SOME FACTORS AFFECTING THE TRANSMISSION OF LEAF-ROLL VIRUS BY APHIDS

Datura tatula is a more suitable host than potato for studying the factors influencing the transmission of potato leaf-roll virus by Myzus persicae ; it is more easily infected, provides a better source of virus for feeding aphids, produces symptoms more quickly and over a longer period of the year.
Loughnane's (1943) claim that leaf-roll virus is transmitted by starved aphids that feed for only 5 min. on infected potato plants was not confirmed. The shortest infection-feeding time in which M. persicae aphids became infective was 2 hr.; such aphids did not infect healthy plants in the first 2 days and, when transferred to a series of healthy plants at intervals, infected only few. The ability to cause infections was increased by increasing the length of infection feeding. Aphids fed for many days on infected plants could infect healthy plants in the first 15 min. of test feeding, and they continued to cause infections for long periods.
Aphids became infective more readily when feeding on recently infected Datura tatula , showing only slight symptoms, than on older plants with pronounced chlorosis; similarly, young potato sprouts showing no symptoms were better sources of virus for aphids than older plants showing severe leaf roll.
The differences in severity of symptoms shown by potato plants with leaf roll in the field mainly occur because of differences in virulence of accompanying strains of potato virus X , but isolates of leaf-roll virus were found that also varied in virulence.     

STUDIES ON TWO APHID-TRANSMITTED VIRUSES OF LEGUMINOUS CROPS

Pea mosaic virus was transmitted by Myzus persicae Sulz., Macrosiphum pisi Kalt., M. solanifolii Ash. and Aphis fabae Scop., but not by Hyperomyzus staphyleae Koch. It is a 'non-persistent' virus (Watson & Roberts, 1939), and is most readily transmitted when vectors are fasted and then given a short infection feeding. Vector efficiency was not increased by increases in preliminary fasting beyond 15 min. or with increasing infection feeding beyond 1 hr. Most aphids became non-infective within 15 min. when feeding, but fasting aphids remained infective for 3 hr. Species that fed readily on the infected plants were less efficient vectors than those which did not. Seed set by infected plants produced healthy seedlings.
Pea enation mosaic virus persisted in Myzus persicae and Macrosiphum pisi for more than 140 hr.; its transmission was unaffected by preliminary treatments of aphids. No transmission was obtained until at least 4 hr. after aphids had left infected plants; usually the 'latent' period exceeded 1 day and its duration was apparently unaffected by the length of the infection feeding.     

THE DISTRIBUTION OF VIRUSES IN DIFFERENT LEAF TISSUES AND ITS INFLUENCE ON VIRUS TRANSMISSION BY APHIDS

Exposing both surfaces of leaves systemically infected with cabbage black ring spot virus (CBRSV) or henbane mosaic virus to ultra-violet radiation decreases the infectivity of expressed sap to about one-fifth. As irradiation probably inactivates virus mainly in the epidermis, which occupies about one-quarter the volume of the leaves, these viruses seem to occur at much higher concentrations in sap from the epidermis than in sap from other cells. By contrast, tobacco mosaic virus seems not to occur predominantly in the epidermis.
CBRSV and henbane mosaic virus are normally transmitted most frequently by previously fasted aphids that feed for only short periods on infected leaves, but aphids treated like this transmit rarely from leaves that have been exposed to ultraviolet radiation. Irradiation has relatively little effect on the proportion of aphids that transmit after long infection feedings. Fasting seems to increase transmission by increasing the probability that aphids will imbibe sap from the epidermis of leaves they newly colonize. With longer periods on infected leaves, the ability of fasted aphids to transmit probably decreases because they then feed from deeper cells and their stylets contain sap with less virus. Only virus contained in the stylets seems to be transmitted, not virus taken into the stomach. About half the transmissions of henbane mosaic virus by aphids that have colonized tobacco leaves for hours may be caused by insects that temporarily cease feeding on the phloem and newly penetrate the epidermis.
Irradiating infected leaves affected the transmission of sugar-beet mosaic virus in the same way as that of henbane mosaic virus, but had little effect on the transmission of beet yellows virus, whose vectors become more likely to transmit the longer they feed on infected plants.     

QUANTITATIVE STUDIES ON THE TRANSMISSION OF CABBAGE BLACK RING SPOT VIRUS BY MYZUS PERSICAE (SULZ.)

