Biomedical subjects

Stephan S Monroe

Publications and source records attributed to Stephan S Monroe.

26 records · Page 2Linked to original sources

PubMed2003

Development of a rapid method using nucleic acid sequence-based amplification for the detection of astrovirus.

We have developed a rapid method to detect astrovirus in fecal specimens utilizing nucleic acid sequence-based amplification (NASBA) and several detection methodologies, including a sandwich hybridization assay based on DNA-tagged liposomes (liposome-strip detection assay). RNA was extracted from 65 stool specimens that were positive for astrovirus by enzyme immunoassay and was amplified by both NASBA and reverse transcriptase PCR (RT-PCR). Also extracted and amplified were 19 specimens containing rotavirus, 20 specimens containing norovirus, five specimens containing adenovirus, 15 water negative control specimens, and eight specimens containing astrovirus reference strains. NASBA products were detected by electrochemiluminescence detection (ECL) and by liposome-strip detection; RT-PCR products were detected by ethidium bromide staining following gel electrophoresis and by liquid hybridization assay (LHA). There was no significant difference in the detection rates of NASBA- and RT-PCR-based assays, with one exception in which the NASBA/ECL assay detected astrovirus in eight specimens that tested negative by the RT-PCR/LHA assay. These results suggest that these NASBA-based detection methods have detection rates that are as good as or better than those of RT-PCR-based methods. Both NASBA and liposome-strip detection may be useful for field studies and environmental testing because these methods are rapid and do not require specialized equipment.

Astroviridae Infections↗

PubMed2003

Characterization of a novel coronavirus associated with severe acute respiratory syndrome.

In March 2003, a novel coronavirus (SARS-CoV) was discovered in association with cases of severe acute respiratory syndrome (SARS). The sequence of the complete genome of SARS-CoV was determined, and the initial characterization of the viral genome is presented in this report. The genome of SARS-CoV is 29,727 nucleotides in length and has 11 open reading frames, and its genome organization is similar to that of other coronaviruses. Phylogenetic analyses and sequence comparisons showed that SARS-CoV is not closely related to any of the previously characterized coronaviruses.

Amino Acid Sequence↗

PubMed2003

International collaborative study to compare reverse transcriptase PCR assays for detection and genotyping of noroviruses.

To allow more rapid and internationally standardized assessment of the spread of noroviruses (previously called Norwalk-like viruses [NLVs]) as important food-borne pathogens, harmonization of methods for their detection is needed. Diagnosis of NLVs in clinical diagnostic laboratories is usually performed by reverse transciptase PCR (RT-PCR) assays. In the present study, the performance of five different RT-PCR assays for the detection of NLVs was evaluated in an international collaborative study by five laboratories in five countries with a coded panel of 91 fecal specimens. The assays were tested for their sensitivity, detection limit, and ease of standardization. In total, NLVs could be detected by at least one RT-PCR assay in 69 (84%) of the samples that originally tested positive. Sensitivity ranged from 52 to 73% overall and from 54 to 100% and 58 to 85% for genogroup I and II viruses, respectively. In all, 64% of the false-negative results were obtained with a set of diluted stools (n = 20) that may have lost quality upon storage. Sensitivity was improved when these samples were excluded from analysis. No one single assay stood out as the best, although the p1 assay demonstrated the most satisfactory overall performance. To promote comparability of data, this assay will be recommended for newly starting groups in future collaborative studies.

Caliciviridae Infections↗

PubMed2002

A waterborne outbreak of Norwalk-like virus among snowmobilers-Wyoming, 2001.

In February 2001, episodes of acute gastroenteritis were reported to the Wyoming Department of Health from persons who had recently vacationed at a snowmobile lodge in Wyoming. A retrospective cohort study found a significant association between water consumption and illness, and testing identified Norwalk-like virus (NLV) in 8 of 13 stool samples and 1 well. Nucleotide sequences from the positive well-water specimen and 6 of the positive stool samples were identical. This multistrain NLV outbreak investigation illustrates the importance of NLV as a cause of waterborne illness and should encourage monitoring for NLVs in drinking water.

Caliciviridae Infections↗

PubMed2002

Foodborne viral gastroenteritis: challenges and opportunities.

