Development of PCR markers linked to resistance to wheat streak mosaic virus in wheat

Wheat streak mosaic virus (WSMV), vectored by the wheat curl mite (Acer tulipae), is an important disease of wheat (Triticum aestivum L.) in the North American Great Plains. Resistant varieties have not been developed for two primary reasons. First, useful sources of resistance have not been available, and second, field screening for virus resistance is laborious and beyond the scope of most breeding programs. The first problem may have been overcome by the development of resistance to both the mite and the virus by the introgression of resistance genes from wild relatives of wheat. To help address the second problem, we have developed polymerase chain reaction (PCR) markers linked to the WSMV resistance gene Wsm1. Wsm1 is contained on a translocated segment from Agropyron intermedium. One sequence-tagged-site (STS) primer set (WG232) and one RAPD marker were found to be linked to the translocation containing Wsm1. The diagnostic RAPD band was cloned and sequenced to allow the design of specific PCR primers. The PCR primers should be useful for transferring Wsm1 into locally adapted cultivars.This is Journal Series No. J-4041 of the Montana Agricultural Experiment Station     

The distribution of wheat curl mite (Aceria tosichella) lineages in Australia and their potential to transmit wheat streak mosaic virus

The wheat curl mite (WCM), Aceria tosichella , is an eriophyid pest of cereals, and the vector responsible for the transmission of wheat streak mosaic virus (WSMV). In a previous study, the taxonomic status of A. tosichella in Australia was assessed using molecular markers. A. tosichella was shown to consist of two genetically distinct lineages likely to represent different species. Here we show that both lineages occupy similar distributions, occurring throughout the entire Australian wheat belt, and that the lineages are often found in sympatry. CLIMEX analysis suggests that tolerance to heat and desiccation limit the distribution of A. tosichella . In the laboratory, only one WCM lineage transmitted WSMV virus under controlled conditions. These results have implications for the management of WCM and WSMV within Australia.     

Responses of maize (<Emphasis Type="Italic">Zea mays</Emphasis> L.) near isogenic lines carrying <Emphasis Type="Italic">Wsm1</Emphasis>, <Emphasis Type="Italic">Wsm2,</Emphasis> and <Emphasis Type="Italic">Wsm3</Emphasis> to three viruses in the Potyviridae

Genes on chromosomes six (Wsm1), three (Wsm2) and ten (Wsm3) in the maize (Zea mays L.) inbred line Pa405 control resistance to Wheat streak mosaic virus (WSMV), and the same or closely linked genes control resistance to Maize dwarf mosaic virus (MDMV) and Sugarcane mosaic virus (SCMV). Near isogenic lines (NIL) carrying one or two of the genes were developed by introgressing regions of the respective chromosomes into the susceptible line Oh28 and tested for their responses to WSMV, MDMV, and SCMV in the field and greenhouse. F₁ progeny from NIL × Oh28 were also tested. Wsm1, or closely linked genes, provided resistance to all three viruses, as determined by symptom incidence and severity. Wsm2 and Wsm3 provided resistance to WSMV. Wsm2 and/or Wsm3 provided no resistance to MDMV, but significantly increased resistance in plants with one Wsm1 allele. NIL carrying Wsm1, Wsm2, or Wsm3 had similar SCMV resistance in the field, but NIL with Wsm2 and Wsm3 were not resistant in the greenhouse. Addition of Wsm2 to Wsm1 increased SCMV resistance in the field. For all viruses, symptom incidence was higher in the greenhouse than in the field, and relative disease severity was higher in the greenhouse for WSMV and MDMV. An Italian MDMV isolate and the Ohio SCMV infected the Wsm1 NIL, while the Ohio MDMV and Seehausen SCMV isolates did not. Our results indicate that the three genes, or closely linked loci, provide virus resistance. Resistance conferred by the three genes is influenced by interactions among the genes, the virus species, the virus isolate, and the environment.     

