Development and evaluation of real-time fluorescence recombinase aided amplification assay without extracting nucleic acid for detection of adenovirus type 3 / 中华实验和临床病毒学杂志

Objective@#To establish a real-time fluorescence recombinase acid amplification (RAA) method for the detection of adenovirus type 3(HAdV-3)without extraction nucleic acid.@*Methods@#According to the conserved sequence of adenovirus type 3 gene, a pair of primers and a probe were designed, and a real-time fluorescence RAA without extracting nucleic acid was established and optimizing the condition of DNA-free extraction. The sensitivity of the method was analyzed by a series of dilution and the specificity of the method was evaluated by detecting the original samples of other respiratory viruses. The clinical samples of HAdV-3 were detected and compared with the traditional real-time fluorescence quantitative PCR method for nucleic acid extraction.@*Results@#The sensitivity of the real-time fluorescence RAA method was as high as that of qPCR in the detection of 10 series diluted HAdV-3 strains. The highest corresponding CT value of qPCR was 36.87. The sensitivity of the real-time fluorescence RAA method was similar to that of the real-time fluorescence quantitative PCR method . There was no cross-reaction to other common types of respiratory viruses. The two method were used to detect 56 clinical samples at the same time, and the result were completely consistent.@*Conclusions@#We provide the first report of the real-time fluorescent RAA assays for the detection of HAdV-3 without extracting nucleic acid and it has high sensitivity and specificity. Is suitable for rapid detection of HAdV-3 in clinical laboratories and on-site unite.

Pathogen spectrum of viral encephalitis in children living in Hebei province, China from May to December 2017 / 中华实验和临床病毒学杂志

Objective@#To investigate the pathogenic characteristics of viral encephalitis in children living in Hebei province.@*Methods@#We randomly collected cerebrospinal fluid specimens from a total of 399 children diagnosed with viral encephalitis in Hebei Children′s Hospital from May to December 2017. Real-time fluorescence quantitative PCR and Sanger sequencing were used to detect viral nucleic acids in cerebrospinal fluid by an automatic laboratory station. Statistical analysis was performed on the experimental data using SPSS 21.0 software and the clinical data were analyzed. Comparison of infection rates of EV encephalitis in different months, using line × column chi-square test. The MRI and EEG positive rates of different viral encephalitis and viral encephalitis patients not infected with the virus were analyzed by Fisher′s exact probability test. The positive rate of infection with different viruses and non-virus agents was analyzed by Fisher′s exact probability test.@*Results@#The result showed that 80 of 399 samples were positive, and the positive rate was 20.05%. It included 22 cases of enterovirus, 4 cases of influenza A virus, 3 cases of mumps virus, 2 cases of herpes simplex virus type 1, 1 case of herpes simplex virus type 2, 4 cases of EB virus, 7 cases of cytomegalovirus, 7 cases of herpes zoster virus, 8 cases of adenovirus, 14 cases of human herpesvirus type 6. Eight cases had combined viral infection. Eight cases had concurrent infections: 3 cases had enterovirus and herpesvirus type 6 concurrent infection, 1 case had enterovirus and Japanese encephalitis virus concurrent infection and 1 case had herpes simplex virus type 2 and adenovirus, 1 case had influenza A virus herpesvirus type 6, 1 case had mumps virus and herpesvirus type 6, 1 case had mumps virus and herpesvirus type 6, 1 case had herpes simplex virus type 1 and herpes zoster virus concurrent infections. Children with EV viral encephalitis in Hebei Province were highly prevalent in May and June (P=0.016). HHV6 virus encephalitis was more susceptible to infection than non-HHV6 virus (P=0.016); The rate of MRI positive findings in patients with different viral encephalitis was not statistically significant (P>0.05). The result of EEG of different viral encephalitis were P>0.05, which was not statistically significant.@*Conclusions@#EV was the most common pathogen of children with viral encephalitis in Hebei province. Encephalitis caused by influenza A virus cannot be ignored in clinical practice.

