Crop borders reduce potato virus Y incidence in seed potato

Crop borders of soybean (Glycine max), sorghum (Sorghum bicolor), winter wheat (Triticum aestivum) and potato (Solanum tuberosum) were tested as a means of reducing potato virus Y (PVY) incidence in seed potato. Borders of fallow cultivated ground served as controls. Aphid landing rates were monitored weekly in plots using green tile traps, and PVY incidence was assessed by serologically testing tuber progeny from selected rows in each plot. Average weekly aphid landing rates in fallow-bordered and crop-bordered plots were not significantly different in 1992 (29.4 and 25.2 aphids, respectively) or 1993 (7.3 and 6.6 aphids, respectively). However, crop borders significantly reduced PVY incidence. In 1992, fallow-bordered and soybean-bordered plots averaged 47.8% and 35.0% PVY infection, respectively. In 1993, PVY infection averaged across all crop (soybean, sorghum, and wheat) bordered plots was 2.7% compared to 6.8% in fallow-bordered plots. PVY incidence in the centre rows of fallow-bordered and crop-bordered plots was statistically equivalent, while outer rows of crop-bordered plots had significantly less PVY than outer rows of fallow-bordered plots. Crop borders apparently reduced the number of viruliferous aphids landing on the edge of the plot. The choice of crop species used as a border, or treating the border with a systemic insecticide, did not affect aphid landing rates or PVY incidence. In 1995, PVY incidence in the centre 10 row block of potatoes averaged 2.1% across all crop borders (potato and soybean). PVY infection in the four row potato border averaged 5.7%. Crop borders are readily adaptable to current production practices, although the greatest benefits in reducing PVY incidence would occur in average sized, generation 0 (< 0.2 ha), elite seed potato fields.     

Limited degree of serological variability in pepper strains of potato virus Y as revealed by analysis with monoclonal antibodies

The degree of serological variability among pepper strains of potato virus Y (PVY) was assessed through the analysis of samples of infected pepper collected in three main pepper producing regions of Spain. Samples corresponding to the period 1980–1991 were analysed by ELISA with five different monoclonal antibodies (MAbs) produced against potato strains of the virus. The results obtained show a limited degree of epitope variability among pepper PVY-isolates, since only eight out of 32 possible serological profiles were found. Most isolates are not recognised by a MAb directed towards an epitope reported to be present in all potato-PVY isolates. The overall serological behaviour of pepper isolates with these MAbs places them as closer to the group O, of the three groups into which the potato isolates of PVY have been subdivided.     

Efficacy of pymetrozine against Myzus persicae and in reducing potato virus Y transmission on tobacco plants

Forty‐four parthenogenetic lineages of Myzus persicae s.l. (Sulzer) from tobacco crops and peach orchards located in various regions of Greece were examined to determine their response to the insecticide pymetrozine using leaf‐dip bio‐assays. The results show that the aphid has not developed resistance, as all lineages exhibited resistance factors bellow 6.0. In transmission experiments of potato virus Y (PVY) using a lineage of the tobacco‐adapted subspecies M. persicae nicotianae Blackman on tobacco plants, one foliar application with pymetrozine provided adequate protection for 7 days. Pymetrozine significantly reduced both virus acquisition and inoculation compared with the untreated control and the reduction was comparable to a mineral oil application. These results are discussed in terms of the advantage of incorporating pymetrozine as a compound of pest management strategies against M. persicae s.l. and for control of non‐persistent viruses, especially in crops such as tobacco because of the high selection pressure from neonicotinoids resulting in potential of resistance developing in aphid populations.     

The effect of mineral oil on stylet activities and potato virus Y transmission by aphids

Mineral oil sprayed onto potato virus Y (PVY) infected tobacco plants reduced acquisition of this potyvirus by Myzus persicae (Sulz.). Although the pre-penetration activities of aphids were longer on oil treated leaves, the inhibitory effect of the oil could not be attributed to differences in the duration of stylet penetration. Aphids were therefore made part of a DC circuit in order to investigate their stylet activities during penetration of PVY infected source plants and healthy test plants. Both acquisition and inoculation of the virus were reduced by the presence of oil on the plant surface, but these reductions could not be related to electrically recorded differences in plant penetration behaviour. In particular, stylet punctures of plant cell membranes were not reduced by mineral oil. Non-behavioural reasons are suggested to explain the mode of action of the oil.     

