Human group C rotavirus: completion of the genome sequence and gene coding assignments of a non-cultivatable rotavirus.

Genome segments 1 and 2 of human group C rotavirus 'Bristol' strain were sequenced and their gene-protein coding properties assigned. This work completed the genome sequence of a human group C rotavirus (17,910 bp) and allowed the full gene-protein coding assignment of the 11 segments of dsRNA. Gene 1 is 3309 bp in size and contains a single ORF of 3272 nucleotides, encoding a protein of 1090 amino acids in length with a predicted molecular mass of 125 kDa. Comparison of the translated sequence with cognate published mammalian group A, B and C rotavirus sequences showed 45.2, 26.4 and 92.6% identity, respectively. The sequence contains conserved amino acid motifs including the classic RNA-dependent RNA polymerase motif GDD, indicating that segment 1 encodes the group C rotavirus polymerase protein. Gene 2 is 2736 bp in size and contains a single ORF of 2655 nucleotides encoding a protein of 884 amino acids in length with a calculated molecular mass of 102 kDa. Database searches showed highest homology with VP2, the main structural component of the 'core' from group A rotaviruses (46% identity). Alignment of the human group C and A rotavirus VP2 proteins revealed several characteristics common to nucleic acid binding proteins. However, these features were not shared with group B rotavirus VP2.

Outbreak of hepatitis A in the injecting drug user and homeless populations in Bristol: control by a targeted vaccination programme and possible parenteral transmission.

OBJECTIVE: To study the use of hepatitis A virus (HAV) vaccination in controlling an outbreak of HAV in inner-city Bristol among injecting drug users (IDUs). To study whether hepatitis C virus (HCV) and hepatitis B virus (HBV) co-infection increases morbidity. DESIGN: Community-based cohort study. SETTING: Avon Health Authority area, UK. PARTICIPANTS: All laboratory-confirmed cases of HAV infection notified in 2000. INTERVENTION: Administration of a targeted vaccination, education and liaison programme. MAIN OUTCOME MEASURES: Number of cases of HAV before and after introduction of HAV vaccination programme. Mortality and number of patients requiring hospital admission. Association of HCV and HBV co-infection with hospital admission. RESULTS: Ninety cases of HAV were reported in the first 6 months of 2000, of whom a substantial number were IDUs and/or inner-city hostel residents. In the second 6 months of 2000, following the introduction of a vaccination programme among homeless people, hostel residents, and IDUs, the number of HAV cases fell to 33. Sixteen patients had evidence of HCV co-infection. No patient had chronic HBV infection. Two patients died as a result of HAV, and two subsequently died from drug misuse. Fifty-six per cent of HCV-co-infected patients required admission to hospital compared with 28% non-HCV-co-infected patients. CONCLUSIONS: This is the first reported successful use of vaccination to control an outbreak of HAV in a population of IDUs and to prevent transmission to the wider population. HCV co-infection appears to increase the severity of HAV illness, as demonstrated by increased incidence of hospital admission.

Vulvovaginal-swab or first-catch urine specimen to detect Chlamydia trachomatis in women in a community setting?

Screening for chlamydia in women is widely recommended. We evaluated the performance of two nucleic acid amplification tests for detecting Chlamydia trachomatis in self-collected vulvovaginal-swab and first-catch urine specimens from women in a community setting and a strategy for optimizing the sensitivity of an amplified enzyme immunoassay on vulvovaginal-swab specimens. We tested 2,745 paired vulvovaginal-swab and urine specimens by PCR (Roche Cobas) or strand displacement amplification (SDA; Becton Dickinson). There were 146 women infected with chlamydia. The assays detected 97.3% (95% confidence interval [CI], 93.1 to 99.2%) of infected patients with vulvovaginal-swab specimens and 91.8% (86.1 to 95.7%) with urine specimens. We tested 2,749 vulvovaginal-swab specimens with both a nucleic acid amplification test and a polymer conjugate-enhanced enzyme immunoassay with negative-gray-zone testing. The relative sensitivities obtained after retesting specimens in the negative gray zone were 74.3% (95% CI, 62.8 to 83.8%) with PCR and 58.3% (95% CI, 46.1 to 69.8%) with SDA. In community settings, both vulvovaginal-swab and first-catch urine specimens from women are suitable substrates for nucleic acid amplification tests, but enzyme immunoassays, even after negative-gray-zone testing, should not be used in screening programs.

Epidemiology of human Sapporo-like caliciviruses in the South West of England: molecular characterisation of a genetically distinct isolate.

