Complete genomic RNA sequences of cucumber mosaic virus strain NT9 from Taiwan

Summary The nucleotide sequences of the RNAs 1, 2, and 3 of the cucumber mosaic virus (CMV) Taiwan isolate NT9 were determined and compared at both the nucleotide and amino acid levels with those of CMV strains Fny, Y, O from subgroup I and strain Q from subgroup II. NT9-CMV has an unique feature at the C-terminus of the 3a protein which contains four extra-amino acids. All three RNAs and their encoded proteins, except 2b, of NT9-CMV share more than 90% identity with those of strains in subgroup I, and 72%–85% identity with Q-CMV. The results indicated the conservation of sequences of CMV derived from different geographical locations.The nucleotide sequence data reported in this paper will appear in the GenBank nucleotide sequence databases with the following accession numbers D28778, D28779, and D28780.     

Cloning and sequencing of the coat protein gene of tobacco ringspot virus isolates from UK and Iran

The coat protein (CP) gene and the 3' untranslated region (UTR) of genomic RNA 2 of Tobacco ringspot virus (TRSV, the genus Nepovirus, subgroup a) isolates from the UK and Iran were cloned from total viral RNA and sequenced. Comparison of these isolates with an isolate from the USA revealed a high degree of nucleotide and amino acid identity which extends the knowledge of molecular relationships between these three TRSV isolates. The UK isolate shared the highest nucleotide identity (95%) with the US isolate as compared to a lower identity with the Iranian isolate (92%). The highest identity (98%) was found between the UK and US isolates at amino acid level. Comparative analysis of the Iranian, UK and US isolates revealed some differences concerning some members of other subgroups of nepoviruses. For example, the N-terminal FDAYXR and the C-terminal FYGRXS motifs conserved among some nepoviruses, which occur adjacent to each other in folded CP molecules, were not detected in the Iranian, UK or US TSRV isolates. These isolates shared similarity only with Tomato ringspot virus (TomRSV) belonging to the subgroup c of nepoviruses. Another similarity of these isolates with TomRSV and Raspberry ringspot virus (RRSV) was the presence of a 34 nucleotide (nt) long sequence within the 3'-UTR.     

Hibiscus virus S is a new subgroup II tobamovirus: evidence from its unique coat protein and movement protein sequences

Summary.  The coat protein (CP) and movement protein (MP) sequences of a new tobamovirus infecting Hibiscus rosa-sinensis L were determined. The CP gene encodes 163 amino acid (aa) residues and with a theoretical molecular weight of 18.19 kDa. The MP gene encodes 282 amino acids and its theoretical molecular weight is 30.36 kDa. The nucleotide (nt) and aa sequences of the CP were 46.88 % to 51.63 % and 45.34 % to 57.06 % identical to other tobamoviruses, respectively. The nt and aa sequence identities of MP ranged from 38.81 % to 43.90 % and 30.85 % to 37.88 %, respectively. The predicted virion origin of assembly (OAS) was located in the CP gene. Phylogenetic trees generated based on the nt and aa sequences of both CP and MP genes indicate that this new virus clusters with members of subgroup II of tobamoviruses. Although this hibiscus virus shared a high nt and aa sequence identity with Sunn-hemp mosaic virus (SHMV), Western analysis showed that it is serologically unrelated to SHMV. We propose the name Hibiscus virus S (HVS) for this Singapore isolate. This is the first report on partial nt sequence of a tobamovirus that infects hibiscus. Received November 22, 2001; accepted February 28, 2002 Published online June 21, 2002     

甲型肝炎病毒龙甲株结构蛋白基因cDNA序列与国外毒…

将甲型肝炎病毒龙甲株（ＬＪ）结构蛋白基因（包括部分５端非编码基因ｎｔ６３０－３０４９）ｃＤＮＡ序徇与国外ＨＭ１７５、ＭＢＢ和ＬＡ三个母株相应区域相比较，结果显示，ＬＪ株结构蛋白基因与ＭＢＢ株的同源性最高，为９６．７％，与ＨＭ１７５和ＬＡ的同源率分别为９５．４％和９１．４％。推导的氨基酸变异率分别为０．９１％，０．９１％和２．９８％。ＬＪ与ＬＪ株氨基酸差异在整个结构蛋白区均有分布，而与ＨＭ１７５和Ｍ     

Molecular analysis of outer capsid glycoprotein (VP7) genes from two isolates of human group C rotavirus with different genome electropherotypes.

