Elsevier

Veterinary Microbiology

Volume 196, 30 November 2016, Pages 27-35
Veterinary Microbiology

Whole-genome characterization of a Peruvian alpaca rotavirus isolate expressing a novel VP4 genotype

https://doi.org/10.1016/j.vetmic.2016.10.005Get rights and content

Highlights

Abstract

The SA44 isolate of Rotavirus A (RVA) was identified from a neonatal Peruvian alpaca presenting with diarrhea, and the full-length genome sequence of the isolate (designated RVA/Alpaca-tc/PER/SA44/2014/G3P[40]) was determined. Phylogenetic analyses showed that the isolate possessed the genotype constellation G3-P[40]-I8-R3-C3-M3-A9-N3-T3-E3-H6, which differs considerably from those of RVA strains isolated from other species of the order Artiodactyla. Overall, the genetic constellation of the SA44 strain was quite similar to those of RVA strains isolated from a bat in Asia (MSLH14 and MYAS33). Nonetheless, phylogenetic analyses of each genome segment identified a distinct combination of genes. Several sequences were closely related to corresponding gene sequences in RVA strains from other species, including human (VP1, VP2, NSP1, and NSP2), simian (VP3 and NSP5), bat (VP6 and NSP4), and equine (NSP3). The VP7 gene sequence was closely related to RVA strains from a Peruvian alpaca (K’ayra/3368-10; 99.0% nucleotide and 99.7% amino acid identity) and from humans (RCH272; 95% nucleotide and 99.0% amino acid identity). The nucleotide sequence of the VP4 gene was distantly related to other VP4 sequences and was designated as the reference strain for the new P[40] genotype. This unique genetic makeup suggests that the SA44 strain emerged from multiple reassortment events between bat-, equine-, and human-like RVA strains.

Introduction

Among the South American camelids (SACs), alpacas (Vicugna pacu) and llamas (Llama glama) are domesticated species, whereas vicuñas (Vicugna vicugna) and guanacos (Llama guanicoe) are wild species. A cornerstone of the Andean highland economy is raising alpacas for meat and fiber. Peru has the largest alpaca herd in the world (4–4.5 million alpacas) (Tuckwell, 1994), producing an average of approximately 3.5 million kilograms of alpaca fiber annually (∼90% of the total worldwide production) (Rosadio et al., 2012). Infections, particularly diarrheal infections, caused by various pathogens are a major cause of neonatal death for SACs (Cebra et al., 2003, Whitehead, 2009, Lopez et al., 2011). Previous studies have demonstrated the importance of rotavirus infections among these animals (Parreño et al., 2001, Parreño et al., 2004, Lopez et al., 2011, Papp et al., 2012, Rosadio et al., 2012, Badaracco et al., 2013, Garmendia et al., 2015).

Whole-genome rotavirus sequences in SACs are available for only two strains isolated from a guanaco (Matthijnssens et al., 2009). Partial genome characterization is available for one strain from a vicuña and two strains from alpacas; however, only one or two of the 11 genome segments have been sequenced for the alpaca Rotavirus A (RVA) isolates (Badaracco et al., 2013, Garmendia et al., 2015). Here, we report the complete genome sequence of rotavirus strain RVA/Alpaca-tc/PER/SA44/2014/G3P[40] and describe a novel VP4 genotype.

Rotaviruses are members of the Rotavirus genus of the Reoviridae family, and are classified into eight species (A-H) (Matthijnssens et al., 2012a). RVA is a major cause of dehydrating diarrhea in humans and animals worldwide (Santos and Hoshino, 2005). The RVA genome consists of 11 segments of double-stranded RNA (dsRNA) encoding six structural proteins (VP1-4, VP6, and VP7) and six nonstructural proteins (NSP1-6) (Estes and Greenberg, 2013). The RVA genomic classification nomenclature is based on all 11 segments of dsRNA. The notation Gx-P[x]-Ix-Rx-Cx-Mx-Ax-Nx-Tx-Ex-Hx is used to represent genotypes of the VP7-VP4-VP6-VP1-VP2-VP3-NSP1-NSP2-NSP3-NSP4-NSP5/6–encoding gene segments, respectively, with x indicating the number of the genotype (Matthijnssens et al., 2008, Matthijnssens et al., 2011a). Currently, there are 28 G, 39 P, 21 I, 14 R, 14C, 13 M, 24 A, 14 N, 16 T, 21 E, and 16 H genotypes (https://rega.kuleuven.be/cev/viralmetagenomics/virus-classification/minutes-of-the-7th-rcwg-meeting).

Section snippets

Stool samples and RT-PCR detection of RV

The RVA strain described in this study (SA44) was isolated from a stool sample collected in February 2014 from a neonatal diarrheic alpaca belonging to the herd of Silli. This community is located in the southern highlands of Peru (14°24′45.3′′s, 71°11′32.6′′ W; ∼4000 m ASL) in the province of Canchis in the state of Cusco. The importation of alpacas stool samples was approved by the Brazilian Institute of Environment (IBAMA; Brasília, DF, Brazil) license in 14BR012948/DF 02/20/2014.

The stool

Virus isolation

The RVA strain was successfully propagated in MA-104 cells, as confirmed by the presence of RV particles by TEM (Fig. 1). PAGE analysis of the genome demonstrated a typical (4:2:3:2) RVA electropherotype (data not shown).

Phylogenetic analyses

Sequences generated for all 11 genome segments of alpaca strain SA44 were deposited into GenBank under accession numbers KT935476-KT935485 and KU168341. Comparison of the SA44 sequence with sequences of known RVA strains suggested that the genomic makeup of the newly identified

Discussion

Genome sequence comparison analyses have determined that most human RVA strains belong to three defined genotype constellations of the non-G, non-P genes: namely, I1-R1-C1-M1-A1-N1-T1-E1-H1 (Wa-like), I2-R2-C2-M2-A2-N2-T2-E2-H2 (DS-1-like), and I3-R3-C3-M3-A3-N3-T3-E3-H3 (AU-1-like) (Matthijnssens and Van Ranst, 2012). The AU-1-like constellation presumably originated from canine or feline strains and has been occasionally detected in humans and bats (Matthijnssens et al., 2011a, He et al., 2013

Conflicts of interest

No conflicts of interest are declared.

Acknowledgements

This study was supported, in part, by the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq; Nos. 471063/2012-6 and 303864/2014-1), the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES), and the Fundação Carlos Chagas de Amparo à Pesquisa do Estado do Rio de Janeiro (FAPERJ; Nos. E-26/103.113/2011 and E-26/201.374/2014), Brazil. The funders had no role in study design, data collection, data interpretation, or the decision to submit the work for publication.

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