carried out in Nha Trang city in Vietnam between December 2005 and June 2006. RV and ... The majority of patients with RV and NoV were children younger than 2 years of ... cause of death among children younger than 5 years of age (1) ...
Jpn. J. Infect. Dis., 63, 405-411, 2010
Original Article
Molecular Epidemiological Study of Rotavirus and Norovirus Infections among Children with Acute Gastroenteritis in Nha Trang, Vietnam, December 2005–June 2006 Tsutomu Tamura1,3*, Makoto Nishikawa1,3, Dang Duc Anh2, and Hiroshi Suzuki3 1Department
of Virology, Niigata Prefectural Institute of Public Health and Environmental Sciences, Niigata 950-2144; 3Department of Public Health, Graduate School of Medical and Dental Sciences, Niigata University, Niigata 951-8510, Japan; and 2National Institute for Hygiene and Epidemiology, Hanoi, Vietnam (Received April 30, 2010. Accepted August 25, 2010) SUMMARY: A molecular epidemiological study of rotavirus (RV) and norovirus (NoV) infections was carried out in Nha Trang city in Vietnam between December 2005 and June 2006. RV and NoV were detected in 87 (47.5z) and 12 (6.6z) of the 183 fecal specimens from children hospitalized with acute gastroenteritis, respectively. The majority of patients with RV and NoV were children younger than 2 years of age. The most frequent RV genotypes detected were G3 (n ＝ 37, 42.5z) and G1 (n ＝ 28, 32.2z) for G type, P[8] (n ＝ 61, 70.1z) for P type, and G3P[8] (n ＝ 33, 38.0z) and G1P[8] (n ＝ 18, 20.7z) for the G and P genotype combination. GII.12 was the most common genotype (6/12, 50z) for NoV, followed by GII.4 (4/12, 33.3z), and we also identified a rare type (GII.19). The results of this study highlight the increased incidence of G3P[8] and the presence of many OP354-like P[8] RVs, as well as the GII.4 2003 Asia variant of NoVs. Furthremore, the first case of GII.19 of NoV in Vietnam is reported. became dominant (4,7). As far as P genotypes are concerned, P[8] is responsible for most RV infections in the world (6). Strain OP354, a rare genotype of P[8] RV, was first detected in Malawi in 1998 (10) and was recently suggested to be subtype P[8]b (8). P[8]b RV has been isolated recently in some Asian countries (8,9,11,12). NoV is the second most common cause of severe childhood gastroenteritis, after RV (13), and its prevalence in children with acute gastroenteritis ranges from 6 to 48z (14). NoV can be classified into five genogroups, GI to GV, and subdivided into many genotypes (15–18). Strains GI, GII, and GIV are associated with human gastroenteritis, whereas GIII and GV are only found in animals. Worldwide outbreak surveillance data show a prominent role for GII strains, especially after the emergence of new GII.4 variants between 1995 and 2007 (14,19). In this study we clarified the etiological role of RV and NoV in sporadic diarrhea and analyzed their genetic diversity in Nha Trang, which is located in southern Vietnam.
INTRODUCTION Diarrheal disease remains the second most common cause of death among children younger than 5 years of age (1), with group A rotavirus (RV) and norovirus (NoV) being the main causative agents of acute gastroenteritis in both developed and developing countries. An estimated 527,000 children younger than 5 years of age die of RV-related diarrhea each year, with more than 85z of these deaths occurring in developing countries in Africa and Asia (2). Vietnam is estimated to account for 5,300–6,800 of such children (3). RV infections in this country occur throughout the year, with a slight peak during the winter dry season in both southern and northern Vietnam (4). RVs have two outer shell proteins, VP4 and VP7, against which protective immunity is mainly targeted. RVs are classified into 28 P and 20 G genotypes on the basis of the VP4 and VP7 gene sequences, respectively (5). Five combinations of G and P genotypes (G1P[8], G2P[4], G3P[8], G4P[8], and G9P[8]) make up 90z of the human RV strains worldwide (6), with the G1 strain currently being the most prevalent G genotype strain (6). The predominant G genotype in Vietnam between July 1998 and June 2000 was G2, although G1 then
MATERIALS AND METHODS Samples: A total of 183 fecal specimens were collected from children with gastroenteritis at hospitals in Nha Trang in Vietnam between December 2005 and June C until further 2006. Stool samples were stored at －209 analysis. RNA extraction and reverse transcription: The fecal specimens were diluted with phosphate-buffered saline pH 7.2 to form a 10z suspensions then centrifuged at 8,000 × g for 20 min. Viral RNA genomes were extract-
*Corresponding author: Mailing address: Department of Virology, Niigata Prefectural Institute of Public Health and Environmental Sciences, 314-1 Sowa, Nishi-Ku, Niigata-Shi, Niigata 950-2144, Japan. Tel: ＋81-25-2639414, Fax: ＋81-25-263-9410, E-mail: tamura.tsutomu＠ pref.niigata.lg.jp 405
Table 1. Primer list for rotavirus and norovirus used in this study Primer
Type
Sequence (5?–3?)