Behavior of bird cherry-oat aphid and green peach aphid in relation to potato virus Y transmission

Potato virus Y is transmitted to potato in a nonpersistent manner by many aphid species, some of which do not colonize this crop. The behavior of bird cherry-oat aphid, Rhopalosiphum padi (L.) on potato, Solanum tuberosum L., a plant species that is not colonized by this aphid, was described and compared with that of the potato-colonizing green peach aphid, Myzuspersicae (Sulzer). A higher proportion of winged morph of R. padi than M. persicae left the plant, but aphids that stayed in contact with the plant took the same mean time to initiate the first probe and it lasted the same mean time compared with M. persicae. Electronic penetration graph technique was used to study the probing behavior of the aphids during Potato virus Y (family Potyviridae, genus Potyvirus, PVY) transmission tests. Transmission rate decreased from 29 to 8% when the acquisition time increased from 5 min of continuous probing to 1 h with M. persicae, but it remained low (2 and 1%) with R. padi. Most of the difference in transmission rate between acquisition time with M. persicae and between aphid species was related to the change in the time and behavior taking place between the last cell puncture of the acquisition phase to the first cell puncture of the inoculation phase. Results presented here clearly demonstrated the importance of host plant selection and probing behavior in the transmission of nonpersistent plant viruses. They also stress the need to consider the behavior of the aphid in the design of laboratory tests of virus vector efficacy.     

Determination of aphid transmission efficiencies for N, NTN and Wilga strains of Potato virus Y

Potato virus Y (PVY, genus Potyvirus, family Potyviridae) causes high economic losses worldwide, especially in the production of seed potatoes (Solanum tuberosum). PVY control systems rely on measuring virus pressure and vector pressure in the field. Calculation of the vector pressure is based on the relative efficiency factors (REFs) of aphid species. These REFs express the transmission efficiency of aphid species in relation to the transmission efficiency of Myzus persicae, the most efficient vector of PVY. In this paper, we report on the determination of aphids' relative transmission efficiency factors (REFs) for isolates of the PVY strains PVY^N, PVY^NTN and PVY^N-Wi. Biotype Mp2 of M. persicae was tested for its transmission efficiency for six PVY isolates (one PVY^N, three PVY^NTN and two PVY^N-Wi isolates) and showed comparable average transmission efficiencies for all isolates. The transmission rate of this biotype for the six PVY isolates was set to 1 and Mp2 was used as an internal control in transmission experiments to determine the REFs of three other biotypes of M. persicae and 16 other aphid species (three biotypes per species when available) for the six PVY isolates. Comparing the calculated REFs for PVY^N with the REFs reported in the previous century for PVY^N, we observe overall comparable REFs, except for Aphis fabae, Aphis spp., Hyperomyzus lactucae, Macrosiphum euphorbiae and Rhopalosiphum padi, which have a lower REF in our experiments, and Aphis frangulae and Phorodon humuli, which have now a higher REF. Comparing the new REFs found for the PVY^NTN strains with the new REFs for PVY^N, we observe that they are overall comparable, except for A. frangulae (0.17 compared with 0.53) and Schizaphis graminum (0.05 compared with 0.00). Comparing the REFs calculated for PVY^N-Wi with those calculated for PVY^N, we can observe six aphid species with higher REFs (Acyrthosiphon pisum, A. fabae, Aphis nasturtii, Aphis spp., P. humuli and R. padi). Only the species A. frangulae shows a lower REF for PVY^N-Wi compared with the transmission efficiency of PVY^N. Three aphid species (Aulacorthum solani, Myzus ascalonicus and S. graminum) for which no REF was determined earlier were found to be capable to transmit PVY and their REFs were determined.     

Comparison of transmission efficiency of various isolates of Potato virus Y among three aphid vectors

