Biomedical subjects

S S Monroe

Publications and source records attributed to S S Monroe.

At least 55 records · Page 3Linked to original sources

PubMed1997

Capsid protein composition of reference strains and wild isolates of human astroviruses.

Astroviruses are small RNA viruses that are frequently associated with gastroenteritis in humans and animals. Despite much work on the genetic analysis of astrovirus strains, little progress has been made in the characterization of the proteins composing mature virions. We have analyzed the capsid protein composition of the reference strains and several wild isolates of human astroviruses using high-resolution polyacrylamide gradient gels. For reference strains of the seven serotypes analyzed, a consistent pattern of three infection-specific proteins--designated P1, P2, and P3 -was generally observed. The strains could be divided into two groups, based upon the reactivity of these proteins in immune precipitation assays that used homologous rabbit serum. One group included reference types 1 4 for which all three proteins were precipitated by homologous rabbit sera; for the other group, types 5 7, only proteins P2 and P3 were precipitated. When wild isolates from around the world were compared to the reference strains, a correlation between genetic type and the pattern of protein sizes and immune reactivity was observed for strains of the common types (1-4). Strains of types 2 and 4 consistently exhibited P3 proteins larger than those of types 1 and 3. Unusual patterns of proteins or immune reactivity were detected in strains of types 5-7, indicating that there may be incomplete processing of the capsid precursor during growth in cell culture.

Amino Acids↗

PubMed1997

A one-tube method of reverse transcription-PCR to efficiently amplify a 3-kilobase region from the RNA polymerase gene to the poly(A) tail of small round-structured viruses (Norwalk-like viruses).

Amplification of a 3-kb genome region from the RNA polymerase gene to the 3' poly(A) tail of small round-structured virus (SRSV) by reverse transcription-PCR (RT-PCR) has been difficult to achieve because of a stable secondary structure in a region between the RNA polymerase gene and the 5' end of the second open reading frame. We have developed a one-tube RT-PCR method to efficiently amplify this region. The method comprises three procedures: purification of poly(A)+ RNA from a starting RNA solution by oligo(dT)30 covalently linked to latex particles, buffer exchange, and continuous RT and PCR in a single tube containing all reaction components. The key elements of this method are (i) first-strand cDNA synthesis with the Superscript II version of RNase H- Moloney murine leukemia virus reverse transcriptase at 50 degrees C for 10 min by using the RNA-oligo(dT)30 hybrid on the latex particles as the template and primer, and (ii) PCR by Taq and Pwo DNA polymerases mixed together with a mixture of 12 phased oligo(dT)25 antisense primers. The detection threshold of the one-tube RT-PCR method was as little as 0.2 ng of the crude RNA used as the source of the template. Using this method, we obtained 3-kb products from 24 SRSV strains previously characterized into four genetic groups. These included 5 P1-A, 4 P1-B, 5 P2-A, and 10 P2-B strains. Because SRSVs have not yet been cultivated in vitro, this novel method should facilitate molecular characterization of SRSVs to provide a firm scientific foundation for improvements and refinements of SRSV diagnostics.

Base Sequence↗

PubMed1997

Parkville virus: a novel genetic variant of human calicivirus in the Sapporo virus clade, associated with an outbreak of gastroenteritis in adults.

This report describes the characterization of Parkville virus, the etiologic agent of an outbreak of foodborne gastroenteritis, that has the morphology of a calicivirus and genetic properties that distinguish it from previously identified strains in the Sapporo/Manchester virus clade. Sequence analysis of the Parkville virus genome showed it contained the RNA-dependent RNA polymerase motifs GLPSG and YGDD characteristic of members of the family Caliciviridae with an organization identical to that reported for the Manchester virus where the capsid region of the polyprotein is fused to the RNA polymerase. Parkville virus however, demonstrates considerable sequence divergence from both the Manchester and Sapporo caliciviruses, providing the first indications that genetic diversity exists within caliciviruses of this previously homogeneous clade. On the basis of recent advances in the genetic characterization of members of the family Caliciviridae, we propose a new interim phylogenetic classification system in which Parkville virus would be included with Manchester and Sapporo virus as a separate group distinct from the small round-structured viruses (Norwalk-like viruses) that also cause diarrhea in humans.

