Anders Kvarnheden

Cotton leaf curl disease – an emerging threat to cotton production worldwide

Cotton leaf curl disease (CLCuD) is a serious disease of cotton which has characteristic symptoms, the most unusual of which is the formation of leaf-like enations on the undersides of leaves. The disease is caused by whitefly-transmitted geminiviruses (family Geminiviridae, genus Begomovirus) in association with specific, symptom-modulating satellites (betasatellites) and an evolutionarily distinct group of satellite-like molecules known as alphasatellites. CLCuD occurs across Africa as well as in Pakistan and north-western India. Over the past 25 years, Pakistan and India have experienced two epidemics of the disease, the most recent of which involved a virus and satellite that are resistance breaking. Loss of this conventional host-plant resistance, which saved the cotton growers from ruin in the late 1990s, leaves farmers with only relatively poor host plant tolerance to counter the extensive losses the disease causes. There has always been the fear that CLCuD could spread from the relatively limited geographical range it encompasses at present to other cotton-growing areas of the world where, although the disease is not present, the environmental conditions are suitable for its establishment and the whitefly vector occurs. Unfortunately recent events have shown this fear to be well founded, with CLCuD making its first appearance in China. Here, we outline recent advances made in understanding the molecular biology of the components of the disease complex, their interactions with host plants, as well as efforts being made to control CLCuD.

The impact of global warming on plant diseases and insect vectors in Sweden

Cold winters and geographic isolation have hitherto protected the Nordic countries from many plant pathogens and insect pests, leading to a comparatively low input of pesticides. The changing climate is projected to lead to a greater rise in temperature in this region, compared to the global mean. In Scandinavia, a milder and more humid climate implies extended growing seasons and possibilities to introduce new crops, but also opportunities for crop pests and pathogens to thrive in the absence of long cold periods. Increased temperatures, changed precipitation patterns and new cultivation practices may lead to a dramatic change in crop health. Examples of diseases and insect pest problems predicted to increase in incidence and severity due to global warming are discussed.

Sequence characterization of Tomato leaf curl Sinaloa virus and Tomato severe leaf curl virus: Phylogeny of New World begomoviruses and detection of recombination

Summary.Diseases caused by begomoviruses (family Geminiviridae, genus Begomovirus) constitute a serious constraint to tomato production in Nicaragua. In this study, the complete nucleotide (nt) sequences of the DNA-A and DNA-B components were determined for the first time for Tomato leaf curl Sinaloa virus (ToLCSinV). In addition, the complete nt sequence was determined for the DNA-A component of two isolates of Tomato severe leaf curl virus (ToSLCV). The genome organization of ToLCSinV and ToSLCV was identical to the bipartite genomes of other begomoviruses described from the Americas. A phylogenetic analysis of DNA-A including 45 begomovirus species showed that the indigenous begomoviruses of the New World can be divided into three major clades and an intermediate group: AbMV clade, SLCV clade, “Brazil clade”, and BGYMV group. Phylogenetic analyses of the DNA-A and DNA-B components and their open reading frames indicated that ToLCSinV and ToSLCV belong to different clades: ToLCSinV to the AbMV clade, and ToSLCV to the SLCV clade. The two Nicaraguan isolates of ToSLCV showed a close relationship with ToSLCV from Guatemala (ToSLCV-[GT96-1]) and Tomato chino La Paz virus (ToChLPV), but differed significantly in the AV1 and AC1 regions, respectively. Computer-based predictions indicated that recombination with another begomovirus had taken place within AV1 of ToSLCV dividing this species into two strains. A high probability was also found that ToChLPV is involved in the evolution of ToSLCV.

The complete genome sequence for a Turkish isolate of Wheat dwarf virus (WDV) from barley confirms the presence of two distinct WDV strains

The complete genome for a barley isolate of Wheat dwarf virus (WDV) from Tekirdağ, Turkey, WDV-Bar[TR], was isolated and sequenced. The genome was found to be 2739 nucleotides long, which is shorter than wheat-infecting WDV isolates, and with a genome organization typical for mastreviruses. The complete genome of WDV-Bar[TR] showed 83–84% nucleotide identity to wheat isolates of WDV, with the non-coding regions SIR and LIR least conserved (72–74% identity). The deduced amino acid sequences for Rep and RepA were most conserved (92–93%), while CP and MP were less conserved (87% and 79–80%, respectively). The identity to other mastrevirus species was significantly lower. In phylogenetic analyses, the WDV isolates formed a distinct clade, well separated from the other mastreviruses with the wheat isolates grouping closely together. Phylogenetic analyses of WDV-Bar[TR], the partial sequence for another Turkish barley isolate (WDV-Bar[TR2]) and published WDV sequences further supported the division of WDV into two distinct strains. The barley strain could also be divided into three subtypes based on relationships and geographic origin. This study shows the first complete published sequence for a barley isolate of WDV.