Molecular characterization of the genome composition of partial amphiploids derived from Triticum aestivum×Thinopyrum ponticum and T. aestivum×Th. intermedium as sources of resistance to wheat streak mosaic virus and its vector, Aceria tosichella

 Wheat streak mosaic virus (WSMV), vectored by the wheat curl mite (WCM), is one of the most important viral diseases of wheat (Triticum aestivum) in the world. Genetic resistance to WSMV and the WCM does not exist in wheat. Resistance to WSMV and the WCM was evaluated in five different partial amphiploids namely Agrotana, OK7211542, ORRPX, Zhong 5 and TAF 46, which were derived from hybrids of wheat with decaploid Thinopyrum ponticum or with hexaploid Th. intermedium. Agrotana was shown to be immune to WSMV and the WCM; the other four partial amphiploids were susceptible to the WCM. Genomic in situ hybridization (GISH) using genomic DNA probes from Th. elongatum (EE, 2n=14), Th. bessarabicum (JJ, 2n=14), Pseudoroegneria strigosa (SS, 2n=14) and T. aestivum (AABBDD, 2n=42) demonstrated that three of the partial amphiploids, Agrotana, OK7211542 and ORRPX, have almost identical alien genome constitutions: all have 16 alien chromosomes, with 8 chromosomes being closely related to the J^s genome and 8 chromosomes belonging to the E or J genomes and no evidence of any S-genome chromosomes. GISH confirmed that the alien genomes of Agrotana and OK7211542, like ORRPX, were all derived from Th. ponticum, and not from Th. intermedium. However, in contrast to Agrotana, ORRPX and OK7211542 were susceptible to the WCM and WSMV. The partial amphiploid Zhong 5 had a reconstituted alien genome composed of 4 S-and 4 J^s-genome chromosomes of Th. intermedium with 6 translocated chromosomes between the S and J^s genomes. This line was highly resistant to WSMV, but was susceptible to the WCM. TAF 46, which contained a synthetic genome consisting of 3 pairs of S-genome chromosomes and 4 pairs of E- or J-genome chromosomes in addition to the 21 pairs of wheat chromosomes, was susceptible to the WCM, but moderately resistant to WSMV. Agrotana offers great potential for transferring WSMV and WCM resistance into wheat. Received: 27 January 1998 / Accepted: 10 February 1998     

Distribution and influence of grazing on wheat curl mites (Aceria tosichella Keifer) within a wheat field

Wheat streak mosaic virus (WSMV) is a serious disease of wheat and is primarily transmitted from infected to healthy plants by the wheat curl mite, Aceria tosichella Keifer. Although wheat is the primary plant host of A. tosichella, wheat curl mites have been recorded on more than 60 different plant hosts; this broad host range allows mites to survive outside the wheat‐growing season by providing a ‘green bridge’. Despite the fact that A. tosichella can only crawl short distances, the mites can disperse via wind and thus have the capacity to readily infest wheat crops from neighbouring refuges. In this study, we undertook field trials to investigate the temporal movement of A. tosichella, as well as the importance of wind and livestock grazing on mite dispersal late in the cropping season. We demonstrate there is a window in spring when A. tosichella undergo significant movement in south‐eastern Australia, and this is likely related to the development stage of wheat plants, and may also be influenced by wind direction. We found that grazing wheat crops reduced mite numbers, suggesting that any increase in WSMV issues in ‘grain and graze’ crops is likely due to the longer season wheat varieties used in these systems rather than the direct effects of grazing. These results emphasize the importance of crop management strategies in the control of A. tosichella.     