Integrating host response analysis and VSITA metagenomic detection in cerebrospinal fluid from suspected encephalitis cases / 中华实验和临床病毒学杂志

Objective@#To detect pathogens in clinical cerebrospinal fluid samples from 5 suspected encephalitis cases.@*Methods@#Five cerebrospinal samples were treated with both random and virus sequence independent targeted amplification(VSITA) method , followed by next-generation sequencing. Host trascriptome profiling combined with quantitative PCR were conducted to verify the detected pathogen.@*Results@#Cytomegalovirus(CMV) was detected in 3 cases. Quantitative PCR showed weakly positive result (Ct value: 30.23, 32.83, 34.08). The enhancement result of host infection pathway indicated CMV infection (P=0.213). Sequencing reads analysis showed better result in VSITA (P=0.096).@*Conclusions@#This is the first application of VSITA for cerebrospinal fluid samples. This enrichment method showed great potential while it needs further improvement. Consistency in gene expression detection between VSITA and random method makes it possible for host transcriptome profiling. Analysis result detected increase of CMV infection pathway in 3 positive cases, which further identified CMV infection in the 3 cases.

Comparison of detection rates of herpes virus and enterovirus in paired cerebrospinal fluid and serum specimens of patients with viral encephalitis / 中华实验和临床病毒学杂志

Objective@#To compare the detection rate of herpes virus and enterovirus (EV) in paired cerebrospinal fluid and serum samples of patients with viral encephalitis.@*Methods@#A total of 109 paired cerebrospinal fluid and serum specimens were collected from patients who were clinically diagnosed with suspected viral meningitis in Children′s Hospital of Hunan from December 2017 to February 2018. One-step nested real-time fluorescence quantitative reverse transcription polymerase chain reaction (RT-PCR) and real-time fluorescence quantitative PCR were used to detect enterovirus and herpes virus respectively and the detection rates of different virus and sample types were analyzed. SPSS 17.0 was used for statistical analysis of the test result .@*Results@#Among the 109 pairs of specimens, the positive rates of human herpes virus type 6 (HHV6), herpes simplex virus-1 (HSV1), Epstein-Barr virus (EBV), cytomegalovirus (CMV) and enterovirus group A type 71(EV-A71) in serum were 7.34%, 4.59%, 7.34%, 9.17% and 10.09%, respectively, and in cerebrospinal fluid were 5.50%, 2.75%, 0, 5.50%, and 6.42%, respectively. The result showed that there were statistically significant differences between the two types of specimens for herpes virus and enterovirus (P<0.05). In cerebrospinal fluid and serum samples, the longest time for EV-A71 positive detection was 2 and 7 days after onset, respectively; the longest time for CMV positive detection was 3 and 26 days after onset, respectively; the longest time for HHV6 positive detection was 7 and 8 days after onset, respectively; the longest time for HSV1 positive detection was both 12 days after the onset; in serum samples, the longest time for EBV positive detection was10 days after onset, but in cerebrospinal fluid, no EBV was detected within 10 days of onset.@*Conclusions@#EV-A71 is the most prevalent pathogen causing viral encephalitis in hunan, the overall positive rate of virus in serum samples was higher than that in cerebrospinal fluid samples. Virus stays longer in serum than in cerebrospinal fluid. It is suggested that the time is of great significance for the pathogen detection of children with viral encephalitis, the specimen type can be selected reasonably according to the time of onset.

Design and analysis of two groups interrupt time series / 中华流行病学杂志

Interrupted time-series (ITS) is a quasi-experimental design which evaluates the effectiveness of an intervention based on time-series outcome variables. Compared with the single group of ITS, the two groups of ITS can better control the influence of pre-interventional confounding factors and evaluate the effectiveness of the intervention. This paper summarizes the principles and statistical methods of two groups of ITS by an example of evaluating vaccine effect on the incidence of a disease in two cities. The regression model is fitted by Prais-Winsten method and Newey-West method and the results are explained and compared in detail. When the intervention is performed with other confounding interventions at the same time, the two groups of ITS can be more effective to balance the existing trends before the intervention, and evaluate the effectiveness of intervention. The method of two groups of ITS has important practical significance, providing new insights in program evaluation.

Application and trend of development of digital PCR in virus detection / 中华实验和临床病毒学杂志

In recent years, digital PCR (dPCR), as a novel nucleic acid amplification detection technique, contrasts real-time fluorescent quantitative PCR (qPCR) without the need to establish a standard curve, "single molecule template amplification" absolute quantification, insensitive to inhibitors that affect PCR efficiency, and excellent sensitivity, specificity, extremely strong repeatability and the development of the existing commercial platform technology that has great application advantages in virology and nucleic acid quantification. In this review, we address the application advantages of digital PCR in virus detection and describe its future development.

