Toll-like receptor 4 (TLR4), which recognizes lipopolysaccharide from Gram-negative bacteria, plays a major role in resistance of mice and humans to Salmonella infection. In chickens, Salmonella may establish a carrier state whereby bacteria are able to persist in the host organism without triggering clinical signs. Based on cellular morphological parameters, we developed a method, without using antibodies, to separate three cecal cell subpopulations: lymphocytes, enterocytes, and a population encompassing multiple cell types. We analyzed the mRNA expression of TLR4, interleukin-1 beta (IL-1 beta), IL-8, IL-12, and lipopolysaccharide-induced tumor necrosis factor alpha factor (LITAF) in cecal subpopulations of chicks from inbred lines resistant or susceptible to the carrier state infected with Salmonella enterica serovar Enteritidis. The results showed that resistance to the carrier state in chicks is associated with a larger percentage of lymphocytes and with higher levels of expression of TLR4 and IL-8 at homeostasis in the three cell subpopulations, as well as with a higher level of expression of LITAF in lymphocytes during the carrier state. In contrast to the early phase of infection, the carrier state is characterized by no major cell recruitment differences between infected and noninfected animals and no significant modification in terms of TLR4, IL-1 beta, IL-8, IL-12, and LITAF expression in all cell subpopulations measured. However, TLR4 expression increased in the lymphocytes of chicks from the susceptible line, reaching the same level as that in infected chicks from the resistant line. These observations suggest that the carrier state is characterized by a lack of immune activation and highlight the interest of working at the level of the cell population rather than that of the organ.

Dose–response experiments characterize the relationship between infectious agents and their hosts. These experiments are routinely used to estimate the minimum effective infectious dose for an infectious agent, which is most commonly characterized by the dose at which 50 per cent of challenged hosts become infected—the ID50. In turn, the ID50 is often used to compare between different agents and quantify the effect of treatment regimes. The statistical analysis of dose–response data typically makes the assumption that hosts within a given dose group are independent. For social animals, in particular avian species, hosts are routinely housed together in groups during experimental studies. For experiments with non-infectious agents, this poses no practical or theoretical problems. However, transmission of infectious agents between co-housed animals will modify the observed dose–response relationship with implications for the estimation of the ID50 and the comparison between different agents and treatments. We derive a simple correction to the likelihood for standard dose–response models that allows us to estimate dose–response and transmission parameters simultaneously. We use this model to show that: transmission between co-housed animals reduces the apparent value of the ID50 and increases the variability between replicates leading to a distinctive all-or-nothing response; in terms of the total number of animals used, individual housing is always the most efficient experimental design for ascertaining dose–response relationships; estimates of transmission from previously published experimental data for Campylobacter spp. in chickens suggest that considerable transmission occurred, greatly increasing the uncertainty in the estimates of dose–response parameters reported in the literature. Furthermore, we demonstrate that accounting for transmission in the analysis of dose–response data for Campylobacter spp. challenges our current understanding of the differing response of chickens with respect to host-age and in vivo passage of bacteria. Our findings suggest that the age-dependence of transmissibility between hosts—rather than their susceptibility to colonization—is the mechanism behind the ‘lag-phase’ reported in commercial flocks, which are typically found to be Campylobacter free for the first 14–21 days of life.

Foot-and-mouth disease vaccine potency testing involving live virus challenge can be problematical in pigs. Alternative methods of assessing vaccine efficacy are therefore desirable. Here we investigate the link between IL-6 in blood at time of challenge and protection against challenge by carrying out statistical analyses utilising data from six separate potency tests performed in swine with the aim of assessing whether IL-6 could be exploited as an additional parameter for confirming vaccine efficacy in pigs. These analyses confirmed that systemic IL-6 levels increased when the administered vaccine dose increased and that the odds of protection against challenge increased as IL-6 levels increased. The link between increased protection and increased antibody was reaffirmed and a significant link between IL-6 levels and antibody levels was shown. We therefore conclude that quantifying the levels of IL-6 in serum could provide additional means of qualifying whether a vaccine will afford clinical protection or not in pigs, in the absence of an actual challenge, and thus offer the possibility of improved vaccine potency testing in pigs both in terms of animal welfare as well as cost.

Targeting dendritic cells (DC) is key to driving effective immune responses. Lymphatic cannulation provides access to the heterogeneous populations of DC draining peripheral sites in rodents and ruminants. Afferent lymph DEC-205(+) CD11c(+) SIRP alpha(+) DC were preferentially infected ex vivo with three vaccine viral vectors: recombinant human replication-defective human adenovirus 5 (rhuAdV5), recombinant modified vaccinia virus Ankara (rMVA), and recombinant fowlpox virus (rFPV), all expressing green fluorescent protein (GFP). The rhuAdV5-infected cells remained viable, and peak GFP expression was observed 16 to 24 h posttransduction. Increasing the incubation period of DC with rhuAdV5 enhanced GFP expression. In contrast, DC infected with rMVA-GFP or rFPV-GFP became rapidly apoptotic and GFP expression peaked at 6 h postinfection. Delivery of foot-and-mouth disease virus (FMDV) A(22) antigen to DC by rhuAdV5-FMDV-A(22) ex vivo resulted in significantly greater CD4(+) T cell proliferation than did delivery by rFPV-FMDV-A(22). Delivery of rhuAdV5-GFP in oil adjuvant in vivo, to enhance DC-vector contact, resulted in increased GFP expression in migrating DC compared to that with vector alone. Similarly, CD4(+) T cell responses were significantly enhanced when using rhuAdV5-FMDV-A(22) in adjuvant. Therefore, the interaction between viral vectors and afferent lymph DC ex vivo can predict the outcome of in vivo immunization and provide a means of rapidly assessing the effects of vector modification.