Hairpin RNA derived from viral NIa gene confers immunity to wheat streak mosaic virus infection in transgenic wheat plants

Wheat streak mosaic virus (WSMV), vectored by Wheat curl mite, has been of great economic importance in the Great Plains of the United States and Canada. Recently, the virus has been identified in Australia, where it has spread quickly to all major wheat growing areas. The difficulties in finding adequate natural resistance in wheat prompted us to develop transgenic resistance based on RNA interference (RNAi). An RNAi construct was designed to target the nuclear inclusion protein ‘a’ (NIa) gene of WSMV. Wheat was stably cotransformed with two plasmids: pStargate‐NIa expressing hairpin RNA (hpRNA) including WSMV sequence and pCMneoSTLS2 with the nptII selectable marker. When T₁ progeny were assayed against WSMV, ten of sixteen families showed complete resistance in transgenic segregants. The resistance was classified as immunity by four criteria: no disease symptoms were produced; ELISA readings were as in uninoculated plants; viral sequences could not be detected by RT‐PCR from leaf extracts; and leaf extracts failed to give infections in susceptible plants when used in test‐inoculation experiments. Southern blot hybridization analysis indicated hpRNA transgene integrated into the wheat genome. Moreover, accumulation of small RNAs derived from the hpRNA transgene sequence positively correlated with immunity. We also showed that the selectable marker gene nptII segregated independently of the hpRNA transgene in some transgenics, and therefore demonstrated that it is possible using these techniques, to produce marker‐free WSMV immune transgenic plants. This is the first report of immunity in wheat to WSMV using a spliceable intron hpRNA strategy.     

Population genetics of the wheat curl mite (Aceria tosichella Keifer) in Australia: implications for the management of wheat pathogens

The wheat curl mite (WCM), Aceria tosichella Keifer, is a polyphagous eriophyoid mite and the primary vector of wheat streak mosaic virus (WSMV) and five other viral pathogens in cereals. Previous research using molecular markers and a series of laboratory experiments found A. tosichella in Australia to consist of two genetically distinct lineages, which have broad overlapping distributions and differ in their ability to transmit WSMV under controlled conditions. This pattern of transmission also appears to be apparent in the field, whereby a strong association between WSMV detection and a single WCM lineage has been detected. In this study, we conduct a population genetic analysis and provide information on the genetic structure of the Australian viruliferous WCM lineage. We assessed genetic differentiation of 16 WCM populations using nine microsatellite markers. Strong evidence for extensive gene flow and low genetic structuring throughout the Australian wheatbelt was evident, with an exception for Western Australian and far north Queensland populations that appear to be genetically isolated. The data also indicate genetic patterns consistent with an arrhenotokous parthenogenetic mode of reproduction. Implications of these findings are discussed with reference to the management of WCM and associated cereal pathogens in Australia and overseas.     

Global spread of wheat curl mite by its most polyphagous and pestiferous lineages

The wheat curl mite (WCM), Aceria tosichella, is an important pest of wheat and other cereal crops that transmits wheat streak mosaic virus and several other plant viruses. Wheat curl mite has long been considered a single polyphagous species, but recent studies in Poland revealed a complex of genetically distinct lineages with divergent host‐acceptance traits, ranging from highly polyphagous to host‐specific. This diversity of WCM genotypes and host‐acceptance phenotypes in Europe, the presumed native range of WCM, raises questions about the lineage identities of invasive WCM populations on other continents and their relationships to European lineages. The goals of this study were to examine the global presence of WCM and determine the relatedness of lineages established in different continents, on the basis of phylogenetic analyses of mitochondrial and nuclear DNA sequence data. Host‐range bioassays of a highly polyphagous WCM lineage were performed to supplement existing data on this lineage's ability to colonise graminaceous and non‐graminaceous hosts. Invasive WCM populations in North and South America and Australia assorted with the only three known polyphagous and pestiferous WCM lineages (‘MT‐1’, ‘MT‐7’ and ‘MT‐8’) from a total of eight currently described lineages. These results show that the most polyphagous lineages were more successful colonisers and reflect a need for extensive surveys for WCM on both crops and wild grass species in invaded continents. The most invasive lineage (‘MT‐1’) was shown to successfully colonise all 10 plant species tested in three families and has spread to North and South America and Australia from its presumed origins in Eurasia.     