Factors affecting the transmission of cabbage black ring spot virus by Mysus persicae (Sulz.) were studied quantitatively using the local lesions produced on tobacco leaves. Aphids prevented from feeding for 15 min. or more, before feeding for a few minutes on an infected plant, caused more infections than unfasted aphids. Fasted aphids acquired virus from infected plants in feeding times as short as 10 sec., and infected healthy plants in test-feeding times of 5 sec. Increasing test-feeding times to 30 min. increased the numbers of infections. Increasing infection- feeding times from 10 sec. to 5 min. had little effect, but increasing to more than 5 min. greatly reduced the number of transmissions. This reduction was partly offset if the aphids were prevented from feeding continuously while on the infected plants. With undisturbed infection-feeding periods of 15 min. or longer, previously fasted aphids caused no more infections than unfasted aphids.
Infective aphids lost their ability to produce lesions more rapidly when feeding than when fasting.
Winged and wingless aphids were equally efficient vectors.     

THE MANNER OF TRANSMISSION OF SOME BARLEY YELLOW-DWARF VIRUSES BY DIFFERENT APHID SPECIES

Some barley yellow-dwarf (BYD) viruses isolated from cereal crops in Great Britain were transmitted by Rhopalosiphum padi , L. and others were not. Sitobion fragariae (Walker), S. avenae (Fabricius), and Metopolophium dirhodum (Walker) all transmitted viruses of both types, but they usually transmitted those of which Rhopalosiphum was a vector less readily than did R. padi. The transmissibility of a virus by a given aphid species was not affected by transmission with another, less efficient, vector species. Neomyzus circumflexus (Buckt.) and Rhopalosiphum maidis (Fitch) transmitted the few viruses with which they were tested.
A few R. padi acquired virus from infected leaves during 30 min. feeding and inoculated healthy seedlings during 15 min. feeding, but the minimum total time taken to acquire and transmit was 10 hr. and 32 hr. were needed for about half the aphids that were able to acquire and transmit virus to do so. This may indicate the existence of a short latent period of the virus in the vector, although the evidence is not conclusive. The times spent on infected plants influenced the results more than those spent on healthy ones; many transmissions occurred with short feeding times on healthy plants so long as the time spent on infected leaves was long, but the reverse was not true. Nymphs of R. padi that moulted after they left infected plants on which they fed long enough to become infective, infected slightly fewer plants than adults fed for the same times.     

THE TRANSMISSION BY MITES, HOST-RANGE AND PROPERTIES OF RYEGRASS MOSAIC VIRUS

A virus that causes chlorotic streaks on ryegrass leaves was transmitted by the eriophyid mite Abacarus hystrix (Nalepa). Virus-free mites acquired the virus in 2 hr. feeding on infected ryegrass and the proportion that became infective increased with increased feeding time up to 12 hr.; vectors lost infectivity within 24 hr. of leaving the infected leaves. All instars of A. hystrix transmitted the virus.
The virus was transmitted by manual inoculation of sap to other species of Gramineae, including oats, rice, cocksfoot and meadow fescue, but none of these hosts seemed to contain as much virus as ryegrass; their saps did not precipitate specifically with antiserum prepared against the virus in ryegrass, whereas sap from infected ryegrass precipitated up to a dilution of 1/32. Infective sap of S22 Italian ryegrass contained flexuous rod-shaped particles; the dilution end-point of the virus was about 1 in 1000; the virus was inactivated when held for 10 min. at 60°C. and most of its infectivity was lost after 24 hr. at room temperature.     

Resolution of Strawberry Virus Complexes

Aphids ( Capitophorus fragariae Theob.) allowed to feed for several days on a strawberry plant infected with yellow-edge transmitted two virus fractions. The isolation and properties of one (virus 1) have been described previously. The other (virus 2) was separated by transferring the aphids to fresh indicators after 24 hr.
Virus 2 was retransmitted after infection feeding periods of 24 hr. or more and persisted in the vector for several days. There is some evidence that it is itself a complex of viruses which can be separated further. On Fragaria vesca virus 2 produced chlorotic spotting, slight marginal chlorosis of the leaves and slight cupping of the leaflets. On Royal Sovereign strawberry it produced slight chlorosis of the young leaves.
On Royal Sovereign viruses 1 and 2 together produced symptoms of yellow-edge which is thus shown to be caused by a virus complex which can be resolved by means of the aphis vector.     