Amplicon
GGC TTT AAA AGA GAG AAT TTC CGT CTG G GGT CAC ATC ATA CAA TTC TAA TCT AAG
1062
20 20
CAA GTA CTC AAA TCA ATG ATG G CAA TGA TAT TAA CAC ATT TTC TGT G CGT TTG AAG AAG TTG CAA CAG CGT TTC TGG TGA GGA GTT G GTC ACA CCA TTT GTA AAT TCG CTA GAT GTA ACT ACA ACT AC GGT CAC ATC ATA CAA TTC T
749 652 374 583 885 306
20 20 20 20 20 20 20
ACG AAC TCA ACA CGA GAG G
813
22
TGG CTT CGC CAT TTT ATA GAC A ATT TCG GAC CAT TTA TAA CC TCT ACT TGG ATA ACG TGC CTA TTG TTA GAG GTT AGA GTC TGT TGA TTA GTT GGA TTC AA TGA GAC ATG CAA TTG GAC ATC ATA GTT AGT AGT CGG
876 345 483 267 391 573
21 21 21 21 21 21 21
TCT ACT GGR TTR ACN TGC
345
23
TGG ATC AGA AAA AAC TCA AG CGG TAT TAT GTA AAA CTC AGA G
550
This paper This paper
F R F R
CTG CCC GAA CCA ACC CAR CAR GAR BCN CCR CCN GCA
330
25 25 26 25
F R
TTC CCY ATY CCT TTG GAA AAG TTG CAC TCA AAY AGA ACC CT
F/R
Rotavirus primer Detection of RV Beg9 common F End9 common R VP7 typing aBT1 G1 F aCT2 G2 F aET3 G3 F aDT4 G4 F aAT8 G8 F aFT9 G9 F RVG9 common R Alternative primer for G3 typing G3 G3 F VP4 typing con3 common F con2 common R 1T-1 P[8] R 2T-1 P[4] R 3T-1 P[6] R 4T-1 P[9] R 5T-1 P[10] R Alternative primer for P[8] typing 1T-1D P[8] Detection of P[8]b VP4-F103G P[8]b F VP4-R612C P[8]b R Norovirus primer Detection of NoV G1SKF G1SKR COG2F G2SKR Analysis of P2 region L1F L7R
TTY GTA AAT GA CCA TTR TAC A ATG TTY AGR TGG ATG AG TRH CCR TTR TAC AT
ed from these fecal suspensions using Extragen II (Tosoh, Tokyo, Japan) according to the manufacturer's instructions. The extracted RNA was reverse-transcribed using M-MLV reverse transcriptase (Promega, Madison, Wis., USA) and random hexamer (GE Healthcare, Piscataway, N.J., USA), with preincubaC for 5 min, followed by treatment at 379 C tion at 259 for 50 min and 709C for 15 min. RV detection and genotyping: The PCR assay was performed with Beg9 and End9 primers, as described elsewhere (20). Representative amplified PCR products were sequenced to ensure that only desired the PCR product was analyzed. The primers used in this study are shown in Table 1. G and P typing: RV-positive PCR samples were subjected to G and P genotyping by nested PCR assay with type-specific primers, as described previously (19,20). For G typing, 0.5 ml of the 1st round PCR products was added to 24.5 ml of the 2nd round PCR product containing the primer mix with all six serotype-specific forward primers (aBT1, aCT2, aET3, aDT4, aAT8, and aFT9) and a common reverse primer (RVG9) (20).
