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. 2018 Jun 18;15(1):101.
doi: 10.1186/s12985-018-1010-y.

Chinese woodchucks with different susceptibility to WHV infection differ in their genetic background exemplified by cytochrome B and MHC-DRB molecules

Bin Zhu  1 , Zhenni Zhu  1   2 , Junzhong Wang  1 , Shunmei Huang  1 , Fanghui Li  1 , Lu Wang  1 , Yanan Liu  1 , Qi Yan  1 , Shunchang Zhou  3 , Mengji Lu  4 , Dongliang Yang  1 , Baoju Wang  5
Affiliations

Chinese woodchucks with different susceptibility to WHV infection differ in their genetic background exemplified by cytochrome B and MHC-DRB molecules

Bin Zhu et al. Virol J. .

Abstract

Background: Chinese woodchucks (M. himalayana) were recently found to be susceptible to woodchuck hepatitis virus (WHV) infection. In this study, we aimed to determine the susceptibility to WHV infection of M. himalayana from different areas and their association with the animal genetic background exemplified by cytochrome B and MHC-DRB molecules.

Methods: Animals from four different areas in Qinghai province were inoculated with WHV59 strains. The virological markers including WHV surface antigen (WHsAg), WHV core antibody (WHcAb), and WHV DNA in serum were measured by ELISA and Real-time PCR, respectively. The sequences of cytochrome B gene and MHC-DRB molecules were obtained and sorted with Clustalx software. The nucleotide variation sites were identified using MEGA5 software.

Results: The animals from four different areas had different susceptibility to WHV infection. Animals from TR and TD areas had a high level of long-lasting viremia, while those from GD and WL areas had a low level of transient viremia after WHV inoculation. All of the animals belong to the same subspecies M. himalayana robusta identified by cytochrome B gene sequences. Based on their nucleotide variation pattern, 8 alleles of cytochrome B gene were identified, and 7 MHC-DRB alleles were identified. Allele A of cytochrome B and Allele Mamo-DRB1*02 of MHC-DRB was found to be frequent in animals from TR and TD areas, while Allele H of cytochrome B and Allele Mamo-DRB1*07 of MHC-DRB was predominant in animals from GD and WL areas.

Conclusion: Chinese woodchucks from different areas differed in their susceptibility to WHV infection, though they belong to the same subspecies M. himalayana robusta. The genetic background exemplified by cytochrome B and MHC-DRB differed in Chinese woodchucks with different susceptibility to WHV infection.

Keywords: Chinese woodchuck; Cytochrome B; Hepatitis B virus; Major histocompatibility complex class II DRB gene; Woodchuck hepatitis virus.

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Conflict of interest statement

Ethical approval

All the animal experiments were performed in accordance with the protocol approved by Animal Ethics Committee of Tongji Medical College, Huazhong University of Science & Technology, China). (IACUC Number: S659).

Competing interests

The authors declare that they have no competing interests.

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Figures

Fig. 1
Fig. 1
Kinetics of WHV DNA in WHV-infected Chinese woodchucks from the TR (a), TD (b), GD (c), and WL (d) areas in Qinghai province of China. Four animals from each area were randomly selected, and were inoculated with WHV59 strains. WHV DNA in serum was measured by real-time PCR at one-week interval
Fig. 2
Fig. 2
Kinetics of WHsAg in WHV-infected Chinese woodchucks from the TR (a), TD (b), GD (c), and WL (d) areas in Qinghai province of China. Four animals from each area were randomly selected, and were inoculated with WHV59 strains. WHsAg in serum was measured by ELISA at one-week interval. The results for WHsAg are presented as S/N values = OD of sample / OD of negative control. The cut-off value is presented by a dotted horizontal line
Fig. 3
Fig. 3
WHcAb in WHV-infected Chinese woodchucks from the TR, TD, GD, and WL areas in Qinghai province of China. Four animals from each area were randomly selected, and were inoculated with WHV59 strains. WHcAb in serum was measured by ELISA at the end of follow up (12 weeks post WHV inoculation). The results for WHcAb are presented as the percentage of inhibition = [((OD of negative control – OD of sample) / OD of negative control) × 100]. The cut-off value is 50%, and presented by a dotted horizontal line
Fig. 4
Fig. 4
The neighbor-joining phylogenetic tree of the Chinese woodchuck species. The published cytochrome B sequences of the Chinese woodchuck species (M. himalayana, M. sibirica, M. caudata, M. baibacina, and M. bobak) were downloaded at least two sequences for one species. The 8 cytochrome B alleles were obtained in 37 animals from TR, TD, GD, and WL areas, Qinghai province, China. The neighbor-joining phylogenetic tree was conducted using MEGA5 software
Fig. 5
Fig. 5
A phylogenetic tree constructed using 789 bp fragments of MHC class II DRB1 genes from Chinese woodchucks. The sequences used in constructing the phylogenetic tree are as following: Sper-DRB from Spermophilus tridecemlineatus (XM005338933), Scur-DRB from Sciurus aberti aberti (M97616.1), Mamu-DRB1 from Macaca mulatta (EF362437.1), Ovar-DRB1 from Ovis aries (KM588646.1), Susc-DRB1 from Sus scrofa (EU431221.1), HLA-DRB1 from Homo sapiens (M33600.1)
Fig. 6
Fig. 6
Deduced amino acid sequences of partial Mamo-DRB1 alleles. The HLA-DRB1*1 sequence is chosen as reference. Identity to the consensus is illustrated by spots
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