Nigel A. Cunliffe

Effect of Human Rotavirus Vaccine on Severe Diarrhea in African Infants

BACKGROUND Rotavirus is the most common cause of severe gastroenteritis among young children worldwide. Data are needed to assess the efficacy of the rotavirus vaccine in African children. METHODS We conducted a randomized, placebo-controlled, multicenter trial in South Africa (3166 infants; 64.1% of the total) and Malawi (1773 infants; 35.9% of the total) to evaluate the efficacy of a live, oral rotavirus vaccine in preventing severe rotavirus gastroenteritis. Healthy infants were randomly assigned in a 1:1:1 ratio to receive two doses of vaccine (in addition to one dose of placebo) or three doses of vaccine--the pooled vaccine group--or three doses of placebo at 6, 10, and 14 weeks of age. Episodes of gastroenteritis caused by wild-type rotavirus during the first year of life were assessed through active follow-up surveillance and were graded with the use of the Vesikari scale. RESULTS A total of 4939 infants were enrolled and randomly assigned to one of the three groups; 1647 infants received two doses of the vaccine, 1651 infants received three doses of the vaccine, and 1641 received placebo. Of the 4417 infants included in the per-protocol efficacy analysis, severe rotavirus gastroenteritis occurred in 4.9% of the infants in the placebo group and in 1.9% of those in the pooled vaccine group (vaccine efficacy, 61.2%; 95% confidence interval, 44.0 to 73.2). Vaccine efficacy was lower in Malawi than in South Africa (49.4% vs. 76.9%); however, the number of episodes of severe rotavirus gastroenteritis that were prevented was greater in Malawi than in South Africa (6.7 vs. 4.2 cases prevented per 100 infants vaccinated per year). Efficacy against all-cause severe gastroenteritis was 30.2%. At least one serious adverse event was reported in 9.7% of the infants in the pooled vaccine group and in 11.5% of the infants in the placebo group. CONCLUSIONS Human rotavirus vaccine significantly reduced the incidence of severe rotavirus gastroenteritis among African infants during the first year of life. (ClinicalTrials.gov number, NCT00241644.)

Serotype Diversity and Reassortment between Human and Animal Rotavirus Strains: Implications for Rotavirus Vaccine Programs

The development of rotavirus vaccines that are based on heterotypic or serotype-specific immunity has prompted many countries to establish programs to assess the disease burden associated with rotavirus infection and the distribution of rotavirus strains. Strain surveillance helps to determine whether the most prevalent local strains are likely to be covered by the serotype antigens found in current vaccines. After introduction of a vaccine, this surveillance could detect which strains might not be covered by the vaccine. Almost 2 decades ago, studies demonstrated that 4 globally common rotavirus serotypes (G1-G4) represent >90% of the rotavirus strains in circulation. Subsequently, these 4 serotypes were used in the development of reassortant vaccines predicated on serotype-specific immunity. More recently, the application of reverse-transcription polymerase chain reaction genotyping, nucleotide sequencing, and antigenic characterization methods has confirmed the importance of the 4 globally common types, but a much greater strain diversity has also been identified (we now recognize strains with at least 42 P-G combinations). These studies also identified globally (G9) or regionally (G5, G8, and P2A[6]) common serotype antigens not covered by the reassortant vaccines that have undergone efficacy trials. The enormous diversity and capacity of human rotaviruses for change suggest that rotavirus vaccines must provide good heterotypic protection to be optimally effective.

Rotavirus G and P types in children with acute diarrhea in Blantyre, Malawi, from 1997 to 1998: predominance of novel P[6]G8 strains.

One hundred rotavirus strains detected in children with acute diarrhea in Blantyre, Malawi, between July 1997 and January 1998 were characterized for G (VP7) and P (VP4) types by using multiplex, heminested, reverse transcription–polymerase chain reaction. A novel P[6]G8 rotavirus strain was identified in 42% of the specimens. The remaining strains comprised P[8]G3 (20%), P[6]G3 (10%), P[4]G8 (9%), P[6]G9 (3%), P[8]G4 (2%), P[6]G4 (2%), and P[4]G3 (1%). Rotavirus strains with mixed G or P types were identified in 2% of the specimens. Nine percent of the strains were nontypeable with the primers used. The P[6] genotype was identified in 57% of strains overall. This first description of serotype G8 rotavirus as a predominant strain has important implications for vaccine development in Africa. The finding of novel P/G combinations (P[6]G8 and P[4]G8) highlights the extraordinary diversity of rotaviruses in some countries. J. Med. Virol. 57:308–312, 1999.