387
743
Reference
Shinohara (personal communication) 27
P genotyping was performed with the 1st round PCR product using con2 and con3 primers to amplify the VP4 gene, and the 2nd round PCR using primers 1T-1 (P[8]), 2T-1 (P[4]), 3T-1 (P[6]), 4T-1 (P[9]), 5T-1 (P[10]), and a common primer (con3), as described previously (21). Alternative primers (22,23) were used when the G and P type could not be determined using the aforementioned primers. G and P types were determined on the basis of the size of the amplicon after electrophoresis. When coexistence of types was suspected in a sample by multiplex typing PCR, singleplex PCR was performed using primers for the suspected types. We designed a pair of primers to detect P[8]b as VP4F103G and VP4-R612C (Table 1), based on strain MMC38 G9P8/05/Bangladesh (EU979379) from GenBank, for P[8]a and P[8]b subtyping. This primer pair was used after first PCR with the con3 and con2 primer pair (product size, 550 bp). Representative amplified PCR products were sequenced to ensure that only the desired PCR products had been analyzed. The PCR assay was conducted in the presence of 1.5 mM MgCl2, 1 mM/ml of each primer, and Taq polymerase (Biotech 406
International, Sydney, Australia) in a final volume of 25 ml. The thermal cycling conditions were 949C for 3 min, followed by 30 cycles of 949C, 559 C, and 729C for C for 7 min. 1 min each, and final incubation at 729 NoV detection and typing: The presence of NoV in the fecal specimens was determined by multiplex PCR with two primer sets: G1SKF/G1SKR for GI NoV and COG2F/G2SKR for GII NoV (Table 1), as described previously (23,25,26). Since GI and GII NoV were the prevalent genogroups of gastroenteritis, only these two genogroups were included in the analysis. Analysis of the VP1 gene of GII.4 NoV: The VP1 gene of GII.4 strains was analyzed further to determine the variant type. A PCR assay was performed using L1 and L7 (24) primers to detect the P2 region of the VP1 gene of GII.4 NoV (Table 1). The thermal cycling conditions were 949C for 3 min, followed by 40 cycles of C, 509C, and 729 C for 1 min, 1 min, and 2 min, re949 spectively, and final incubation at 729C for 7 min. Nucleotide sequencing and phylogenetic analysis: Direct sequencing of the VP7 gene of G3 strains and the VP4 gene of P[8] strains of RV, and sequencing of the 5? region of NoV ORF2 and the P2 region of the VP1 gene of GII.4 NoV, was performed using ABI Prism 310
and 3130 Genetic Analyzer and Big Dye Terminator Cycle Sequencing kit version 3.1 (Applied Biosystems, Foster City, Calif., USA) according to the manufacturer's instructions. Sequence alignment and phylogenetic analysis were carried out by the neighborjoining method using MEGA 3.1 (28). NoV genotyping was based on Kageyama and Okada's scheme using the nucleotide sequence of the N-terminal region of the capsid gene (17,18). The nucleotide sequence accession numbers in GenBank for the samples sequenced in this study are AB525797–AB525815 and AB536802– AB536810. Reference sequences for the phylogenetic analysis of NoV and RV were obtained from GenBank. RESULTS RV and NoV were detected in 87 (47.5z) and 12 (6.6z) of 183 fecal specimens from children with gastroenteritis collected in Nha Trang, Vietnam between November 2005 and June 2006, respectively. Dual infection with NoV and RV was detected in only one sample. The age distribution of the patients ranged from 1 month up to 44 months, although 88z of patients were younger than 24 months old. The majority of patients
Table 2. Monthly distribution of rotavirus and norovirus infections in patients with acute gastroenteritis Month Virus
Genotype
Rotavirus
Norovirus
G1 G2 G3 G4 G1/G3 G1/G2 Subtotal GII.4 GII.6 GII.12 GII.19 Subtotal
Total 1):
2005 Dec.
2006 Jan.
3
13 2
3
Total (z) Feb.
