Serotype G1 Research Articles

Sustained Decline in Cases of Rotavirus Gastroenteritis Presenting to the Children's Hospital of Philadelphia in the New Rotavirus Vaccine Era

A dramatic diminution in the number of rotavirus gastroenteritis cases during the 2007 to 2008 rotavirus season in the United States was likely attributable to the availability of an effective rotavirus vaccine for infants since February 2006. To exclude the possibility that factors other than vaccination accounted for the declining case frequency, we examined the 2008 to 2009 experience at the Children's Hospital of Philadelphia (CHOP). Infants with acute gastroenteritis presenting to CHOP have been monitored for the presence of rotavirus antigen in the stool by enzyme-linked immunosorbent assay (followed by serotyping if enzyme-linked immunosorbent assay-positive) since the 1994 to 1995 epidemic season. The number of community-acquired cases during the last full rotavirus season before licensure of a vaccine was 271 in 2005 to 2006, followed by 167 cases in 2006 to 2007 and 36 in 2007 to 2008. Between 2008 and 2009, 73 community-acquired cases were identified. Almost half of the cases were seen among children older than 2 years. Unlike the late-appearing 2007 to 2008 season, the 2008 to 2009 season paralleled the typical time course observed in the prevaccine era. G9P[8] strains caused 64% of the cases. The sustained decline in the frequency of community-acquired rotavirus gastroenteritis has likely resulted from the use of the new rotavirus vaccines. The age distribution of children hospitalized for rotavirus gastroenteritis has shifted toward older children with the introduction of effective vaccines. The G9 serotype (not included in either vaccine) emerged as the most common cause of rotavirus gastroenteritis at CHOP during the 2008 to 2009 season.

Molecular and biological characterization of the 5 human-bovine rotavirus (WC3)-based reassortant strains of the pentavalent rotavirus vaccine, RotaTeq®

RotaTeq is a pentavalent rotavirus vaccine that contains five human-bovine reassortant strains (designated G1, G2, G3, G4, and P1) on the backbone of the naturally attenuated tissue culture-adapted parental bovine rotavirus (BRV) strain WC3. The viral genomes of each of the reassortant strains were completely sequenced and compared pairwise and phylogenetically among each other and to human rotavirus (HRV) and BRV reference strains. Reassortants G1, G2, G3, and G4 contained the VP7 gene from their corresponding HRV parent strains, while reassortants G1 and G2 also contained the VP3 gene (genotype M1) from the HRV parent strain. The P1 reassortant contained the VP4 gene from the HRV parent strain and all the other gene segments from the BRV WC3 strain. The human VP7s had a high level of overall amino acid identity (G1: 95-99%, G2: 94-99% G3: 96-100%, G4: 93-99%) when compared to those of representative rotavirus strains of their corresponding G serotypes. The VP4 of the P1 reassortant had a high identity (92-97%) with those of serotype P1A[8] HRV reference strains, while the BRV VP7 showed identities ranging from 91% to 94% to those of serotype G6 HRV strains. Sequence analyses of the BRV or HRV genes confirmed that the fundamental structure of the proteins in the vaccine was similar to those of the HRV and BRV references strains. Sequences analyses showed that RotaTeq exhibited a high degree of genetic stability as no mutations were identified in the material of each reassortant, which undergoes two rounds of replication cycles in cell culture during the manufacturing process, when compared to the final material used to fill the dosing tubes. The infectivity of each of the reassortant strains of RotaTeq, like HRV strains, did not require the presence of sialic acid residues on the cell surface. The molecular and biologic characterization of RotaTeq adds to the significant body of clinical data supporting the consistent efficacy, immunogenicity, and safety of RotaTeq.