Nucleotide sequences for the VP7 gene of human group C rotavirus were determined for two strains isolated in Okayama, Japan, during a 1988-1990 epidemic. These isolates, OK118 and OK450, were selected as prototypes of two different electropherotypes, patterns I and II, respectively. The genes were identical in size (1,063 bp), and both contained singled open reading frames encoding 332 amino acids. The alignment of two sequences revealed 46 nucleotide substitutions, 11 of which were predicted to give amino acid changes. The deduced amino acid sequence of VP7 from OK118 was similar to published sequences of a Japanese isolate and three foreign isolates (more than 98.4% identity), whereas the VP7 sequence of OK450 revealed around 96% identity with these isolates and had nine unique amino acid substitutions. The VP7 genes of nine Okayama isolated were than analyzed by dot blot hybridization with the VP7 probes of OK118 and OK450. Under highly stringent conditions, the OK118 probe produced strong hybridization signals with the genes of five pattern I strains and one pattern II strain, while the OK450 probe strongly reacted only with those of three pattern II strains. Our results concluded that relative sequence diversity in the VP7 gene was observed between two different electropherotypes prevalent in a limited area.     

Nucleotide sequence variation of the VP7 gene of two G3-type rotaviruses isolated from dogs

The sequence of the VP7 gene of two rotaviruses isolated from dogs in southern Italy was determined and the inferred amino acid sequence was compared with that of other rotavirus strains. There was very high nucleotide and amino acid identity between canine strain RV198/95 and other canine strains, and to the human strain HCR3A. Strain RV52/96, however, was found to have about 95% identity to the G3 serotype canine strains K9, A79-10 and CU-1 and 96% identity to strain RV198/95 and to the simian strain RRV. Therefore both of the canine strains belong to the G3 serotype. Nevertheless, detailed analysis of the VP7 variable regions revealed that RV52/96 possesses amino acid substitutions uncommon to the other canine isolates. In addition, strain RV52/96 exhibited a nucleotide divergence greater than 16% from all the other canine strains studied; however, it revealed the closest identity (90.4%) to the simian strain RRV. With only a few exceptions, phylogenetic analysis allowed clear differentiation of the G3 rotaviruses on the basis of the species of origin. The nucleotide and amino acid variations observed in strain RV52/96 could account for the existence of a canine rotavirus G3 sub-type.     

Molecular characterization of G11P[25] and G3P[3] human rotavirus strains associated with asymptomatic infection in South India

Rotaviruses are the major etiological agents of diarrhea in children less than 5 years of age. Two unusual rotavirus strains not previously reported in India, G11P[25] (CRI 10795) and G3P[3] (CRI 33594) were isolated from faecal samples of asymptomatic children in India. The strains were characterized by sequence analysis of the genes encoding the VP7, VP4, VP6, and NSP4. The G11P[25] strain was closely related to the human G11P[25] strains from Bangladesh (with 98% identity at the nucleotide [nt] level and the amino acid [aa] level for the VP7 gene and 96% identity at the nt and 98% at the aa level for the VP4 gene). The G3P[3] strain was found to be related to a G3P[3] strain isolated in Thailand (CMH222; 88% identity at the nt level and 97% at aa level for the VP7 gene and 84% identity at the nt level and 90% at the aa level for the VP4 gene). Phylogenetic analysis of the VP6 and the NSP4 genes revealed that the Vellore G11P[25] strain was of VP6 subgroup II and NSP4 genotype B. The G3P[3] strain was identified as NSP4 genotype C and the VP6 gene showed 97% identity at the deduced amino acid level with strain CMH222 (Thailand) strain but did not cluster with sequences of SGI, SGII, SGI+II or SG-nonI/nonII. Both strains had gene segments of animal rotavirus origin suggesting inter-species transmission of rotavirus, and in the case of G11P[25] possibly underwent reassortment subsequently with human strains resulting in an animal-human hybrid strain.     

Variation in the spike protein of the 793/B type of infectious bronchitis virus, in the field and during alternate passage in chickens and embryonated eggs.

The degree of variation exhibited within the 793/B serotype (also known as 4/91 and CR88 serotypes) was investigated with nine French and 10 British isolates, collected between 1985 and 1994. The S1 part (1644 nucleotides) of the spike protein gene of the first known isolate of this serotype, FR/CR85131/85, had 95.9% to 97% nucleotide identity with the other isolates. Partial sequencing of isolates from Iran and Saudi Arabia, isolated in 2000, revealed approximately 95% nucleotide identity with European isolates, including the two live 793/B vaccinal strains, showing that they were not re-isolations of vaccinal virus. The data indicates that strains within the 793/B serotype have > or =96% nucleotide identity within the whole S1 gene and > or =93% nucleotide identity within the first 560 nucleotides, and > or =92% and > or =86% amino acid identities in the corresponding protein regions. This is similar to the identities exhibited within the Massachusetts serotype. Sequence analysis of a 793/B field isolate after passage in embryonated eggs, then in chickens and then again in eggs revealed selection for a serine and alanine at S1 amino acid position 95 in chicken-passaged and egg-passaged virus, respectively. There was no change in pathogenicity. This is the first demonstration at gene sequence level of host-driven selection for infectious bronchitis virus.     

