Jacqueline S. Noel

Genetic Classification of “Norwalk-like Viruses”

Reverse transcription-polymerase chain reaction has been used worldwide for the diagnosis of Norwalk-like virus (NLV) infection, yet a commonly accepted genetic classification scheme has not been established. Amino acid sequences from four regions of open-reading frame 2 (ORF2) were used to analyze 101 NLV strains, including 2 bovine strains. On the basis of this analysis, a genetic classification scheme is proposed that differentiates 99 human strains into 2 major genetic groups consisting of 5 and 10 genetic clusters, respectively. The 2 bovine strains constitute a newly defined third major genetic group composed of 2 putative clusters represented by each strain. This classification scheme is well supported by the analysis of the entire ORF2 sequences from 38 strains selected to represent the genetic diversity of the human strains used above. This scheme should provide a firm scientific basis for the unified classification of NLV strains detected around the world.

The Epidemiology of Enteric Caliciviruses from Humans: A Reassessment Using New Diagnostics

In the United States, acute gastroenteritis is one of the most commonly noted illnesses on hospital discharge records and death certificates, yet few of these cases have an etiologic diagnosis. The application of new molecular diagnostic methods has shown caliciviruses (previously referred to as the Norwalk family of viruses or small round structured viruses) to be the most common cause of acute gastroenteritis (AGE) outbreaks in the United States, and they may emerge as a common cause of sporadic cases of AGE among both children and adults. Novel molecular methods have permitted outbreak strains to be traced back to their common source and have led to the first identification of virus in implicated vehicles of infection-water, shellfish, and foods contaminated both at their source and by food handlers. The broad application of these methods to routine diagnosis in hospitals and public health laboratories is advancing our appreciation of the full burden of calicivirus-associated diarrhea, and it is opening new avenues for its prevention and control.

Cold Weather Seasonality of Gastroenteritis Associated with Norwalk-like Viruses

Norwalk-like viruses (NLVs) are the most common cause of acute nonbacterial gastroenteritis in adults, but little is known about their seasonality. The lack of specific diagnostic tools impeded study of these viruses in the past, and surveys using electron microscopy often grouped NLVs with other unrelated viruses. A search of the scientific literature found eight surveys of gastroenteritis, which were conducted for at least 1 year, that specifically identified NLVs. Unpublished data from laboratories of 4 NLV researchers were also used. These surveys, which were conducted in eight countries, reported sporadic cases and outbreaks of NLV-associated gastroenteritis among all age groups. The monthly occurrence of these cases and outbreaks was plotted, and while transmission occurred year-round in most surveys, a cold weather peak was demonstrated in 11 of the 12 studies. This key epidemiologic feature of the viruses has important implications concerning their mode of transmission and for understanding the etiology of acute gastroenteritis in adults.

Correlation of patient immune responses with genetically characterized small round-structured viruses involved in outbreaks of nonbacterial acute gastroenteritis in the United States, 1990 to 1995

Small round‐structured viruses (SRSVs) are a genetically and antigenically diverse group of caliciviruses that are the most common cause of outbreaks of acute nonbacterial gastroenteritis. We have applied both molecular techniques to characterize SRSVs in fecal specimens and serologic assays using four different expressed SRSV antigens to examine the distribution of outbreak strains in the United States and determine if the immune responses of patients were strain specific. Strains from 23 outbreaks of SRSV gastroenteritis were characterized by reverse transcription‐PCR and nucleotide sequencing of a 277‐base region of the capsid gene. These strains segregated into two distinct genogroups, I and II, comprising four and six clusters of strains respectively, each representing a distinct phylogenetic lineage. Serum IgG responses in patients were measured by enzyme immunoassay using expressed capsid antigens of Norwalk virus (NV), Toronto virus (TV), Hawaii virus (HV), and Lordsdale virus (LV), representing four of the 10 clusters. While strains in genogroups I and II were antigenically distinct, within genogroups, the specificity of the immune response varied greatly. Patients infected with genogroup I strains which had as much as 38.5% aa divergence from NV demonstrated relatively homologous seroresponses to the single NV antigen. In contrast, in genogroup II, homologous seroresponses to TV and HV were only present when the infecting strains showed less than 6.5% aa divergence from these antigens. These results suggest that TV and HV represent not only separate genetic clusters in genogroup II but also separate antigenic groups, each of which is related but distinguishable. In addition, two genetically distinct SRSV strains were identified for which we have no homologous antigen. This study suggests that while current molecular diagnostics are capable of detecting the full range of SRSVs, additional expressed antigens will be required to detect an immune response to SRSV infection caused by all the antigenically diverse strains. J. Med. Virol. 53:372–383, 1997.

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.

