Low Genetic Diversity of the Coat Protein Gene among Blueberry Red Ringspot Virus Isolates

Blueberry red ringspot virus (BRRSV) isolates have been investigated for genetic diversity. Nucleotide sequences of the coat protein (CP) gene of 19 isolates from Poland, Czech Republic, Slovenia and the United States were analysed. The nucleotide and amino acid sequence identity were 92–100% and 89–100%, respectively. Estimations of the distribution of synonymous and non‐synonymous changes indicated negative selection within the analysed CP gene and confirmed the genetic stability of the virus. At a capsid protein level, our results revealed BRRSV to be distinct from other, recombination‐prone pararetroviruses.     

Sequence analysis of the Czech Potato mop-top virus (PMTV) isolate Korneta-Nemilkov

The nucleotide sequence was determined for Czech potato mop-top virus (PMTV) isolate Korneta-Nemilkov, found in the potato field situated in South Bohemia. The nucleotide and amino acid sequences were compared with other PMTV isolates available in databases. The sequence identity was always >99% when Czech isolate RNA 2 and RNA 3 sequences were compared with each of the 3 Danish isolates and with Sw isolate, and slightly lower when compared to Scottish isolates. Similarity of deduced proteins was 100% for 5 out of 6 proteins used in comparison of Czech isolate with Danish isolate 54-15. The only difference between 2 isolates was found in coat protein (CP) gene. Interestingly, the CP of the Czech isolate seems to be 100% identical to the one of Sw, while many changes were found in the region encoding TGBp2, TGBp3 and cysteine-rich protein (CRP) for these 2 isolates. The lowest similarity scores were found when comparing the Czech isolate CRP with CRP of Scottish isolates.     

Partial Sequence Analysis of Geographically Close Grapevine virus A Isolates Reveals their High Regional Variability and an Intra‐Isolate Heterogeneity

To evaluate the genetic diversity of Grapevine virus A (GVA), the genomic region encompassing the partial capsid protein gene and ORF5 was analysed from 10 GVA isolates. Phylogenetic study showed a broad variability of the GVA isolates recovered from a limited geographical area (Slovakia and Czech Republic) and further confirmed the absence of geographical structuration within GVA. Moreover, assessment of structure and intra‐isolate variability revealed that grapevine samples infected with SK13 and SK29 isolates have harboured a population of different sequence variants.     

PCR detection and sequence analysis of duck circovirus in sick Muscovy ducks

The duck circovirus (DuCV) infection in sick ducks from Fujian Province was investigated. The liver samples of 43 sick Muscovy ducks with infectious serositis were collected from 12 duck farms in Fujian Province.Based on the published sequences of DuCV, two primers were designed for the detection of DuCV and four pairs of primers were designed to amplify four overlapping fragments that cover the complete genome of DuCV. The specific PCR products were amplified from positive samples. The fragments were then cloned into pMD18-T vector and sequenced, and the full length genomic sequence of the FJ0601 isolate of DuCV was obtained. PCR analysis showed that the proportion of ducks which were positive for circovirus was 79% and 10 out of the 12 farms were positive. Sequence analysis showed that the complete genome of DuCV-FJ0601 was 1988 bp and possessed features common to the family Circoviridae which included a stem-loop structure and the Rep protein motifs. Homology analysis showed that FJ0601 isolate of DuCV had 97.3%～97.5% nucleotide sequence identity to all the four Taiwan isolates (TC1/2002, TC2/2002, TC3/2002, TC4/2002), 82.9% identity to the America (33753-52) isolate and 82.3% identity to the Germany isolate. Phylogenetic analysis with Clustal W, however,showed that FJ0601 isolate of DuCV was on a common branch with Taiwan isolates, and Germany and America isolates belonged to the other branch.     

ITS rDNA sequences of Pomphorhynchus laevis (Zoega in Müller, 1776) and P. lucyi Williams & Rogers, 1984 (Acanthocephala: Palaeacanthocephala)

The internal transcribed spacers (ITS-1 and ITS-2) of the ribosomal RNA gene of Pomphorhynchus laevis (Zoega in Müller, 1776) (Acanthocephala) isolated from various fish species across Central and Southern Europe were compared with those of P. lucyi Williams & Rogers, 1984 collected from the largemouth bass Micropterus salmonoides Boulenger from the USA. The nucleotide sequences of ITS regions of P. laevis from minnows Phoxinus phoxinus (L.) and chub Leuciscus cephalus (L.) from two distant localities in the Slovak Republic were found to be 100% identical. The ITS-1 and ITS-2 of P. laevis from chub from the Czech Republic and Italy were also mutually identical, but significantly different from Slovak worms (88.7% identity for ITS-1, 91.3% identity for ITS-2). A fifth sample collected from Barbus tyberinus Bonaparte from Italy was very similar to the sympatric Italian isolate from chub, possessing four nucleotide substitutions in ITS-1 (98.4% identity). The ITS rDNA sequences of P. lucyi differed significantly from those of P. laevis; the values of identity were 51.8–56.1% for ITS-1 and 63.1–65.3% for ITS-2, and were significantly higher than the range of P. laevis within-species variability. The results based on the ITS sequences confirmed the occurrence of strains in P. laevis from Continental Europe which are well defined by molecules but reveal only slight differences in their morphology.     

