Manual Inoculation Research Articles

Beetle Transmission of Plant Viruses

Although the majority of plant viruses are transmitted by insects with sucking mouthparts, some viruses are transmitted by insects with biting mouthparts. Of the insects with biting mouthparts that transmit viruses, the beetles are the most important vectors. Viruses transmitted by beetles apparently cannot be transmitted by insects with sucking mouthparts. Until recently it has been generally assumed that when a biting insect behaves as a vector, it only does so by carrying infective sap from diseased to healthy plants on its mouthparts. However, when a beetle is an efficient vector and retains virus up to 20 days as the cucumber beetle does with squash mosaic virus, some additional mechanism of transmission probably is involved. This chapter brings together the various aspects of beetle transmission of plant viruses. Viruses transmitted by beetles have many properties in common. Usually, they are relatively stable, develop a high titer in infected plants, can be readily transmitted by manual inoculation, have spherical or polyhedral particles from 25 to 30 mp in diameter, and are highly antigenic. Host ranges and symptoms are usually not correlated with any specific features of the virus particles or with their vectors. The beetle-transmitted viruses can be divided into two groups depending on the length of time the viruses are retained by their vectors. The larger group consists of the viruses that are retained by their vectors for a prolonged period of time. The other group is made up of those viruses that are retained by their vectors for short periods of time, usually 24-48 hours. Only beetles in the family, Chrysomelidae, commonly called leaf beetles, have been shown to transmit plant viruses. All but two of the beetles belong in two subfamilies-Galerucinae (Galerucine leaf beetles) and Halticinste (flea beetles).

Reactions of some old and new British potato cultivars to tobacco rattle virus

Fourteen potato cultivars were grown on sandy land in Norfolk infested withTrichodorus pachydermas carrying tobacco rattle virus (TRV). Four out of five old cultivars showed few or no symptoms of spraing, whereas six out of nine modern cultivars were moderately or severely affected. In cultivars that showed severe spraing, TRV rarely passed from affected tubers to progeny plants and tubers, but did so more often in cultivars that showed slight spraing. Haulm symptoms in progeny plants consisted of stem-mottle, sometimes including figure aucuba. Some of the tubers produced by plants with stem-mottle showed internal markings whose severity was related to that of the primary spraing in that cultivar.Arran Pilot showed neither spraing nor stem-mottle; its roots resisted infection by vector-borne TRV, but its leaves were infected by manual inoculation with infective sap. TRV was transmitted by inoculation of sap to indicator plants from more than 90% of freshly dug tubers showing spraing, and of shoots showing stem-mottle.

Groundnut rosette and its assistor virus

SUMMARYChlorotic rosette from Malawi (isolate CR1), passed through Stylosanthes gracilis and S. juncea, was not subsequently transmissible from groundnuts (Arachis hypogaea) by Aphis craccivora or A. gossypii, but with S. mucronata transmissibility was occasionally regained after a period of time. Aphid transmissibility was similarly lost after passage of two isolates (a chlorotic rosette from Rhodesia, CR2, and a green rosette from Nigeria, GR) through soybean (Soja max) and after manual inoculation to groundnuts. Groundnut plants that remained symptomless after exposure to rosette infection by aphids often contained a virus that restored aphid transmissibility when introduced into groundnuts containing the vectorless virus from that isolate. Groundnut rosette disease therefore consists of a symptom‐inducing virus that we call groundnut rosette virus (GRV) and a symptomless assistor virus (GRAV) that must be present for aphid transmission. The interactions between the GRV and GRAV of chlorotic and green rosette, and their transmission by different vector races, are described.

Some properties of cocksfoot mottle virus

SUMMARYCocksfoot mottle virus (CFMV) was transmitted by manual inoculation of sap to cocksfoot (Dactylis glomerata L.), wheat, oats and barley, but not to nineteen other monocotyledonous and thirteen dicotyledonous plant species. The virus was also transmitted by cereal leaf beetles (Lema melanopa L.). Adult beetles infected plants more frequently than larvae, and remained infective for up to 2 weeks after they had fed on infected plants. Seed from infected cocksfoot and oat plants produced virus‐free seedlings.The infectivity of sap was lost during 10 min. at 65° C., and 2 weeks at 20° C., but survived many months at — 15° C.Purified virus preparations, made by various methods, contained numerous nearly spherical particles, about 30 mμ in diameter. In electron micrographs some of the particles were penetrated by negative stain though most appeared intact. However, all the particles migrated together in a centrifugal (sedimentation coefficient = 118 S) or electrophoretic field. The ultraviolet absorption spectrum, and the phosphorus and nitrogen contents of the virus preparations, were typical of a nucleoprotein containing about 25 % nucleic acid.Serological tests failed to show any relationship between CFMV and eleven other viruses with particles of similar shape and size.

