Quantitation of group A rotavirus by real‐time reverse‐transcription‐polymerase chain reaction: Correlation with clinical severity in children in South India

Kang, Gagandeep; Iturriza‐Gómara, Miren; Wheeler, Jeremy G; Crystal, Premila; Monica, Bindhu; Ramani, Sasirekha; Primrose, Beryl; Moses, Prabhakar D.; Gallimore, Christopher; Brown, David; Gray, Jim

doi:10.1002/jmv.20053

Cited by 129 publications

(100 citation statements)

References 11 publications

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“…Further, several viruses in our panel, such as enterovirus, adenovirus, and parechovirus, are known to also cause infections with diverse other complaints, such as respiratory complaints, sepsis-like illness, or meningitis. Another explanation for these findings can be asymptomatic or long-lasting carriage of these viruses, as has been described in other publications (7). Carriage or shedding of these viruses during a period of diarrhea may give the clinicians a false clue of a correct diagnosis.…”

Section: Discussionmentioning

confidence: 73%

Replacing Traditional Diagnostics of Fecal Viral Pathogens by a Comprehensive Panel of Real-Time PCRs

et al. 2011

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Molecular DNA-based diagnostics are increasingly being used for diagnosis of viral infections. For enteric viruses, PCR assays have also been developed. The aims of this study were to compile and evaluate a comprehensive panel of PCR assays for diagnosis of viruses causing diarrheal disease and to evaluate its use in a largely pediatric population in a 750-bed university medical center. The PCR panel was designed to include assays for detection of adenovirus, astrovirus, enterovirus, norovirus, parechovirus, rotavirus, and sapovirus. The results of the PCR panel were evaluated in relation to conventional viral diagnostics consisting of viral culture and/or rotavirus and adenovirus rapid antigen tests on samples that were taken for routine diagnostics. Comparing conventional with PCR-based testing, the number of viruses detected increased dramatically from 25 to 106 when PCR assays were used. This increase was due mainly to detection of previously undetected viruses, i.e., astrovirus, norovirus, and sapovirus. In 24% of the samples, norovirus was detected. Also, the lower detection limit of PCR-based adenovirus, enterovirus, parechovirus, and rotavirus diagnostics further increased the detection rate. By focusing on samples from patients with complaints of gastroenteritis, detection of a causative agent was increased from 49% by conventional tests to 97% by molecular diagnostics. However, many samples containing low viral loads were found in patients with complaints other than intestinal complaints. In conclusion, the proposed comprehensive PCR panel with appropriate cutoff values can be used for sensitive, rapid, and clinically relevant diagnosis of gastrointestinal viruses.

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Section: Discussionmentioning

confidence: 73%

Replacing Traditional Diagnostics of Fecal Viral Pathogens by a Comprehensive Panel of Real-Time PCRs

et al. 2011

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“…Thirteen samples remained completely non-typeable and none of these samples gave amplicon with a broadly reactive primer pair specific for the VP6 gene [14]. To further investigate if these samples were non-typeable due to the possibly low amount of template we obtained with column based RNA extraction, they were re-amplified after utilization of an alternative RNA extraction method (TRIzol).…”

Section: Resultsmentioning

confidence: 99%

“…Alternatively, an oligonucleotide primer mixture containing G1 to G4 specific typing primers described by Gouvea et al [13] was used in the second-round PCR. cDNAs that gave no amplicons in any of these reactions were subjected to VP6 gene-specific PCR using consensus primers VP6F and VP6R in the reverse transcription and the same primers were utilized in the amplification step [14].…”

Section: Rna Extraction and Genotypingmentioning

confidence: 99%

Molecular characterization of rotaviruses in mid-western Turkey, 2006–2007

et al. 2010

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AbstractVaccines against rotaviruses are now available in numerous countries, including Turkey. As the vaccines may show various efficiencies against different type specificities and routine vaccination in infants might result in selection and immune escape of wild-type rotavirus strains, strain surveillance has been initiated before and during the vaccine introduction. We aimed to provide corresponding information on local strain prevalence in Anatolia, mid-western Turkey during the introduction of rotavirus vaccines. Stool samples positive for group A rotavirus by commercial enzyme immunoassay were subjected to reverse transcription-polymerase chain reaction based genotyping of the outer capsid antigens, VP7 and VP4, determining G and P type specificities respectively. Among 36 fully and 5 partially typeable strains we detected genotype G1, G2, and G9 VP7 specificities and genotype P[4], P[6] and P[8] VP4 specificities in 5 individual and 4 mixed combinations. The most common strain was G2P[4] (n=17), followed by G9P[8] (n=9). Other strains were G1P[8] (n=2), G2P[8] (n=2), G1+2P[8] (n=2), G9P[4] (n=1), G2+9P[8] (n=1), G4+9P[6] (n=1), and G2P[4+8] (n=1). Partially typed strains included 2 G1P[NT] and 3 G2P[NT] strains. Our data may help determine a baseline of the rotavirus genotype prevalence in Turkey and see if changes in the incidence of individual strains will be observed after routine use of vaccine.

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“…The enhanced sensitivity and a lower risk for cross-contamination in rRT-PCR makes it a useful tool for the detection of rotavirus in various situations including sporadic gastroenteritis, outbreaks, and environmental investigations [48] . A competitive rRT-PCR [49] and a semi-quantitative rRT-PCR assays [50] using SYBR Green I dye was developed to detect and to quantitate the RNA of group A rotaviruses, and to estimate the rotavirus load in diarrhoeal samples, respectively. These assays were very efficient as it could detect even small amounts of viral RNA of group A rotaviruses in clinical samples obtained from humans.…”

Section: Negative-stranded Rna Virusesmentioning

confidence: 99%

Applications of Real-time Reverse Transcription Polymerase Chain Reaction in Clinical Virology Laboratories for the Diagnosis of Human Diseases

Manojkumar¹,

Mrudula²

2006

American J. of Infectious Diseases

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Early diagnosis that prevents further expansion of the causative agent and thereby aids control and eradication of infectious agents is critical for countries trying to obtain a particular disease free status. In these diagnostics speed is paramount, including the assay's applicability, sensitivity, specificity, cost effectiveness and patentability. Real-time Reverse Transcription PCR (rRT-PCR) has revolutionized the field of molecular biology and is being utilized increasingly in novel clinical diagnostic assays. This technique has been applied for rapid detection of point mutation, drug resistant variant and genotyping in several viruses. The combination of excellent sensitivity, specificity, low contamination risk, and speed has made this technique an appealing alternative to cell culture or immunoassay-based testing methods for diagnosing infectious diseases. rRT-PCR assays are most established for the detection of viral load and therapy monitoring. In this review, the usefulness or applications of rRT-PCR assays in the diagnosis of some of the important human viral infections have been summarized.

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Quantitation of group A rotavirus by real‐time reverse‐transcription‐polymerase chain reaction: Correlation with clinical severity in children in South India

Cited by 129 publications

References 11 publications

Replacing Traditional Diagnostics of Fecal Viral Pathogens by a Comprehensive Panel of Real-Time PCRs

Replacing Traditional Diagnostics of Fecal Viral Pathogens by a Comprehensive Panel of Real-Time PCRs

Molecular characterization of rotaviruses in mid-western Turkey, 2006–2007

Applications of Real-time Reverse Transcription Polymerase Chain Reaction in Clinical Virology Laboratories for the Diagnosis of Human Diseases

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