侵染人参果的马铃薯M病毒基因组全序列分析

测定了侵染人参果的马铃薯M病毒(PVMCh)基因组全序列。线状、单链正义RNA全长8526bp,含6个开读框(ORF),具麝香石竹潜隐病毒属(Genus Carlavirus)典型结构特征。序列比较表明它与其他PVM分离物各基因核苷酸和编码蛋白氨基酸序列同源性分别为62.5%～9702%和60.9%～97.4%,其中CP基因最保守,而TGB3基因变异最大。系统进化树分析表明美国爱达荷州马铃薯分离物(PVMId) (AF023877)为PVM的一个远缘株系,而其他4个PVM分离物的成簇在外壳蛋白(Coat protein, CP)和核酸结合蛋白(Nucleic acid binding protein, NABP)区域略有差异。这是PVM在人参果上侵染的首次报道。     

Low Genetic Diversity Suggests a Single Introduction and Recent Spread of Tomato chlorosis virus in Brazil

By comparing the partial nucleotide sequences of the heat shock protein HSP70 homologue gene, we assessed the genetic diversity of Brazilian tomato isolates of Tomato chlorosis virus (ToCV), as well as their relationship with other ToCV isolates found worldwide. The Brazilian ToCV isolates shared 99.9–100% nucleotide identity, which indicates low genetic diversity. Brazilian ToCV isolates showed a closer evolutionary relationship to those from Mediterranean countries. Based on these results, the origin of Brazilian ToCV isolates and the possible number of introductions of the virus into Brazil are discussed.     

Serological and molecular characteristics of pea seed borne mosaic virus-PSbMV from lentil (Lens culinaris Medik L.) in Iran

Abstract

During 2016 growing season, samples with leaf yellowing and mosaic like symptoms were collected from main lentil (Lens culinaris Medik L.) fields. Specific ELISA positive PSbMV samples were selected for RT-PCR. Using specific pair of primer towards CP gene region of the virus, PCR product of approx.235?bp was amplified. Based on the nucleotide sequences of the CP gene PSbMV isolates were classified in two major groups. In which, isolates in group I divided into two subgroups of Iranian isolates (B2) and (G1), along with Czech Republic, Australia, China, Greece, Pakistan and Egypt were placed in the subgroup-I. Isolate (V18) from Iran placed independently in group II. In the grouping based on the amino acid sequencing the isolates divided into two phylogenetic groups. Iranian isolates along with an isolate from Australia categorized in group-II, however world isolates were all clustered in group-I.     

Begomoviruses Associated with Yellow Leaf Curl Disease of Tomato in Iran*

The occurrence of Tomato yellow leaf curl virus (TYLCV; genus Begomovirus, family Geminiviridae) in the major tomato‐growing areas of Iran was determined using TAS‐ELISA and PCR. The nucleotide sequences of the coat protein (CP) gene and intergenic region (IR) of eight Iranian isolates were determined. CP nucleotide identities among the Iranian isolates were 96–98%, and showed 94–96% identity with TYLCV‐IR [IR:Ira:98] and TYLCV‐IL [IL:Reo:86]. However, they showed low identity (68–69%) with ToLCIRV‐[IR:Ira]. Sequence analyses of IR indicated that seven Iranian isolates had sequence identity of 93–100% with each other, and 76% identity with the Jiroft isolate; identities of 75–79% with TYLCV‐IR[IR:Ira:98] were observed in every case, and 59–62% identity with ToLCIRV‐[IR:Ira]. The IR nucleotide sequences of Iranian isolates showed 92–93% identity with TYLCV‐IL[IL:Reo:86], except the Jiroft isolate (75%). The CP and IR sequence analyses suggested that eight Iranian TYLCV isolates probably differ from ToLCIRV‐[IR:Ira]. Based on IR sequence comparisons and phylogenetic analyses, the Iranian isolates were divided into two groups. The first major group (A), consists of seven virus isolates, was most closely related to TYLCV‐IL[IL:Reo:86], and relatively divergent from TYLCV‐IR [IR:Ira:98] and ToLCIRV‐[IR:Ira]. However, the Jiroft isolate from group B did not show high similarity with TYLCV‐IR[IR:Ira:98], ToLCIRV‐[IR:Ira], and TYLCV‐IL[IL:Reo:86], suggesting that the isolate may be a divergent variant. The differences are in a range that suggests different strains or species from TYLCV‐IR[IR:Ira:98] and ToLCIRV‐[IR:Ira] are probably associated with tomato yellow leaf curl disease in Iran.     