Multiplex real time polymerase chain reaction technique in detection of respiratory syncytial virus and adenovirus: a Meta analysis / 中华实验和临床病毒学杂志

Objective@#To systematically review the diagnostic accuracies of multiplex real time polymerase chain reaction (MRT-PCR) technique for detection of respiratory syncytial virus (RSV) and adenovirus (ADV).@*Methods@#PubMed, EMBASE, Cochrane, Wanfang and CNKI databases were searched from January1 2010 to January1 2018, to collect reports on MRT-PCR for detection of common respiratory viruses. Then two authors independently exacted the data and assessed the risk of bias of included studies by using the QUADAS-2 tool. Meta-disc 1.4.@*Results@#Ten articles with 2528 cases were eligible for analysis. The result of meta-analysis showed that, the pooled Sen, Spe and area under SROC curve, for detecting RSV were 0.87 (95% CI 0.83 to 0.90), 0.98 (95% CI 0.97 to 0.98) and 1.00. The pooled Sen, Spe, and area under SROC curve of MRT-PCR for detecting ADV were 0.64 (95% CI 0.56 to 0.71), 0.99 (95% CI 0.98 to 0.99) and 0.99. Deeks test indicated that no publication bias was found.@*Conclusions@#The sensitivity of MRT-PCR in RSV and ADV detection is still to be improved, but the overall detection ability is good which deserves to be recommended for clinical use.

Application of nanopore sequencing technology in viral genome research / 中华实验和临床病毒学杂志

MinION is a commercial nanopore sequencer developed by Oxford Nanopore Techno-logies. Compared with the second-generation sequencing platform, MinION is characterized by ultra-long reads, simple and fast sequencing, high portability and real-time data analysis. MinION has been widely applied to the fields of virus identification, whole genome sequencing, new virus discovery, quasispecies and virus evolution. In this article, we review the application of nanopore sequencing technology in the research of the viral genome in recent years, and discuss the existing problems. We also present the prospect for the development trend of nanopore sequencing in the future.

A study of establishment of non-standard methods in qualification accreditation of virological laboratories / 中华实验和临床病毒学杂志

The laboratories of National Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention, have developed a series of experimental method . These method have unique advantages over the national standard method and industry standard method . How to make these method become public products through legal procedures to serve disease prevention and control more extensively is an obligatory task for national virological laboratories. This article explores the establishment, verification, validation, and examination of virological non-standard method under laboratory certification and accreditation conditions through empirical research on the " RT-RAA method for detecting EV71/CA16 and other viruses" .

Detection of human Torque teno virus in serum sample of patients with fever of unknown origin by next generation sequencing / 中华实验和临床病毒学杂志

Objective@#To clarify the potential pathogen for fever of unknown origin (FUO) in serum samples for which pathogenic agents were hardly identified with conventional exainatins.@*Methods@#Random capturing the nucleic acid of pathogen was performed by utilizing the property of sequence non-dependence of next generation sequencing (NGS), followed by enrichment of nucleic acid with multiple displacement amplification (MDA). After sequencing, metagenomic analysis was applied to the raw data and the phylogenetic tree was built to identify the potential pathogen.@*Results@#The result did not indicate common pathogens for FUO but showed the existence of Torque teno Viurs (TTV). Assembly was carried out to all sequencing reads. The coverage of consensus sequence on reference was calculated. Phylogenetic result indicated that all confidence sequences belonged to 3 genera (α TTV, β TTV and γ TTV).@*Conclusions@#The characteristics of genome, phylogenesis of TTV and TTV as signal at immunology level were analyzed and clarified. The possible explanation for detection of TTV in 3 genera may be that TTV itself or non-infectious factors caused the immunosuppression, which finally result e in rise of TTV detection.