Potato virus Y (PVY) strains are transmitted by different aphid species in a non‐persistent, non‐circulative manner. Green peach aphid (GPA), Myzus persicae Sulzer, is the most efficient vector in laboratory studies, but potato aphid (PA), Macrosiphum euphorbiae Thomas (both Hemiptera: Aphididae, Macrosiphini), and bird cherry‐oat aphid (BCOA), Rhopalosiphum padi L. (Hemiptera: Aphididae, Aphidini), also contribute to PVY transmission. Studies were conducted with GPA, PA, and BCOA to assess PVY transmission efficiency for various isolates of the same strain. Treatments included three PVY strains (PVY^O, PVY^N:O, PVY^NTN) and two isolates of each strain (Oz and NY090031 for PVY^O; Alt and NY090004 for PVY^N:O; N4 and NY090029 for PVY^NTN), using each of three aphid species as well as a sham inoculation. Virus‐free tissue‐cultured plantlets of potato cv. Russet Burbank were used as virus source and recipient plants. Five weeks post inoculation, recipient plants were tested with quantitative DAS‐ELISA to assess infection percentage and virus titer. ELISA‐positive recipient plants were assayed with RT‐PCR to confirm presence of the expected strains. Transmission efficiency (percentage infection of plants) was highest for GPA, intermediate for BCOA, and lowest for PA. For all aphid species, transmission efficiency did not differ significantly between isolates within each strain. No correlations were found among source plant titer, infection percentage, and recipient plant titer. For both GPA and BCOA, isolates of PVY^NTN were transmitted with greatest efficiency followed by isolates of PVY^O and PVY^N:O, which might help explain the increasing prevalence of necrotic strains in potato‐growing regions. Bird cherry‐oat aphid transmitted PVY with higher efficiency than previously reported, suggesting that this species is more important to PVY epidemiology than has been considered.     

Bacillus thuringiensis and neem seed oil (Azadirachta indica) effects on the potato tuber moth Phthorimaea operculella zeller in the field and stores

A comparative evaluation for the efficacy of Bacillus thuringiensis and neem seed oil on Phthorimaea operculella has been carried out in the field and store. These two preparations were almost equally effective on the potato tuber moth infestation. The percentage of infestation was reduced through successive application of either preparations in the field up to harvest. No synergism was observed upon using combination of the two preparations. In the store, neem seed oil (500 ppm) was highly protective and was as effective as sevin. A combination of both neem and B.t. (Delfin) significantly protects the tubers. This suggests the possible use of either neem seed oil or B.t. in combating the insect pest in the field or during storage.     

Forecasting virus disease in seed potatoes using flight activity data of aphid vectors

We compiled data from the Swiss seed certification programme for the country‐wide incidence of viruses in seed potato crops for the years 1989–2012. Model selection techniques were used to regress year‐to‐year variation in the incidence of potato viruses – largely dominated by Potato virus Y (PVY) – in three susceptible varieties against the abundance of virus vectors (winged aphids), obtained in a suction trap, to identify the most important vector species. The ultimate aim of this study was to develop a decision‐support system capable of forecasting virus spread during the current season using trap data of aphid flights. The average virus incidence in the varieties Bintje, Sirtema and Charlotte varied considerably among years, ranging from 1.0% in 2009 to 13.6% in 1989 (N = 150–611 seed lots per year). A linear regression model including the cumulative sums (until mid‐June) of two aphid species (Brachycaudus helichrysi and Phorodon humuli) as predictor variables for virus disease was remarkably well supported by the data (R² = 0.86). Similarly, using counts of B. helichrysi alone resulted in a good model fit (R² = 0.81). Cross‐validation revealed high predictive accuracy of the model. Although prediction root mean squared errors (RMSE) calculated for different timings of forecasts were high for extremely early forecasts, they rapidly declined for forecasts conducted by the end of May (i.e. 2–4 weeks after potato emergence). Winter temperature (January–February) was positively correlated with the abundance of B. helichrysi in early summer as well as with post‐harvest virus incidence. Remarkably, the abundance of Myzus persicae, often considered the main vector of PVY, was not correlated with virus incidence. Taken together, our analysis suggests that the early migrating aphid B. helichrysi, rather than M. persicae, is the main vector of PVY in Switzerland, and that suction trap data are useful for the design of decision‐support systems aimed to optimise virus control in seed potato production.     

A natural recessive resistance gene against potato virus Y in pepper corresponds to the eukaryotic initiation factor 4E (eIF4E)