RESOLUTION OF STRAWBERRY VIRUS COMPLEXES IV. THE LATENT PERIOD OF VIRUS 3 IN THE VECTOR

From the time of first feeding on plants infected with strawberry virus 3, 10–19 days elapsed before Capitophorus fragariae became infective, a longer 'latent period' than any previously recorded for an aphid-transmitted virus. The time taken for aphids to develop infectivity after leaving infected plants decreased with increasing duration of the infection feed. Aphids which had fed for 16 days on an infected plant caused infection in the first day of test feeding.     

Transmission of potato leafroll virus in relation to the honeydew excretion of Myzus persicae

Honeydew excretion of single Myzus persicae nymphs on potato leafroll virus (PLVR)-infected Physalis floridana was studied during the acquisition access period (AAP) in relation to the efficiency of virus transmission.
With increasing length of the AAP, the percentage of nymphs that transmitted the virus increased. These nymphs produced significantly more honeydew droplets during the AAP on PLRV-infected P. floridana plants than nymphs which failed to transmit the virus. However, the number of honeydew droplets excreted during the AAP by transmitting nymphs did not affect the length of the latency period. Nymphs which infected the first test plant after a short latency period produced a similar amount of honeydew during the AAP to those with a longer latency period.
Honeydew excretion recorded on plants of varied age, showed that nymphs feeding on bottom leaves of infected plants produced more honeydew droplets than on comparable leaves of healthy plants. On infected plants, nymphs produced more honeydew droplets on bottom leaves with pronounced symptoms than on top leaves that hardly showed any symptom of PLRV infection.
The concentration of viral antigen measured by ELISA was lower in top leaves than in bottom leaves of infected plants. Nevertheless, nymphs feeding on top leaves transmitted the virus more efficiently than those which used bottom leaves as virus source. When bottom leaves were used as a virus source, the percentage of viruliferous nymphs decreased with plant age. These results indicate that the availability of virus for acquisition by aphids declines with increasing plant age and symptom severity.     

Transmission characteristics and some other properties of bean yellow vein-banding virus, and its association with pea enation

Bean yellow vein-banding virus (BYVBV) has been found occasionally in mixed infection with pea enation mosaic virus (PEMV) in spring-sown field beans (Vicia faba minor) in southern England. Glasshouse tests confirmed that, like PEMV, BYVBV is transmissible by manual inoculation and by aphids in the persistent manner. However, BYVBV can be transmitted by aphids only from plants that are also infected with a helper virus, usually PEMV. Thus after separation from PEMV by passage through Phaseolus vulgaris it was no longer aphid-transmissible. It became aphid-transmissible again only after re-mixing in plants with PEMV or with a substitute helper, bean leaf roll virus (BLRV). It was not transmitted by aphids that fed sequentially on plants singly infected with PEMV and BYVBV. Thus the interaction between BYVBV and PEMV (or BLRV) that enables BYVBV to be transmitted by aphids seems to occur only in doubly infected plants. However, it was not transmitted by aphids from plants doubly infected with BYVBV and broad bean wilt virus (BBWV). BYVBV and PEMV were transmitted more readily by Acyrthosiphon pisum than by Myzus persicae; neither virus was transmitted by Aphis fabae. Phenol extracts of BYVBV-infected leaves were more infective than phosphate buffer or bentonite-clarified extracts and were sometimes infective when diluted to 1/1000. The infectivity of BYVBV in phosphate buffer extracts of leaves singly infected with BYVBV, unlike that in extracts of leaves doubly infected with BYVBV and PEMV (or BLRV), was destroyed by treatment with organic solvents. BYVBV infected 11 of 28 plant species that were inoculated with phenol extracts; seven of the infected species were legumes. No transmission of BYVBV was detected through seed harvested from infected field bean plants. Isometric particles c. 30 nm in diameter were seen in extracts of plants doubly infected with BYVBV and PEMV but not in extracts of plants infected with BYVBV alone. Leaves of plants infected with BYVBV, alone or with PEMV, contained membrane-bound structures c. 50–90 nm in diameter associated with the tonoplast in cell vacuoles. These structures were not found in healthy leaves. BYVBV has several properties in common with other known aphid-borne viruses that are helper-dependent and transmitted in a persistent manner. Possibly, as suggested for some of them, aphid transmission of BYVBV depends on the coating of its nucleic acid with helper virus coat protein.     