Detection of Vibrio parahaemolyticus virulence gene by cross primer thermostatic amplification technology / 中华实验和临床病毒学杂志

Objective@#To establish a CPA-nucleic acid test strip method for rapid detection of Vibrio parahaemolyticus thermostable direct hemolysin (TDH) and heat-labile hemolysin (TLH).@*Methods@#Primers and probes were designed according to the sequence of TDH and TLH virulence genes of Vibrio parahaemolyticus, The optimal reaction temperature and time were established by optimizing the reaction system. The CPA method was compared with the fluorescence quantitative PCR.@*Results@#The optimal temperature and time for the CPA-nucleic acid test strip method established in this study were 60 ℃ and 40 min, which were highly specific to the TDH and TLH virulence genes of Vibrio parahaemolyticus. The TLH And TDH detection limit of 1.8 cfu/mL and 16.0 cfu/mL; consistent with the method of fluorescence quantitative PCR, and has good stability. The positive rates of TLH and TDH genes were 100% (464/464) and 72.6% (337/464) in 464 isolates, respectively. The positive rate of TDH virulence genes in isolates of aquatic products was 7.81 % (10/128). The positive rate of TDH virulence gene in clinical isolates was 97.32% (327/336).@*Conclusions@#The CPA-nucleic acid test strip method for detection of Vibrio parahaemolyticus TDH and TLH genes is rapid, efficient, sensitive, specific and easy to operate. It can be used in the rapid field detection of Vibrio parahaemolyticus.

Loop-mediated isothermal amplification technique in the diagnosis of hepatitis B virus infection: a Meta-analysis / 中华预防医学杂志

Objective@#To evaluate the diagnostic performance of Loop-mediated isothermal amplification (LAMP) in the diagnosis of Hepatitis B Virus (HBV) infection using Meta-analysis.@*Methods@#Literatures about LAMP in the diagnosis of HBV throughPubMed database of the National Library of Medicine, the EMBASE database of the Dutch Medical Digest, the Cochrane Clinical Trials Database, China Science Periodical Database, CSPD and the China National Knowledge Infrastructure (CNKI) were searched from 2000 to 2016, and the Language limited to Chinese and English. English search terms include: LAMP, Loop-mediated isothermal amplification, HBV, hepatitis B virus; Chinese search terms include: loop-mediated isothermal amplification technology, HBV, hepatitis B virus. The keywords and free words are combined to search the literature, and the references mentioned in the retrieval literature are searched twice. The pooled sensitivity, specificity, positive likelihood ratio, negative likelihood ratio, diagnostic odds ratio (DOR), Q index as well as area under summary receiver operating characteristic curve (SROC) were calculated with Stata 12.0 software.@*Results@#A total of 12 literatures with 1 494 cases were included. The pooled sensitivity, specificity, positive likelihood ratio and negative likelihood ratio were 0.922 (95%CI: 0.905-0.937), 0.860 (95%CI: 0.818-0.896), 0.093 (95%CI: 0.048-0.182), and 15.400 (95%CI: 2.003-118.380), respectively. The DOR, area under SROC and Q index were 311.090 (95%CI: 95.841-1 009.800), 0.986 (95%CI: 0.974-0.998) and 0.949 (95%CI: 0.922-0.976), respectively. Deek's test indicates that no publication bias were found (P=0.140).@*Conclusion@#LAMP is worth to be popularized in field tests and primary-level hospitals tests.

Medical service and management of pregnant women during the Ebola epidemic / 中华实验和临床病毒学杂志

The Ebola epidemic occurrence during 2014-2016 is the largest-ever Ebola virus outbreak with an unprecedented scale and impact. The difficulties faced by pregnant women living within the epidemic area are complex and multi-factorial which merits our concern. In this review, we aimed to summarize some of the guidelines published by the World Health Organization during the outbreak, and to provide a broad overview of the issues that arise from pregnant women. Based on our own experience in Sierra Leone, we also made an analysis of the complex interaction between the Ebola virus disease, pregnancy, medical staff, public health systems and the society, and intended to afford peers useful information. The management strategies should be prepared in advance to against the potential epidemic threats and ensure the reliability of life-saving maternity services in China.

Distinguish vaccine strain and wild type strain of yellow fever virus imported to China using high-throughput sequencing technology / 中华实验和临床病毒学杂志

