Molecular analysis of G3P[6] rotavirus in the Amazon region of Brazil: evidence of reassortment with equine-like strains

Aim: To perform a molecular analysis of rotavirus A (RVA) G3P[6] strains detected in 2012 and 2017 in the Amazon region of Brazil. Materials & methods: Eighteen RVA G3P[6] strains were collected from children aged under 10 years hospitalized with acute gastroenteritis, and partial sequencing of each segment genome was performed using Sanger sequencing. Results: Phylogenetic analysis showed that all G3P[6] strains had a DS-1-like genotype constellation. Two strains had the highest nucleotide identities with equine-like G3P[6]/G3P[8] genotypes. Several amino acid alterations in VP4 and VP7 neutralizing epitopes of equine-like RVA G3P[6] strains were observed in comparison with vaccine strains. Conclusion: These findings suggest that equine-like RVA G3P[6] strains have been circulating in the Amazon region of Brazil as a result of direct importation, and support natural RVA evolutionary mechanisms.

Molecular characterization of group A rotavirus strains detected in alpacas (Vicugna pacos) from Peru

The alpaca is a very important social and economic resource for the production of fibre and meat for Andean communities. Peru is the main producer of alpacas. Group A rotavirus (RVA) has been sporadically detected in alpacas. In this study, a total of 1423 faecal samples from alpacas from different locations of the Puno department in Peru were collected and analysed by an antigen-capture ELISA in order to detect RVA. Four per cent of the samples were RVA-positive (57/1423). The genotype constellation of three selected alpaca RVA strains were G3/8 P[1/14]-I2-R2/5-C2/3-M2/3-A17-N2/3-T6-E3-H3. Two of the analysed strains presented a bovine-like genotype constellation, whereas the third strain presented six segments belonging to the AU-1-like genogroup (G3, M3, C3, N3, T3 and E3), suggesting reassorting events. Monitoring of the sanitary health of juvenile alpacas is essential to reduce the rates of neonatal mortality and for the development of preventive health strategies.

Comparative genomic analysis of genogroup 1 and genogroup 2 rotaviruses circulating in seven US cities, 2014-2016

For over a decade, the New Vaccine Surveillance Network (NVSN) has conducted active RVA strain surveillance in the USA. The evolution of RVA in the post-vaccine introduction era and the possible effects of vaccine pressure on contemporary circulating strains in the USA are still under investigation. Here we report the whole-gene characterization (11 ORFs) for 157 RVA strains collected at 7 NVSN sites during the 2014 through 2016 seasons. The sequenced strains included 52 G1P[8], 47 G12P[8], 18 G9P[8], 24 G2P[4], 5 G3P[6], as well as 7 vaccine strains, a single mixed strain (G9G12P[8]), and 3 less common strains. The majority of the single and mixed strains possessed a Wa-like backbone with consensus genotype constellation of G1/G3/G9/G12-P[8]-I1-R1-C1-M1-A1-N1-T1-E1-H1, while the G2P[4], G3P[6], and G2P[8] strains displayed a DS-1-like genetic backbone with consensus constellation of G2/G3-P[4]/P[6]/P[8]-I2-R2-C2-M2-A2-N2-T2-E2-H2. Two intergenogroup reassortant G1P[8] strains were detected that appear to be progenies of reassortment events between Wa-like G1P[8] and DS-1-like G2P[4] strains. Two Rotarix® vaccine (RV1) and two RV5 derived (vd) reassortant strains were detected. Phylogenetic and similarity matrices analysis revealed 2-11 sub-genotypic allelic clusters among the genes of Wa- and DS-1-like strains. Most study strains clustered into previously defined alleles. Amino acid (AA) substitutions occurring in the neutralization epitopes of the VP7 and VP4 proteins characterized in this study were mostly neutral in nature, suggesting that these RVA proteins were possibly under strong negative or purifying selection in order to maintain competent and actual functionality, but 14 radical (AA changes that occur between groups) AA substitutions were noted that may allow RVA strains to gain a selective advantage through immune escape. The tracking of RVA strains at the sub-genotypic allele constellation level will enhance our understanding of RVA evolution under vaccine pressure, help identify possible mechanisms of immune escape, and provide valuable information for formulation of future RVA vaccines.

