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NSP4 gene analysis of rotaviruses recovered from infected children with and without diarrhea.

ABSTRACT: The transmembrane glycoprotein NSP4 functions as a viral enterotoxin capable of inducing diarrhea in young mice. It has been suggested that NSP4 may be a key determinant of rotavirus pathogenicity and a target for vaccine development. Twenty two G1P[6] rotaviruses from babies with and without diarrhea were comparatively analyzed along with reference strains and another 22 Taiwanese human rotaviruses of G and P combination types different from the G1P[6] type. The sequence variations in the NSP4 genes were studied by direct sequencing analysis of the amplicons of reverse transcription-PCR. Two genetic groups could be identified in this analysis. While the majority of these strains were closely related to the Wa strain, the G2 viruses were closely related to the S2 strain. Furthermore, phylogenetic analysis of the NSP4 gene among the G2 rotaviruses revealed three distinct lineages associated with DS-1, S2, and E210, respectively, as has been reported previously for the VP7 gene. However, we found no apparent correlation in the deduced amino acid sequences corresponding to the proposed enterotoxic peptide region between the rotaviruses recovered from individuals with and without diarrhea. The NSP4 gene product being a pathogenic determinant may not be a generalized phenomenon.

SUBMITTER: Lee CN

PROVIDER: S-EPMC87623 | biostudies-literature | 2000 Dec

REPOSITORIES: biostudies-literature

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NSP4 gene analysis of rotaviruses recovered from infected children with and without diarrhea.

Lee C N CN Wang Y L YL Kao C L CL Zao C L CL Lee C Y CY Chen H N HN

Journal of clinical microbiology 20001201 12

The transmembrane glycoprotein NSP4 functions as a viral enterotoxin capable of inducing diarrhea in young mice. It has been suggested that NSP4 may be a key determinant of rotavirus pathogenicity and a target for vaccine development. Twenty two G1P[6] rotaviruses from babies with and without diarrhea were comparatively analyzed along with reference strains and another 22 Taiwanese human rotaviruses of G and P combination types different from the G1P[6] type. The sequence variations in the NSP4 ...[more]

PMID: 11101582

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Project description:All over the world, children and adults are severely affected by acute gastroenteritis, caused by one of the emerging enteric pathogens, rotavirus C (RVC). At present, no extensive surveillance program is running for RVC in India, and its prevalence is largely unknown except cases of local outbreaks. Here, we intended to detect the presence of RVC in diarrheic children visiting or admitted to hospitals in Haldwani (state of Uttarakhand, India), a city located in the foothills of the Himalayas. During 2010-2013, we screened 119 samples for RVC by an RVC VP6 gene-specific RT-PCR. Of these, 38 (31.93%) were found positive, which is higher than the incidence rates reported so far from India. The phylogenetic analysis of the derived nucleotide sequences from one of the human RVC (HuRVC) isolates, designated as HuRVC/H28/2013/India, showed that the study isolate belongs to genotype I2, P2 and E2 for RVC structural genes 6 and 4 (VP6, and VP4) and non-structural gene 4 (NSP4), respectively. Furthermore, the VP6 gene of HuRVC/H28/2013/India shows the highest similarity to a recently-reported human-like porcine RVC (PoRVC/ASM140/2013/India, KT932963) from India suggesting zoonotic transmission. We also report a full-length NSP4 gene sequence of human RVC from India. Under the One-health platforms there is a need to launch combined human and animal RVC surveillance programs for a better understanding of the epidemiology of RVC infections and for implementing control strategies.Reoviridae, possess 11 double-stranded segments of RNA that encode six structural viral proteins (VP1, VP2, VP3, VP4, VP6, VP7) and five/six non-structural proteins (NSP1-NSP5/6) [7]. Based on the antigenic properties of the major inner capsid protein (VP6), RVs are subdivided into eight well-characterized species (A-H) and two putative species viz. I and J [8-10]. Humans and other mammalian species are affected by species A, B, C and H rotaviruses and birds by species D, F and G, and species E has been reported exclusively in pigs [7,8,11-17]. The newly-proposed species I is reported in dogs [18] and cats [19], whereas species J is found in bats [10].

Project description: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.

Project description:Feces and serum specimens were collected from three farms in Michigan on which approximately 50-lb (8- to 9-week-old) pigs experienced diarrhea just after placement into all-in-all-out finishing barns. The clinical signs (profuse watery diarrhea lasting about 2 weeks and no vomiting) were similar on all farms, and the morbidity rate was high (ranging from 60 to 80%) but without mortality. Eleven diarrheic fecal samples from the farms were tested for group A and C rotaviruses by immune electron microscopy (IEM) and various assays. IEM indicated that the fecal samples reacted only with antiserum against group C rotaviruses, and polyacrylamide gel electrophoresis indicated that the samples had characteristic genomic electropherotypes for group C rotavirus. Group C rotavirus was detected by cell culture immunofluorescence (CCIF) tests in nine fecal samples, but no group A rotavirus was detected by enzyme-linked immunosorbent assay or CCIF. By reverse transcription (RT)-PCR, all 11 fecal samples were positive for group C rotaviruses, with only 2 samples positive for group A rotaviruses. However, a second amplification of RT-PCR products using nested primers detected group A rotaviruses in all samples. Analysis of nucleotide and deduced amino acid sequences of the RT-PCR product (partial-length VP7) of the group C rotavirus showed 87.2 to 91% nucleotide identity and 92.6 to 95.9% amino acid identity among two strong samples from the different farms and the Cowden strain of porcine group C rotavirus. All nine convalescent-phase serum samples tested had neutralizing antibodies to the Cowden strain, and the majority of them had neutralizing antibody against group A rotaviruses (OSU or/and Gottfried strains) by fluorescent focus neutralization tests. Although group C rotaviruses have been reported as a cause of sporadic diarrhea in suckling or weanling pigs, to our knowledge, this is the first report of epidemic diarrhea outbreaks associated with group C rotavirus in older pigs.

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NSP4 gene analysis of rotaviruses recovered from infected children with and without diarrhea.

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NSP4 gene analysis of rotaviruses recovered from infected children with and without diarrhea.

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