Molecular cytogenetic discrimination and reaction to wheat streak mosaic virus and the wheat curl mite in Zhong series of wheat--Thinopyrum intermedium partial amphiploids.

Thinopyrum intermedium (2n = 6x = 42, JJJsJsSS) is potentially a useful source of resistance to wheat streak mosaic virus (WSMV) and its vector, the wheat curl mite (WCM). Five partial amphiploids, namely Zhong 1, Zhong 2, Zhong 3, Zhong 4, and Zhong 5, derived from Triticum aestivum x Thinopyrum intermedium crosses produced in China, were screened for WSMV and WCM resistance. Zhong 1 and Zhong 2 had high levels of resistance to WSMV and WCM. The other three partial amphiploids, Zhong 3, 4, and 5, were resistant to WSMV, but were susceptible to WCM. Genomic in situ hybridization (GISH) using a genomic DNA probe from Pseudoroegneria strigosa (SS, 2n = 14) demonstrated that two partial amphiploids, Zhong 1 and Zhong 2, have almost the identical 10 Th. intermedium chromosomes, including four Js, four J, and two S genome chromosomes. Both of them carry two pairs of J and a pair of Js genome chromosomes and two different translocations that were not observed in the other three Zhong lines. The partial amphiploids Zhong 3, 4, and 5 have another type of basic genomic composition, which is similar to a reconstituted alien genome consisting of four S and four Js genome chromosomes of Th. intermedium (Zhong 5 has two Js chromosomes plus two Js-W translocations) with six translocated chromosomes between S and Js or J genomes. All three lines carry a specific S-S-Js translocated chromosome, which might confer resistance to barley yellow dwarf virus (BYDV-PAV). The present study identified a specific Js2 chromosome present in all five of the Zhong lines, confirming that a Js chromosome carries WSMV resistance. Resistance to WCM may be linked with J or Js chromosomes. The discovery of high levels of resistance to both WSMV and WCM in Zhong 1 and Zhong 2 offers a useful source of resistance to both the virus and its vector for wheat breeding programs.     

Influence of volunteer wheat plant condition on movement of the wheat curl mite, Aceria tosichella, in winter wheat

The wheat curl mite (WCM), Aceria tosichella Keifer, is the vector of wheat streak mosaic virus and high plains virus which cause significant crop loss in winter wheat throughout the western Great Plains. Volunteer wheat emerging before harvest, as a result of severe hail, is the primary source of mites and virus that infect fall-planted winter wheat. Wind-borne movement of the WCM is of key importance in the spread and infection of the virus complex. Significant movement of WCM from wheat has been thought to be closely tied to the senescence or deterioration of the host. Results from field and greenhouse studies indicated that movement from un-vernalized winter wheat was not closely associated with the deterioration of the wheat host. Greenhouse studies showed no correlation between WCM movement and plant condition, but there was a highly significant relationship between WCM movement and mite population on the host plant. Field studies did not demonstrate increased movement associated with deteriorating un-vernalized winter wheat. However, healthier hosts which were able to support a larger population of mites were associated with increased movement. The main influence on the level of mite movement relates to the size of the source population and not the condition of the host plant, but plant condition appears to be a factor in limiting the increase of the WCM population.     

Response of maize (Zea mays L.) lines carrying Wsm1, Wsm2, and Wsm3 to the potyviruses Johnsongrass mosaic virus and Sorghum mosaic virus