Whole Genome Analysis of African G12P[6] and G12P[8] Rotaviruses Provides Evidence of Porcine-Human Reassortment at NSP2, NSP3, and NSP4

Group A rotaviruses (RVA) represent the most common cause of pediatric gastroenteritis in children <5 years, worldwide. There has been an increase in global detection and reported cases of acute gastroenteritis caused by RVA genotype G12 strains, particularly in Africa. This study sought to characterize the genomic relationship between African G12 strains and determine the possible origin of these strains. Whole genome sequencing of 34 RVA G12P[6] and G12P[8] strains detected from the continent including southern (South Africa, Zambia, Zimbabwe), eastern (Ethiopia, Uganda), central (Cameroon), and western (Togo) African regions, were sequenced using the Ion Torrent PGM method. The majority of the strains possessed a Wa-like backbone with consensus genotype constellation of G12-P[6]/P[8]-I1-R1-C1-M1-A1-N1-T1-E1-H1, while a single strain from Ethiopia displayed a DS-1-like genetic constellation of G12-P[6]-I2-R2-C2-M2-A2-N2-T2-E2-H2. In addition, three Ethiopian and one South African strains exhibited a genotype 2 reassortment of the NSP3 gene, with genetic constellation of G12-P[8]-I1-R1-C1-M1-A1-N1-T2-E1-H1. Overall, 10 gene segments (VP1–VP4, VP6, and NSP1–NSP5) of African G12 strains were determined to be genetically related to cognate gene sequences from globally circulating human Wa-like G12, G9, and G1 strains with nucleotide (amino acid) identities in the range of 94.1–99.9% (96.5–100%), 88.5–98.5% (93–99.1%), and 89.8–99.0% (88.7–100%), respectively. Phylogenetic analysis showed that the Ethiopian G12P[6] possessing a DS-1-like backbone consistently clustered with G2P[4] strains from Senegal and G3P[6] from Ethiopia with the VP1, VP2, VP6, and NSP1–NSP4 genes. Notably, the NSP2, NSP3, and NSP4 of most of the study strains exhibited the closest relationship with porcine strains suggesting the occurrence of reassortment between human and porcine strains. Our results add to the understanding of potential roles that interspecies transmission play in generating human rotavirus diversity through reassortment events and provide insights into the evolutionary dynamics of G12 strains spreading across selected sub-Saharan Africa regions.

First isolation and whole-genome characterization of a G9P[14] rotavirus strain from a diarrheic child in Egypt

An unusual group A rotavirus (RVA) strain (RVA/Human-tc/EGY/AS997/2012/G9[14]) was isolated for the first time in a faecal sample from a 6-month-old child who was hospitalized for treatment of acute gastroenteritis in Egypt in 2012. Whole-genome analysis showed that the strain AS997 had a unique genotype constellation: G9-P[14]-I2-R2-C2-M2-A11-N2-T1-E2-H1. Phylogenetic analysis indicated that the strain AS997 had the consensus P[14] genotype constellation with the G9, T1 and H1 reassortment. This suggests either a mixed gene configuration originated from a human Wa-like strain and a P[14]-containing animal virus, or that this P[14] could have been acquired via reassortment of human strains only. The study shows the possible roles of interspecies transmission and multiple reassortment events leading to the generation of novel rotavirus genotypes and underlines the importance of whole-genome characterization of rotavirus strains in surveillance studies.