We show here that the pvr2 locus in pepper, conferring recessive resistance against strains of potato virus Y (PVY), corresponds to a eukaryotic initiation factor 4E (eIF4E) gene. RFLP analysis on the PVY-susceptible and resistant pepper cultivars, using an eIF4E cDNA from tobacco as probe, revealed perfect map co-segregation between a polymorphism in the eIF4E gene and the pvr2 alleles, pvr2(1) (resistant to PVY-0) and pvr2(2) (resistant to PVY-0 and 1). The cloned pepper eIF4E cDNA encoded a 228 amino acid polypeptide with 70-86% nucleotide sequence identity with other plant eIF4Es. The sequences of eIF4E protein from two PVY-susceptible cultivars were identical and differed from the eIF4E sequences of the two PVY-resistant cultivars Yolo Y (YY) (pvr2(1)) and FloridaVR2 (F) (pvr2(2)) at two amino acids, a mutation common to both resistant genotypes and a second mutation specific to each. Complementation experiments were used to show that the eIF4E gene corresponds to pvr2. Thus, potato virus X-mediated transient expression of eIF4E from susceptible cultivar Yolo Wonder (YW) in the resistant genotype YY resulted in loss of resistance to subsequent PVY-0 inoculation and transient expression of eIF4E from YY (resistant to PVY-0; susceptible to PVY-1) rendered genotype F susceptible to PVY-1. Several lines of evidence indicate that interaction between the potyvirus genome-linked protein (VPg) and eIF4E are important for virus infectivity, suggesting that the recessive resistance could be due to incompatibility between the VPg and eIF4E in the resistant genotype.     

Alfalfa mosaic and cucumber mosaic virus infection in chickpea and lentil: incidence and seed transmission

Samples collected in 1994 and 1995 from commercial crops of chickpeas and lentils growing in the agricultural region of south-west Western Australia were tested for infection with alfalfa mosaic (AMV) and cucumber mosaic (CMV) viruses, and for members of the family Potyviridae using enzyme-linked immunosorbent assay (ELISA). In 1994 no virus was detected in the 21 chickpea crops tested but in 1995, out of 42 crops, AMV was found in two and CMV in seven. With lentils, AMV and/or CMV was found in three out of 14 crops in 1994 and 4 out of 13 in 1995, both viruses being detected in two crops in each year. Similar tests on samples from chickpea and lentil crops and plots growing at experimental sites, revealed more frequent infection with both viruses. No potyvirus infection was found in chickpeas or lentils in agricultural areas either in commercial crops or at experimental sites. However, bean yellow mosaic virus (BYMV) was detected along with AMV and CMV in irrigated plots of chickpeas and lentils at a site in Perth. When samples of seed from infected crops or plots of chickpeas and lentils were germinated and leaves or roots of seedlings tested for virus infection by ELISA, AMV and CMV were found to be seed-borne in both while BYMV was seed-borne in lentils. The rates of transmission found through seed of chickpea to seedlings were 0.1–1% with AMV and 0.1–2% with CMV. Seed transmission rates with lentil were 0.1–5% for AMV, 0.1–1% for CMV and 0.8% for BYMV. Individual seed samples of lentil and chickpea sometimes contained both AMV and CMV. With both species, infection with AMV and CMV was sometimes found in commercial seed stocks or seed stocks from multiplication crops of advanced selections nearing release as new cultivars. Seed-borne virus infection has important practical implications, as virus sources can be re-introduced every year to chickpea and lentil crops or plots through sowing infected seed stocks leading to spread of infection by aphid vectors, losses in grain yield and further contamination of seed stocks.     

Properties of cocksfoot streak and cocksfoot cryptic, two viruses infecting cocksfoot (Dactylis glomerata) in Scotland

A Scottish isolate of cocksfoot streak virus (CSV-S) was found to have flexuous filamentous particles which, in sap of infected cocksfoot plants, had a modal length of 712 nm. It was transmitted from infected to healthy cocksfoot plants in a non-persistent manner by Myzus persicae and by mechanical inoculation of infective sap extracts containing an anti-oxidant. Apart from cocksfoot, mechanical inoculation of infective sap succeeded in infecting only four of 22 plant species tested. The infectivity of sap extracts containing 0.2% thioglycerol was lost after heating for 10 min at 55^oC but not 50^oC, storage at room temperature for 48 but not 24 hours, and after diluting 10^-2to 10^-3. Highly purified preparations of CSV-S particles sedimented as a single component with a sedimentation coefficient of 139S and had a buoyant density in rubidium bromide of 1.31 g/cm³. Virus particles were composed of one protein and one ssRNA species with estimated M_r of 31 000 and 3.2 times 10⁶respectively. In ELISA, an antiserum prepared to CSV-S detected the virus in all aerial parts of infected cocksfoot plants and, when present in the ratio of 1 infected leaf: 1000 healthy leaves. Both CSV-S-infected and -uninfected cocksfoot also contained a previously undescribed virus with isometric particles c. 30 nm in diameter. This virus, named cocksfoot cryptic virus (CCV), was seed-borne in two cvs of cocksfoot tested and its particles contained two dsRNA species of estimated M_r of 1.14 times 10⁶and 1.27 times 10⁶. Despite the fact that particles of CSV-S were largely free from CCV particles following exclusion chromatography on agarose beads prior to immunisation, immunoelectron microscopy (IEM) showed that the antiserum prepared to CSV-S also contained some antibodies to CCV. Evidence from IEM suggested a possible distant serological relationship of CCV to ryegrass and beet (BCV 1 or BCV 2, or both) cryptoviruses, all members of sub-group A of crypto viruses.     