Biophysical characterization of rotavirus serotypes G1, G3, and G4

The stability of attenuated virus vaccines has traditionally been assessed by a plaque assay to measure the virus’s loss of replication competency in response to a variety of environmental perturbations. Although this method provides information regarding the impact of the vaccine formulation, it involves an empirical approach to evaluate stability. Biophysical studies on the other hand have the potential to provide insight into the mechanisms of inactivation of a viral vaccine in response to a variety of stressed conditions. Herein, we have employed a variety of spectroscopic techniques (i.e., circular dichroism, fluorescence spectroscopy, and dynamic light scattering) for a comprehensive examination of the thermal stability of three live-attenuated human-bovine reassortant rotavirus strains (G1, G3, and G4) in the 5-8 pH range. The spectroscopic methods employed are not specific and response changes reflect an average change over the entire virus structure. The present work, however, suggests the utility of these methods in early formulation of rotaviral vaccines due to their ability to identify regions of marginal stability over which high throughput excipient screening assays can be designed. We have further shown that these methods are sufficiently sensitive to differentiate the stability of the three homologous G-subtypes differing only in the composition of their surface antigenic proteins. The data from these spectroscopic methods are also compared to biological activity using a tissue culture viral infectivity assay. Partial correlation between the structural alterations and losses in activity are observed, further suggesting the utility of biophysical studies in early formulation studies of rotavirus vaccines.

Reanálise da vacina tetravalente (RRV-TV) no contexto da prevenção das gastrenterites por rotavírus

Rotaviruses are considered the leading cause of severe gastroenteritis in children under five years of age, especially in developing countries. Vaccination in the first months is the most effective public health action for the control and prevention of infections by such agents. Despite the recent licensing of two vaccines for use in infants (Rotarix® and RotaTeq®), researchers continue to seek new alternatives for prevention and treatment. Herein, we provide a review of the RhesusHuman Reassortant Rotavirus Tetravalent Vaccine (RRV-TV), with an emphasis on its clinical efficacy as regards clinical parameters, the most prevalent serotypes in the region, the occurrence of severe adverse events (e.g., intussusception), and selective protection in the most severe cases. The clinical and epidemiological data were obtained from medical records pertaining to 91 episodes of diarrhea among children in a previous investigation conducted in Belem, Para State, Brazil. Clinical patterns and a scoring system commonly used in studies on the efficacy of RRV-TV were considered as indicators of severity. The most impressive results of this study, such as a significant protection (p 14 related to serotype G2, a 75% efficacy rate against severe episodes, and the non-occurrence of intussusception, are discussed in the context of current knowledge on this issue.

Analysis on epidemiologic characteristics of viral diarrhea among infants in Taiyuan, Shanxi province,2007-2008

To study the epidemiologic characteristics of virus-induced acute diarrhea in children under 5 years old in Taiyuan, Shanxi province. Stool specimens and clinical data were collected from 346 inpatients with acute diarrhea from children less than 5 years old. Rotavirus-positive specimens were identified by ELASA kit. Calicivirus and astrovirus were detected by reverse transcription-polymerase chain reaction (RT-PCR). Adenovirus was done by polymerase chain reaction (PCR). Of the 346 specimens, the percentage of samples with Rotavirus, Calicivirus, Astrovirus, and Adenovirus was 40.8%, 7.5%, 6.4% and 3.2%. Among 141 rotavirus positive samples, serotype G1 (42.6%) was the predominant strain. More than 95% of viral diarrhea patients under hospitalization occurred among children younger than 2 years. Rotavirus is the major pathogen contributing to the acute diarrhea. The disease generally peaks at autumn/winter. The predominant rotavirus strain circulated was G1P[8].

The burden of hospitalised rotavirus infections in Fiji

Rotavirus is the most common cause of acute severe dehydrating diarrhoea in young children worldwide. We describe the burden of rotavirus disease and the rotavirus types causing it in the largest city in Fiji. During 2006 and 2007, 592 children under 5 years of age were admitted to hospital in Suva, Fiji with acute diarrhoea. Of the 454 children for whom a stool specimen was tested, 39% were positive for rotavirus and the predominant strain found was the serotype G3[P8]. There is a significant burden of disease due to rotavirus in Fiji and the introduction of rotavirus vaccines into the national immunization schedule may drastically reduce inpatient diarrhoeal disease.

Surveillance of rotavirus in a rural diarrhoea treatment centre in Bangladesh, 2000–2006

Rotavirus was detected in 33% of 4519 children less than 5 years of age admitted with diarrhoea to treatment centres at Matlab in rural Bangladesh from 2000 to 2006. Highest rotavirus detection rates were in children aged 6–11 months with 56% being less than 1 year old. The peak seasonal detection was in July–September and December–February. The population-based incidence rates of rotavirus ranged from 10.8 to 19.6/1000 children less than 5 years of age. G1 serotype predominated between June 2002–May 2005 and June 2005–May 2006 the predominant type was G2 (41%) followed by G1 (22%) and G9 (22%). Rotavirus is an important cause of childhood diarrhoea in rural Bangladesh and this burden may be reduced with a rotavirus vaccination programme.