Molecular evidence and sequence analysis of a natural reassortant between <Emphasis Type="Italic">Cucumber mosaic virus</Emphasis> subgroup IA and II strains

Cucumber mosaic virus (CMV) is a tripartite RNA virus and has been divided into three subgroups, named IA, IB, and II. Some studies have found a few natural reassortants between CMV subgroups, although reassortment between CMV subgroups is infrequent. In our present work, a CMV reassortant, named CMV-Tsh, was obtained from a tomato plant. The complete sequence of CMV-Tsh genomic RNAs has been determined and analyzed. The results of sequence comparisons and phylogenetic analyses revealed that CMV-Tsh RNAs 1 and 3 are derived from one or two CMV subgroup II strain(s), while RNA2 is derived from a CMV subgroup IA strain. A PCR and restriction enzyme analysis-based method was developed to analyze the possibility of mixed infection by CMV strains of different subgroup in the CMV-Tsh-infected tomato plant. The results of the restriction enzyme analysis proved that CMV-Tsh is the unique strain in the tomato plant. Taken together, CMV-Tsh is a natural reassortant having CMV subgroup IA RNA2 and subgroup II RNAs 1 and 3. The GenBank Accession numbers of the sequences reported in this paper are EF202595- EF202597.     

Modification of rotavirus multiplex RT-PCR for the detection of G12 strains based on characterization of emerging G12 rotavirus strains from South India

Rotaviruses are the major etiological agents of diarrhea in children less than 5 years of age. The commonest G types in humans are G1-4 and G9. G12 is a rare human rotavirus (HRV) strain first reported in the Philippines. In this study, 13 G12 strains obtained from a community-based cohort and a hospital-based surveillance system in 2005 were characterized by phylogenetic analysis of partial nucleotide sequences of VP7, VP6, and NSP4 genes. Sequence and phylogenetic analysis of VP7 gene sequences showed that these southern Indian strains had the greatest homology with G12 strains recently reported from eastern India (97-99% identity both at the nucleotide level and deduced amino acid level) and less homology with the prototype G12 strain, L26 (89-90% identity at the nucleotide level and 90-94% at the deduced amino acid level). Phylogenetic analysis of the VP6 and the NSP4 genes revealed that the Vellore G12 strains belonged to VP6 subgroup II and NSP4 genotype B. The P types associated with these strains were P[6] and P[8]. A G12 type-specific primer was designed for inclusion in an established VP7 G-typing multiplex RT PCR, and tested against a panel of known G types and untyped samples and was found to detect G12 strains in the multiplex-PCR. Close homology of the South Indian G12 strains to those from Kolkata suggests that G12 HRV strains are emerging in India. Methods for characterization of rotaviruses in epidemiological studies need to be updated frequently, particularly in developing countries.     

Comparison of the RNA polymerase genes of marine birnavirus strains and other birnaviruses

Summary.  The cDNA nucleotide sequence of the genome segment B encoding the VP1 protein, the putative RNA-dependent RNA polymerase (RdRp), was determined for 5 marine birnavirus (MABV) strains from different host or geographic origins and 1 infectious pancreatic necrosis virus (IPNV) strain AM-98. Segment B of the IPNV AM-98 strain and 4 MABV strains, Y-6, YT-01A, H1 and NC1, contained a 2535 bp ORF, which encoded a protein of 845 amino acid residues with a predicted MW of 94.4 kDa. Only the MABV AY-98 RdRp had 1 amino acid shorter RdRp. Pairwise comparisons were made among our data and 4 other known IPNV sequences. The nucleotide sequences of the 5 MABV strains were very similar each other, with identities of 98.3–99.7%. The highest divergence of the nucleotide level was between MABV strains and IPNV SP strain (serotype A2), with 20.4–20.8% divergences in the coding region, which gave 10.1–11.3% divergence in the amino acid level. The aquabirnavirus RdRp was noticeably conserved in amino acid sequences. Though the identities of the nucleotide sequences of encoding region were 85.1–85.9% between MABV strains and IPNV serotype A1 strains, they shared as high as 95.1–95.9% identities in amino acid level. A phylogenetic tree was constructed based on the amino acid sequences of the RdRp gene from different birnaviruses including avibirnavirus and entomobirnavirus. Ten aquabirnavirus strains were clustered into 3 Genogroups. The Genogroup I consisted of four IPNV A1 serotype strains. All MABV strains were clustered into Genogroup II. Only IPNV SP strain was clustered into an independent Genogroup III. Received August 19, 2002; accepted October 30, 2002     