1 1 17
Mar.
Apr.
1 1 5 1 1
5 3 151)
2
3
1
3
12
2
1
2
9
9
24
7
24
3
1 1 4/10
0 17/29
May
Jun. (32.2) ( 6.9) (42.5) ( 1.1) (16.1) ( 1.1)
3
28 6 37 1 14 1 87
(33.3) ( 8.3) (50.0) ( 8.3)
0
4 1 6 1 12
3/5
99/183
1
5
1 11)
8
2
17/39
1
26/45
8/23
0 24/32
One sample of mixed infection with G3 rotavirus and GII.12 norovirus was included. Table 3. Distribution of rotavirus G and P genotypes among patients with acute gastroenteritis P type
G type
P[4]
P[6]
Mixed1)
P[8] P[4]/P[6]
G1 G2 G3 G4 G1/G2 G1/G3 Total (z)
2 6 2 1
1
P[4]/P[8]
P[4]/P[6]/P[8]
18
NT P[6]/P[8] 7
33
2
1
28 6 37 1 1 14
9 (10.3)
1 (1.1)
87
2 1
10 11 (12.6)
1 (1.1)
61 (70.1)
1 1 (1.1)
1 (1.1)
1): co-infected with two or three G or P genotypes. NT, non-typable samples.
407
2 (2.3)
Total (z) (32.2) ( 6.9) (42.5) ( 1.1) ( 1.1) (16.1)
with RV or NoV infection were also younger than 24 months old. RV was detected throughout the study period, with a peak observed in March (24/87, 27.6z) (Table 2). G types among the 87 RV-positive samples were G3 (n ＝ 37, 42.5z), G1 (n ＝ 28, 32.2z), G2 (n ＝ 6, 6.9z), and G4 (n ＝ 1, 1.1z) (Table 3). Mixed G-type RV infections (G1/G3 and G1/G2) were seen in 14 (16.1z) and 1 (1.1z) of the 87 cases, respectively. P[8] represented was the major P type (61/87, 70.1z), with a further 11 cases being P[4] (12.6z) and one P[6] (1.1z). Mixed P-type RV infections were seen in 9 (P[6]/P[8]), 2 (P[4]/P[6]/P[8]), one (P[4]/P[6]), and one (P[4]/P[8]) cases. G3P[8] and G1P[8] were the most common G and P genotype combinations, accounting for 33 (38z) and 18 (21z) of the RV infections, respectively. The predominant G type was found to vary with time, with G1 predominating from December 2005 to January 2006 and G3 from February to June 2006 (Table 2). As G3 was the most predominant type in this study, we sequenced the VP7 gene from five randomly selected G3 strains, all of which were grouped in the same clade as Asian reference strains, including other Vietnamese strains collected from the database (Fig. 1). Likewise, as P[8] was the most common P genotype, we conducted a genetic analysis of the VP4 gene of 10 randomly selected samples of G1 or G3 genotypes. These were grouped into two clusters: an OP601-like cluster for P[8]a and an OP354-like cluster for P[8]b (Fig. 2). Our design of a specific primer for P[8]b allowed 14 further strains to be differentiated from 49 tested samples out of 73 P[8] strains. These were grouped with G1 of the VP7 gene and classified into G1P[8]b. Only one case of NoV was detected in December
Fig. 1. Phylogenetic analysis of VP7 nucleotide sequences of rotavirus G3 strains. The phylogenetic tree was constructed based on 1,007 nucleotide sequences of the G3 VP7 genes. Percentage bootstrap values above 70z were shown at the branch nodes. , collected in Nha Trang, Vietnam, 2005–2006; #, collected during 2001–2004; , collected in Haiphong, Vietnam, September 2006.
Fig. 2. Phylogenetic analysis of partial VP4 nucleotide sequences of the rotavirus P[8] strains. The phylogenetic tree was constructed based on 645 nucleotide sequences of the VP4 genes. Percentage bootstrap values above 70z were shown at the branch nodes. , collected in Nha Trang, Vietnam; $, reference strains collected in Vietnam.