Isolation and characterization of a rare group A rotavirus G13P[18] strain from a diarrhoeic foal in Japan

A rare genotype G13P[18] group A rotavirus (RVA/Horse-tc/JPN/MK9/2019/G13P[18]) was isolated from a diarrhoeic foal for the first time in 28 years. The genotype constellation of the virus was assigned to G13-P[18]-I6-R9-C9-M6-A6-N9-T12-E14-H11 and was the same as that of the first isolated strain, RVA/Horse-tc/GBR/L338/1991/G13P[18]. Phylogenetic analysis suggests that the virus is related to RVA/Horse-tc/GBR/L338/1991/G13P[18] and is distant from typical equine rotaviruses of the G3P[12] and G14P[12] genotypes.

High prevalence and genomic characteristics of G6P[1] Bovine Rotavirus A in yak in China

Yak is an iconic species of the Qinghai–Tibet Plateau, which is the world's highest plateau. Here, a total of 541 yak diarrhoeic samples were collected from 69 farms in four provinces in the Qinghai–Tibet Plateau from April 2015 to June 2018, and 73.6 % of samples were detected as Bovine Rotavirus A (BRVA) positive by RT-PCR assay. Two G genotypes (G6, G10) and two P genotypes (P[1], P[11]) were determined, in which G6P[1] BRVA was the predominant strain. Moreover, VP7 and VP4 of these G6P[1] strains showed unique amino acid mutations, such that they clustered into an independent branch in the phylogenetic tree. A strain of BRVA designated as RVA/Yak-tc/CHN/QH-1/2015/G6P[1] was isolated successfully using MA104 cells, and the virus titre was determined as 105.84 TCID50 ml–1. The genome of strain QH-1 had a G6-P[1]-I2-R2-C2-M2-A3-N3-T6-E2-H3 genotype constellation. QH-1 was identified as a reassortment strain of BRVA, human RVA and ovine RVA based on the nucleotide identity and phylogenetic tree of 11 gene segments, indicating its public health significance. To the best of our knowledge, this is the first report on the molecular prevalence and genome characteristics of BRVA in yak, contributing to further understanding of the epidemic and genetic evolution of BRVA.

Reference Human Rotavirus A Genome Sequence from a Previously Vaccinated Child with Diarrhea in Nigeria

In 2018, a 26-month-old girl, fully vaccinated with Rotarix in 2016, presented with fever, diarrhea, and vomiting. A rapid test showed that her feces contained rotavirus A (RVA). VP7 reverse transcription-PCR (RT-PCR) and Illumina sequencing showed that a G1P[8] strain with a Wa-like genotype constellation was the etiologic agent. This is the first near-complete RVA genome sequence from Nigeria.

Full genomic characterization of a porcine rotavirus strain detected in an asymptomatic piglet in Accra, Ghana

Background The introduction of rotavirus A vaccination across the developing world has not proved to be as efficacious as first hoped. One cause of vaccine failure may be infection by zoonotic rotaviruses that are very variable antigenically from the vaccine strain. However, there is a lack of genomic information about the circulating rotavirus A strains in farm animals in the developing world that may be a source of infection for humans. We therefore screened farms close to Accra, Ghana for animals sub-clinically infected with rotavirus A and then sequenced the virus found in one of these samples. Results 6.1% of clinically normal cows and pigs tested were found to be Rotavirus A virus antigen positive in the faeces. A subset of these (33.3%) were also positive for virus RNA. The most consistently positive pig sample was taken forward for metagenomic sequencing. This gave full sequence for all open reading frames except segment 5 (NSP1), which is missing a single base at the 5′ end. The virus infecting this pig had genome constellation G5-P[7]-I5-R1-C1-M1-A8-N1-T7-E1-H1, a known porcine genotype constellation. Conclusions Farm animals carry rotavirus A infection sub-clinically at low frequency. Although the rotavirus A genotype discovered here has a pig-like genome constellation, a number of the segments most closely resembled those isolated from humans in suspected cases of zoonotic transmission. Therefore, such viruses may be a source of variable gene segments for re-assortment with other viruses to cause vaccine breakdown. It is recommended that further human and pig strains are characterized in West Africa, to better understand this dynamic.