Adult↗

PubMed1996

The changing epidemiology of astrovirus-associated gastroenteritis: a review.

Our understanding of the epidemiology of astrovirus-associated gastroenteritis has changed markedly with each improvement in detection method. In early surveys based on electronmicroscopy (EM), astroviruses appeared to be a rare cause of gastroenteritis, being found in fewer than 1% of children with diarrhea, usually in small outbreaks of disease and primarily during the winter season. The development and use of monoclonal antibodies and enzyme immunoassays (EIA) to detect astroviruses led to reports of a higher prevalence (2.5%-9%) of astrovirus infection among patients hospitalized with diarrhea. Astroviruses appeared second only to rotaviruses as a cause of hospitalization for childhood viral gastroenteritis. Studies based on EIA detection of astroviruses indicate that astroviruses are common causes of diarrhea in children worldwide, and that most children are infected during their first two years of life. The elderly and the immunocompromised represent high-risk groups as well. The observations that newborns monitored prospectively rarely have repeat disease and that the rate of detection decreases with increasing age suggest that immunity to astroviruses, as immunity to rotaviruses, may develop early in life. The cloning and sequencing of astroviruses have led to more sensitive assays to detect the viruses by reverse transcription, polymerase chain reaction (RT-PCR). Application of RT-PCR for detection of astroviruses in children in day-care centers showed a marked increase in the detected prevalence of astrovirus-associated diarrhea, the rate of asymptomatic infection, and the duration of shedding of virus among those infected, when compared with studies that used other methods. As with rotaviruses, neither the mode of transmission nor the reservoir of astrovirus infection has been identified. Both immune and molecular-based assays to detect astrovirus serotypes indicate that serotype 1 is most common worldwide, although the predominant serotypes may vary by region and time. In the absence of obvious strategies to prevent astrovirus-associated diarrhea, vaccines might be considered if further studies establish that the disease burden would render such a vaccine cost-effective.

Animals↗

PubMed1996

Characterization of Toronto virus capsid protein expressed in baculovirus.

Toronto virus (TV), previously called "minireovirus", a human calicivirus classified as genogroup 2 and phylogenetic type P2-A, was originally described in association with diarrhea in children. The second open reading frame, encoding the capsid protein of TV24, was expressed in a baculovirus recombinant. The recombinant baculovirus produced a protein (rTV) with an apparent molecular mass of 58 kDa that self-assembled into virus-like particles approximately 30 nm in diameter with a density of 1.29 g/ml. Antigenic and immunogenic characteristics of these particles were determined by protein immunoblot, immunoprecipitation, and enzyme immunoassay. Seroconversion to the rTV protein was detected in 6 of 8 (75%) patients from a recent outbreak of gastroenteritis associated with a virus of similar phylogenetic type. These results confirm and extend the previous reports of the expression of the Norwalk and Mexico virus capsid proteins.

Animals↗

PubMed1996

An outbreak of viral gastroenteritis in a nursing home: importance of excluding ill employees.