Prevalence of diarrhea viruses in hospitalized children in Hong Kong in 2008

A total of 209 stool samples were collected from pediatric patients admitted for acute gastroenteritis in a hospital in Hong Kong, during an 8-month period from January to August 2008, and were tested for the presence of rotavirus, norovirus, sapovirus, adenovirus, and astrovirus using a multiplex RT-PCR assay. The most common virus was rotavirus group A (59 of 209, 28%, mainly serotypes G1, G2, G3, and G9), followed by norovirus group II (48 of 209, 23%), adenovirus (7 of 209, 3%, serotypes 2, 3, and 41), and sapovirus (2 of 209, 1%). Interestingly, none of the specimens in this study were positive for astrovirus. One sample was found to have a dual infection with both norovirus group II and adenovirus. The results support the importance of norovirus as a causative agent of diarrhea in children, which may be underestimated by the current routine diagnostic testing.

Molecular and epidemiological trend of rotavirus infection among infants and children in Japan

This study investigated the prevalence of group A, B, and C rotavirus (RAV, RBV, RCV) and adenovirus (AdV) infections in infants and children with acute gastroenteritis in Japan from July 2006 to June 2007. A total of 628 fecal specimens collected from infants and children with acute gastroenteritis in five different places (Maizuru, Tokyo, Sapporo, Saga and Osaka) in Japan during the period of July 2006–June 2007 were examined for RAV, RBV, RCV and AdV by RT-multiplex PCR. RAV was further characterized to G-typing and P-typing by RT-multiplex PCR and sequencing method. It was found that 123 (19.6%) fecal specimens were positive for RAV followed by AdV of 4.5%. RBV and RCV could not be detected in this study. Serotype G1 (58.5%) was identified at high levels followed by G9 (20.3%), G2 (11.4%), and G3 (7.3%). P genotyping revealed P[8] as predominant (84.6%) followed by P[4] (13.8%) and P[6] (1.6%). During the 2006/2007 rotavirus season, G1P[8] strains were most common with G9P[8], G2P[4], G3P[8], G1P[4] and G9P[6] also detected. It is clear from this study that RAV is still the cause of diseases in Japan. To our knowledge, this is the first report of RV P[6] strain in humans in Japan.

Molecular analysis of G3 rotavirus among infants and children in Dhaka City, Bangladesh after 1993

Between 2004 and 2005, 917 fecal specimens were collected from children below age 5 who presented to Child Health Institute for treatment of diarrhoea in Dhaka City, Bangladesh. The specimens were screened by RT-PCR for the presence of group A rotavirus and positive stools genotyped. Group A rotavirus was detected in 307 stools and serotype G3P[8] strains were detected in nine specimens. Sequence analysis clustered the G3 strains into one distinct lineage (lineage I) with other Asian G3 strains. In addition, one amino acid change at position 96 in antigenic region A, similar to lineage II G3 Chinese strains, was noted. To our knowledge this is the first report of serotype G3 strains in Bangladesh since 1993 and the first report of the molecular characterization of these strains.

RotaTeq™: a three-dose oral pentavalent reassortant rotavirus vaccine

Rotavirus (RV) is a leading cause of diarrhea worldwide, and will infect every child by their 5th birthday. The 3-dose oral pentavalent RV vaccine (RotaTeq™), consisting of serotypes G1, G2, G3, G4 and P1A[8], was licensed in the USA in 2006. The first dose should be administered between the ages of 6 and 12 weeks and the third dose should be given before the age of 32 weeks (USA) or 26 weeks (Europe). In a large clinical trial, the vaccine prevented 74% of cases of RV gastroenteritis, and 98% of cases of severe RV gastroenteritis (based on a symptomatology scoring scale) in the first rotavirus season. Vaccine efficacy in the second season was 62.6% against any RV gastroenteritis and 88% against severe disease. Vaccine efficacy in reducing hospitalizations and emergency department visits was 94.5% up to 2 years after vaccination. Results from a large clinical trial, as well as ongoing postmarketing surveillance, indicate that the vaccine is safe and well tolerated, without evidence of increased risk of intussusception. Further research is needed to establish vaccine efficacy in developing countries, particularly in Africa and Asia. Dissemination efforts should focus on ensuring the vaccine reaches the poorest children in whom disease morbidity and mortality is the highest.