Cauliflower mosaic virus protein P6-TAV plays a major role in alteration of aphid vector feeding behaviour but not performance on infected Arabidopsis

Emerging evidence suggests that viral infection modifies host plant traits that in turn alter behaviour and performance of vectors colonizing the plants in a way conducive for transmission of both nonpersistent and persistent viruses. Similar evidence for semipersistent viruses like cauliflower mosaic virus (CaMV) is scarce. Here we compared the effects of Arabidopsis infection with mild (CM) and severe (JI) CaMV isolates on the feeding behaviour (recorded by the electrical penetration graph technique) and fecundity of the aphid vector Myzus persicae. Compared to mock-inoculated plants, feeding behaviour was altered similarly on CM- and JI-infected plants, but only aphids on JI-infected plants had reduced fecundity. To evaluate the role of the multifunctional CaMV protein P6-TAV, aphid feeding behaviour and fecundity were tested on transgenic Arabidopsis plants expressing wild-type (wt) and mutant versions of P6-TAV. In contrast to viral infection, aphid fecundity was unchanged on all transgenic lines, suggesting that other viral factors compromise fecundity. Aphid feeding behaviour was modified on wt P6-CM-, but not on wt P6-JI-expressing plants. Analysis of plants expressing P6 mutants identified N-terminal P6 domains contributing to modification of feeding behaviour. Taken together, we show that CaMV infection can modify both aphid fecundity and feeding behaviour and that P6 is only involved in the latter.     

Biochemical and physiological mechanisms underlying effects of Cucumber mosaic virus on host‐plant traits that mediate transmission by aphid vectors

The transmission of insect‐vectored diseases entails complex interactions among pathogens, hosts and vectors. Chemistry plays a key role in these interactions; yet, little work has addressed the chemical ecology of insect‐vectored diseases, especially in plant pathosystems. Recently, we documented effects of Cucumber mosaic virus (CMV) on the phenotype of its host (Cucurbita pepo) that influence plant‐aphid interactions and appear conducive to the non‐persistent transmission of this virus. CMV reduces host‐plant quality for aphids, causing rapid vector dispersal. Nevertheless, aphids are attracted to the elevated volatile emissions of CMV‐infected plants. Here, we show that CMV infection (1) disrupts levels of carbohydrates and amino acids in leaf tissue (where aphids initially probe plants and acquire virions) and in the phloem (where long‐term feeding occurs) in ways that reduce plant quality for aphids; (2) causes constitutive up‐regulation of salicylic acid; (3) alters herbivore‐induced jasmonic acid biosynthesis as well as the sensitivity of downstream defences to jasmonic acid; and (4) elevates ethylene emissions and free fatty acid precursors of volatiles. These findings are consistent with previously documented patterns of aphid performance and behaviour and provide a foundation for further exploration of the genetic mechanisms responsible for these effects and the evolutionary processes that shape them.     

Incidence of potato virus Y infection in seed and ware tubers of the potato cv. Record

The incidence of potato virus Y (PVY) infection was assessed in samples of potato tubers, cv. Record, taken from Scottish seed stocks and English ware crops grown from some of these seed stocks. PVY was readily detected by ELISA of tuber sprouts. PVY-infected tubers were found in 10 seed stocks of 84 tested. The mean level of virus infection was 0.23%, 0.76% and 0.56% in Super Elite, Elite and AA stocks respectively. In 46 commercial ware crops grown from some of these seed stocks, a substantial proportion of the harvested tubers in all but one of the crops were infected with PVY, the mean percentage of infected tubers was 58.5%. Ware crops grown from seven seed stocks in which PVY had been detected (mean 6.2% infection in seed) contained a mean of 70% infected tubers, compared with 56% infection in crops grown from 39 stocks in which PVY was not detected in the seed tubers. The predominant PVY strain detected in the ware crops was the veinal necrosis strain (PVY^vn).     

