Avian metapneumovirus subtype B vaccination in commercial broiler chicks: heterologous protection and selected host transcription responses to subtype A or B challenge.

Avian metapneumovirus (aMPV) causes respiratory disease and drops in egg production in chickens, and is routinely controlled by vaccination. However, the host's immune response to virulent challenge in vaccinated or unvaccinated broiler chickens is poorly characterized. We show that subtype B vaccination offers heterologous (subtype A challenge) and homologous (subtype B challenge) protection. Subtype B challenge caused significantly greater humoral antibody titres in vaccinated and unvaccinated chickens. In turbinate and lung tissues of unvaccinated-challenged chickens, IgA and IgY mRNA transcription was significantly up-regulated after subtype B challenge compared to subtype A. Cellular immunity (CD8-α and CD8-ß) gene transcripts were significantly up-regulated during early and later stages of infection from subtype B or subtype A, respectively. Immune gene transcriptional responses (IL-1ß, IL-6 and IL-18) were significantly up-regulated after challenge. Gene transcription results showed that mRNA expression levels of CD8-α, CD8-ß, TLR3 and IL-6, particularly in turbinate and trachea tissues, are useful parameters to include in future aMPV vaccination-challenge studies.

Host immune response to infectious bronchitis virus Q1 in two commercial broiler chicken lines.

This study investigated the pathogenesis of infectious bronchitis virus (Gammacoronavirus) strain Q1 in two commercial broiler chicken lines, and the host immune response to infection. Chicks from each line were grouped into either infected or control. Following Q1 infection at day-old, fast (Line-A) and slow (Line-B) growing chicks were monitored for clinical signs and body weights. At 3, 7, 9, 14, 21 and 28 days post infection (dpi), five birds were humanely euthanised, and trachea, kidney and proventriculus tissues were collected for quantitative RT-PCR and histopathology. Blood was collected weekly to determine IBV-specific ELISA antibody titres. Q1 infection significantly reduced the body weights of Line-A chicks at 14 and 21 dpi, but there were no significant differences in Line-B. Through qRT-PCR, significantly higher viral loads were found in the trachea, proventriculus and kidney tissues of Line-A chicks at 7-9 dpi. At day-old and at 28 dpi, the mean antibody titre in Line-B was notably higher than Line-A. Significant IFN-α mRNA expression was noted in the trachea and kidneys of Line-A, whereas no change occurred in Line-B. Chicks in Line-B, compared to those in Line-A, demonstrated a tissue-dependent increase of IFN-ß, TLR3, IL-1ß and IL-6 and LITAF gene transcription responses to IBV Q1. It appears that the level of maternal antibodies, growth rates, and other inherent host genetic factors could have influenced the differences in viral loads and immune responses.

Evaluation of full S1 gene sequencing of classical and variant infectious bronchitis viruses extracted from allantoic fluid and FTA cards.

Sequence variability in the S1 gene determines the genotype of infectious bronchitis virus (IBV) strains. A single RT-PCR assay was developed to amplify and sequence the full S1 gene for six classical and variant IBVs (M41, D274, 793B, IS/885/00, IS/1494/06 and Q1) enriched in allantoic fluid (AF) or the same AF inoculated onto Flinders Technology Association (FTA) cards. Representative strains from each genotype were grown in specific-pathogen-free eggs and RNA was extracted from AF. Full S1 gene amplification was achieved using primer A and primer 22.51. Products were sequenced using primers A, 1050+, 1380+ and SX3+ to obtain short sequences covering the full gene. Following serial dilutions of AF, detection limits of the partial assay were higher than those of the full S1 gene. Partial S1 sequences exhibited higher-than-average nucleotide similarity percentages (79%; 352 bp) compared to full S1 sequences (77%; 1756 bp), suggesting that full S1 analysis allows greater strain differentiation. For IBV detection from AF-inoculated FTA cards, four serotypes were incubated for up to 21 days at three temperatures, 4°C, room temperature (approximately 24°C) and 40°C. RNA was extracted and tested with partial and full S1 protocols. Through partial sequencing, all IBVs were successfully detected at all sampling points and storage temperatures. In contrast, using full S1 sequencing it was not possible to amplify the gene beyond 14 days or when stored at 40°C. Data presented show that for full S1 sequencing, a substantial amount of RNA is needed. Field samples collected onto FTA cards are unlikely to yield such quantity or quality. ABBREVIATIONS: AF: allantoic fluid; CD50: ciliostatic dose 50; FTA: Flinders Technology Association; IB: infectious bronchitis; IBV: infectious bronchitis virus.