STUDIES ON THE TRANSMISSION OF CASSAVA MOSAIC VIRUS BY BEMISIA SPP. (ALEYRODIDAE)

Whiteflies, which had originated from a mixed culture of Bemisia spp. collected from cassava ( Manihot utilissima Pohl.) in the field, needed to feed for at least 4 hr. on the young leaves of a cassava plant with mosaic before they acquired the virus. Whiteflies that acquired virus in 4–6 hr. required another 4 hr. to become viruliferous. Once viruliferous they could infect healthy plants in a feeding period of 15 min., but longer periods gave more infections. Adult whiteflies remained infective for more than 48 hr. after ending their infection feed. Cassava fed upon by only one viruliferous fly sometimes became infected.
The virus-vector relationships of cassava mosaic virus resemble those of cotton leaf-curl virus but the first could not be transmitted to cotton or the second to cassava.     

Cucurbit aphid‐borne yellows virus (CABYV) modifies the alighting,settling and probing behaviour of its vector Aphis gossypii favouring its own spread

Plant pathogens are able to influence the behaviour and fitness of their vectors in such a way that changes in plant–pathogen–vector interactions can affect their transmission. Such influence can be direct or indirect, depending on whether it is mediated by the presence of the pathogen in the vector's body or by host changes as a consequence of pathogen infection. We report the effect that the persistently aphid‐transmitted Cucurbit aphid‐borne yellows virus (CABYV, Polerovirus) can induce on the alighting, settling and probing behaviour activities of its vector, the cotton aphid Aphis gossypii. Only minor direct changes on aphid feeding behaviour were observed when viruliferous aphids fed on non‐infected plants. However, the feeding behaviour of non‐viruliferous aphids was very different on CABYV‐infected than on non‐infected plants. Non‐viruliferous aphids spent longer time feeding from the phloem in CABYV‐infected plants compared to non‐infected plants, suggesting that CABYV indirectly manipulates aphid feeding behaviour through its shared host plant in order to favour viral acquisition. Viruliferous aphids showed a clear preference for non‐infected over CABYV‐infected plants at short and long time, while such behaviour was not observed for non‐viruliferous aphids. Overall, our results indicate that CABYV induces changes in its host plant that modifies aphid feeding behaviour in a way that virus acquisition from infected plants is enhanced. Once the aphids become viruliferous they prefer to settle on healthy plants, leading to optimise the transmission and spread of this phloem‐limited virus.     

Studies on the transmission of groundnut rosette virus by Aphis craccivora Koch

Four strains of groundnut rosette virus were transmitted by a race of Aphis craccivora (Koch) from groundnut in Nigeria. Two of these strains, both from East Africa, were transmitted only by A. craccivora from Kenya. A fifth isolate, from Nigeria, was not transmissible by either race. The two races of aphids have been shown elsewhere to be distinct biotypes. Most A. craccivora needed longer than 24 h feeding on infected groundnuts to acquire virus, and many needed 2–3 days of feeding on healthy plants to cause infection, even after several days on infected plants. The delays partly reflect the slow uptake of virus and possibly a period needed for virus multiplication in aphid tissue but some is lost through resistance of the test plants to infection. In consecutive feeding experiments Natal Common variety could be infected soon after aphids had left the source of virus, but a more resistant Nigerian variety sometimes needed several more days. The frequency of inoculation by aphids, or the concentration of virus in the inocula or both, increased with time, but the times at which aphids were able to infect plants was also dependent on variety.     