Human enteric caliciviruses have been assigned to two distinct genera: the Norwalk-like viruses (NLVs) and the Sapporo-like viruses (SLVs). During a 3-year surveillance of gastroenteritis in the South West of England during November 1997-2000, a total of 27 clinical samples containing SLVs were collected. PCR amplicons covering a region of the RNA polymerase gene were obtained from 18 of the SLV samples. Sequence analysis of the PCR products indicated that the SLV isolates could be assigned to one of the two major genetic groups represented by Sapporo and London/92 caliciviruses. One of these isolates belonging to the London/92 group (Bristol/98) was subjected to a complete genome sequence analysis. The full genomic sequence of the Bristol/98 isolate was determined from RNA extracted from a single stool sample and consists of 7490 nucleotides, excluding the poly(A) tail. The genome is organised into two open reading frames (ORFs), similar to that of Manchester SLV although the small ORF overlapping the region encoding the capsid protein observed in Manchester SLV is absent in Bristol/98 SLV. The polyprotein (ORF1) of Bristol/98 SLV consists of 2,280 amino acids and, as observed in all SLVs, the structural protein is encoded in frame and contiguous with the 3' terminus of the ORF1. Phylogenetic studies based on complete capsid sequences and genome arrangements within the SLVs indicate that the human enteric viruses within the "Sapporo-like" virus clade should be divided into two distinct genetic groups analogous to the assignment of the Norwalk-like viruses.

Surveillance of norovirus infection in a study of sporadic childhood gastroenteritis in South West England and South Wales, during one winter season (1999-2000).

Reverse transcriptase polymerase chain reaction (RT-PCR), electron microscopy (EM) and a genotype II specific antigen capture enzyme immunoassay (EIA), (Lordsdale strain) were used to establish the prevalence of Norwalk-like viruses (NLV) among sporadic cases of childhood gastroenteritis in South West England over a winter season. Samples of 3,172 stools from cases of gastroenteritis in children aged under 7 years sent to the Bristol Public Health Laboratory over the 1999/2000 winter 'season' were tested prospectively by EM, EIA and RT-PCR. The results from sporadic cases were compared with 1,360 samples from 285 outbreaks of gastroenteritis which were sent to the laboratory over the same period. In total NLV was established as the causal agent in 326 cases (10.3%) of sporadic gastroenteritis by one or more of the tests (EM 30 (0.9%), EIA 132 (4.2%) and RT-PCR 276 (8.7%)). The presence of other enteric viruses was established using EM and rotavirus EIA. Rotaviruses were the most common cause of viral gastroenteritis with 684 cases (21.6%). Other viruses detected included, adenovirus 124 cases (3.9%), astrovirus 97 cases (3.1%) and calicivirus in 7 cases (0.2%). NLV was the second most common viral agent indicating a significant role in cases of sporadic childhood gastroenteritis.

Molecular characterization of human group C rotavirus genes 6, 7 and 9.

Genes 6, 7 and 9 of human group C rotavirus 'Bristol' strain, encoding non-structural proteins (NSP) 3, 1 and 2, respectively, were cloned and sequenced. Human group C rotavirus genome segment 6 is 1350 bp and contains a single ORF of 1231 nucleotides (encoding 402 amino acids). Genome segment 7 is 1270 bp and encodes a protein of 394 amino acids and genome segment 9 is 1037 bp and encodes a 312 amino acid protein. The human group C rotavirus genes 6, 7 and 9 showed 78, 67 and 88% sequence identity, respectively, to the corresponding porcine group C rotavirus genes. The derived protein sequences were compared with those of the porcine 'Cowden' group C and mammalian group A rotavirus strains. The human group C rotavirus NSP1 protein sequence is one amino acid longer than the porcine group C equivalent. In common with group A and porcine group C rotaviruses, the human group C rotavirus NSP1 protein has a zinc finger motif. Human group C rotavirus NSP2 has two hydrophobic heptad repeat regions, a basic, RNA-binding domain and a basic, proline-rich region. Human group C rotavirus NSP3 has both single- and double-stranded RNA-binding domains and several hydrophobic heptad repeat regions, one of which forms a leucine zipper. This work completes the molecular characterization of the non-structural proteins of a human group C rotavirus. Phylogenetic analysis of all the non-structural genes of group A, B and C rotaviruses suggests that these viruses have diverged at a constant rate from a common ancestor.

Tubal damage in infertile women: prediction using chlamydia serology.

BACKGROUND: The study explores the relationship between serum chlamydia antibody titres (CATs) and detection of tubal damage in infertile women. METHODS: The tubal status and pelvic findings in 1006 women undergoing laparoscopy for infertility were related to CAT, which was measured using the whole-cell inclusion immunofluorescence test. RESULTS: A negative correlation between CAT and age was noted. A linear trend between serum CAT and the likelihood of tubal damage, including severe damage, was observed (P < 0.001). Titres in women with tubal damage (median 1:1024; range <1:64-1:4096) were significantly (P < 0.001) higher than in women with endometriosis alone (median <1:64; range <1:64-1:512) or those with a normal pelvis (median <1:64; range <1:64-1:1024). Women with positive titres were more likely to have pelvic adhesions than tubal occlusion unless titres were very high, when tubal damage was likely to be more severe. CONCLUSIONS: CATs are of predictive value in the detection of tubal damage and are quantitatively related to the severity of damage. For practical clinical purposes, Chlamydia serology is useful mainly as a screening test for the likelihood of tubal damage in infertile women and may facilitate decisions on which women should proceed with further investigations without delay.