Inter- and Intragenus Structural Variations in Caliciviruses and Their Functional Implications

ABSTRACT The family Caliciviridae is divided into four genera and consists of single-stranded RNA viruses with hosts ranging from humans to a wide variety of animals. Human caliciviruses are the major cause of outbreaks of acute nonbacterial gastroenteritis, whereas animal caliciviruses cause various host-dependent illnesses with a documented potential for zoonoses. To investigate inter- and intragenus structural variations and to provide a better understanding of the structural basis of host specificity and strain diversity, we performed structural studies of the recombinant capsid of Grimsby virus, the recombinant capsid of Parkville virus, and San Miguel sea lion virus serotype 4 (SMSV4), which are representative of the genera Norovirus (genogroup 2), Sapovirus, and Vesivirus, respectively. A comparative analysis of these structures was performed with that of the recombinant capsid of Norwalk virus, a prototype member of Norovirus genogroup 1. Although these capsids share a common architectural framework of 90 dimers of the capsid protein arranged on a T=3 icosahedral lattice with a modular domain organization of the subunit consisting of a shell (S) domain and a protrusion (P) domain, they exhibit distinct differences. The distally located P2 subdomain of P shows the most prominent differences both in shape and in size, in accordance with the observed sequence variability. Another major difference is in the relative orientation between the S and P domains, particularly between those of noroviruses and other caliciviruses. Despite being a human pathogen, the Parkville virus capsid shows more structural similarity to SMSV4, an animal calicivirus, suggesting a closer relationship between sapoviruses and animal caliciviruses. These comparative structural studies of caliciviruses provide a functional rationale for the unique modular domain organization of the capsid protein with an embedded flexibility reminiscent of an antibody structure. The highly conserved S domain functions to provide an icosahedral scaffold; the hypervariable P2 subdomain may function as a replaceable module to confer host specificity and strain diversity; and the P1 subdomain, located between S and P2, provides additional fine-tuning to position the P2 subdomain.

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. J. Med. Virol. 52:173–178, 1997.

A Prospective Case-Control Study of the Role of Astrovirus in Acute Diarrhea among Hospitalized Young Children

This study examines the importance of astroviruses as a cause of acute diarrhea in hospitalized children <10 years old during a 5-year period. Stools were screened by electron microscopy and were tested for astrovirus, rotavirus, and enteric adenovirus by EIA. During the study, 14.6% of hospitalized children had diarrhea. Astroviruses were second only to rotaviruses as etiologic agents of both community-acquired and nosocomial diarrhea. Community-acquired astrovirus infection occurred in 6.8% of patients, and nosocomial disease occurred in 16.2%. Most cases occurred from March through June, and astrovirus type 1 was the most common. The symptoms of astrovirus-infected children were similar to those of children with rotavirus infection. However, astrovirus-infected children had a lower median age, less dehydration, and lower symptom severity scores and were less likely to have been admitted for gastroenteritis than were children with rotavirus. Astrovirus, for which only rehydration therapy is required, should be considered as another common diarrheal pathogen in children <2 years old.

Characterization of Toronto virus capsid protein expressed in baculovirus

SummaryToronto 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 ∼ 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.

A viral gastroenteritis outbreak associated with person-to-person spread among hospital staff.

OBJECTIVE To identify the etiologic agent and risk factors associated with a hospital ward outbreak of gastroenteritis. SETTING A regional referral hospital in upstate South Carolina. METHODS We reviewed patient charts, surveyed staff, and tested stool from acutely ill persons. A case was defined as diarrhea and vomiting in a staff member or patient from January 5 to 13, 1996. RESULTS The initial case occurred on January 5 in a staff nurse who subsequently was hospitalized on the ward and visited by many staff colleagues. The staff were at a significantly greater risk for gastroenteritis than were patients (28/89 [31%] vs 10/91 [11%]; relative risk [RR], 2.9; 95% confidence interval [CI95], 1.5-5.5). All 10 case-patients had been exposed to case-nurses (assigned nurses who were primary caretakers), and eight had documented exposure to case-nurses 1 to 2 days before their illness. Patients exposed to case-nurses had a significantly increased risk of illness (8/57 [14%] vs 0/32; RR, >4.5; CI95, undefined). Neither staff nor patients had significantly increased risk from food, water, ice, or exposure to case-patients. Electron microscopy identified small round-structured viruses (SRSVs) in nine of nine stool samples. CONCLUSION This nosocomial outbreak of gastroenteritis was likely caused by SRSVs introduced by a staff member and spread via person-to-person transmission from and among staff. The potential for spread of SRSV-associated gastroenteritis from and among staff should be considered in developing strategies to prevent similar outbreaks in hospital settings.