Host range and some properties of groundnut rosette virus

SUMMARYTwo isolates of groundnut rosette virus from East Africa (GRVE1 and GRVE2) and from West Africa (GRVW1 and GRVW2) were transmitted by Aphis craccivora obtained from West Africa. A third isolate from West Africa (GRVW3) was not transmitted by A. craccivora from three widely separated sources. GRVW1, GRVW2 and GRVW3 caused leaf‐symptoms in groundnut of a mosaic pattern in light and dark green. GRVE1 and GRVE2 caused chlorosis or chlorosis and leaf distortion as well as mosaic symptoms. Groundnut plants with GRVW1 could not be infected by means of aphids with GRVE1, and GRVE1 gave similar protection against GRVW1, which suggests that they are strains of the same virus.All isolates were transmissible manually from groundnut to groundnut (Arachis hypogea), Trifolium incarnatum and T. repens, and caused systemic infection. Inoculated Nicotiana clevelandii and N. rustica developed symptoms but virus could not be recovered from them. Chenopodium amaranticolor, C. hybridum and C. quinoa showed local lesions on inoculated leaves. Virus could be acquired by aphids from groundnut or Trifolium repens infected by means of aphids, but not from those infected by manual inoculation. Virus could not be recovered from T. incarnatum manually or by aphids, but was transmitted by cleft‐grafting from clover to groundnut.Saps extracted in borax buffer plus zinc sulphate at pH 9 from plants infected with GRVW1 and GRVE1 remained infective at 18° C. for 1 week, and at — 20° C. for up to 4 weeks. Virus could be recovered from frozen leaves. Buffered saps lost infectivity when heated above 50° C. for 10 min.; most were still infective when diluted 1/10 and some at 1/100.Electron micrographs of partially purified preparations contained spherical particles 25–28 mμ in diameter. There were usually only about five per microscope field and they resembled those of some other viruses.

Carrot motley dwarf and parsnip mottle viruses

SUMMARY Carrots that show symptoms of carrot motley dwarf contain two viruses, carrot mottle virus (CMV) and red-leaf virus (RLV). CMV cannot be manually inoculated to carrot, but can be to some other members of the Umbelliferae, as well as to some species of the Solanaceae, Leguminoseae and Chenopodiaceae. The host range of RLV is limited to the Umbelliferae, and it is not manually transmissible but was transmitted by grafting. Cavariella aegopodiae Scop, transmits RLV alone but will transmit CMV only from plants infected with both viruses. Thus aphids were unable to transmit from coriander plants manually inoculated with CMV, but after these plants were infected with RLV by aphids, virus-free aphids acquired and transmitted both viruses from them. Aphids remain infective with both viruses for 1–2 weeks, and retain infectivity through the moult. A minimum total of about 9 hr. is needed for acquisition and transmission; vector-efficiency increases with increasing feeding times up to severaj days. The viruses causing motley dwarf become attenuated in the glasshouse after continued aphid-transmission; avirulent isolates protect their hosts against infection by virulent ones. Infectivity of saps from CMV infected plants was increased by extraction at high pH in the presence of a trace of Zn, and was associated in carrot with particles 30 mμ in diameter. Water-phenol extracts are almost as infective as extracts in buffer, but are inactivated by 0·02 μg./l. pancreatic ribonuclease. Parsnip mottle virus (PMV) resembles CMV in many ways, but also differs in some important respects. Unlike CMV it infects celery and parsnip, and it is transmitted by C. pastinacae as well as C. aegopodiae, the only vector of CMV (in combination with RLV). PMV is transmitted by aphids from plants infected with it alone, whether these plants were infected by aphids or by manual inoculation. Carrot plants infected first with PMV and then with the motley dwarf virus complex developed symptoms of motley dwarf, but in coriander the reverse happened. CMV and PMV appear to interfere with each other's multiplication in the hosts, but CMV is dominant in carrot and PMV in coriander. They have some properties of distantly related strains.

