Lined seahorse, Hippocampus erectus, is an important aquatic animal due to its medicinal and ornamental purposes. However, our understanding of the viral spectrum in H. erectus is still limited. Here, we studied the viruses in H. erectus using meta-transcriptomic sequencing. A total of 213,770,166 reads were generated and assembled de novo into 539 virus-associated contigs. Three novel RNA viruses from the Astroviridae, Paramyxoviridae, and Picornaviridae families were finally identified. In addition, we identified a strain of nervous necrosis virus from H. erectus. In particular, the unhealthy group showed a higher viral diversity and abundance than the normal group. These results revealed the diversity and cross-species transmission of viruses in H. erectus and highlighted the threat of viral infections to H. erectus.

Reticuloendotheliosis is an immunosuppressive disease caused by avian reticuloendotheliosis virus (REV). It is commonly found in poultry farms and has caused a notable economic loss worldwide. Despite this, there is currently no effective vaccine available to protect against REV infection. In this study, gp90 protein derived from an REV isolated from a contaminated vaccine was co-administered with cytosine-phosphate-guanine oligodeoxynucleotide (CpG-ODN) adjuvant to hens to determine if it protects their chicks against REV infection. To synthesize the gp90 protein, the gp90 gene was amplified using polymerase chain reaction, expressed in Escherichia coli, and purified. The resulting recombinant protein was injected intramuscularly into breeder hens along with CpG-ODN adjuvant and then serum antibody levels were regularly evaluated. After the fertilized eggs from these vaccinated hens had hatched, the resulting chicks were challenged with a 102.7 50% tissue culture infectious dose (TCID50) of REV at 1 day old and the REV antibody levels in these hatched chickens were evaluated before and after the challenge. Viremia and growth rate were measured weekly and statistically analyzed. The results suggest that the gp90 recombinant protein was successfully prepared and, when used with CpG-ODN adjuvant to immunize breeder hens, induced serological antibody production against REV in both hens and their hatched chicks. In addition, the maternal antibodies induced by the gp90 protein vaccine effectively protected majority of the chicks from REV infection. Overall, we found the gp90 protein obtained in this study may be a potential vaccine candidate that had good immunogenicity and could be an auxiliary measure to accelerate the eradication of REV.

Goose astrovirus (GoAstV) has frequently been isolated in China since it was first identified as the etiological agent of visceral gout in goslings in 2017. However, the actual prevalence of GoAstV infection and its economic impact on commercial goose production remain poorly characterized. Here, virus detection and serological testing were conducted to determine the extent of GoAstV infection in commercial goose flocks. We detected GoAstV RNA in 2 (6/300) of dead-in-shell embryos and day-old hatched goslings by RT-PCR, indicating vertical transmission under natural conditions. Using a virus neutralization test, GoAstV antibodies were detected in 41.761.1 of serum samples from four commercial goose flocks, indicating that infections were common. To determine the virus types circulating in the commercial flocks, we isolated 15 GoAstVs from goose tissue samples from farms located in five provinces during 20182022. Genomic sequence analysis showed that all sequences were corresponded to GoAstV group 2 (GoAstV-2) but were assigned into three capsid subgroups based on sequence variations in the capsid protein. Representative isolates of capsid subgroups were also antigenically evaluated using cross-neutralization tests in LMH cell cultures. The antigenic relatedness values (R) calculated using the Horsfall formula were between 62 and 86, indicating that no significant antigenic differences exist between the isolates. Our findings indicate that GoAstV-2 viruses are an important cause of fatal gout in goose flocks, as well as hatchery contamination in China.

Both Chicken anemia virus (CAV) and Gyrovirus galga 1 (GyVg1) belong to the family Anelloviridae and genus Gyrovirus. They are non-enveloped, single-stranded circular DNA viruses that can infect chicken populations, leading to similar clinical symptoms such as growth retardation and immunosuppression, making clinical differentiation challenging. This study established a duplex TaqMan real-time PCR assay to simultaneously detect CAV and GyVg1. Specific primers and probes were developed to target the VP3 gene of CAV (labeled with VIC) and the VP1 gene of GyVg1 (labeled with FAM). The optimization process involved fine-tuning reaction conditions, such as primer/probe concentrations and annealing temperature, through a systematic matrix approach. The assay demonstrated remarkable linearity under optimized conditions (0.4 μmol/L primers, 0.2 μmol/L probes, and an annealing temperature of 52 °C), yielding high R² values of 0.9973 for CAV and 0.9981 for GyVg1. The method exhibited high specificity, showing no cross-reactivity with other common avian pathogens. The limits of detection were 1.0 × 10⁰ copies/μL for CAV and 1.0 × 10¹ copies/μL for GyVg1, making it 10-fold more sensitive than conventional PCR. Intra-assay and inter-assay coefficients of variation were below 1.5% and 2.5%, respectively, indicating excellent reproducibility. When tested on 411 clinical samples from Shandong Province, CAV showed a positivity rate of 48.66% (200/411), GyVg1 18.73% (77/411), and a co-detection rate of 17.52% (72/411). Moreover, 25 complete genome sequences of GyVg1 (2376 bp each) were acquired from positive samples. These sequences exhibited a nucleotide identity ranging from 93.6% to 99.3% among the isolates and 92.2% to 99.6% with reference strains. The phylogenetic analysis, which was based on whole-genome sequences, categorized the Shandong isolates into four clusters (I–IV). Notably, the strains were dispersed across these clusters and were intertwined with references from various regions and hosts. This observation suggests the absence of distinct geographic clustering and implies frequent cross-regional and cross-species transmission. In summary, this duplex real-time PCR assay offers a sensitive, specific, and dependable method for concurrently detecting and distinguishing CAV and GyVg1. The study’s findings reveal a significant co-infection rate and genetic variation in GyVg1, underscoring the importance of increased surveillance and deeper exploration of its pathogenicity and transmission patterns.