Objective@#To identify whether the three imported yellow fever cases in China in March 2016 were infections by wild type strain of yellow fever virus in Angola in 2016, vaccine-associated disease or co-infection of both.@*Methods@#Sequences of three yellow fever virus strains were obtained by high-throughput sequencing with IonTorrent PGM platform from blood or urine samples of three yellow fever cases, and their genomic characteristics were analyzed. Then the regions with relatively great difference between the wild type strain and 17D vaccine strain were identified, and then served as the reference sequences when mapping the reads obtained by high-throughput sequencing.@*Results@#Partial yellow fever virus genomes were obtained from three samples of yellow fever patients, among them a full length coding region sequence was gained in sample 2. Comparing the genome sequences, the three newly obtained strains of yellow fever virus were highly similar to strain CNYF01R / 2016 which was isolated from the first imported yellow fever case to China in 2016 and strain Angola 71 from Angola in 1971, and they all belonged to Angola genotype of yellow fever virus. In this study, we found five regions in yellow fever virus genomes with great diversity between the vaccine strain and the wild type strain. In these five regions, a number of short reads obtained by high-throughput sequencing of the three samples were mapped to the sequence of wild type virus, while no short reads matched the vaccine strain.@*Conclusions@#There were no viral nucleic acid of 17D vaccine strain in the blood or urine samples of these three cases of yellow fever. They are all infected by wild type strains of Angola in 2016.

Research advances in the etiology of viral encephalitis meningitis syndromes / 中华实验和临床病毒学杂志

Encephalitis meningitis syndrome is a disease of the central nervous system which is caused by bacteria, viruses, parasites and other pathogens. Viral encephalitis meningitis is more common, and the major pathogens include arbovirus, enterovirus, paramyxovirus, herpes simplex virus, rhabdovirus, adenovirus, etc. Here we briefly review the literature regarding the development of the etiology of viral encephalitis meningitis syndrome, meanwhile the development trend of the research on viral meningitis syndrome is also discussed.

A improved tetra-primer amplification refractory mutation system PCR based assay to detect four single nucleotide polymorphisms associated with folate metabolism in a single reaction / 中华检验医学杂志

Objective To develop a tetra-primer amplification refractory mutation system PCR(T-ARMS-PCR)assay for detecting four single nucleotide polymorphisms(SNPs): rs1801133,rs1801131, rs1805087 and rs1801394 associated with folate metabolism in a single reaction tube.Methods Methodology was developed.Then, it applied the established method to analyze 150 physical examination children′s anticoagulant blood samples admitted to the Department of Clinical Laboratory, Children′s Hospital of Hebei Province from January 2017 to April 2017.Four sets of chimeric primers consisting of a universal Tag sequence and targeting rs 1801133,rs1801131,rs1805087 and rs1801394 fused to the specific sequence were designed according to the T-ARMS-PCR principle and the multiplex chimeric primers strategy.A single tube PCR was then conducted after optimizing the primer concentrations and the reaction conditions.The amplified products were analyzed by QIAxcel capillary electrophoresis, and the corresponding SNP genotypes of 150 samples were identified.Furthermore,all samples were verified by direct sequencing.And the Hardy-Weinberg Equilibrium(HWE)testing of four SNPs from 150 samples were conducted by SPSS17.0 Chi-square test.Results The improved T-ARMS-PCR combined with capillary electrophoresis can accurately verify eight different alleles of the four SNPs associated with folate metabolism at one time in 3 hours.Four SNPs of 150 whole blood samples were accurately classified and the results were completely consistent with direct sequencing.All the genotype frequencies of these four SNPs were in HWE (χ2rs1801133=0.69, Prs1801133=0.40; χ2rs1801131=0.21, Prs1801131=0.64; χ2rs1805087=3.32, Prs1805087=0.07;χ2rs1801394=1.91, Prs1801394=0.17).Conclusions The proposed improved T-ARMS-PCR combined with capillary electrophoresis in this study can accurately verify four SNPs associated with folate metabolism in a single reaction tube.This method might be a valuable tool to specifically guide the folate supplement in general population.

Detection of the Zaire Subtype of the Ebola Virus by Isothermal Multiple Self-matching Initiated Amplification / 病毒学报

Given the Ebola outbreak in West Africa and the risks of spread to other regions, a rapid, sensitive and simple method for the detection of the Ebola virus (EBOV) is of great significance for the prevention and control of Ebola. We developed a simple colorimetric isothermal multiple self-matching initiated amplification (IMSA) for rapid detection of the Zaire subtype of the Ebola virus (EBOV-Z). This method employed six primers that recognized seven sites of the EBOV-Z nucleoprotein gene for amplification of nucleic acids under isothermal conditions at 63 degrees C for 1 h. Amplification products were detected through visual inspection of color change by pre-addition of hydroxyl naphthol blue dye. Relative sensitivity was validated by detection of serial tenfold dilutions of virus-like particles containing the partial EBOV-Z nucleoprotein gene and mock clinical sample. Specificity of IMSA was validated by detection of the plasma of 30 healthy volunteers, the dengue virus, and Japanese encephalitis virus. IMSA had comparable sensitivity to Reverse transcription-quantitative polymerase chain reaction (RT-qPCR), and cross-reaction with human plasma or other viruses was not observed. Reverse transcription-isothermal multiple self-matching initiated amplification (RT-IMSA) was also evaluated and compared in parallel with the commercial RT-qPCR kit for detection of EBOV-suspected samples of human blood in Sierra Leone. Sensitivity and specificity of the RT-IMSA was 91.4% and 100%, respectively. These data suggest that RT-IMSA is a valuable tool for the detection of the EBOV with the distinct advantages of simplicity and low cost compared with RT-qPCR.

