Astrovirus infections pose a significant problem in the poultry industry, leading to multiple adverse effects such as a decreased egg production, breeding disorders, poor weight gain, and even increased mortality. The commonly observed chicken astrovirus (CAstV) was recently reported to be responsible for the “white chicks syndrome” associated with an increased embryo/chick mortality. CAstV-mediated pathogenesis in chickens occurs due to complex interactions between the infectious pathogen and the immune system. Many aspects of CAstV–chicken interactions remain unclear, and there is no information available regarding possible changes in gene expression in the chicken spleen in response to CAstV infection. We aim to investigate changes in gene expression triggered by CAstV infection. Ten 21-day-old SPF White Leghorn chickens were divided into two groups of five birds each. One group was inoculated with CAstV, and the other used as the negative control. At 4 days post infection, spleen samples were collected and immediately frozen at −70 °C for RNA isolation. We analyzed the isolated RNA, using RNA-seq to generate transcriptional profiles of the chickens’ spleens and identify differentially expressed genes (DEGs). The RNA-seq findings were verified by quantitative reverse-transcription PCR (qRT-PCR). A total of 31,959 genes was identified in response to CAstV infection. Eventually, 45 DEGs (p-value < 0.05; log₂ fold change > 1) were recognized in the spleen after CAstV infection (26 upregulated DEGs and 19 downregulated DEGs). qRT-PCR performed on four genes (IFIT5, OASL, RASD1, and DDX60) confirmed the RNA-seq results. The most differentially expressed genes encode putative IFN-induced CAstV restriction factors. Most DEGs were associated with the RIG-I-like signaling pathway or more generally with an innate antiviral response (upregulated: BLEC3, CMPK2, IFIT5, OASL, DDX60, and IFI6; downregulated: SPIK5, SELENOP, HSPA2, TMEM158, RASD1, and YWHAB). The study provides a global analysis of host transcriptional changes that occur during CAstV infection in vivo and proves that, in the spleen, CAstV infection in chickens predominantly affects the cell cycle and immune signaling.

The only knowledge of the molecular structure of European turkey coronaviruses (TCoVs) comes from France. These viruses have a quite distinct S gene from North American isolates. The aim of the study was to estimate the prevalence of TCoV strains in a Polish turkey farm during a twelve-year period, between 2008 and 2019, and to characterize their full-length S gene. Out of the 648 flocks tested, 65 (10.0%, 95% CI: 7.9–12.6) were positive for TCoV and 16 of them were molecularly characterized. Phylogenetic analysis showed that these strains belonged to two clusters, one formed by the early isolates identified at the beginning of the TCoV monitoring (from 2009 to 2010), and the other, which was formed by more recent strains from 2014 to 2019. Our analysis of the changes observed in the deduced amino acids of the S1 protein suggests the existence of three variable regions. Moreover, although the selection pressure analysis showed that the TCoV strains were evolving under negative selection, some sites of the S1 subunit were positively selected, and most of them were located within the proposed variable regions. Our sequence analysis also showed one TCoV strain had recombined with another one in the S1 gene. The presented investigation on the molecular feature of the S gene of TCoVs circulating in the turkey population in Poland contributes interesting data to the current state of knowledge.

Introduction: Due to their immunostimulatory properties TLR ligands are used prophylactically to protect against a variety of viral and bacterial pathogens in mammals. Knowledge of the molecular and functional aspects of TLRs is essential for a better understanding of the immune system and resistance to diseases in birds. For that reason, this study attempted to determine the impact of TLR21 stimulation by its synthetic ligand (CpG ODN, class B) on the chicken immune system. Material and Methods: Sixty embryonated chicken eggs were randomly allocated into three groups (control and two experimental groups). On day 18 of embryonic development, chickens in one experimental group were administered in ovo a low dose of CpG ODN and the birds of the second experimental group were given a high dose of the ligand. Spleens were collected at 1, 2, 5, and 10 days post-hatching (dph) for analysis of IFN-α, IFN-β, IFN-γ, IL-6, and IL-10 expression using qRT-PCR. Results: Significant differences were observed in mRNA expression levels of all the measured cytokines associated with the modulation and regulation of the immune response at different time points. Conclusion: The obtained data clearly demonstrate that immune response induction takes place after in ovo administration of class B CpG ODN, and that the ligand has the ability to induce cytokine responses in neonatal chicken spleen.

