Can a combination of vaccination, probiotic and organic acid treatment in layer hens protect against early life exposure to Salmonella Typhimurium and challenge at sexual maturity?

Day old layer chicks were challenged with Salmonella Typhimurium using a seeder bird technique. Treatment groups were untreated control, administration of a probiotic in drinking water weekly, vaccination by intramuscular injection of a live aro-A deletion mutant vaccine at 10 weeks of age (woa) followed by an oral dose at 16 woa, probiotic administration plus vaccination, vaccination plus the administration of an organic acid preparation in feed from 16 woa and a combination of probiotic, vaccine and organic acid. Faecal shedding was monitored by culture at 1, 2, 3, 4, 8, 12, 15, 17, 20, 21, 23 and 25 woa and in dust from settle plates by PCR at intervals from 8 woa. Birds from each group were separated at 17 and 18 woa and challenged orally with 106 CFU of S. Typhimurium. Both untreated and probiotic groups shed Salmonella until 56 days. Salmonella was also detected in dust from 8 until 12 woa but little after this. After vaccination, from sexual maturity (18 woa) all groups except those that were vaccinated with and without probiotic re-excreted Salmonella. The probiotic alone was ineffective against this re-excretion and all groups receiving organic acids shed Salmonella. At 17 woa, unchallenged controls were fully susceptible to caecal colonization, however all other groups showed reduced susceptibility, including the untreated challenged group. However, at 18 woa (sexual maturity) only the groups that were vaccinated with or without probiotic showed reduced susceptibility to colonization. The organic acid treated groups (including the vaccinated group) did not show a difference to the untreated controls. S. Typhimurium demonstrated an ability to re-emerge at sexual maturity, similar to other serovars. The vaccine assisted in limiting the re-excretion at sexual maturity and decreased susceptibility to subsequent challenge. Use of a probiotic augmented the vaccine's protective capacity.

A practical method for assessing infectious laryngotracheitis vaccine take in broilers following mass administration in water: Spatial and temporal variation in viral genome content of poultry dust after vaccination.

Infectious laryngotracheitis is an important disease of chickens caused by infectious laryngotracheitis virus (ILTV). Outbreaks commonly occur in meat chicken flocks and mass vaccination with live attenuated vaccines, usually in water, is used to control the disease in these populations. Vaccination with live virus via water and nipple drinkers requires stringent adherence to protocols to ensure success, but vaccine administration monitoring is not currently assessed due to a lack of economically viable methods. Vaccinal ILTV has been shown to be detectable in dust in experimental studies and has potential as a method of assessing vaccination success. However, the pattern of vaccinal ILTV detection in dust following vaccination under commercial conditions has not been defined. We report the longitudinal profile of ILTV genome copies (GC) in poultry house dust collected on settle plates following vaccination of 8 flocks of commercial meat chickens on four farms. ILTV GC was enumerated using quantitative real-time polymerase chain reaction (qPCR). There was considerable variation between flocks in the levels of ILTV GC detected post vaccination and this variation was significantly associated with vaccine take measured in individual birds in a companion study. There was no effect of sampling location on ILTV GC in dust but the amount of dust collected was greater in locations closer to the exhaust fans in artificially ventilated houses. Results indicate that measurement of ILTV GC in single or pooled dust samples at 7-8 days post vaccination enables detection of poor vaccine takes and provides a practical means of monitoring ILT vaccination.

Uptake and spread of infectious laryngotracheitis vaccine virus within meat chicken flocks following drinking water vaccination.

Vaccination against infectious laryngotracheitis virus (ILTV) in commercial broiler flocks in the field, which is only undertaken in the face of a local outbreak, requires mass administration techniques, usually via drinking water. This is often fraught with difficulties such as variable vaccination "reactions" and sometimes, vaccination failure. Laboratory testing of the outbreak strains however invariably shows the vaccines in use to be protective. To investigate this paradox, the dynamics of an ILT vaccine virus was examined within broiler flocks during a natural outbreak. In an initial flock, 70 birds were individually identified and had tracheal swabs collected sequentially at intervals from 1 to 26 days after vaccination and submitted for ILTV detection using qPCR. This evaluation was extended by collection of tracheal swabs from 40 to 45 random birds at 4, 7-8, 12-13 and 25-26 days post vaccination (pv) across a further 7 flocks. The results showed a very variable early uptake of vaccine virus from the drinking water (between 3% and 52% of tested birds with detectable virus in trachea at 4 days pv) and revealed that actual vaccination of the flocks relied on bird to bird transmission of the vaccine virus. In flocks with very low (<10%) initial bird uptake, successful exposure of vaccine virus to the majority of the flock can be delayed, leaving a large proportion of birds as susceptible at the likely time of possible exposure to wild virus. This may explain the cases of apparent failure of vaccination in the field. The variable bird to bird spread can be associated with reversion to virulence, this may explain the rolling vaccine reactions often observed. The variation in initial vaccine uptake may be affected by some factors involved with the administration technique and this requires further study in a larger sample size.

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, application, and results of routine monitoring of Marek's disease virus in broiler house dust using real-time quantitative PCR.

Results are presented from four studies between 2002 and 2011 into the feasibility of routinely monitoring Marek's disease virus serotype 1 (MDV-1) in broiler house dust using real-time quantitative PCR (qPCR) measurement. Study 1 on two farms showed that detection of MDV-1 occurred earlier on average in dust samples tested using qPCR than standard PCR and in spleen samples from five birds per shed assayed for MDV-1 by qPCR or standard PCR. DNA quality following extraction from dust had no effect on detection of MDV-1. Study 2 demonstrated that herpesvirus of turkeys (HVT) and MDV serotype 2 (MDV-2) in addition to MDV-1 could be readily amplified from commercial farm dust samples, often in mixtures. MDV-2 was detected in 11 of 20 samples despite the absence of vaccination with this serotype. Study 3 investigated the reproducibility and sensitivity of the qPCR test and the presence of inhibitors in the samples. Samples extracted and amplified in triplicate showed a high level of reproducibility except at very low levels of virus near the limit of detection. Mixing of samples prior to extraction provided results consistent with the proportions in the mixture. Tests for inhibition showed that if the template contained DNA in the range 0.5-20 ng/microl no inhibition of the reaction was detectable. The sensitivity of the tests in terms of viral copy number (VCN) per milligram of dust was calculated to be in the range 24-600 VCN/mg for MDV-1, 48-1200 VCN/mg for MDV-2, and 182-4560 VCN/mg for HVT. In study 4 the results of 1976 commercial tests carried out for one company were analyzed. Overall 23.1% of samples were positive for MDV-1, 26.1% in unvaccinated and 16.4% in vaccinated chickens. There was marked regional and temporal variation in the proportion of positive samples and the MDV-1 load. The tests were useful in formulating Marek's disease vaccination strategies. The number of samples submitted has increased recently, as has the incidence of positive samples. These studies provide strong evidence that detection and quantitation of MDV-1, HVT, and MDV-2 in poultry house dust using qPCR is robust, sensitive, reproducible, and meaningful, both biologically and commercially. Tactical vaccination based on monitoring of MDV-1 rather than routine vaccination may reduce selection pressure for increased virulence in MDV-1.