BACKGROUND: In May 1994, 43 persons in a nursing home were reported with gastroenteritis. An outbreak investigation was conducted to determine risk factors for gastroenteritis among residents and staff. METHODS: Data were analyzed using contingency tables; relative risks (RR) and statistical significance were determined with Fisher's Exact Test. The chi-squared statistic to perform a goodness of fit test for the binomial distribution was used to determine whether cases occurred randomly and independently of each other. Stools were tested for bacterial enteric pathogens, ova, and parasites and were examined by electron microscopy, Southern hybridization, and reverse transcription-polymerase chain reaction. Paired sera were collected to detect fourfold rises in antibody titer by enzyme immunoassay against Norwalk viruses. RESULTS: Of 121 residents, 62 (51%) had gastroenteritis, as did 64 (47%) of the 136 staff. The index case was a nurse who became ill at work and continued to work, while symptomatic, for another 2 days. Only residents who had received medications from this nurse between May 17 and May 20 became ill on the first day of the outbreak (13 of 35 versus 0 of 5). Nurses and nurse aides were more likely than employees without direct resident contact to be cases (46 of 68 versus 18 of 58; RR, 2.18; P < .001). Bacterial stool cultures and parasite examinations were negative. Results of electron microscopy, polymerase chain reaction with Southern hybridization, and enzyme immunoassay indicated the causative agent was a small, round, structured virus similar to the Snow Mountain Agent. CONCLUSION: To minimize outbreaks in nursing homes, we recommend that ill staff be excluded from work until symptoms resolve.

Adult↗

PubMed1996

A university outbreak of gastroenteritis due to a small round-structured virus. Application of molecular diagnostics to identify the etiologic agent and patterns of transmission.

An epidemiologic investigation of a gastroenteritis outbreak in December 1994 indicated that salad consumption during lunch was linked with illness on 2 days (5 December: odds ratio [OR]=3.1, 95% confidence interval [CI]=2.0-5.0; 6 December: OR=3.1, 95% CI=1.9-4.9). Single stool or vomitus specimens from ill students and staff (case-patients) were examined for bacterial and viral pathogens. Small round-structured viruses (SRSVs) were detected by electron microscopy in stool specimens from 9 of 19 case-patients and in vomitus specimens from 3 of 5 case-patients. By reverse transcription-polymerase chain reaction (RT-PCR), the SRSVs were shown to be G-2/P2-B type strain. The nucleotide sequences of RT-PCR products from vomitus and stool specimens of ill students were identical to stool specimens from the ill salad chef. These findings suggest that a single SRSV strain was the etiologic agent in the outbreak that was possibly transmitted to students through consumption of contaminated salad. Epidemiologic investigation in conjunction with molecular diagnostics may enable early identification of sources of infection and improve outbreak control.

Caliciviridae↗

PubMed1996

Detection and analysis of a small round-structured virus strain in oysters implicated in an outbreak of acute gastroenteritis.

Outbreaks of shellfish-transmitted viral disease occur periodically, but frequently the causative agent is not identified. In November 1993, during investigation of a multistate outbreak of acute gastroenteritis, incriminated lots of oysters were collected. Oyster tissues (stomachs and digestive diverticula) were processed for virus extraction and nucleic acid purification. Human calicivirus sequences were sought by reverse transcriptase PCR using different primer sets. Amplicons were obtained from 9 of 10 shellfish samples from four different lots when primers specific for the outbreak virus strain were used. The specificity of the amplification was confirmed by hybridization. The amplicons from the nine positive oysters were cloned and sequenced. The sequence of each of the clones was identical to the others but showed some variation (7 of 81 bp) from the sequences obtained from the stools of three persons made III by the outbreak.

Animals↗

PubMed1995

An outbreak of Norwalk virus gastroenteritis associated with eating raw oysters. Implications for maintaining safe oyster beds.