Nucleotide sequences of 5'-terminal parts of coat protein genes of various isolates of NTN strain of potato virus Y

Sequences of the first 300 nucleotides of coat protein (CP) genes of 7 isolates of NTN strain of potato virus Y (PVY, PVY(NTN)) were determined and compared with analogous published sequences of various isolates and strains of PVY. The sequence identity among the sequenced isolates ranged from 96 to 100%. The differences were found at different positions. The nucleotide sequence of this part of CP gene seems to be very conservative among the isolates tested that means that PVY(NTN) is the evolutionary youngest among all PVY strains.     

Induction of tobacco chlorosis by certain cucumber mosaic virus satellite RNAs is specific to subgroup II helper strains

Two satellite (sat) RNAs of cucumber mosaic virus (CMV), B2- and WL3-sat RNAs, which induce systemic chlorosis on tobacco, were inoculated onto tobacco with a number of CMV strains. Systemic chlorosis was observed only when these satellite RNAs were associated with subgroup II CMV strains. Infection of tobacco with various pseudorecombinants of subgroup I and II CMV strains, together with WL3- or B2-sat RNA, suggests that chlorosis is associated with RNA 2 of subgroup II CMV strains. Chlorosis was not induced when B2- or WL3-sat RNAs were inoculated onto tobacco with tomato aspermy virus. In contrast, the induction of chlorosis on tomato by B1-sat RNA did not show any clear dependence on the subgroup of its CMV helper strain although chlorosis did tend to be more severe in association with subgroup II CMV strains.     

Cactus mild mottle virus is a new cactus-infecting tobamovirus

Summary. A new cactus-infecting tobamovirus, Cactus mild mottle virus (CMMoV), was isolated from diseased grafted cactus, Gymnocalycium mihanovichii and its molecular properties were characterized. CMMoV is distantly related to known species of the genus Tobamovirus on the basis of serological and sequence analyses. Western blot analysis showed that CMMoV is serologically unrelated to Sammon’s Opuntia virus, which is the only known species of the genus Tobamovirus found in cactus plants. The 3′-terminal 2,910 nucleotides of CMMoV have been sequenced. The coat protein (CP) and movement protein (MP) genes encode 161 and 306 amino acids residues, respectively, and the 3′ untranslated region (UTR) consists of 229 nucleotides long. The nucleotide and amino acid sequences of the CP of CMMoV were 39.6% to 49.2% and 25.8% to 40.3% identical to other seventeen tobamoviruses, respectively. The MP shared 34.9% to 40.6% and 16.3% to 27.0% and 44.6% to 63.4% identities, respectively, at the amino acid and nucleotide levels with other members of the genus. Percentage identities of nucleotides of the 3′ UTR ranged from 42.5% to 63.4%. Phylogenetic tree analyses of the CP and MP suggest the existence of the fifth cactus-infecting subgroup in the genus Tobamovirus. Sequence analyses of these two viral proteins revealed that the highest amino acid sequence identity between the virus and seventeen other tobamoviruses was 40.6%, supporting the view that CMMoV is a new definite species of the genus Tobamovirus.     

中国部分甲肝病毒流行株结构蛋白VP3-VP1区基因特点分析

目的 分析中国部分甲肝病毒流行株结构蛋白VP3-VP1区基因特点.方法 收集42份甲肝患者急性期血清标本,经核酸提取、逆转录及巢氏PCR,测得结构-非结构蛋白VP3-VP1-2A区序列,进行序列同源性比较并分析其基因特点.结果 42株HAV病毒株在VP1-2A连接处核苷酸和氨基酸序列同源性分别为89.1%～100%和97.3%～100%;在全长结构蛋白VP3-VP1区的核苷酸和氨基酸序列同源性分别为87.6%～100%和98.8%～100%.VP1-2A连接处序列相同的病毒株在全长结构蛋白VP3-VP1区的核苷酸同源性为98.4%～100%,0～2个氨基酸位点不同.本实验所得序列在中和抗原位点处氨基酸序列均未变异.结论 42株病毒株均属于I型,40株是IA亚型,2株IB亚型.本实验所用HAV流行株在结构蛋白VP3-VP1区的核苷酸存在差异但是氨基酸序列高度保守且没有中和抗原位点处的变异.VP1-2A结合处核苷酸序列相同的分离株在全长结构蛋白VP3-VP1区核苷酸序列相同或相近,氨基酸序列保守.     