Maize dwarf mosaic disease is one of the most important viral diseases of maize (Zea mays L.) throughout the world. It is caused by several virus species in the family Potyviridae, genus Potyvirus, including Maize dwarf mosaic virus (MDMV), Sugarcane mosaic virus (SCMV), Johnsongrass mosaic virus (JGMV) and Sorghum mosaic virus (SrMV). Resistance to another member of the family Potyviridae, Wheat streak mosaic virus (WSMV), is conferred by three alleles (Wsm1, Wsm2, Wsm3) in the maize inbred line Pa405, and these or closely linked genes were previously shown to confer resistance to the potyviruses MDMV and SCMV. In this study, we assessed whether Wsm alleles are linked to resistance to JGMV and SrMV. Near isogenic lines (NILs) carrying one or two of the Wsm alleles introgressed into the susceptible line Oh28 and F1 progeny from NIL × Oh28 were tested for their response to JGMV and SrMV. Our results indicate that Wsm1 provides resistance to both JGMV and SrMV in a dose-dependent manner. Wsm2 and Wsm3 each provide limited resistance, and combining Wsm alleles enhances that resistance.     

Mapping and KASP marker development for wheat curl mite resistance in “TAM 112” wheat using linkage and association analysis

Wheat curl mite (WCM, Aceria tosichella Keifer) and WCM-transmitted wheat streak mosaic virus (WSMV, genus Tritimovirus) are devastating production constraints for wheat in the US Great Plains. Breeding wheat cultivars with genetic resistance to WCM and WSMV is a viable and economically feasible way to reduce yield loss. The objectives of this study were to (a) identify tightly linked markers for WCM resistance in the wheat cultivar TAM 112 (PI 643143) using linkage and association analysis with the 90K Infinium iSelect SNP array and genotyping-by-sequencing, respectively and (b) develop and test kompetitive allele specific PCR (KASP) single-nucleotide polymorphisms (SNPs) for marker-assisted selection (MAS) of WCM resistance. We tested 124 F_5:7 recombinant inbred lines (RILs) derived from the cross of TAM 112 and the WCM-susceptible cultivar TAM 111 (PI 631352). All lines were infested with a Texas WCM collection 2 (TWCMC2) that is virulent to resistance found on the wheat-rye 1AL.1RS translocation at the two-leaf stage and were rated for symptoms on the first and second week after infestation. Linkage maps were constructed with 4890 markers, including SNPs, simple sequence repeats (SSRs), and sequence-tagged site (STS) markers covering 21 chromosomes. A WCM resistance gene present in TAM 112 (Cmc_TAM112) was mapped onto chromosome arm 6DS. A genome-wide association study of wheat streak mosaic (WSM) symptoms from a separate experiment in Colorado showed significant marker-trait associations at the target regions on 6DS where Cmc_TAM112 was located, which demonstrated the effectiveness of this gene to reduce symptom severity. Four SNPs flanking Cmc_TAM112 were mapped within 3.6 cM in the biparental mapping population. We developed two KASP markers that are within 1.3 cM distal to Cmc_TAM112 and tested in diverse germplasm. These two markers can be used in MAS for improving WCM resistance in some wheat genetic backgrounds.     

Wheat curl mite, Aceria tosichella, and transmitted viruses: an expanding pest complex affecting cereal crops

The wheat curl mite (WCM), Aceria tosichella, and the plant viruses it transmits represent an invasive mite-virus complex that has affected cereal crops worldwide. The main damage caused by WCM comes from its ability to transmit and spread multiple damaging viruses to cereal crops, with Wheat streak mosaic virus (WSMV) and Wheat mosaic virus (WMoV) being the most important. Although WCM and transmitted viruses have been of concern to cereal growers and researchers for at least six decades, they continue to represent a challenge. In older affected areas, for example in North America, this mite-virus complex still has significant economic impact. In Australia and South America, where this problem has only emerged in the last decade, it represents a new threat to winter cereal production. The difficulties encountered in making progress towards managing WCM and its transmitted viruses stem from the complexity of the pathosystem. The most effective methods for minimizing losses from WCM transmitted viruses in cereal crops have previously focused on cultural and plant resistance methods. This paper brings together information on biological and ecological aspects of WCM, including its taxonomic status, occurrence, host plant range, damage symptoms and economic impact. Information about the main viruses transmitted by WCM is also included and the epidemiological relationships involved in this vectored complex of viruses are also addressed. Management strategies that have been directed at this mite-virus complex are presented, including plant resistance, its history, difficulties and advances. Current research perspectives to address this invasive mite-virus complex and minimize cereal crop losses worldwide are also discussed.     