OBJECTIVE: To determine the characteristics and the cause of an outbreak of gastroenteritis associated with eating raw oysters. DESIGN: Survey of groups of persons reporting illness to the health department after eating oysters; survey of convenience sample of oyster harvesters; and tracing of implicated oysters. SETTING: General community. MAIN OUTCOME MEASURES: Relative risk for illness after oyster consumption, source bed of contaminated oysters, presence of antibodies to Norwalk virus in serum, presence of a Norwalk virus in stool by direct electron microscopy and reverse transcription-polymerase chain reaction (RT-PCR), and DNA sequences of RT-PCR products. RESULTS: Seventy (83%) of 84 persons who ate raw oysters became ill vs three (7%) of 43 people who did not eat raw oysters (relative risk, 11.9; 95% confidence interval, 4.0 to 34.2). Eleven (79%) of 14 serum pairs had at least a fourfold increase in antibody to Norwalk virus. All 12 stool samples tested were positive by electron microscopy and/or RT-PCR for Norwalk virus. The RT-PCR products from all seven stool samples tested had identical DNA sequences. Implicated oysters were harvested November 9 through 13, 1993, from a remote oyster bed. Crews from 22 (85%) of 26 oyster harvesting boats working in this area reported routine overboard disposal of sewage. One harvester with a high level of antibodies to Norwalk virus reported having gastroenteritis November 7 through 10 and overboard disposal of feces into the oyster bed. CONCLUSIONS: This outbreak was caused by contamination of oysters in the oyster bed, probably by stool from one or more ill harvesters. Education of oyster harvesters and enforcement of regulations governing waste disposal by oyster harvesting boats might prevent similar outbreaks.

Adolescent↗

PubMed1995

Epidemiologic applications of novel molecular methods to detect and differentiate small round structured viruses (Norwalk-like viruses).

The molecular epidemiology of a large, multistate outbreak of oyster-associated gastroenteritis [Kohn et al. (1995): Journal of the American Medical Association 273:466-471. Dowell et al. (1995): Journal of Infectious Diseases 171:1497-1503.] was examined using new methods to detect small round structured viruses (SRSVs) by reverse transcription-polymerase chain reaction (RT-PCR) and to characterize strains by Southern hybridization and nucleotide sequencing of 81-bp of a PCR product amplified from the RNA polymerase gene. Of 37 stool specimens examined from patients in eight clusters of the multistate outbreak, 32 (86%) gave RT-PCR products specific for SRSVs of P1-A phylogenetic group. Nineteen PCR products from the eight clusters were confirmed to have the identical sequence, indicating that this large outbreak was attributed to a single strain of SRSV. In one of the eight clusters, five (63%) of eight patients had a mixed infection with a second SRSV strain that belonged to P2-B phylogenetic group. Of 12 specimens from patients in five other outbreaks and one sporadic case which occurred at the same time as the multistate outbreak, 10 (83%) gave products specific for SRSVs representing four phylogenetic groups (P1-A, P1-B, P2-A, and P2-B). The sequences of the P1-A products from two outbreaks and that of the P2-B product from another outbreak were identical to the P1-A sequence from the eight clusters and the P2-B sequence from the one cluster of the multistate outbreak, respectively. These results demonstrate the first application of these methods to enhance our understanding of the molecular epidemiology of SRSVs and provide answers of public health interest that could not have been obtained using classical epidemiologic methods alone.

Animals↗

PubMed1995

A multistate outbreak of oyster-associated gastroenteritis: implications for interstate tracing of contaminated shellfish.

In November 1993, clusters of gastroenteritis in six states following oyster consumption were investigated to identify common features, and stool samples were obtained to identify a pathogen. Efforts were made to account for all potentially contaminated oysters using harvest tags and the interstate recall system. Consumption of oysters was associated with illness in 10 clusters; no other food was implicated. A Norwalk-like virus was detected by electron microscopy in 9 of 18 samples and by reverse transcription-polymerase chain reaction in 20 of 26 samples from 6 clusters. Nucleotide sequences of a 123-bp fragment from all specimens were identical, consistent with a common source outbreak. Implicated oysters were harvested from the Louisiana coast between 9 and 12 November. Although some were recalled and destroyed, most oysters harvested from the area during this time remain unaccounted for. Current regulations and commercial practices need to be revised to permit thorough tracing and recall of contaminated oysters and to improve control of future epidemics.

Animals↗

PubMed1995

Virologic features of an astrovirus diarrhea outbreak in a day care center revealed by reverse transcriptase-polymerase chain reaction.