Geminiviruses Infecting Tomato Crops in Nicaragua

Geminiviruses transmitted by whiteflies are believed to be responsible for the devastating epidemic in tomato crops in Nicaragua, as well as in other Central American countries. Polymerase chain reaction with degenerate primers was used to amplify partial sequences of the geminivirus coat protein gene from samples of diseased tomato plants collected from the major tomato-growing areas of Nicaragua. The data indicated the presence of geminiviruses in all tested regions of the country. DNA sequence analysis and phylogenetic analysis of the amplified sequences showed that they corresponded to four different geminiviruses related to the other begomoviruses native to the Americas. One of the viruses, which was detected in three regions of Nicaragua, is probably Sinaloa tomato leaf curl virus. The sequences of two of the other detected viruses showed close relationships with several geminiviruses, including Tomato mottle virus, Tomato leaf crumple virus, and Sida golden mosaic virus, all of which previously have been reported from Central America. The fourth virus is closely related at sequence level to a tomato-infecting geminivirus from Honduras, putatively designated Tomato mild mottle virus. This virus seems to be different from the other known American begomoviruses because it groups separately in the phylogenetic analysis.

Molecular diversity of the coat protein-encoding region of Barley yellow dwarf virus-PAV and Barley yellow dwarf virus-MAV from Latvia and Sweden ∗

Summary.The sequence variability of Barley yellow dwarf virus-PAV (PAV) and Barley yellow dwarf virus-MAV (MAV) was studied by comparing 502 nucleotides from the coat protein-encoding region of six isolates from Latvia and four from Sweden. The diversity within MAV was low (>97% sequence identity), also when compared to isolates from USA and China. In contrast, the variability among PAV isolates was greater and phylogenetic analysis including isolates of a wide geographic origin detected two major clusters, of which both contained isolates from Latvia and Sweden. A new distinct variant of BYDV-PAV was discovered in Latvia, and because of the sequence difference it is proposed to belong to a new species (BYDV-OYV).

Begomovirus disease complex: emerging threat to vegetable production systems of West and Central Africa

Vegetables play a major role in the livelihoods of the rural poor in Africa. Among major constraints to vegetable production worldwide are diseases caused by a group of viruses belonging to the genus Begomovirus, family Geminiviridae. Begomoviruses are plant-infecting viruses, which are transmitted by the whitefly vector Bemisia tabaci and have been known to cause extreme yield reduction in a number of economically important vegetables around the world. Several begomoviruses have been detected infecting vegetable crops in West and Central Africa (WCA). Small single stranded circular molecules, alphasatellites and betasatellites, which are about half the size of their helper begomovirus genome, have also been detected in plants infected by begomoviruses. In WCA, B. tabaci has been associated with suspected begomovirus infections in many vegetable crops and weed species. Sequencing of viral genomes from crops such as okra resulted in the identification of two previously known begomovirus species (Cotton leaf curl Gezira virus and Okra yellow crinkle virus) as well as a new recombinant begomovirus species (Okra leaf curl Cameroon virus), a betasatellite (Cotton leaf curl Gezira betasatellite) and new alphasatellites. Tomato and pepper plants with leaf curling were shown to contain isolates of new begomoviruses, collectively referred to as West African tomato-infecting begomoviruses (WATIBs), new alphasatellites and betasatellites. To study the potential of weeds serving as begomovirus reservoirs, begomoviruses and satellites in the weed Ageratum conyzoides were characterized. Sequence analyses showed that they were infected by isolates of a new begomovirus (Ageratum leaf curl Cameroon virus) that belong to the WATIBs group, a new betasatellite (Ageratum leaf curl Cameroon betasatellite), an alphasatellite and two types of defective recombinants between a begomovirus and an alphasatellite. Putative recombinations were detected in begomovirus genomes for all four plant species studied, indicating that recombination is an important mechanism for their evolution. A close relationship between the begomoviruses infecting pepper and tomato and A. conyzoides and the detection of the same alphasatellite in them support the idea that weeds are important reservoirs for begomoviruses and their satellites. With this high diversity, recombination potential and transmission by B. tabaci, begomoviruses and ssDNA satellites pose a serious threat to crop production in West and Central Africa.

Genetic diversity of Wheat dwarf virus.

Summary. The complete sequence of a new wheat-infecting isolate of Wheat dwarf virus from Sweden (WDV-[Enk1]) was determined, as well as a 726-nt region covering part of rep and the long intergenic region (LIR) of six other wheat-infecting Swedish isolates and a barley-infecting isolate from Hungary (WDV-Bar[HU]). Analyses including these and previously published sequences showed that the wheat-infecting isolates of WDV displayed less than 3% of divergence. Most of the nucleotide changes were silent and the largest variation was detected in LIR. In contrast, the barley-infecting isolate was clearly different with 16% of sequence divergence compared to the wheat isolates. The WDV isolates infecting barley or wheat seem to represent two differentiated strains.