408
Fig. 3. Phylogenetic analysis of norovirus capsid gene. (A) Phylogenetic analysis of partial capsid nucleotide sequences of norovirus strains. The phylogenetic tree was constructed based on 218 nucleotide sequences of capsid gene. Percentage bootstrap values above 70z are shown at the branch nodes. Reference strains and their accession numbers used in Fig. 3(A) are as follows: GII.4: Bristol/93/UK (X76716), VN915/2003/VNM (DQ377174), VN800/2003/VNM (DQ377172), Hokkaido/194/2004/JP (AB240180), Sakai2/2006/JP (AB447448), Beijing/ 274/2005/CHN (EU839585), Chiba/040974/2004/JP (AB294782), Hunter 284E/04O/AU (DQ078794), Terneuzen70/2006/NL (EF126964), Farmington Hills/2002/USA (AY502023), Nijmegen115/2006/NL (EF126966); GII.13: M7/99/US (AY130761); GII.19: NLV/J23/1999/US (AY130762), Shaibah-2/2007/IRQ (EU138878), 299/JPN (EF630529), Hiroshima/66-1110/2006/JP (AB360387), CMH148/01/2001/THA (EU363866); GII.6: SaitamaU3/97/JP (AB039776), Miami/292/94/US (AF414410), HCMC204/2006/VNM (EU137732), HCMC311/2006/VNM (EU137733); GII.10: Mc37/99/Thai (AY237415); GII.15: SaitamaKU80aGII/99/JP (AB058582); GII.12: SaitamaU1/97/JP (AB039775), Hiroshima/25-583/2002/JP (AB354292), NZ587/2006/ NZL (EF187600), HCMC91/2006/VNM (EU137734), NSW021E/2006/AUS (EF187525), CMH145/05/2005/ THA (EU872289), NSW330F/2006/AUS (EF187578); GI.1: Norwalk/68/US2 (M87661). , collected in NhaTrang, Vietnam; ( ), number of strains. (B) Phylogenetic analysis of capsid amino acid sequences include P2 region of the norovirus GII.4 strains. The phylogenetic tree was constructed based on 200 amino acids sequences of the capsid gene include P2 region. Percentage bootstrap values above 70z are shown at the branch nodes. Reference strains and their accession numbers used in Fig. 3(B) are as follows: Beijing/79/2004/CHN (EU839583), Chiba/040974/2004/JP (AB294782), Sakai/04–179/2005/JP (AB220922), Sakai2/2006/JP (AB447448), Beijing/274/2005/CHN (EU839585), Chiba/040110/2004/JP (AJ844477), Chiba/031038/2003/JP (AJ844476), Nijmegen115/2006/NL (EF126966), Kobe034/2006/JP (AB291542), Terneuzen70/2006/NL (EF126964), Hunter 284E/04O/AU (DQ078794), Farmington Hills/2002/USA (AY502023), Bristol virus/B493/ 93 (X76716). , collected in NhaTrang, Vietnam.
2005, with a further 8 in February, 2 in March, and one in April (Table 2). All these cases were identified as GII (12/12, 100z). Phylogenetic analysis based on the nucleotide sequences of the N-terminal region of the capsid gene showed that half (6/12) these GII specimens were GII.12, with a further 4 being GII.4 and one each for was GII.6 and GII.19 (Fig. 3). A phylogenetic analysis of the P2 region of the VP1 amino acid sequences
allowed us to classify two GII.4 samples into the GII.4 2003 Asian clade. DISCUSSION RV has been detected essentially year round throughout the country, with a slight peak in the dry winter season, in a six-hospital surveillance in four cities in Viet409
nam (Hanoi, Haiphong, Nha Trang, and Ho Chi Minh) (4). Although the present study in Nha Trang, which has a tropical climate, was conducted during the dry season, and RVs were also detected throughout the study period, with a peak in March. In accordance with previous reports, our results indicate that RV is one of the leading viruses responsible for childhood gastroenteritis in Vietnam (7,29–35). The predominant G type for RV in Vietnam was G2 from 1998 to 2000 and G1 from 2000 to 2003, whereas genotypes G3 and G3P[8] RV were rare (7,31). However, G3 was isolated as the predominant