Norwalk-like viruses (NLVs) are estimated to be the most common causes of foodborne disease in the United States, accounting for two-thirds of all food-related illnesses. The epidemiologic features and disease burden associated with NLVs have, until recently, been poorly understood because of the lack of sensitive detection assays and the underuse of available diagnostic tools. However, the application of molecular techniques to diagnose and investigate outbreaks of infection during recent years has led to a growing appreciation of the importance of these agents. NLVs are a principal cause of outbreaks of acute-onset vomiting and diarrhea in all age groups-most commonly, via contamination of uncooked foods by infected foodhandlers, but also via foods contaminated at their sources, such as oysters and raspberries. NLVs may also account for >10% of sporadic cases of gastroenteritis in children and adults. Future research will focus on the development of easy-to-use diagnostic assays based on antigen and antibody detection as well as vaccine development. Implementation of simple prevention measures, including correct food-handling practices, will continue to be a priority.

Caliciviridae Infections↗

PubMed2002

Epidemiologic and molecular trends of "Norwalk-like viruses" associated with outbreaks of gastroenteritis in the United States.

Between July 1997 and June 2000, fecal specimens from 284 outbreaks of nonbacterial gastroenteritis were submitted to the Centers for Disease Control and Prevention for testing for "Norwalk-like viruses" (NLVs). Specimens were examined by reverse-transcription polymerase chain reaction and direct electron microscopy for the presence of NLVs. Adequate descriptive data were available from 233 of the outbreaks, and, of these, 217 (93%) were positive for NLVs. Restaurants and events with catered food were the most common settings, and contaminated food was the most common mode of transmission. Genogroup II (GII) strains were the predominant type (73%), with genogroup I strains causing 26% of all NLV-positive outbreaks. Certain GII clusters (GII/1,4,j) were more commonly associated with outbreaks in nursing home settings than with outbreaks in other settings. Strain diversity was great: one potential new sequence cluster was implicated in multiple outbreaks, and strains belonging to a tentative new genogroup were identified.

Adolescent↗

PubMed2002

Detection and characterisation of human astroviruses in children with acute gastroenteritis in Blantyre, Malawi.

In a 2-year hospital-based study of paediatric gastroenteritis in Blantyre, Malawi, astroviruses were detected by enzyme immunoassay in 15 (1.9%) of 786 inpatients and in 9 (2.3%) of 400 outpatients. Greater disease severity was noted in children coinfected with human immunodeficiency virus (HIV). Six human astrovirus (HAstV) genotypes were identified, including HAstV-1 (25%), HAstV-2 (21%), HAstV-3 (25%), HAstV-4 (13%), HAstV-5 (4%), and HAstV-8 (13%). Although astroviruses are not major causes of gastroenteritis among children admitted to hospital in Blantyre, concomitant HIV infection appears to be a risk factor for increased severity of disease.

Acute Disease↗

PubMed2002

Determination of a universal nucleic acid extraction procedure for PCR detection of gastroenteritis viruses in faecal specimens.

Four nucleic acid extraction protocols were examined for their suitability for extraction of the ssRNA, dsRNA and dsDNA genomes of gastroenteritis viruses, for PCR detection. Protocol (A), employed specimen lysis with guanidinium thiocyanate, extraction with phenol-chloroform-isoamyl alcohol and nucleic acid purification by size-fractionated silica particles. Protocol (B), utilised specimen lysis with guanidinium thiocyanate and nucleic acid purification by silica, followed by phenol-chloroform-isoamyl alcohol extraction. Protocol (C), employed specimen lysis with guanidinium thiocyanate and nucleic acid purification by RNAID glass powder. Protocol (D), employed specimen lysis with sodium dodecyl sulphate, proteinase K digestion and extraction with phenol-chloroform-isoamyl alcohol. Of the four protocols, (B) appeared to be a suitable candidate 'universal' nucleic acid extraction procedure for PCR detection of different viral agents of gastroenteritis in a single nucleic acid extract of a faecal specimen, irrespective of genome composition. Omission of the phenol-chloroform extraction step did not affect negatively the ability of protocol (B) to allow PCR detection of gastroenteritis viruses in faecal specimens. PCR detection of NLVs, astroviruses, rotaviruses and adenoviruses, in single nucleic acid extracts of faecal specimens obtained from the field, confirmed the universality of the modified protocol (B). We propose the modified protocol (B) as a 'universal' nucleic acid extraction procedure, for monoplex PCR detection of gastroenteritis viruses in single nucleic acid extracts of faecal specimens and for development of multiplex PCR for their simultaneous detection.

Adenovirus Infections, Human↗