Astroviruses cause outbreaks of diarrhea in children attending day care centers (DCCs). Reverse transcriptase-polymerase chain reaction (RT-PCR) was compared with EIA detection of astrovirus in stool specimens to characterize further the molecular epidemiology of an outbreak of astrovirus-associated gastroenteritis. Three hundred sixty-eight stool specimens collected prospectively from 36 children enrolled in a DCC during an 11-week outbreak of diarrhea were evaluated by EIA and RT-PCR. Astrovirus was detected in 32% of specimens by RT-PCR versus 10% by EIA (P < .001) and in 89% of children by RT-PCR versus 50% by EIA. The median duration of astrovirus excretion episodes detected by EIA was 1.5 days versus 4 days by RT-PCR (P = .06). Astrovirus was excreted for prolonged periods by immunocompetent children during this outbreak. RT-PCR was more sensitive than EIA for detection of astrovirus in stool specimens and redefined the epidemiology of astrovirus infection in this setting.

Age Factors↗

PubMed1995

Detection and differentiation of antigenically distinct small round-structured viruses (Norwalk-like viruses) by reverse transcription-PCR and southern hybridization.

Application of reverse transcription (RT)-PCR to detect small round-structured viruses (SRSVs) from fecal specimens of patients with gastroenteritis has been insensitive because of the tremendous sequence heterogeneity between strains. We have designed two RT-PCR primer sets (G-1 and G-2) based on the nucleotide sequence diversity in the RNA polymerase gene of SRSVs belonging to two distinct genogroups represented by Norwalk virus (primers G-1) and Snow Mountain agent (primers G-2). All 22 SRSV strains examined that had been classified previously by solid-phase immune electron microscopy into four antigenic types (UK1, UK2, UK3, and UK4) could be detected by RT-PCR with these two primer sets. The G-1 primer set detected 6 UK2 strains, and the G-2 primers detected 16 strains, including 7 UK1, 5 UK3, and 4 UK4 strains. On the basis of nucleotide sequences of 81-bp fragments of the RT-PCR products from 13 strains determined in this study, together with those previously reported for 17 SRSV strains, we designed four sets of internal oligonucleotide probes (P1-A, P1-B, P2-A, and P2-B) for Southern hybridization, using chemiluminescent detection. The P1-A probe hybridized with PCR products from the UK2 strains; the P1-B probe, with products from two of the seven UK1 strains; the P2-A probe, with four of the remaining five UK1 strains; and the P2-B probe, with products from both UK3 and UK4 strains, as well as with one strain originally typed as UK1 which showed cross-reactivity with UK4 upon retesting by solid-phase immune electron microscopy. RT-PCR with both the G-1 and the G-2 primer sets can increase the detection rate of the many antigenically distinct SRSVs and, when combined with Southern hybridization, may predict the antigenic type of the SRSV associated with infection.

Adult↗

PubMed1995

Use of solid-phase immune electron microscopy for classification of Norwalk-like viruses into six antigenic groups from 10 outbreaks of gastroenteritis in the United States.

Norwalk-like viruses observed in fecal specimens from 10 outbreaks of gastroenteritis investigated in the United States between 1987 and 1992 were analyzed by solid-phase immune electron microscopy. Outbreak virus strains were classified into six antigenic groups: the four types (UK1 to UK4) previously defined in the United Kingdom, Norwalk virus, and the Oklahoma agent that was newly defined in this study. The diversity of antigenic types demonstrated in these outbreaks was greater than previously recognized and will serve as a basis for characterization of these strains at the molecular level.

Antibodies, Viral↗

PubMed1995

Typing of human astroviruses from clinical isolates by enzyme immunoassay and nucleotide sequencing.