THE DIFFERENT DISTRIBUTION OF TWO BRASSICA VIRUSES IN THE PLANT AND ITS INFLUENCE ON SPREAD IN THE FIELD

Previous knowledge provided no explanation for the greater prevalence of cauliflower mosaic than of cabbage black ring spot in field crops of cauliflower. Both viruses are spread principally by Myzus persicae and Brevicoryne brassicae , and both are transmitted equally readily from infected seedlings. Cabbage black ring spot virus has a much wider host range, and sap from infected leaves has a higher dilution end-point than sap from leaves infected with cauliflower mosaic virus.
At least part of the difference between the rate at which the two viruses spread in the field may be accounted for by the different manner in which they are distributed in old infected plants, and the effect this has on transmission by aphids. Cauliflower mosaic virus occurs in high concentration in all the new leaves produced by infected plants. Cabbage black ring spot virus, on the other hand, occurs mainly in the older leaves, and even there is localized in parts that show symptoms. Only in recently infected plants does cabbage black ring spot virus occur in young leaves.
After flying, most aphids alight on the upper parts of plants; they are therefore less likely to acquire cabbage black ring spot virus than cauliflower mosaic virus. It may be significant that cabbage, a host in which old leaves are in a more favourable position for alighting aphids than are those of cauliflower, is also often extensively infected with cabbage black ring spot virus.     

Resolution of strawberry virus complexes by means of the aphis vector Capitophorus fragariae Theob

Aphides ( Capitophorus fragariae ) were fed for periods of up to 24 hr. on strawberry plants infected with mild crinkle, severe crinkle or yellow-edge and then transferred to plants of the wild strawberry, Fragaria vesca , or of the cultivated strawberry, variety Royal Sovereign. On F. vesca the symptoms produced were chlorotic speckling, distortion and dwarfing of the leaves, varying in intensity', and on Royal Sovereign scattered, inconspicuous, diffuse, chlorotic spots.
The symptoms from all three sources of infection were similar and were indistinguishable from those of mild crinkle of Harris & King. The virus thus selectively transmitted is tentatively concluded to be the mild crinkle virus.
The virus was transmitted after feeding periods of 1 hr. or more and did not generally persist in the vector for more than 3 hr.     

TRANSMISSION OF GROUNDNUT ROSETTE VIRUS

The rosette virus was transmitted to groundnut plants, if previously etiolated, bymechanical inoculation of juice with 'Celite' addition; but only a small proportion of the inoculations succeeded.
Aphis craccivora (Koch), the known vector, transmitted the virus by feeding on germinating groundnut seeds; and from this fact we developed an experimental technique that is convenient and flexible.
Different races of this species appeared to vary in inherent efficiency in transmission, and one failed ever to transmit. Within races that could transmit, all larval forms and alate and apterous adults might transmit; but alatae were sometimes significantly more efficient than apterae, and at other times the converse held. A field experiment showed that wingless forms, moving over the soil surface, might play a predominant part in secondary spread around a rosetted plant.
Comparative tests with groups of I, 2, 3 and 4 infective aphids supported the hypothesis that infections by this vector are individual and independent.
Single aphids, tested for 24 hr. on 10 successive days without access to an outside source of virus, might infect on any day up to the tenth. Similar results were obtained in a succession of I hr. tests on a single day. In the infected seed the virus rapidly became available to feeding aphids; previously non-infective aphids acquired the virus by feeding on a seed during the third day from the first exposure of this seed to infective aphids.     

Aphid-injection experiments with carrot mottle virus and its helper virus, carrot red leaf

Carrot mottle virus (CMotV) and its helper virus, carrot red leaf (CRLV), were not transmitted by aphids (Cavariella aegopodii) that had fed through membranes on, or had been injected with, sap from mixedly infected chervil plants or partially purified preparations of CMotV. However, the viruses were transmitted by recipient aphids injected with haemolymph from donor aphids that had fed on mixedly infected plants but not by a second series of recipients injected with haemolymph from the first series. Some of the first series of recipients transmitted both viruses for up to 11 days but others transmitted erratically and many lost ability to transmit after a few days. The results confirm that both viruses are circulative but provide no evidence for multiplication in the vector. Non-viruliferous aphids, or aphids that had acquired CRLV by feeding, did not transmit CMotV when they were injected with haemolymph from aphids that had fed on a source of CMotV alone, confirming that they can only transmit CMotV when they acquire it from a mixedly infected plant. When extracts from donor aphids were treated with ether before injection, recipient aphids transmitted both CRLV and CMotV, although the infectivity of CMotV grown in Nicotiana clevelandii in the absence of CRLV is destroyed by ether treatment. CMotV particles acquired by aphids from mixedly infected plants therefore differed in some way from those in singly infected plants. A plausible explanation of these results, and of the dependence of CMotV on CRLV for aphid transmission, is that doubly infected plants contain some particles that consist of CMotV nucleic acid coated with CRLV protein.     