Changing distribution of human rotavirus serotypes during two epidemic outbreaks of gastroenteritis in Campinas, São Paulo, Brazil, 2003–2004: Detection of G6 strains

Background Rotavirus serotypes G1–G4 and G9 are the most important agents of severe diarrhea in children worldwide. Objective To characterize rotavirus serotypes/genotypes causing two large outbreaks of diarrhea in Campinas, São Paulo, during 2003–2004. Study Rotavirus infection was investigated in 328 stool specimens collected from children and adults with diarrhea by PAGE and RT-PCR and further characterized by semi-nested PCR-typing assays. Results G3P[8] (26.1%), G9P[8] (18.7%) and G1P[8] (17.9%) were the most frequently detected serotypes/genotypes. G1P[8] was predominant in 2003, but significantly decreased the following year when G3P[8] and G9P[8] prevailed. G5P[8] was identified in about 9% of the typed specimens from each year consistent with its endemic nature in Brazil for over two decades. The other globally common serotypes (G4P[8] and G2P[4]), uncommon G-P combinations, and multiple G serotypes were also found. Rarely found in humans, and not previously reported in Brazil, serotype G6 was identified in three specimens obtained from children in 2004. Conclusion Multiple rotavirus serotypes were observed co-circulating in the city with serotype predominance changing between the two-year study. This study provides pre-vaccine baseline information on locally endemic strains that might help analysis of post-vaccine data.

Genomic characterization of a novel group A lamb rotavirus isolated in Zaragoza, Spain

An ovine rotavirus (OVR) strain, 762, was isolated from a 30-day-old lamb affected with severe gastroenteritis, in Zaragoza, Spain, and the VP4, VP7, VP6, NSP4, and NSP5/NSP6 genes were subsequently characterized molecularly. Strain OVR762 was classified as a P[14] rotavirus, as the VP4 and VP8* trypsin-cleavage product of the VP4 protein revealed the highest amino acid (aa) identity (94% and 97%, respectively) with that of the P11[14] human rotavirus (HRV) strain PA169, isolated in Italy. Analysis of the VP7 gene product revealed that OVR762 possessed G8 serotype specificity, a type common in ruminants, with the highest degree of aa identity (95-98%) shared with serotype G8 HRV, bovine rotavirus, and guanaco (Lama guanicoe) rotavirus strains. Moreover, strain OVR762 displayed a bovine-like NSP4 (genotype E2) and NSP5/NSP6 (genotype H3), and a VP6 genotype I2, as well as a long electropherotype pattern. This is the first report of a lamb rotavirus with P[14] and G8 specificities, providing additional evidence for the wide genetic and antigenic diversity of group A rotaviruses.

Acrylamide concentration affects the relative position of VP7 gene of serotype G2 strains as determined by polyacrylamide gel electrophoresis

Background During the course of development and characterization of various rotavirus reassortants, we found that the relative position of the gene encoding neutralization and protective antigen VP7 of certain rotavirus strains in a PAGE gel was influenced by the concentration of acrylamide. Objectives We investigated systematically various factors that affected the relative position of the rotavirus VP7 gene in a PAGE gel. Study design We analyzed dsRNAs of selected rotavirus strains bearing G1, G2, G3 or G9 specificity by PAGE at varying concentrations of acrylamide. Results We demonstrated that the relative position of the VP7 gene of three G2 strains varied depending upon the concentration of acrylamide in a PAGE gel, which occurred not only in a homologous G2 virus gene background but also in a heterologous G3 virus gene background; and the VP7 gene bearing G1, G3, G4 or G9 specificity did not display this phenomenon when the PAGE running conditions were varied. Conclusions The concentration of acrylamide in a PAGE gel was the major factor that influenced the relative position of the VP7 gene of G2 rotavirus strains (i.e., VP7 gene coding assignment by PAGE).