genotype as of February 2006 in this study, as well as in Haiphong in north Vietnam during the 2006–2007 season (36). These observations therefore suggest that G3 infections occur throughout the country. Phylogenetic analysis of the VP7 gene of RV showed that the G3 strains in this study clustered with the strains collected from Ho Chi Minh (Vietnam), Kunming (China), Khabarovsk (Russia), Chiang Mai (Thailand) and Japan between 2001 and 2003 (37), Wuhan (China) in 2003–2005 (38), and Malaysia in 2004. Thus, these observations also suggest that isolation of the G3 strain has increased in frequency both in Vietnam and in neighboring countries, although the reasons for this increase remain unclear. Since a clinical trial of RotaTeq} (Merck, Whitehouse Station, N.J., USA) vaccine has been underway in Vietnam since 2007, continuous monitoring of the change of RV genotype will be required to understand the epidemiology of RV infection and to evaluate the efficacy of this vaccine in the future. The P[8] strain is the most predominant VP4 gene in the world (7), and this situation was mirrored in our study. Furthermore, phylogenetic analysis showed that P[8] could be grouped into two subclusters: an OP601like cluster for P[8]a and an OP354-like cluster for P[8]b. Strain OP354 was first detected in Malawi in 1998 (10) as a rare genotype of P[8] RV and was named as subtype P[8]b (8). This genotype was also isolated recently in some Asian countries, including Vietnam (8,9,11,12). All P[8]b strains in our study were classified into G1P[8]b and are therefore closely related to the CU20 strain, which was reported in Thailand during 2004–2005. Thus, our observations suggest that the G1P[8]b strain was circulating in Vietnam and neighboring countries during the same period. We designed a specific primer for P[8]b to detect more such strains and found 14 (28.6z) from a total of 49 P[8] strains. This result suggests that our specific primer for P[8]b may be effective for identifying and monitoring P[8]b, although this aspect warrants further study. The prevalence of NoV infection in our study was 7z, with GII.4 and GII.12 being predominant genotypes. These findings are consistent with a previous study performed in Ho Chi Minh, Vietnam during 2005 and 2006 (39) and suggest that these NoV genotypes were prevalent in southern Vietnam in that period. In accordance with previous reports (33,39), NoV GI was not detected in our study, although we identified one rare NoV GII.19 using partial capsid sequence analysis, which was the first case in Vietnam. This genotype had previously only been reported in the USA (15) and Japan (40).
Phylogenetic analysis of the P2 region of GII.4 strains allowed us to classify two representative samples into GII.4 2003 Asia variant, suggesting that an epidemic of this GII.4 variant occurred throughout Asia, including Vietnam. In conclusion, we have analyzed the prevalence of RV and NoV and shown that these viruses are important causal agents of acute viral gastroenteritis in children in Vietnam. Furthermore our detection of numerous genotypes of these viruses suggests that continuous surveillance and genetic analysis of gastroenteritis viruses is required in order to monitor and characterize the epidemiological correlation or possible antigenic changes in epidemic strains. Acknowledgments We thank Dr. Shinohara of Saitama Institute of Public Health, Japan, who kindly provided the primers for the nucleotide sequence analysis of P2 region of norovirus.