Dissemination and detection of peanut clump virus in groundnut seed

Experiments were done to assess the role of seed-transmission in the dissemination of peanut clump virus (PCV) in groundnut (Arachis hypogea L.), and the usefulness of enzyme-linked immunosorbent assay (ELISA) for detecting the virus in infected groundnut seed. The virus was present in 7.5% of seedling progeny from infected plants and could be detected in 16.5% of the seeds by ELISA. When groundnut seeds were grown in a field contaminated by the virus, it was shown that by roguing the infected plants, only 0.1% of the seeds from the remaining plants contained the virus. It was also established that the level of contamination of seeds by the virus was inversely proportional to the seed size.     

The role of flying aphid vectors in the transmission of cucumber mosaic virus and potato virus Y to peppers in Israel

More than 44 species of aphids were trapped by suction during the spring seasons of 1981, 1982 and 1983 over a pepper field at Bet Dagan, Israel. Nineteen species transmitted cucumber mosaic virus (CMV), while seven transmitted potato virus Y (PVY) at least once. Over 80% of the CMV and of the PVY infection among test plants (Capsicum annuum cv. Weindale) exposed to trapped aphids was caused by Aphis citricola and two or three other Aphis species, Myzus persicae and Macrosiphum euphorbiae. Landing rate was determined by comparing the proportion of each species found on green tiles or pepper plants with that found in suction traps. A. citricola was the most common but was found in a much lower proportion on plants than either in flight or on green tiles. Aphis spp. and M. persicae were more than 2–5 times more frequent (relative to other species) on green tiles than in flight. M. persicae and M. euphorbiae, which colonise peppers, were found on peppers at a proportion several times higher than either on green tiles or in the air. The relative importance of the different vector species was calculated by multiplying abundance by the proportion of transmitters and the landing rate. A. citricola and Aphis spp. were responsible for more than 50% of the total transmission of either CMV in 1981 and 1982 and of PVY in 1981. Peaks of CMV infection of bait plants coincided with peaks of transmitters of A. citricola and Aphis spp. caught in suction traps. The significance of these findings in primary infection of peppers with CMV and PVY is discussed.     

Properties of cocksfoot streak and cocksfoot cryptic, two viruses infecting cocksfoot (Dactylis glomerate) in Scotland

A Scottish isolate of cocksfoot streak virus (CSV-S) was found to have flexuous filamentous particles which, in sap of infected cocksfoot plants, had a modal length of 712 nm. It was transmitted from infected to healthy cocksfoot plants in a non-persistent manner by Myzus persicae and by mechanical inoculation of infective sap extracts containing an anti-oxidant. Apart from cocksfoot, mechanical inoculation of infective sap succeeded in infecting only four of 22 plant species tested. The infectivity of sap extracts containing 0.2% thioglycerol was lost after heating for 10 min at 55^oC but not 50^oC, storage at room temperature for 48 but not 24 hours, and after diluting 10-2 to 10-3. Highly purified preparations of CSV-S particles sedimented as a single component with a sedimentation coefficient of 139S and had a buoyant density in rubidium bromide of 1.31 g/cm3. Virus particles were composed of one protein and one ssRNA species with estimated Mr of 31 000 and 3.2 times 106 respectively. In ELISA, an antiserum prepared to CSV-S detected the virus in all aerial parts of infected cocksfoot plants and, when present in the ratio of 1 infected leaf: 1000 healthy leaves. Both CSV-S-infected and -uninfected cocksfoot also contained a previously undescribed virus with isometric particles c. 30 nm in diameter. This virus, named cocksfoot cryptic virus (CCV), was seed-borne in two cvs of cocksfoot tested and its particles contained two dsRNA species of estimated Mt of 1.14 times 106 and 1.27 times 106. Despite the fact that particles of CSV-S were largely free from CCV particles following exclusion chromatography on agarose beads prior to immunisation, immunoelectron microscopy (IEM) showed that the antiserum prepared to CSV-S also contained some antibodies to CCV. Evidence from IEM suggested a possible distant serological relationship of CCV to ryegrass and beet (BCV 1 or BCV 2, or both) cryptoviruses, all members of sub-group A of cryptoviruses.