Effects of the mutation of selected genes of cotton leaf curl Kokhran virus on infectivity, symptoms and the maintenance of cotton leaf curl Multan betasatellite

Cotton leaf curl Kokhran virus (CLCuKoV) is a cotton-infecting monopartite begomovirus (family Geminiviridae). The effects of mutation of the coat protein (CP), V2, C2 and C4 genes of CLCuKoV on infectivity and symptoms in Nicotiana benthamiana were investigated. Each mutation introduced a premature stop codon which would lead to premature termination of translation of the gene. Mutation of the CP gene abolished infectivity. However, transient expression of the CLCuKoV CP gene under the control of the Cauliflower mosaic virus 35S promoter (35S-Ko(CP)), at the point of inoculation, led to a small number of plants in which viral DNA could be detected by PCR in tissues distal to the inoculation site. Mutations of the V2, C2 and C4 genes reduced infectivity. The V2 and C2 mutants did not induce symptoms, whereas the C4 mutation was associated with attenuated symptoms. Infections of plants with the C4 mutant were associated with viral DNA levels equivalent to the wild-type virus, whereas viral DNA levels for the V2 mutant were low, detectable by Southern blot hybridisation, and for the C2 mutant were detectable only by PCR. Significantly, transient expression of the CLCuKoV C2 gene at the point of inoculation, raised virus DNA levels in tissues distal to the inoculation site such that they could be detected by Southern hybridisation, although they remained at well below the levels seen for the wild-type virus, but reduced the infectivity of the virus. These findings are consistent with earlier mutation studies of monopartite begomoviruses and our present knowledge concerning the functions of the four genes suggesting that the CP is essential for long distance spread of the virus in plants, the C4 is involved in modulating symptoms, the C2 interferes with host defence and the V2 is involved in virus movement. The results also suggest that the V2, C2 and C4 may be pathogenicity determinants. Additionally the effects of the mutations of CLCuKoV genes on infections of the virus in the presence of its cognate betasatellite, Cotton leaf curl Multan betasatellite (CLCuMuB), were investigated. Mutation of the C4 gene had no effect on maintenance of the betasatellite, although the betasatellite enhanced symptoms. Inoculation of the C2 mutant with CLCuMuB raised the infectivity of the virus to near wild-type levels, although the numbers of plants in which the betasatellite was maintained was reduced, in comparison to wild-type virus infections with CLCuMuB, and viral DNA could not be detected by Southern hybridisation. Transient expression of the C2 gene at the point of inoculation raised virus DNA levels in tissues distal to the inoculation site but also reduced the infectivity of the virus and the numbers of plants in which the betasatellite was maintained. CLCuMuB restored the infectivity of the V2 mutant to wild-type levels but only in a small number of plants was the satellite maintained and infections were non-symptomatic. Although inoculation of the CP mutant with CLCuMuB did not restore infectivity, co-inoculation with 35S-Ko(CP) increased the number of plants in which the virus could be detected, in comparison to plants inoculated with the mutant and 35S-Ko(CP), and also resulted in two plants (out of 15 inoculated) in which the betasatellite could be detected by PCR. This indicates that the V2, C2 and almost certainly the CP are important for the maintenance of betasatellites by monopartite begomoviruses. The significance of these findings is discussed.

Ageratum conyzoides: A host to a unique begomovirus disease complex in Cameroon

Ageratum conyzoides (goat weed) is a widespread uncultivated species in Cameroon that exhibits leaf curl disease (LCD) symptoms suggestive of begomovirus infection. In Asia, different begomovirus-satellite complexes have been identified in A. conyzoides. The objective of this study was to determine the identity of the suspect begomoviruses and their associated satellites in A. conyzoides in Cameroon. The results indicated that all three symptomatic A. conyzoides plants examined were infected with a new begomovirus species, herein named Ageratum leaf curl Cameroon virus (ALCCMV). The ALCCMV genome sequences shared their highest identity, at 84.3-88.5%, with a group of tomato-infecting begomoviruses from West Africa. In addition, a betasatellite and an alphasatellite were cloned from the same symptomatic A. conyzoides plants. The betasatellite sequences shared limited sequence identity at 37% or less with the betasatellite Cotton leaf curl Gezira betasatellite, and the new betasatellite species is herein named Ageratum leaf curl Cameroon betasatellite (ALCCMB). The alphasatellite shared 80% nt identity with Tomato leaf curl Cameroon alphasatellite (ToLCCMA), and the new alphasatellite species is herein named Ageratum leaf curl Cameroon alphasatellite (ALCCMA). In addition, two fragments containing begomovirus-alphasatellite sequences were cloned from sample AGLI4, and they were related to the defecting interfering molecule (Y14167) associated with Ageratum yellow vein virus from Asia. These results suggest that the begomoviral-satellite complexes infecting A. conyzoides in Cameroon may be as complex or more so, to species and strains reported thus far from Asia.