similar profile of enterotoxigenicity.