Molecular Analysis of ORF6, ORF7 and ORF8 of Beet yellows virus Iranian Isolate

Beet yellows virus (BYV), a member of the Closteroviridae family, is one of the most important sugar beet yellowing viruses. The nine ORFs of BYV genome encode different proteins required for BYV life cycle. We sequenced a part of the genome of BYV Iranian isolate consisting of ORF6, ORF7 and ORF8. The primer pair BYVA/Z was used for amplification of this region in RT‐PCR. The amplicon (1615 bp) was cloned and sequenced. Comparisons showed the amplified segment is corresponding to ORF6, ORF7 and ORF8 of BYV genome encoding coat protein, p20 and p21 proteins, respectively. The ORF7 of BYV Iranian isolate overlaps with ORF6 and ORF8 in four and 26 nucleotides at 5′ and 3′ ends, respectively. The ORF7 of Iranian isolate of BYV was sequenced completely. However, approximately 24 nt. from the beginning of ORF6 and 23 nt. from end of ORF8, including the stop codon, were not determined. ORF6, ORF7 and ORF8 showed the highest similarity at nucleotide (98.3, 99.4 and 99.2%) and amino acid (97.4, 98.9 and 100%) sequence levels, with BYV Ukrainian isolate. Phylogenetic analysis of the deduced amino acid sequences of ORF6, ORF7 and ORF8 revealed closer relationship of Iranian isolate of BYV with BYV Ukrainian isolate than other BYV isolates available at GenBank.     

Mixed Infection of Two Begomoviruses in Malvastrum coromandelianum in Fujian, China

Virus isolates were obtained from three Malvastrum coromandelianum plants showing vein thickening symptoms in Fujian Province, China. A fragment of approximately 500 bp was amplified from all the samples by PCR using the special degenerate primer pair PA/PB for begomoviruses. Sequence differences among the partial DNA-A fragments revealed that all three samples contained two virus isolates. Isolate I and isolate II share the highest nucleotide sequence identity (98–99%), respectively, with Malvastrum leaf curl Guangdong virus (MLCuGdV) and Ageratum yellow vein virus (AYVV). The complete nucleotide sequences of Fs1 and Fs2 isolates representing each virus were determined to be 2741 and 2756 nucleotides, respectively. Alignment and phylogenetic analysis showed that the complete DNA-A sequences of Fs1 and Fs2 were most closely to those of MLCuGdV (AM503104) and AYVV (AB100305), with 90.4% and 93.3% nucleotide sequence identity, respectively. Fs1 and Fs2 are considered therefore to be isolates of MLCuGdV and AYVV, respectively. This is the first report of AYVV in M. coromandelianum.     

我国水稻条纹病毒一个强致病性分离物的RNA4序列测定与分析

对我国水稻条纹病毒(Rice Stripe Virus,RSV)一个强致病性分离物(辽宁PJ分离物)的RNA4区段进行扩增、克隆和测序,其核苷酸序列全长2157bp。与已报道的日本T和M分离物及我国云南CX分离物的RNA4序列进行比较分析,结果表明,这4个分离物可分为两组,其中,PJ、T和M分离物为一组,组内分离物之间,RNA4的毒义链(vRNA4)及RNA4的毒义互补链(vcRNA4)上的ORF的核苷酸一致性分别为970%和970%～975%,5′末端和3′末端非编码区的序列则完全一致。但PJ分离物与T分离物的亲缘关系更为密切,其基因间隔区(IR)与T分离物的等长,核苷酸一致性为930%,比M分离物的IR多了一段长19bp的插入序列,核苷酸一致性仅为850%。另一组为我国CX分离物,组与组之间,vRNA4及vcRNA4上的ORF的核苷酸一致性分别为940%和925%～935%,但在氨基酸水平上则没有明显的差异。CX分离物的IR与PJ分离物相比有一段长84bp的插入序列,组间,IR的核苷酸一致性仅为720%～750%,5′末端非编码区的序列完全一致,但3′末端非编码区有两个碱基的差异。这些结果表明,RSV在自然界的分子变异与其地理分布具有密切的关系。此外,非编码区序列的高度保守性暗示着它们在病毒基因转录和复制的调控方面具有重要的功能。本文还讨论了RSV的分子流行学。     

Molecular Characterization of Whole Genomic RNA2 From Iranian Isolates of Grapevine Fanleaf Virus