Molecular characterization of unusual human G5P[6] rotaviruses identified in China

Background We found an unusual human rotavirus, LL36755 of G5P[6] genotype, in a stool sample collected in Lulong County, Hebei Province, China. This is the first detection of rotavirus serotype G5 in Asia. Objectives To identify and characterize G5 rotaviruses in 988 stool samples collected from children under 5 years old with acute gastroenteritis. Study design We analyzed 459 rotavirus-positive samples with RT-PCR using G5 genotype-specific primers. The G5 strains were sequenced. Results Two additional G5-positive samples (LL3354 and LL4260) were identified. VP7, VP4, VP6 and NSP4 genes of LL3354, LL4260 and LL36755 strains were sequenced. The VP4 sequences formed a group with porcine P[6] strains. The VP6 sequences of strains LL3354 and LL36755 were phylogenetically close to the major clusters of SGI and SGII rotaviruses, respectively. The deduced VP6 protein of strain LL4260 had characteristics of both SGI and SGII strains, but best fit with a cluster of atypical SGI viruses. In addition, based on NSP4 sequences, the three G5 strains belonged to genogroup B and were closest to human strain Wa. Conclusion These results indicate a dynamic interaction of human and porcine rotaviruses and suggest that reassortment could result in the stable introduction and successful spread of porcine gene alleles into human rotaviruses.

Prevalence of enteropathogenic viruses and molecular characterization of group A rotavirus among children with diarrhea in Dar es Salaam Tanzania

BackgroundDifferent groups of viruses have been shown to be responsible for acute diarrhea among children during their first few years of life. Epidemiological knowledge of viral agents is critical for the development of effective preventive measures, including vaccines.MethodsIn this study we determined the prevalence of the four major enteropathogenic viruses – rotavirus, norovirus, adenovirus and astrovirus – was determined in 270 stool samples collected from children aged 0 – 60 months who were admitted with diarrhea in four hospitals in Dar es Salaam, Tanzania, using commercially available ELISA kits. In addition, the molecular epidemiology of group A rotavirus was investigated using reverse transcriptase multiplex polymerase chain reaction (RT-PCR).ResultsAt least one viral agent was detected in 87/270 (32.2%) of the children. The prevalence of rotavirus, norovirus, adenovirus and astrovirus was 18.1%, 13.7%, 2.6% and 0.4%, respectively. In most cases (62.1%) of viruses were detected in children aged 7–12 months. The G and P types (VP7 and VP4 genotypes respectively) were further investigated in 49 rotavirus ELISA positive samples. G9 was the predominant G type (81.6%), followed by G1 (10.2%) and G3 (0.2%). P[8] was the predominant P type (83.7%), followed by P[6] (0.4%) and P[4] (0.2%). The following G and P types were not detected in this study population; G2, G4, G8 G10, P[9], P[10] and P[11]. The dominating G/P combination was G9P[8], accounting for 39 (90.7%) of the 43 fully characterized strains. Three (6.1%) of the 49 rotavirus strains could not be typed.ConclusionNearly one third of children with diarrhea admitted to hospitals in Dar es Salaam had one of the four viral agents. The predominance of rotavirus serotype G9 may have implication for rotavirus vaccination in Tanzania.

Serological and molecular diversity of human rotavirus in São Paulo, Brazil

From a total of 187 fecal samples from children with ages between 0 and 5 years, collected in the Hospital Universitario -USP, Brazil, from 1994 to 1996, 54 (28.9%) were positive for rotavirus. Positive samples were characterized by electropherotyping, subgrouping, G serotype and genotype and P genotype. Rotavirus electropherotypes were characterized in four different long genome patterns (38.9%), one short genome pattern (34.0%) and 18.0% were characterized as an unusual pattern. Subgroup I was found in 38.9% strains, subgroup II in 50.0% and 7.7% was subgroup nonI-nonII. For G serotypes, G2 was found in 59.3%, G1 was identified in 33.3% of strains, two samples showed mixtures of G1+G2 and one sample was G1+G3. Ten samples characterized as serotype G2 showed a long eletropherotype. Genotype G2 was the most frequently and was found in 37 (44.0%) samples (23 samples as a single genotype and 14 as mixtures of genotypes). G1 was found in 15 samples. G3 and G4 was detected mainly in mixtures of genotypes and G5, G6 and G9 were identified only in mixtures. A total of 20 (38.5%) samples were characterized as G genotype mixtures and P mixtures were found in 16 (29.6%) samples. P[4] was found in 55.6% of samples, P[8] in 51.9% and P[6-M37 like] in 22.3% of cases. P[6-Gottfried like] and P[11] were detected only in mixtures. One sample with G6 specificity, mixed with a G2 rotavirus and a P[11] strain, mixed with P[4] and P[8]strain was described for the first time in Latin America.

