A hypervariable region in VP1 of chicken infectious anemia virus mediates rate of spread and cell tropism in tissue culture

Abstract

Chicken infectious anemia virus (CIAV) is a unique infectious agent with an amino acid composition that has been found to be remarkably conserved even in isolates from different parts of the world. We have characterized field isolates of CIAV which vary significantly in terms of their abilities to replicate in culture, demonstrating a biological difference between isolates. Two sublines of MDCC-MSB1 cells that differ in their abilities to support CIAV were identified. In the MSB1(S) subline the CIA-1 isolate of CIAV was found to be less cytopathogenic than the prototype Cux-1(C) isolate; the MSB1(L) subline, which supports Cux-1(C) replication, was found to be nonpermissive for CIA-1. Alignments of the VP1 sequences of previously examined isolates with those of the field isolates CIA-1 and L-028 and the culture-adapted ConnB isolate revealed a previously unreported hypervariable region spanning amino acid positions 139 to 151. Chimeras of Cux-1(C) and CIA-1 were constructed to examine the potential for this region to affect cytopathogenicity. Transfer of a 316-bp region of Cux-1(C) open reading frame 1 into CIA-1 produced a virus with a cytopathogenic profile typical of Cux-1(C), indicating that one or both of the amino acid differences at positions 139 and 144 affect the rate of replication or the spread of infection. Transfection experiments with additional chimeras indicated that the inability of CIA-1 to replicate in MSB1(L) cells is mediated by a larger region of the genome which contains the hypervariable region in addition to upstream amino acid differences. Analysis of chimeras excluding the entire region of open reading frame 1 suggested the presence of a secondary mediator in the progression of infection in culture that was localized to a region containing a single nucleotide difference which results in amino acid differences in both VP2 (V-153) and the nuclear localization signal of VP3 (C-118). Immunofluorescence assays indicated an increased cytoplasmic distribution of VP3 and a general lack of VP3-associated apoptotic bodies in infections of CIA-1 and chimeras containing V-153 or C-118, as opposed to a primarily nuclear distribution and association with well-formed apoptotic bodies in Cux-1(C)-infected cells.

Sequence Analysis of the Full‐Length Cloned DNA of a Chicken Anaemia Virus (CAV) Strain from Bangladesh: Evidence for Genetic Grouping of CAV Strains Based on the Deduced VP1 Amino Acid Sequences

“…Although IBDV had been isolated and characterized from the same sample (Islam et al, 2001), repeated attempts to isolate BD‐3 CAV in MDCC‐MSB1 cells were not successful. Field isolates of CAV can vary significantly in their ability to replicate in MDCC‐MSB1 cells and different sublines of MDCC‐MSB1 cells also may vary in their susceptibility to CAV (Renshaw et al, 1996; Calnek et al, 2000). BD‐3 CAV could be one of those strains which do not grow in the MDCC‐MSB1 cell line used in this study.…”

“…VP3 is an apoptosis inducing protein (Noteborn et al, 1994). On the basis of comparison between Cux‐1 and CIA‐1, Renshaw et al (1996) suggested that an amino acid substitution at position 118 of the VP3 could influence nuclear localization of the protein and the development of distinct apoptotic bodies in cell culture. However, our alignment revealed that R at position 118 is present only in Cux‐1 (N), but Cux‐1 (M) and all other CAV strains, including BD‐3 CAV, have C at this position (data not shown).…”

“…The analysis of the deduced amino acid sequences of VP1 of 13 strains identified 16 positions at which more than one strain had amino acid substitutions (Table 1). Renshaw et al (1996) identified a hypervariable region spanning from amino acid position 139 to 151 in VP1. Brown et al (2000) aligned this 13‐amino acid region of 12 strains and further confirmed the hypervariability of this region.…”

“…VP3 is a non‐structural protein, which is also called apoptin for its ability to induce apoptosis (Noteborn and Koch, 1995). However, Renshaw et al (1996) identified a hypervariable region in VP1 and suggested that certain amino acid exchanges in this region could influence the rate of virus replication and spread of CAV strains in cell culture. In the present paper, we report the full‐length cloning and sequencing of the genome of a CAV strain which was found along with very virulent IBDV in a sample of the bursa of Fabricius of a chicken in Bangladesh.…”

Molecular epidemiology of chicken anaemia virus in sick chickens in China from 2014 to 2015

“…In the case of predicted amino acid sequences, minor differences have been noted particularly in the VP1 amino acid sites 139-151. The observed differences in the hypervariable region had impact on the predicted protein structure of VP1 [48][49][50]. In this study, the phylogenetic analysis of VP1 embodied the diversity of CAV, while the VP2 and VP3 embodied the conservatism.…”