Grapevine fanleaf virus (GFLV) is the major causal agent of the grapevine degeneration disease. To characterize the genomic RNA2 segment from Iranian isolates of GFLV, leaf samples were collected from infected vineyards in different locations with a long history of vine cultivation. Four isolates were selected for cloning and sequencing on the basis of the restriction profiles of RT‐PCR products. The sequencing data revealed that the RNA2 of the Iranian GFLV isolates were the shortest compared with that of all previously described GFLV isolates. The sizes were 3730 nucleotides (nt) for Shir‐Amin and Urmia isolates and 3749 nt for Takestan and Bonab isolates (excluding the poly (A) tail), due to deletion events in both 5′ and 3′ non‐coding regions. In the phylogenetic tree based on the full‐length nucleotide sequences of GFLV RNA2, all the GFLV isolates clustered into two groups with the exception of the Hungarian isolate (GHu). The Iranian isolates grouped as a distinct cluster. Recombination analyses showed that GFLV‐NW (Germany), GFLV‐F13 (reference isolate), GFLV isolate Shir‐Amin (Iran) and Arabis mosaic virus isolate Lv were recombinant isolates and one of their parents belonged to the same lineage as the Iranian isolates. These findings suggest that these isolates originated from a common ancestor.     

Characterization and Complete Nucleotide Sequence of Two Isolates of Tomato mosaic virus

Two virus isolates, designated S1 and TL, were obtained from tomato and camellia root in China, respectively, and their host ranges, symptomatology, serological reactions and complete nucleotide sequences were determined. Isolate TL systemically infected Chenopodium amaranticolor causing leaf chlorosis, but the isolate S1 induced only local necrotic lesions. The complete nucleotide sequences of S1 and TL were determined and consisted of 6384 and 6383 nucleotides (Genbank accessions AJ132845 and AJ417701 ), respectively. Sequence analysis revealed that both isolates have the highest nucleotide sequence identity (over 92%) with Tomato mosaic virus (ToMV), but less (80%) with other tobamoviruses. Phylogenetic analyses based on the amino acid sequences of 30‐kD and 17.5‐kD proteins also indicated that both the isolates form a cluster with the isolates of ToMV. These data suggest that S1 and TL are isolates of ToMV. The possible reasons that TL infected C. amaranticolor systemically but S1 induced only local necrotic lesions are discussed.     

Molecular Analysis of Zucchini yellow mosaic virus Isolates from Hangzhou, China

Isolates of Zucchini yellow mosaic virus were obtained from different cucurbit crops in Hangzhou city, China. The complete nucleotide sequences of four isolates and the 3′‐terminal sequences, including the coat protein coding region, of four others were determined and then compared with other available sequences. Phylogenetic analysis of the coat protein nucleotide sequences showed that these isolates fell into three significant groups, one of which (designated group III) consisted exclusively of Chinese isolates and is reported for the first time. Comparisons over the completely sequenced genomes showed that, typically for potyviruses, the 5′‐end of the genome was usually the most variable but that the group III isolate differed from the others most significantly in the N‐terminal part of the coat protein. Partially sequenced group III isolates also varied from other isolates in this region. Group III isolates appear to differ biologically from the other isolates because they do not cause symptoms in watermelon fruit but induce more severe symptoms on the watermelon leaves.     

Isolation and partial characterization of equine herpesvirus type 1 in Czechia

Equine herpesvirus type 1 was determined as the etiological cause of an abortion storm in Czechia in 2003 after the virus strain was isolated from aborted fetus and identified by serological means and by PCR technique. Cloning and sequencing of the glycoprotein D confirmed the identity of the isolates and showed molecular relationships to known EHV-1 strains. Comparison of glycoprotein D sequences with corresponding sequence of EHV-1 reference strains (Kentucky-A and Ab1) revealed high nucleotide homology. The Czech isolate of EHV-1 virus does not differ significantly from the Ab1 strain regarding the glycoprotein D gene and does not bear the frameshift in the 3' terminus which occurs in the Kentucky-A strain.     

Sequence analysis of a soilborne wheat mosaic virus isolate from Italy shows that it is the same virus asEuropean wheat mosaic virus andSoilborne rye mosaic virus

The complete sequence of the two RNAs of a furovirus isolate from durum wheat in Italy was determined. Sequence comparisons and phylogenetic analysis were done to compare the Italian virus withSoilborne wheat mosaic virus (SBWMV) from the USA and with furovirus sequences recently published asEuropean wheat mosaic virus (EWMV), from wheat in France, andSoilborne rye mosaic virus (SBRMV), from rye and wheat in Germany. Over the entire genome, the Italian isolate RNA1 and RNA2 had respectively 97.5% and 98.6% nucleotide identity with EWMV, 95.5% and 85.8% with SBRMV-G and 70.6% and 64.5% with SBWMV. The Italian isolate was therefore clearly distinct from SBWMV The European isolates all appear to belong to the same virus and the nameSoilborne cereal mosaic virus may resolve earlier ambiguities.