A plant virus vector for systemic expression of foreign genes in cereals

Inserts bearing the coding sequences of NPT II and beta-glucuronidase (GUS) were placed between the nuclear inclusion b (NIb) and coat protein (CP) domains of the wheat streak mosaic virus (WSMV) polyprotein ORF. The WSMV NIb-CP junction containing the nuclear inclusion a (NIa) protease cleavage site was duplicated, permitting excision of foreign protein domains from the viral polyprotein. Wheat, barley, oat and maize seedlings supported systemic infection of WSMV bearing NPT II. The NPT II insert was stable for at least 18-30 days post-inoculation and had little effect on WSMV CP accumulation. Histochemical assays indicated the presence of functional GUS protein in systemically infected wheat and barley plants inoculated with WSMV bearing GUS. The GUS constructs had greatly reduced virulence on both oat and maize. RT-PCR indicated that the GUS insert was subject to deletion, particularly when expressed as a GUS-NIb protein fusion. Both reporter genes were expressed in wheat roots at levels comparable to those observed in leaves. These results clearly demonstrate the utility of WSMV as a transient gene expression vector for grass species, including two important grain crops, wheat and maize. The results further indicate that both host species and the nature of inserted sequences affect the stability and expression of foreign genes delivered by engineered virus genomes.     

Evidence that soilborne wheat mosaic virus moves long distance through the xylem in wheat

Summary Soilborne wheat mosaic virus (SBWMV) is a member of the genusFurovirus of plant viruses. SBWMV is transmitted to wheat roots by the plasmodiophorid vectorPolymyxa graminis. Experiments were conducted to determine the path for SBWMV transport from roots to leaves. The results of immunogold labeling suggest that SBWMV enters and moves long distance through the xylem. SBWMV may enter primary xylem elements before cell death occurs and then move upward in the plant after the xylem has matured into hollow vessels. There is also evidence for lateral movement between adjacent xylem vessels.Abbreviations SBWMV Soilborne wheat mosaic virus - TMV Tobacco mosaic virus - BMV Brome mosaic virus - PMTV Potato mop-top virus - BNYVV Beet necrotic yellow vein virus - WSSMV Wheat spindle streak mosaic virus - WSMV Wheat streak mosaic virus     

High Levels of Resistance in Agropyron Species to Barley Yellow Dwarf and Wheat Streak Mosaic Viruses

Several Agropyron species were tested for new sources of resistance to barley yellow dwarf virus (Bydv ) and wheat streak mosaic virus (WSMV). With BYDV strain PAV, 11 of the 17 Agropyron species showed no virus transmission when plants were given access feed by viruliferous Rhopalosiphum padi. Similar trials with BYDV strain RMV (vectored by R. maidis) indicated that all plants, except susceptible control plants, remained virus free. Virus status was confirmed by enzyme-linked immunosorbent assays. When plants were mechanically inoculated with WSMV, 11 Agropyron species failed to express symptoms, while five other species showed a segregating response or had some accessions segregating and some resistant. Test results suggest that resistance to BYDV and WSMV in Agropyron species does not appear to be correlated with any specific genome of Agropyron species although most of the Agropyron species containing S genome were resistant to BYDV and WSMV.     