Effectiveness of monovalent rotavirus vaccine (rotarix) against severe diarrhea caused by serotypically unrelated G2P[4] strains in Brazil.

BACKGROUND. In a Latin American trial, a monovalent G1P[8] rotavirus vaccine showed high efficacy against severe rotavirus diarrhea. Protection was lower against serotypically unrelated G2P[4] strains, which circulated infrequently. This case-control study was undertaken to assess the effectiveness of this monovalent G1P[8] rotavirus vaccine against G2P[4] strains in Brazil. METHODS. Case patients were children with severe G2P[4] rotavirus diarrhea who presented at a hospital in Recife, Brazil, from March 2006 through September 2008. Vaccination rates among case patients were compared with rates among 2 groups of control participants-children with rotavirus-negative diarrhea and children admitted for acute respiratory tract infection (ARI)-to calculate vaccine effectiveness, after controlling for the birth month and year. RESULTS. We enrolled 70 G2P[4] rotavirus-positive case patients with severe diarrhea, 484 rotavirus-negative control participants with diarrhea, and 416 control participants with ARI, aged 6 months. Among children aged 6-11 months, the effectiveness of the vaccine against G2P[4] diarrhea was 77% (95% confidence interval [CI], 42%-91%) and 77% (95% CI, 43%-90%) among the rotavirus-negative control participants with diarrhea and control participants with ARI, respectively. Vaccine effectiveness in children aged 12 months decreased to -24% (95% CI, -190% to 47%) and 15% (95% CI, -101 to 64) among the rotavirus-negative control groups with diarrhea and ARI, respectively. CONCLUSIONS. This monovalent G1P[8] rotavirus vaccine was effective against severe G2P[4] rotavirus diarrhea among children aged 6-11 months. Effectiveness declined among children aged 12 months, which suggests waning immunity.

Molecular epidemiology of noroviruses associated with acute sporadic gastroenteritis in children: global distribution of genogroups, genotypes and GII.4 variants

Noroviruses are a leading cause of epidemic and sporadic acute gastroenteritis worldwide. The development of sensitive molecular diagnostic techniques has revolutionized our understanding of norovirus epidemiology over the past two decades, but norovirus strain types associated with sporadic gastroenteritis remain poorly described. Therefore, we conducted a systematic review of studies performed after 2000 to clarify the genotypic distribution of noroviruses in children (≤18 years of age) with sporadic acute gastroenteritis. Genogroup GII norovirus was the most prevalent, accounting for 96% of all sporadic infections. GII.4 was the most prevalent genotype, accounting for 70% of the capsid genotypes and 60% of the polymerase genotypes, followed by the capsid genotype GII.3 (16%) and the polymerase genotype GII.b (14%). The most common ORF1/ORF2 inter-genotype recombinants were GII.b, GII.12, and GII.4 polymerase genotypes combined with the capsid genotype GII.3, accounting for 19% of all genotyped strains. The distribution of GII.4 variants over the last decade was dominated by successive circulation of GII.4/2002, GII.4/2004, GII.4/2006b, and GII.4/2008 with GII.4/2006b continuing to date. Genotypes GII.4 and GII.3 have predominated in children during the past decade; this is most notable in the global emergence of GII.4 variant noroviruses. As the burden of rotavirus disease decreases following the introduction of childhood immunization programs, the relative importance of norovirus in the etiology of acute childhood gastroenteritis will likely increase. In order for a successful norovirus vaccine to be developed, it should provide immunity against strains with capsid genotypes GII.4 and GII.3.

Predominance of Rotavirus P(4)G2 in a Vaccinated Population, Brazil

We identified 21 rotaviruses in 129 patients with diarrhea in a Brazilian city with high rotavirus vaccine coverage. All rotaviruses were genotype P[4]G2 with 1 mixed infection with P[NT]G9. Although virus predominance could have occurred randomly, the vaccine may be less protective against P[4]G2. Prospective surveillance is urgently needed.

Rotavirus strain diversity in Blantyre, Malawi, from 1997 to 1999.