MiRNAs represent ubiquitous regulators of gene expression and play an important and pivotal regulatory role in viral disease pathogenesis and virus–host interactions. Although previous studies have provided basic data for understanding the role of miRNAs in the molecular mechanisms of viral infection in birds, the role of miRNAs in the regulation of host responses to chicken astrovirus (CAstV) infection in chickens is not yet understood. In our study, we applied next-generation sequencing to profile miRNA expression in CAstV-infected chickens and to decipher miRNA-targeted specific signaling pathways engaged in potentially vital virus-infection biological processes. Among the 1354 detected miRNAs, we identified 58 mature miRNAs that were significantly differentially expressed in infected birds. Target prediction resulted in 4741 target genes. GO and KEGG pathway enrichment analyses showed that the target genes were mainly involved in the regulation of cellular processes and immune responses.

Infectious bronchitis virus (IBV) of the GI-23 lineage, which first emerged in the Middle East in the late 1990s, has since spread worldwide. The factors driving its expansion, whether human involvement, wild bird migration, or the virus’s biological traits, are still unclear. This study aimed to trace the genome evolution of GI-23 IBV in chickens and its adaptability to quails, which are susceptible to both gamma- and deltacoronaviruses. Thirty specific-pathogen-free (SPF) birds, aged between two and three weeks, were used. Initially, three birds were inoculated with the G052/2016 IBV via the oculo-nasal route. On the third day post-infection (dpi), oropharyngeal swabs were collected from the whole group, pooled, and subsequently used to infect three next birds. This process was repeated nine more times during consecutive IBV passages (P-I–P-X), and eventually, virus sequencing was performed using Next-Generation Sequencing (NGS). The obtained results showed that quails were not susceptible to the IBV GI-23 lineage, as the virus RNA was detected in low amounts only during the first passage (QP-I) with no further detections in later rounds of IBV passaging. In chickens, only mild diarrhea symptoms appeared in a few individuals. The NGS analysis identified sixty-two single nucleotide variants (SNVs), thirty of which caused amino acid changes, twenty-eight were synonymous, and one SNV introduced a stop codon. Three SNVs were found in untranslated regions. However, none of these SNVs lasted beyond seven passages, with forty-four being unique SNVs. The Shannon entropy values measured during passages varied for pol1a, pol1b, S, 5a, 5b, and N genes, with overall genome complexity peaking at CP-VI and CP-X. The highest complexity was observed in the pol1a (CP-X) and S genes (CP-IV, CP-VI, CP-VIII, and CP-X). Along with the S gene that was under positive selection, eight codons in pol1a were also positively selected. These findings suggest that even in an adapted host, IBV variability does not stabilize without immune pressure, indicating continuous molecular changes within its genome.

Highly pathogenic avian influenza viruses (HPAIVs) of the H5N8 subtype have caused severe outbreaks in poultry and wild birds worldwide. Disease outcome differs markedly between avian species: chickens typically develop acute infection with high mortality, whereas ducks often exhibit milder or delayed disease. The molecular basis underlying these species-specific differences in early host responses remains incompletely understood. To characterize early transcriptional responses to HPAIV infection, RNA sequencing (RNA-seq) was performed on lung tissues from chickens and ducks experimentally infected with H5N8 HPAIV at 24 hours post-inoculation (hpi). Differential gene expression analysis and functional enrichment analyses were conducted, and selected host genes were validated by quantitative real-time PCR. At 24 hpi, a comparable number of differentially expressed genes (DEGs) was identified in chickens and ducks; however, the organization of the transcriptional response differed markedly between species. Chickens exhibited a pathway-centered transcriptional profile dominated by innate immune and inflammatory signaling, whereas ducks displayed a more distributed and balanced transcriptional response spanning multiple functional categories. Functional enrichment analyses revealed strong activation of interferon- and cytokine-associated pathways in chickens, while transcriptional changes in ducks were dispersed across processes related to RNA metabolism, antiviral regulation, and cellular homeostasis. Among species-specific differentially expressed genes, MOV10L1 showed opposite regulation in these bird species. These findings demonstrate that early host responses to H5N8 HPAIV infection differ between chickens and ducks not only in gene identity but also in transcriptional organization. The distributed response architecture observed in ducks may contribute to controlled antiviral responses with limited immunopathology. This study provides a comparative transcriptomic framework for understanding species-specific host responses to HPAIV infection in poultry.