However, it should be noted that at several of these sites, only small sample numbers were obtained, which lowers the confidence that this population is truly negative. in New Zealand. The serological survey indicated that both RCV and RHDV are widespread in New Zealand wild rabbits, with antibodies detected in 10 out of 14 and 12 out of 14 populations, respectively. Two closely related RCV strains were identified in the duodenal tissue from a New Zealand wild rabbit (RCV Gore-425A and RCV Gore-425B). Both variants are most closely related to Australian RCV strains, but with 88% nucleotide identity, they are genetically distinct. Phylogenetic analysis revealed that the New Zealand RCV strains fall within the genetic diversity of the Australian RCV isolates, indicating a relatively recent movement of RCVs between Australia and New Zealand. IMPORTANCE Wild rabbits are important and Salicin (Salicoside, Salicine) damaging introduced vertebrate pests in Australia and New Zealand. Although RHDV1 is used as a biological control agent, some nonpathogenic RCVs can provide partial immunological cross-protection against lethal RHDV infection and thus interfere with its effectiveness for rabbit control. The presence of nonpathogenic RCVs in New Zealand wild rabbits has been long hypothesized, but earlier attempts to isolate a New Zealand RCV strain have been unsuccessful. Therefore, it is important to determine if such nonpathogenic viruses exist in New Zealand rabbits, especially considering the proposed introduction of new RHDV strains into New Zealand as biocontrols. within the genus (1). RHDV was first reported in China in 1984 and causes acute necrotizing hepatitis with a high mortality rate Salicin (Salicoside, Salicine) in European rabbits (= 0.036) and sex (= 0.019) on the seroprevalence of RCV, which was best explained using a multiterm model (i.e., age + sex). The model coefficients were age = ?0.01 (standard error [SE] = 0.05) and sex = ?0.73 (SE = 0.31). This model had the lowest Akaike’s information criterion value (245.5) of all models, and the addition of age as a quadratic term showed no evidence of strengthening the model based on likelihood ratio test (= 0.70). There was also no evidence of an age-by-sex interaction. Salicin (Salicoside, Salicine) Based on the most parsimonious model, the predicted probability of RCV antibody status can be calculated using the age and sex of a rabbit (Fig. 2). There was clearly a difference in the predicted probability between males and females, despite the overlap between 95% confidence intervals. Also, younger rabbits have a higher probability of Salicin (Salicoside, Salicine) being RCV seropositive than do older rabbits. Open in a separate window FIG 2 Predicted probability of a rabbit exhibiting RCV-A1-seropositive antibodies based on the most parsimonious general linear model with age (in weeks) and sex as factors. Shaded areas for both male and females show the 95% confidence intervals around the mean. Detection of nonpathogenic rabbit calicivirus. Other nonpathogenic RCVs have been predominantly detected in the small intestine (10, 11, 26), so duodenal tissue was targeted for the detection of a putative SMARCA4 nonpathogenic calicivirus. Approximately 100 duodenal RNA samples were screened using universal lagovirus PCR primers. Initially, young rabbits were preferentially selected because other RCV viruses are more commonly isolated from younger animals (10, 11). As there were few very young rabbits in the RCV antibody-positive populations, 143 rabbits up to 35 weeks of age were subsequently screened. A positive PCR product was obtained from a 33-week-old male rabbit. Other samples from the same location were weakly positive (= 9), but good-quality sequencing product was obtained from only one rabbit (Gore-425). This sample was collected on a property near Gore in Southland and was seropositive for RCV antibodies at a 1:40 dilution using the RCV-blocking ELISA, and it was also seropositive for RHDV antibodies at a 1:40 dilution. Initial sequencing of the 320-nucleotide product revealed 86% nucleotide identity with RHDV Czech v351 isolate (GenBank accession no. “type”:”entrez-nucleotide”,”attrs”:”text”:”KF594473″,”term_id”:”674785306″KF594473) and 93% nucleotide identity with the Australian RCV-A1 MIC3-3 (GenBank accession no. “type”:”entrez-nucleotide”,”attrs”:”text”:”GU368892.1″,”term_id”:”311294018″GU368892.1). The initial sequenced product matched the sequence from RCV Gore-425A. Inoculum production trial. To reproduce RCV infection and generate inocula for future studies, two New Zealand White rabbits were infected orally with material from rabbit Gore-425 (inoculum production trial). Dosed animals appeared healthy and were euthanized at 4 days postinfection. RNA was extracted from the duodenum of each rabbit and the presence of RCV confirmed by universal lagovirus PCR and Sanger sequencing. RNA extracts from the duodenum were used for the sequencing of the viral capsid gene (VP60). VP60 sequencing. Two variants of RCV were detected, each selectively amplified by Salicin (Salicoside, Salicine) a different primer set, indicating a mixed infection with two strains of RCV in rabbit.