ABSTRACT In a 2-year study of viral gastroenteritis in children in Blantyre, Malawi, the diversity of rotavirus strains was investigated by using electropherotyping, reverse transcription-PCR amplification of the VP7 and VP4 genes (G and P genotyping), and nucleotide sequencing. Of 414 rotavirus strains characterized, the following strain types were identified: P[8], G1 (n = 111; 26.8%); P[6], G8 (n = 110; 26.6%); P[8], G3 (n = 93; 22.5%); P[4], G8 (n = 31; 7.5%); P[8], G4 (n = 21; 5.1%); P[6], G3 (n = 12; 2.9%); P[6], G1 (n = 7; 1.7%); P[6], G9 (n = 3; 0.7%); P[6], G4 (n = 3; 0.7%); P[4], G3 (n = 1; 0.2%); and mixed (n = 15; 3.6%). While all strains could be assigned a G type, seven strains (1.7%) remained P nontypeable. The majority of serotype G8 strains and all serotype G9 strains had short electropherotype profiles. All remaining typeable strains had long electropherotypes. Divergent serotype G1 rotaviruses, which contained multiple base substitutions in the 9T-1 primer binding site, were commonly identified in the second year of surveillance. Serotype G2 was not identified. Overall, G8 was the most frequently identified VP7 serotype (n = 144; 34.8%) and P[8] was the most frequently detected VP4 genotype (n = 227; 54.8%). Partial sequence analysis of the VP4 gene of genotype P[8] rotaviruses identified three distinct clusters, which predominantly (but not exclusively) comprised strains belonging to a distinct VP7 serotype (G1, G3, or G4). As a result of mutations in the 1T-1 primer binding site, strains belonging to each cluster required a separate primer for efficient typing. One cluster, represented by P[8], G4 strain OP354, was highly divergent from the established Wa and F45 VP4 P[8] lineages. As is the case for some other countries, the diversity of rotaviruses in Malawi implies that rotavirus vaccines in development will need to protect against a wider panel of serotypes than originally envisioned.

Apparent extinction of non-G2 rotavirus strains from circulation in Recife, Brazil, after the introduction of rotavirus vaccine.

The introduction of a G1P[8] rotavirus vaccine in Recife, Brazil, caused a decrease in rotavirus detection from 27% (March–May, 2006) to 5.0% (March–May, 2007), with all strains becoming G2, against which less protection had been predicted.

Molecular Epidemiology of Cryptosporidiosis in Children in Malawi

ABSTRACT: Few studies have examined the molecular epidemiology of cryptosporidiosis in developing countries. In this study, DNA of 69 microscopy‐positive human fecal samples collected from Malawi were examined by multilocus genetic analyses. From 43, 27 and 28 of the samples, the SSU rRNA, 70 kDa heat shock protein (HSP70) and 60 kDa glycoprotein (GP60) genes, respectively, were successfully PCR‐amplified. Restriction analysis of the SSU PCR products showed that 41 of the 43 PCR‐positive samples had C. hominis and 2 had C. parvum. Sequence analysis of the HSP70 and GP60 gene contirmed the species identification by SSU rRNA PCR‐RFLP analysis, but also revealed high intraspecific variations. Altogether, six HSP70 subtypes and six GP60 subtypes (belonging to lour subtype alleles) of C. hominis were found. Linkage diseyuilibrum analysis of the two genetic loci showed possible intraspecitic recombination. Thus, cryptosporidiosis in the study area was largely caused by anthroponotic transmission. The high intraspecitic variation and existence of genetic recombination were probably results of high transmission of cryptosporidiosis in this area.

Molecular Epidemiology of Rotavirus Diarrhea among Children and Adults in Nepal: Detection of G12 Strains with P[6] or P[8] and a G11P[25] Strain

ABSTRACT In anticipation of a rotavirus vaccine in Nepal, this study was undertaken to determine the distribution of the G and P serotypes and electropherotypes of rotaviruses in order to examine if there is any emerging serotype or unusual strain circulating in children and adults in Nepal. Of 1,315 diarrheal stool specimens, rotavirus was detected by an enzyme-linked immunosorbent assay in 116 (17%) of 666 patients less than 5 years of age, in 18 (7%) of 260 patients 5 to 14 years of age, and in 19 (5%) of 358 patients 15 years of age and older. Approximately 75% of rotavirus diarrhea occurred in children less than 5 years of age. Approximately 70% of rotaviruses found in each of the three age groups belonged to serotype G1P[8]. Interestingly, there were 29 (20%) G12 rotaviruses carrying either P[8] or P[6] and one (0.7%) G11 rotavirus carrying an unusual P[25] genotype. RNA polyacrylamide gel electrophoresis discriminated 19 strains (electropherotypes), among which there were three codominant strains carrying G1P[8] and long RNA patterns. Five electropherotypes were discriminated among G12 rotaviruses, all of which had long RNA patterns. The fact that 20% of rotaviruses were G12 strains carrying either P[8] or P[6] and had multiple electropherotypes suggest that G12 strains are not more rare strains but that they pose an emerging challenge to current and future vaccines. The presence of multiple strains as defined by electropherotypes suggests the richness of the rotavirus gene pool in Nepal, where unusual strains may continue to emerge.