Colorimetric detection of coxsackievirus A6 by reverse transcription loop mediated isothermal amplification / 中华预防医学杂志

<p><b>OBJECTIVE</b>To develop a simple, rapid and sensitive colorimetric reverse-transcription loop-mediated isothermal amplification (RT-LAMP) for rapid detection of coxsackievirus A6 (CV-A6) based on the colour chang of hydroxy naphthol blue (HNB).</p><p><b>METHODS</b>The method employed a set of six primers that recognized sequences of VP1 gene for amplification of nucleic acid under isothermal conditions at 63 °C for 50 min. The products were detected through visual inspection of color change by the pre-addition of HNB dye. The specificity was validated by detecting a collection of different human enteroviruses. The sensitivity of this assay was evaluated by serial dilutions of RNA molecules from in vitro transcription of CV-A6 VP1 gene, and compared with real-time RT-PCR (rRT-PCR) in parallel. This assay was evaluated with 92 clinical specimens from patients with hand-foot-mouth disease.</p><p><b>RESULTS</b>A positive color (sky blue) was only observed in the preparation of CV-A6, whereas none of the other 23 kinds of human enteroviruses showed a color change. The HNB based RT-LAMP showed a sensitivity of 100 copies/reaction, which was at the same level as that of the rRT-PCR. The result of RT-LAMP in analysis of 92 clinical specimens was consistent with that of the rRT-PCR. The kappa correlation between the two methods was 1 and both of the sensitivity and specificity of the RT-LAMP assay were 100%.</p><p><b>CONCLUSION</b>The established RT-LAMP assay had good specificity and sensitivity and thus demonstrated to be a promising screening tool for CV-A6 infection. It also has the potential to be used in resource-limited clinical sites and field study.</p>

Detection of Middle East Respiratory Syndrome Coronavirus by Reverse-transcription Loop-Mediated Isothermal Amplification / 病毒学报

A simple, rapid and sensitive colorimetric reverse-transcription loop-mediated isothermal amplification (RT-LAMP) was developed for rapid detection of Middle East respiratory syndrome coronavirus (MERS-CoV). The method employed six primers that recognized sequences of a nucleocapsid gene for amplification of nucleic acids under isothermal conditions at 63 degrees C for 60 min. Products were detected through a LA-320c Loopamp Turbidimeter (real-time RT-LAMP) or visual inspection of color change by pre-addition of Hydroxynaphthol Blue dye (visual RT-LAMP). Specificity of RT-LAMP was validated by detection of several human coronaviruses and common respiratory viruses. MERS-CoV real-time RT-LAMP had a linear correlation (R2) of 0.995 at 10(3)-10(6) copies. The limit of detection of real-time RT-LAMP, visual RT-LAMP and quantitative real-time PCR was 500, 1000 and 100 copies/reaction, respectively. The established RT-LAMP assay was demonstrated to be a rapid screening tool for MERS-CoV infection, and could be suitable in resource-limited clinical sites and for field studies.

Advance in Deep Sequencing of Small RNAs for Virus Identification and Discovery / 病毒学报

Small RNAs (sRNA) are produced abundantly in either plants or animals and function in regulating gene expression or in defense against virus infection. Deep sequencing of small RNAs is an emerging technology in virus identification and de novo assembly of virus genomes and is demonstrated to be an effective method to discover new viruses and monitor virus variation. A significant number of viruses from plants, invertebrates and human cells has been successfully identified using this technology. In this paper, we summarized the principle, operation process and latest advances of sRNA deep sequencing We also showed the feasibility of sRNA deep sequencing by bioinformatic analysis using sRNA deep sequencing dataset public available for the detection of viruses.