Detection of a human rotavirus with G12 and P[9] specificity in Thailand

G12 rotavirus has not been detected anywhere in the world since the first detection of a human strain, L26 (G12, P1B[4]), in the Philippines in 1990. In this study, we isolated a human rotavirus (strain T152) with a VP7 of G12 specificity from the stool of an 11-month-old diarrheic patient in Thailand. The strain T152 exhibited a long RNA pattern and subgroup I specificity. In the comparison of the nucleotide and amino acid sequences of the VP7 gene of strain T152 with those of rotaviruses with different G type specificities, strain T152 showed the highest identity, 90.9 and 93.9%, respectively, to G12 prototype strain L26. In contrast, the VP4 gene of strain T152 showed the highest identity with P[9] specificity of human strains K8 and AU-1 and feline strains Cat2 and FRV-1, with homologies of 89.3 to 90.6% at the nucleotide level and 93.9 to 95.6% at the amino acid level. Thus, strain T152 was found to be a natural reassortant strain with G12 and P[9] specificities.     

中国部分甲肝病毒流行株结构蛋白VP3-VPl区基因特点分析

目的分析中国部分甲肝病毒流行株结构蛋白VP3-VPl区基因特点。方法收集42份甲肝患者急性期血清标本,经核酸提取、逆转录及巢氏PCR,测得结构一非结构蛋白VP3-VPl-2A区序列,进行序列同源性比较并分析其基因特点。结果42株HAV病毒株在VPl-2A连接处核苷酸和氨基酸序列同源性分别为89．1％～100％和97．3％～100％;在全长结构蛋白VP3-VPl区的核苷酸和氨基酸序列同源性分别为87．6％～100％和98．8％～100％。VPl-2A连接处序列相同的病毒株在全长结构蛋白VP3．VPl区的核苷酸同源性为98．4％～100％,0～2个氨基酸位点不同。本实验所得序列在中和抗原位点处氨基酸序列均未变异。结论42株病毒株均属于I型,40株是IA亚型,2株IB亚型。本实验所用HAV流行株在结构蛋白VP3．VPl区的核苷酸存在差异但是氨基酸序列高度保守且没有中和抗原位点处的变异。VPl-2A结合处核苷酸序列相同的分离株在全长结构蛋白VP3-VPl区核苷酸序列相同或相近,氨基酸序列保守。     

中国部分甲肝病毒流行株结构蛋白VP3-VP1区基因特点分析

目的 分析中国部分甲肝病毒流行株结构蛋白VP3-VP1区基因特点.方法 收集42份甲肝患者急性期血清标本,经核酸提取、逆转录及巢氏PCR,测得结构-非结构蛋白VP3-VP1-2A区序列,进行序列同源性比较并分析其基因特点.结果 42株HAV病毒株在VP1-2A连接处核苷酸和氨基酸序列同源性分别为89.1%～100%和97.3%～100%;在全长结构蛋白VP3-VP1区的核苷酸和氨基酸序列同源性分别为87.6%～100%和98.8%～100%.VP1-2A连接处序列相同的病毒株在全长结构蛋白VP3-VP1区的核苷酸同源性为98.4%～100%,0～2个氨基酸位点不同.本实验所得序列在中和抗原位点处氨基酸序列均未变异.结论 42株病毒株均属于I型,40株是IA亚型,2株IB亚型.本实验所用HAV流行株在结构蛋白VP3-VP1区的核苷酸存在差异但是氨基酸序列高度保守且没有中和抗原位点处的变异.VP1-2A结合处核苷酸序列相同的分离株在全长结构蛋白VP3-VP1区核苷酸序列相同或相近,氨基酸序列保守.     

Transgenic tobacco plants accumulating tobacco rattle virus coat protein resist infection with tobacco rattle virus and pea early browning virus

Transgenic tobacco plants expressing the coat protein (CP) gene of tobacco rattle virus (TRV) strain TCM were found to be resistant to infection with the homologous virus but not to infection with the PLB strain of TRV. The amino acid sequence identity between the CP of TRV strains TCM and PLB is 39% and the two CP genes do not cross-hybridize. On the other hand, there is extensive cross-hybridization between the CP genes of TRV-TCM and a Dutch isolate of pea early browning virus (PEBV). The transgenic plants accumulating TRV-TCM CP showed a considerable resistance to infection with PEBV.