A typing enzyme immunoassay (TYPE-EIA) was used to determine the antigenic types of 64 astrovirus-positive specimens from nine collections from seven countries. Six of the seven known astrovirus types were detected in the collections, with HAstV-1 predominating in all collections for one from the United Kingdom. Selected specimens were analyzed further by reverse transcriptase PCR and nucleotide sequencing of 348 bp within the capsid protein precursor region of the genome. The phylogenetic groupings (genotypes) determined from the sequences were entirely consistent with the antigenic groupings (serotypes) of isolates obtained by using the TYPE-EIA. The genetic variation within genotypes was small compared with the variation between genotypes, allowing unambiguous categorization of all specimens. Although some strains from widely separated geographic areas had identical sequences, in general, within a region most strains of the same type were identical. The TYPE-EIA may help further our understanding of the epidemiology of astrovirus and the possible role of serotype-specific immunity, while further knowledge of sequences could facilitate the development of simpler molecular methods of typing astrovirus strains.

Antigens, Viral↗

PubMed1994

Prevalence of antibodies to Norwalk virus among Amerindians in isolated Amazonian communities.

The seroepidemiology of Norwalk virus infections was examined among Amerindians belonging to eight relatively isolated communities in the Amazon region by means of a new enzyme immunoassay using recombinant Norwalk virus antigen. The seroprevalence of antibodies to Norwalk virus ranged from 39% in the Maiogong to 100% in the Kubenkrankrein. The distribution of antibody levels varied greatly among groups; five of the eight communities had an antibody prevalence greater than 90% with many high values (> 100 units), while three had both a low seroprevalence and a preponderance of low values (< 100 units). While few children less than 5 years of age were sampled, no significant differences in antibody prevalence were noted among age groups, and the prevalence of antibody among children 5-10 years of age approached that of the older age groups. The low prevalence of titers of antibodies to Norwalk virus in several tribes living in these isolated Indian communities suggests that Norwalk virus may have been only recently introduced.

Adolescent↗

PubMed1994

Comparison of the polymerase region of small round structured virus strains previously classified in three antigenic types by solid-phase immune electron microscopy.

We have used a reverse transcription-polymerase chain reaction with nested sets of primers to determine the nucleotide sequences of a 166 base pair segment of the RNA polymerase region of seven strains of small round structured viruses (SRSVs) from the United Kingdom. These SRSV strains were previously classified by solid-phase immune electron microscopy into three antigenic types--UK2, UK3 and UK4, which are comparable to the prototype strains Norwalk virus, Hawaii agent, and Snow Mountain agents, respectively. Based on their sequences, the seven strains from the United Kingdom could be divided into two groups. The first group included two strains of the UK2 type along with Norwalk virus and Southampton virus and the second group included three strains of UK3 and two strains of UK4 types. Viruses in the first group showed 75.3%-77.1% nucleotide and 89.1%-94.6% amino acid identity with Norwalk virus while those of the second group showed 60.8%-63.3% nucleotide and 67.3%-69.1% amino acid identity. Nucleotide and amino acid identity within the second group ranged between 91.6%-99.4% and 96.4%-100%, respectively. These results suggest that the SRSVs antigenically related with Norwalk virus, Hawaii agent, and Snow Mountain agent, can be classified into two genotypes on the basis of their sequences in the RNA polymerase region.

Adult↗

PubMed1994

A large outbreak of acute gastroenteritis associated with astrovirus among students and teachers in Osaka, Japan.

In June 1991, a large outbreak of acute nonbacterial gastroenteritis occurred among students and teachers at 10 primary and 4 junior high schools in Katano City, Osaka, Japan. The outbreak affected > 4700 persons, lasted 5 days, and was believed to have been linked to contaminated food from a common supplier. Astrovirus, identified as the etiologic agent, was detected by direct electron microscopy in 10 of 38 fecal samples obtained from patients with diarrhea. Detection was confirmed by solid-phase immune electron microscopy (IEM), EIA, reverse transcription-polymerase chain reaction, and virus isolation in CaCo-2 cells. Several patients who had astrovirus in their stool also demonstrated a significant antibody response to a reference strain of astrovirus by IEM and EIA and to their own isolate by IEM. Astrovirus can be an important agent of epidemic acute nonbacterial gastroenteritis in school-aged children and adults in Japan.

Acute Disease↗