Conflict of interest None to declare. REFERECES 1. Bryce, J., Boschi-Pinto, C., Shibuya, K., et al. (2005): WHO estimates of the causes of death in children. Lancet, 365, 1147–1152. 2. Centers for Disease, Control and Prevention (2008): Rotavirus surveillance—worldwide, 2001–2008. Morbid. Mortal. Wkly. Rep., 57, 1255–1257. 3. Fischer, T.K., Anh, D.D., Antil, L., et al. (2005): Health care costs of diarrheal disease and estimates of the cost-effectiveness of rotavirus vaccination in Vietnam. J. Infect. Dis., 192, 1720–1726. 4. Van Man, N., Luan, L., Trach, D.D., et al. (2005): Epidemiological profile and burden of rotavirus diarrhea in Vietnam: 5 years of sentinel hospital surveillance, 1998–2003. J. Infect. Dis., 192, S127-S132. 5. Solberg, O.D., Hasing, M.E., Trueba, G., et al. (2009): Characterization of novel VP7, VP4, and VP6 genotypes of a previously untypeable group A rotavirus. Virology, 385, 58–67. 6. Santos, N. and Hoshino, Y. (2005): Global distribution of rotavirus serotypes/genotypes and its implication for the development and implementation of an effective rotavirus vaccine. Rev. Med. Virol., 15, 29–56. 7. Man, N.V., Trang, N.V., Lien, H.P., et al. (2001): The epidemiology and disease burden of rotavirus in Vietnam: sentinel surveillance at 6 hospitals. J. Infect. Dis., 183, 1707–1712. 8. Nagashima, S., Kobayashi, N., Paul, S., et al. (2009): Characterization of full-length VP4 genes of OP354-like P[8] human rotavirus strains detected in Bangladesh representing a novel P[8] subtype. Arch. Virol., 154, 1223–1231. 9. Samajdar, S., Ghosh, S., Dutta, D., et al. (2008): Human group A rotavirus P[8] Hun9-like and rare OP354-like strains are circulating among diarrhoeic children in Eastern India. Arch. Virol., 153, 1933–1936. 10. Cunliffe, N.A., Gondwe, J.S., Graham, S.M., et al. (2001): Rotavirus strain diversity in Blantyre, Malawi, from 1997 to 1999. J. Clin. Microbiol., 39, 836–843. 11. Nguyen, T.A., Hoang, L.P., Pham, L.D., et al. (2008): Use of sequence analysis of the VP4 gene to classify recent Vietnamese rotavirus isolates. Clin. Microbiol. Infect., 14, 235–241. 12. Theamboonlers, A., Bhattarakosol, P., Chongsrisawat, V., et al. (2008): Molecular characterization of group A human rotaviruses in Bangkok and Buriram, Thailand during 2004–2006 reveals the predominance of G1P[8], G9P[8] and a rare G3P[19] strain. Virus Genes, 36, 289–298. 13. Patel, M.M., Widdowson, M.-A., Glass, R.I., et al. (2008): Systematic literature review of role of noroviruses in sporadic gastroenteritis. Emerg. Infect. Dis., 14, 1224–1231. 14. Koopmans, M. (2008): Progress in understanding norovirus epidemiology. Curr. Opin. Infect. Dis., 21, 544–552. 15. Zheng, D.P., Ando, T., Fankhauser, R.L., et al. (2006): Norovirus classification and proposed strain nomenclature. Virology, 346, 312–323.
410
16. Katayama, K., Shirato-Horikoshi, H., Kojima, S., et al. (2002): Phylogenetic analysis of the complete genome of 18 Norwalk-like viruses. Virology, 299, 225–239. 17. Okada, M., Ogawa, T., Kaiho, I., et al. (2005): Genetic analysis of noroviruses in Chiba Prefecture, Japan, between 1999 and 2004. J. Clin. Microbiol., 43, 4391–4401. 18. Kageyama, T., Shinohara, M., Uchida, K., et al. (2004): Coexistence of multiple genotypes, including newly identified genotypes, in outbreaks of gastroenteritis due to Norovirus in Japan. J. Clin. Microbiol., 42, 2988–2995. 19. Siebenga, J.J., Vennema, H., Renckens, B., et al. (2007): Epochal evolution of GGII.4 norovirus capsid proteins from 1995 to 2006. J. Virol., 81, 9932–9941. 20. Gouvea, V., Glass, R.I., Woods, P., et al. (1990): Polymerase chain reaction amplification and typing of rotavirus nucleic acid from stool specimens. J. Clin. Microbiol., 28, 276–282. 21. Gentsch, J.R., Glass, R.I., Woods, P., et al. (1992): Identification of group A rotavirus gene 4 types by polymerase chain reaction. J. Clin. Microbiol., 30, 1365–1373. 22. Iturriza-Gomara, M., Kang, G. and Gray, J. (2004): Rotavirus genotyping: keeping up with an evolving population of human rotaviruses. J. Clin. Virol., 31, 259–265. 23. Iturriza-Gomara, M., Green, J., Brown, D.W.G., et al. (2000): Diversity within the VP4 gene of rotavirus P8 strains: implications for reverse transcription-PCR genotyping. J. Clin. Microbiol., 38, 898–901. 24. Yan, H.N., Yagyu, F., Okitsu, S., et al. (2003): Detection of norovirus (GI, GII), Sapovirus and astrovirus in fecal samples using reverse transcription single-round multiplex PCR. J. Virol. Methods, 114, 37–44. 25. Kojima, S., Kageyama, T., Fukushi, S., et al. (2002): Genogroupspecific PCR primers for detection of Norwalk-like viruses. J. Virol. Methods, 100, 107–114. 26. Kageyama, T., Kojima, S., Shinohara, M., et al. (2003): Broadly reactive and highly sensitive assay for Norwalk-like viruses based on real-time quantitative reverse transcription-PCR. J. Clin. Microbiol., 41, 1548–1557. 27. Shinohara, M., Shimada, S., Uchida, K., et al. (2006): Genetic analysis of P2 subdomain of norovirus strains detected in Saitama prefecture, 2004–2006. Ann. Rep. Saitama Inst. Public Health, 40, 62–66 (in Japanese). 28. Kumar, S., Tamura, K. and Nei, M. (2004): MEGA3: Integrated software for molecular evolutionary genetics analysis and sequence alignment. Brief. Bioinform., 5, 150–163.