Resistance to wheat streak mosaic virus in transgenic wheat expressing the viral replicase (NIb) gene

Wheat (Triticum aestivum L. cv. Hi-Line) immature embryos were transformed with the replicase gene (NIb) of wheat streak mosaic virus (WSMV) by the biolistic method. Six independent transgenic plant lines were analyzed for transgene expression and for resistance to mechanical inoculation of WSMV at R3 or R4 generation. Four out of the six lines showed various degree of resistance to WSMV. These lines had initially milder symptoms than controls, and the new growth ranged from milder symptoms, a substantial delay in symptom development, or asymptomatic. Two lines displayed higher resistance with very mild virus symptoms after inoculation and the new growth of 72% and 32% plants from these lines were asymptomatic and had no detectable virus through the plant life cycle. Interestingly, five out of the six transgenic lines had no detectable transgene mRNA expression by RNA gel blot hybridization. The only line that had detectable transgene mRNA did not show delay in the symptom development but had overall milder symptom to the virus.     

Resistance to wheat streak mosaic virus in transgenic wheat engineered with the viral coat protein gene

Wheat (Triticum aestivum) plants were stably transformed with the coat protein (CP) gene of wheat streak mosaic virus (WSMV) by the biolistic method. Eleven independently transformed plant lines were obtained and five were analyzed for gene expression and resistance to WSMV. One line showed high resistance to inoculations of two WSMV strains. This line had milder symptoms and lower virus titer than control plants after inoculation. After infection, new growth did not show symptoms. The observed resistance was similar to the recovery type resistance described previously using WSMV NIb transgene and in other systems. This line looked morphologically normal but had an unusually high transgene copy number (approximately 90 copies per 2C homozygous genome). Northern hybridization analysis indicated a high level of degraded CP mRNA expression. However, no coat protein expression was detected.     

Wheat curl mite and dry bulb mite: untangling a taxonomic conundrum through a multidisciplinary approach

Two economically important eriophyoid mites, Aceria tosichella (wheat curl mite; WCM) and Aceria tulipae (dry bulb mite; DBM), were frequently confounded in the world literature until the late 20th Century. Their morphological similarity and ambiguous data from plant‐transfer and virus‐transmission trials contributed to this confusion. Until recently, there was a general lack of knowledge about the existence of species complexes and it was not possible to accurately genotype tested mites. In the present study, two WCM genotypes of divergent host specificity (MT‐1 and MT‐2) and one DBM genotype were tested for the acceptance of Poaceae, Amarylidaceae, and Liliaceae species that were reported or suspected as hosts of WCM or DBM. The MT‐1 lineage colonized all tested plants. Onion‐ and garlic‐associated DBM populations did not colonize tulip and wild garlic, suggesting that host‐acceptance variability exists within A. tulipae s.l. Morphometric analysis did not discriminate closely‐related MT‐1 and MT‐2 genotypes but completely separated both WCM genotypes from DBM based on the larger overall body size of the latter. Three morphological traits combined to discriminate between the DBM and MT‐1 genotypes, both of which can infest Amarylidaceae bulbs. In total, these combined DNA sequence, host‐acceptance, morphometrical results unambiguously separated two WCM and one DBM genotypes. Similar studies on additional lineages of both WCM and DBM should ultimately dispel previous taxonomic confusion between these two species. © 2013 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 111 , 421–436.     

Molecular cloning, sequencing, and phylogenetic relationships of a new potyvirus: sugarcane streak mosaic virus, and a reevaluation of the classification of the potyviridae

The nucleic acid of a serologically distinct potyvirus, originally isolated out of sugar cane from Pakistan, was reverse transcribed and the 3' terminal 2000 bp was PCR amplified, cloned, and sequenced. Phylogenetic comparisons of viruses representing each genus of the Potyviridae show that the Pakistani isolate is most closely related to the rymoviruses wheat streak mosaic virus (WSMV) and brome streak mosaic virus. We therefore propose that this new virus species be named sugar cane streak mosaic virus to reflect its similarity to WSMV. The phylogenetic data also show that the genus Rymovirus contains at least two unique evolutionary lineages. Thus the current taxonomy, based on transmission vector, is paraphyletic. We present an analysis of the taxonomic relationships among members of the family and propose a classification that both resolves the paraphyly and more accurately represents the evolutionary history of the Potyviridae.