Diversity in Indian Equine Rotaviruses: Identification of Genotype G10,P6[1] and G1 Strains and a New VP7 Genotype (G16) Strain in Specimens from Diarrheic Foals in India

Rotaviruses causing severe diarrhea in foals in two organized farms in northern India, during the period from 2003 to 2005, were characterized by electropherotyping, serotyping, and sequence analysis of the genes encoding the outer capsid proteins. Of 137 specimens, 47 (34.31%) were positive for rotavirus and exhibited at least five different electropherotypes (E), E1 to E5. Strains belonging to different electropherotypes exhibited either a different serotype/genotype specificity or a lack of reactivity to typing monoclonal antibodies (MAbs) used in this study. Strains belonging to E1, E2, and E5 exhibited genotype G10,P6[1], G3, and G1 specificities and accounted for 19.0, 42.9, and 9.5% of the isolates, respectively. Though they possessed G10-type VP7, the E1 strains exhibited high reactivity with the G6-specific MAb, suggesting that the uncommon combination of the outer capsid proteins altered the specificity of the conformation-dependent antigenic epitopes on VP7. E3 and E4 strains accounted for 28.6% of the isolates and were untypeable. Sequence analysis of VP7 from E4 strains (Erv92 and Erv99) revealed that they represent a new VP7 genotype, G16. The detection of unexpected bovine rotavirus-derived G10,P6[1] reassortants, G1 serotype strains, and a new genotype (G16) strain in two distant farms reveals an interesting epidemiological situation and diversity of equine rotaviruses in India.

Rotavirus G2 Serotypes Are Emerging in Infants

Rotavirus G2 Serotypes Are Emerging in Infants

Concomitant Use of an Oral Live Pentavalent Human-Bovine Reassortant Rotavirus Vaccine With Licensed Parenteral Pediatric Vaccines in the United States

A live pentavalent rotavirus vaccine (PRV) containing 5 human-bovine (WC3) reassortants expressing human serotypes G1, G2, G3, G4 and P1A[8] was evaluated in a blinded, placebo-controlled study. Possible interactions between PRV and concomitantly administered licensed pediatric vaccines were investigated in a United States-based nested substudy (Concomitant Use Study) of the Rotavirus Efficacy and Safety Trial. From 2002 to 2003, healthy infants approximately 6 to 12 weeks of age at entry were randomized to receive either 3 oral doses of PRV or placebo at 4- to 10-week intervals. Subjects were also to receive combined Haemophilus influenzae type b and hepatitis B vaccine (2 doses), diphtheria and tetanus toxoids and acellular pertussis vaccine (3 doses), inactivated poliovirus vaccine (2 doses) and pneumococcal conjugate vaccine (3 doses) on the same day; oral poliovirus vaccine was not administered. Immunogenicity was assessed by measuring antibody responses to PRV and antigens contained in the licensed vaccines. Cases of rotavirus gastroenteritis were defined by forceful vomiting and/or -3 watery or looser-than-normal stools within a 24-hour period, and detection of rotavirus antigen in the stool. Safety was assessed by reporting of adverse events using diary cards. The Concomitant Use Study enrolled 662 subjects in the PRV group and 696 subjects in the placebo group. For the 17 antigens in the concomitantly administered vaccines, antibody responses were similar in PRV and placebo recipients, except for moderately diminished antibody responses to the pertactin component of pertussis vaccine. Efficacy of PRV against rotavirus gastroenteritis of any severity was 89.5% (95% CI = 26.5-99.8%). PRV was generally well tolerated when given concomitantly with the prespecified vaccines. In this study, antibody responses to the concomitantly administered vaccines were generally similar in PRV and placebo recipients. PRV was efficacious and well tolerated when given concomitantly with pediatric vaccines licensed in the United States.