29. Anh, D.D., Thiem, V.D., Fischer, T.K., et al. (2006): The burden of rotavirus diarrhea in Khanh Hoa Province, Vietnam: baseline assessment for a rotavirus vaccine trial. Pediatr. Infect. Dis. J., 25, 37–40. 30. Nishio, O., Matsui, K., Lan, D.T.P., et al. (2000): Rotavirus infection among infants with diarrhea in Vietnam. Pediatr. Int., 42, 422–424. 31. Nguyen, T.V., Le Van, P., Le Huy, C., et al. (2004): Diarrhea caused by rotavirus in children less than 5 years of age in Hanoi, Vietnam. J. Clin. Microbiol., 42, 5745–5750. 32. Nguyen, T.V., Le Van, P., Le Huy, C., et al. (2006): Etiology and epidemiology of diarrhea in children in Hanoi, Vietnam. Int. J. Infect. Dis., 10, 298–308. 33. Nguyen, T.A., Yagyu, F., Okame, M., et al. (2007): Diversity of viruses associated with acute gastroenteritis in children hospitalized with diarrhea in Ho Chi Minh City, Vietnam. J. Med. Virol., 79, 582–590. 34. Nguyen, T.A., Khamrin, P., Takanashi, S., et al. (2007): Evaluation of immunochromatography tests for detection of rotavirus and norovirus among Vietnamese children with acute gastroenteritis and the emergence of a novel norovirus GII.4 variant. J. Trop. Pediatr., 53, 264–269. 35. Bodhidatta, L., Lan, N.T.P., Hien, B.T., et al. (2007): Rotavirus disease in young children from Hanoi, Vietnam. Pediatr. Infect. Dis. J., 26, 325–328. 36. Ngo, T., Nguyen, B., Dang, D., et al. (2009): Molecular epidemiology of rotavirus diarrhoea among children in Haiphong, Vietnam: the emergence of G3 rotavirus. Vaccine, 27 (Suppl. 5), F75–80. 37. Trinh, Q.D., Pham, N.T.K., Nguyen, T.A., et al. (2007): Amino acid substitutions in the VP7 protein of human rotavirus G3 isolated in China, Russia, Thailand, and Vietnam during 2001–2004. J. Med. Virol., 79, 1611–1616. 38. Wang, Y.H., Kobayashi, N., Zhou, D.J., et al. (2007): Molecular epidemiologic analysis of group A rotaviruses in adults and children with diarrhea in Wuhan city, China, 2000–2006. Arch. Virol., 152, 669–685. 39. Nguyen, T.A., Hoang, L., Pham, L.D., et al. (2008): Norovirus and sapovirus infections among children with acute gastroenteritis in Ho Chi Minh City during 2005–2006. J. Trop. Pediatr., 54, 102–113. 40. Ozawa, K., Oka, T., Takeda, N., et al. (2007): Norovirus infections in symptomatic and asymptomatic food handlers in Japan. J. Clin. Microbiol., 45, 3996–4005.
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