OBJECTIVETo establish a one-step four multiplex reverse transcription polymerase chain reaction (RT-PCR) method based on Homo-Tag Assisted Non-Dimer System (HAND) system for simultaneous detection of 4 diarrhea viruses of rotavirus, astrovirus, norovirus and sapovirus.

METHODSPrimers were designed according to the conserved genome sequence of the 4 viruses and the homologous tail sequences were added to the 5' end. The multiplex RT-PCR system was constructed by optimizing the PCR parameters such as the concentration of universal tag primer and genome-specific Homo-Tailed primers. The specificity, stability and sensitivity of the method were evaluated systematically.

RESULTSThe 4 multiplex RT-PCR methods based on HAND system was established successfully. Specificity analysis showed no cross reaction between the 4 diarrhea viruses. The sensitivity analysis showed detection limits for rotavirus, astrovirus, norovirus and sapovirus of 48, 1.92, 9.6 and 48 pg per reaction, respectively.

CONCLUSIONThe established HAND system-based multiplex RT-PCR assay allows simple, rapid, specific, sensitive, and stable for detection of the 4 common diarrhea viruses at low costs and is suitable for application in general medical laboratories.

Astroviridae ; genetics ; isolation & purification ; Diarrhea ; virology ; Feces ; virology ; Humans ; Multiplex Polymerase Chain Reaction ; methods ; Norovirus ; genetics ; isolation & purification ; RNA, Viral ; isolation & purification ; Reverse Transcriptase Polymerase Chain Reaction ; methods ; Rotavirus ; genetics ; isolation & purification ; Sapovirus ; genetics ; isolation & purification

OBJECTIVETo conduct an epidemiological and genotype analysis of sapovirus (SaV) associated with sporadic diarrhea in Shenzhen in the year 2009.

METHODSA total of 852 fecal samples were collected from sporadic cases of diarrhea in Shenzhen in 2009 and detected by reverse-transcription polymerase chain reaction (RT-PCR) using the primers of SLV5317/5749. The PCR products were analyzed with 1.5% agarose gel electrophoresis and sequenced to construct the phylogenetic tree.

RESULTSSixteen samples were found positive for SaV, with a positivity rate of 1.88%. Sequence analysis identified 8 isolates as SaV GI genotype (including 3 SaV GI.1 and 5 SaV GI.2), 7 as SaV GIV genotype, and 1 as GII genotype.

CONCLUSIONSSaV infection is present in Shenzhen with GI as the predominant genotype. This is the first report of SaV GIV strains in China, which differs from the strains of Anhui-A141 and Beijing-CHN99/BJ360, suggesting the genotypic variety of SaV infection in China.

Adult ; China ; epidemiology ; Diarrhea ; epidemiology ; virology ; Female ; Genetic Variation ; Genotype ; Humans ; Infant ; Infant, Newborn ; Male ; Middle Aged ; Phylogeny ; RNA, Viral ; genetics ; Sapovirus ; classification ; genetics ; isolation & purification ; Young Adult

Objective: To understand the epidemiological and etiological characteristics of outbreaks on acute gastroenteritis caused by sapovirus (SaV) worldwide. Methods: Literature about the outbreaks on acute gastroenteritis caused by SaV were retrieved from the databases including WanFang, CNKI, PubMed and Web of Science after evaluation. Time, geography, setting and population distributions of outbreaks, transmission mode, SaV genotype and clinical characteristics of the patients were analyzed. Results: A total of 34 papers about SaV were included, involving 146 outbreaks occurred between October 1976 and April 2016. In these papers, 138 outbreaks were reported on the related months. All these outbreaks occurred in northern hemisphere. SaV outbreaks occurred all year around, but mainly in cold season, the incidence was highest in December (25 outbreaks) and lowest in in August (2 outbreaks). Most outbreaks were reported by Japan, followed by Canada, the United States of America and the Netherlands. There were 141 outbreaks for which the occurring settings were reported, child-care settings were most commonly reported setting (48/141, 34.04%), followed by long-term care facility (41/141, 29.08%) and hospital (16/141, 11.35%). Clinical symptoms of 1 704 cases in 31 outbreaks were reported, with the most common symptom was diarrhea (1 331/1 704, 78.12%), followed by nausea (829/1 198, 69.20%), abdominal pain (840/1 328, 63.25%), vomiting (824/1 704, 48.36%) and fever (529/1 531, 34.53%). Genotypes of SaV were determined for 119 outbreaks. GⅠ(51/119, 42.86%) and GⅣ (45/119, 37.82%) were predominant. The outbreaks of GⅣ SaV increased suddenly in 2007, and the outbreaks of GⅠ SaV mainly occurred in 2008 and during 2011-2013. Conclusions: SaV outbreaks were reported mainly by developed countries, with most outbreaks occurred in cold season, in child-care settings and long term care facility. GⅠ and GⅣ were the most common genotypes of SaV. Prevention and control of SaV outbreak in China seemed relatively weak, and it is necessary to conduct related training and to strengthen the SaV outbreak surveillance in areas where service is in need.
Caliciviridae Infections/virology* ; Child ; China/epidemiology* ; Disease Outbreaks ; Feces/virology* ; Gastroenteritis/virology* ; Genotype ; Humans ; Phylogeny ; RNA, Viral/genetics* ; Sapovirus/isolation & purification* ; Sequence Analysis, DNA

The prevalence of human astroviruses was tested in patients with acute gastroenteritis by using conventional duplex reverse transcription (RT)-PCR and electrophoresis. Diarrheal fecal samples were collected from 9,597 patients at local hospitals in Seoul. The prevalence of astroviruses was 1.0% (94/9,597 patients; mostly infants), and that of sapoviruses was 0.1% (14/9,597 patients). Age- and gender-wise analyses were carried out on 29 astrovirus-positive patients having complete information on file regarding their age, gender, and other particulars. The results were higher in patients of ages 0 to 14 yr, and 69.0% of the astrovirus-positive patients were females, of which 69.2% were infants (0 to 12 months), and 61.5% were 1-4 yr old. Notably, in the case of 5 to 78-yr-old acute gastroenteritis patients, 100% were females.
Acute Disease ; Adolescent ; Adult ; Age Factors ; Aged ; Astroviridae Infections/complications/*epidemiology/virology ; Child ; Child, Preschool ; DNA, Viral/analysis ; Feces/virology ; Female ; Gastroenteritis/complications/*diagnosis ; Humans ; Infant ; Male ; Mamastrovirus/genetics/*isolation & purification ; Middle Aged ; Prevalence ; Reverse Transcriptase Polymerase Chain Reaction ; Sapovirus/genetics/isolation & purification ; Sex Factors ; Young Adult