AGROPYRON MOSAIC AS A DISEASE OF WHEAT IN CANADA

Agropyron mosaic virus (AMV) is common on Agropyron repens (L.) Beauv. in southern Ontario and Quebec and has been found on Prince Edward Island and in southeastern Saskatchewan. It was also found on a few winter wheat plants in many fields in southern Ontario each year since 1957, and on spring wheat in Saskatchewan in 1961. AMV has not appeared to be economically important, but the occasional high incidence of infection in wheat near diseased A. repens and the fact that infected plants were sometimes severely stunted have indicated that it could be important under favorable conditions.Wheat was more susceptible than A. repens, or any of the other hosts tested. Usually all wheat plants became infected when inoculated with sap of diseased plants diluted 1:64, and 50% became infected with dilutions of 1:256. Symptoms appeared in 6 days at temperatures of 20° and 25 °C, but were most pronounced at 15 °C. No symptoms developed on wheat kept at 10° or 35 °C for 19 days after inoculation, but sap from these plants was infectious.All isolates of AMV obtained from A. repens and most from naturally diseased wheat caused only mild symptoms on wheat, but some isolates from wheat caused severe chlorosis and stunting. In field tests, manual inoculation with severe isolates reduced yields of spring wheat 25% to 75%, and of winter wheat up to 42%. Up to 100% of the winter wheat plants in small plots sown near naturally diseased wheat during June to late August became infected, illustrating that AMV can be perpetuated on immature wheat. Less infection developed in plots sown in September or later.Eriophyid mites, including Aceria tulipae (K.), Abacarus hystrix (Nal.), and Vasates mckenziei K. were present on naturally diseased plants, but none of these mites has been proved a vector of AMV.

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.

CHANGES IN THE TRANSMISSIBILITY BY APHIDS OF A STRAIN OF CUCUMBER MOSAIC VIRUS

A strain of cucumber mosaic virus isolated from a spinach plant in 1946 was readily transmitted by Myzus persicae until 1955 when it lost this property, although it was still being propagated in conditions in which other strains remained transmissible. M. circumflexus also transmitted other strains but not this one. It was transmitted as readily as other strains by Aphis gossypii and Myzus ascalonicus. M. ascalonicus transmitted less frequently than Aphis gossypii. Transmission of the spinach strain by other aphids did not make it transmissible by Myzus persicae; nor did propagation in different plant species or several passages through spinach. In 1955 the spinach strain was occasionally transmitted by M. persicae, but the cultures isolated in this way were no more readily transmissible by the aphid than was the bulk culture maintained by manual inoculation of sap, and after a few weeks all cultures ceased to be transmitted by M. persicae.

THE EFFECT OF DIFFERENT HOST PLANTS OF POTATO VIRUS C IN DETERMINING ITS TRANSMISSION BY APHIDS

A stock of potato virus C derived from Edgecote Purple potatoes in 1945 was not then transmitted by aphids, although more than 2000 aphids were used in conditions optimal for transmitting the serologically related potato virus Y. This stock of virus C has been propagated continuously since, by manual inoculation in a series of Nicotiana glutinosa and N. tabacum, and in 1955 it was transmitted by the aphid Myzus persicae (Sulz.): about one in twenty of the aphids transmitted it compared with one in two for potato virus Y.Virus C derived from the Edgecote Purple potatoes in 1955 was not transmitted by aphids; both stocks of virus C produced only local lesions in Majestic potato leaves, and gave similar symptoms in tobacco.When inoculated to Majestic potatoes and then returned to tobacco plants, potato virus C usually ceased to be aphid transmitted and did not recover this property in any of the subsequent subcultures.Transmission from stock by aphids did not isolate a strain of virus C which was any more readily transmitted by aphids, indeed, for the first two or three subcultures, aphids usually transmitted more readily from plants inoculated manually. But the few isolates which remained aphid transmissible, after a second passage through potato, were rather readily transmitted.These results suggest that the ability of a virus to be aphid transmitted is, at least in part, determined by the host plant in which it is multiplying, but the nature of the changes which determine this ability are unknown.