Infection of potato plants with potato leafroll virus changes attraction and feeding behaviour of Myzus persicae

Potato leafroll virus (PLRV; genus Polerovirus, family Luteoviridae) is a persistently transmitted circulative virus that depends on aphids for spreading. The primary vector of PLRV is the aphid Myzus persicae (Sulzer) (Homoptera: Aphididae). Solanum tuberosum L. potato cv. Kardal (Solanaceae) has a certain degree of resistance to M. persicae: young leaves seem to be resistant, whereas senescent leaves are susceptible. In this study, we investigated whether PLRV‐infection of potato plants affected aphid behaviour. We found that M. persicae's ability to differentiate headspace volatiles emitted from PLRV‐infected and non‐infected potato plants depends on the age of the leaf. In young apical leaves, no difference in aphid attraction was found between PLRV‐infected and non‐infected leaves. In fact, hardly any aphids were attracted. On the contrary, in mature leaves, headspace volatiles from virus infected leaves attracted the aphids. We also studied the effect of PLRV‐infection on probing and feeding behaviour (plant penetration) of M. persicae using the electrical penetration graph technique (DC system). Several differences were observed between plant penetration in PLRV‐infected and non‐infected plants, but only after infected plants showed visual symptoms of PLRV infection. The effects of PLRV‐infection in plants on the behaviour of M. persicae, the vector of the virus, and the implications of these effects on the transmission of the virus are thoroughly discussed.     

The role of the helper virus, anthriscus yellows, in the transmission of parsnip yellow fleck virus by the aphid Cavariella aegopodii

Transmission of parsnip yellow fleck virus (PYFV) by the aphid Cavariella aegopodii occurs only when the aphids are also carrying the helper virus, anthriscus yellows (AYV). None of five other viruses tested was able to act as helper. In experiments in which aphids were allowed to feed through membranes on crude or treated extracts from infected plants, aphids already carrying AYV acquired PYFV, but virus-free aphids failed to acquire either AYV or PYFV. PYFV was not transmitted by insects injected with haemolymph from aphids carrying both viruses, or with purified preparations of PYFV. PYFV was transmitted when AYV-carrying aphids, except those whose stylets had been removed, were contaminated externally with PYFV preparations. Ultraviolet irradiation of infected leaves did not prevent aphids from acquiring AYV, presumably because it is confined to deeply-lying tissues. AYV-carrying aphids could acquire PYFV from u.v.-irradiated leaves after acquisition access times of 2 h but not after feeds of only 2 or 15 min (which are adequate on unirradiated leaves), suggesting that PYFV is present in all parts of the leaf. No ‘helper agent’ distinct from AYV itself was detected in these experiments or in experiments on minimum acquisition feeding time or maximum period of persistence in the aphid. U.v.-inactivated PYFV competed with infective PYFV for retention sites in AYV-carrying aphids, whereas AYV apparently did not. It is suggested that there is no helper agent for PYFV, other than AYV particles. The possibility that there is one for AYV is not excluded.     

SOME PROPERTIES OF FOUR STRAINS OF CUCUMBER MOSAIC VIRUS

Different strains of cucumber mosaic virus differ in their host range, symptoms caused, virulence towards different plants, transmissibility by aphids, dilution end-point and thermal inactivation point.
There are seasonal variations in the susceptibility of some host species; French bean is apparently immune during the summer but during the winter produces countable local lesions suitable for quantitative assays.
Different host species differ in the ease with which cucumber mosaic virus is transmitted to and from them; systemic infection in beet rarely occurs unless the virus is introduced into young tissues. Inhibitors of infectivity in sap of sugar beet and Phytolacca sp. make mechanical transmission from these to other hosts difficult; the inhibitors interfere less with the infection of hosts in which they occur than with the infection of tobacco.
Cucumber mosaic virus has a low temperature coefficient of thermal inactivation and much infectivity is destroyed by heating at temperatures below the thermal inactivation point.
Myzus persicae (Sulz.) is a more efficient vector than M. ornatus Laing which is more efficient than Macrosiphum euphorbiae (Thomas); although individual aphids can cause more than one infection, most cease to be infective in feeding periods of from one to five minutes.