BIOLOGIA PLANTARUM (PRAHA) 8 (1) : 73--79, 1966 The Mechanical Transmission and Some Properties of a Virus Disease of Cow-parsnip, Heracleum sphondylium L. ZDENKO POL/[K Department of Plant Pathology, Institute of Experimental Botany, Czechoslovak Academy of Sciences, Praha* Received March 1, 1965 Abstract. The described virus of cow-parsnip, Heracleum 8phondylium L., was found in three ruderal localities of Greater Praha. The symptoms are manifested by dccolorations which consist of bright yellow areas spreading from the centre of the leaf blade along the main veins. These symptoms appear severely in May. Under higher temperatures and in a chronic stage of infec- tion the symptoms are more or less masked. The disease is mechanically transmissible to parsley, coriander, parsnip, dill, sowbane, Chenopodium quinoa and C. giganteum. The author failed to transmit the disease to celery, carrot, caraway and to 27 species of differential host plants, he failed in the transmission of the virus by the dodder, Cuscuta campestri8 Yu~cx., too. Thermal inactivation point of the virus lies between 51 ~ and 55 ~ C. Infectivity of extracted sap was lost after 2 days at room temperature. In 1963, a young plant of Heracleum sphondylium L. found at Praha-Hole- ~ovice showed a type of mosaic disease caused by a virus that preliminarily appeared to differ from the biological point of view from viruses of umbeliferous plants, described in literature. The test showed that this virus is able to infect only a limited number of plant species, including a few umbeliferous crop plants, such as parsley, coriander, parsnip and dill. Principally because of its unusual relationship to obligate differencia! host plants and the limited state of our knowledge concerning European virus diseases of umbeliferous plants, the disease was studied further. The disease is not common and has been found only rarely during routine observations of weed associations at Praha-Dejvice and Praha-Hostiva~ in 1964. Transmission trials gave evidence of identity of causative agents. Results The host range of the virus has not been studied extensively, but most of the species tested proved to be immune from infection. * Address: Na Kar ovce 1, Praha 6. 73 74 z. POL/kK Species of plants found susceptible are as follows: Petroselixu~n hortense HoFr~., parsley; Petroselinura hortense var. crispum (MILL.); Coriandrum sati~uYa L., coriander; Pastinaca sativa L., parsnip; Anethum graveolens L., dill.; Chenopodium quinoa W~LD.; C. giganteum DON., and C. murale L., sowbane. Plants showing no indication of infection are listed below in alphabetical order: Amaranthus caudatus L., lovelies bleeding Apium graveolens L., celery Beta pateUaris Moq. B. vulgaris L., sugar beet Brassica naTus L., rape B. oleracea L., cabbage Capsicum anr, uum L., red pepper Ca~ urn carvi L., caraway Chenopodium polyspern, um L. Cu~urais melo L., musk melon C. sativus L., cucumber Datura innoxia MILL., downy thorn-aple D. stramonium L., jimson-weed Daucus carrota L., carrot Gomphrena globosa L., globe-amaranth Helianthu~ an~uus L., common sunflower I.upinus albus L., white lupine Medicago sativa L., alfalfa Nicandra physaloides (L.) GAERT~., apple-of-Peru Nicotiana glutinosa L. 2V. sylvestris SPEG. et COMES N. tabacum L. var. Samsun, tobacco _IV. taba(ura L. var. Xanthi-nc, tobacco Phaseolus vulgaris L., French bean Physalis floridana RYDB., ground cherry Pi~um sativum L., pea Solanum lycopersicum L., tomato S. luberosum L., potato Tetragonia expansa MURR., ~ew Zealand spinach Zinnia elegans JAcQ., zinnia It seems unlikely that any of the above plants arc symptomless carriers of the virus, as attempts to recover the virus from them have always been unsuccessful. It is necessary to stress the failure to transmit the virus to carrots, caraway and celery. Four varieties of carrots (Amsterodamsks Chantenay, Nantais and Stupicks -- together 150 plants), two of caraway (~esk~ and Moravsk~ -- together 80 plants) and 150 celery plants were tested. Symptoms of the disease in cow-parsnip leaves The first symptoms in nature seem to be a mild chlorosis or flavescence in the terminal leaflets of the first leaf. The leaflets are deformed and often curled downwards. Later decolorations consist of bright yellow areas spread- ing from th3 centre of the leaf blade along the main veins (Fig. 1). The collapse V[P~U.~ DISEASE IN HERACLEUM SPHONDYLIUM 75 of the veins while growth is still proceeding results in a chzck to their growth, relative to that of the interveinal tissue, so that leaves present a curly or crumpled appearance with sinuate margins. Symptoms described appear severally in May. Under higher temperatures and in later stages of infection the symptoms are partly or completely masked. This phenomenon is probably due to the level of virus concentration in the plants which progressively declines. At the end of June it is impossible to distinguish between the diseased and the healthy individuals because of the loss of symptoms. At this stage the transmissibility of the disease from the source is very difficult, if not altogether impossible. There appears to be a distinct difference in the severity and symp- tom response under different substrate conditions. In humid or nitrous locali- ties there are milder changes of habitus, the disease often develops a faint mottling only in the leaflets. Symptoms in susceptible plants tested Petroselinum hortense HOFFM., parsley: The first symptoms to develop under glasshouse conditions are a clearing of veins and veinlets and blister-like elevations of the youngest leaves (symptoms in small seedlings develop 4 to 5 days after inoculation). As the disease progresses, a conspicuous vein-banding occurs. The yellow bands are at first narrow but gradually become broader and diffuse in outline. The interspaces between the veins turn yellow (Fig. 2). This occurs approximately a fortnight after inoculation. These symptoms are followed by severe deformation, leaflets are often narrow and twisted (Fig. 3). At that time, primary chlorotic lesions appear in inoculated leaves. In later stages infection becomes milder; leaves show deformation and slight decoloration only. Plants which have wintered in a glasshouse are in the following year symptomless and apparently free of virus. (Attempts to transmit the virus from wintered plants failed.) Similar symptoms of a varying degree of severity were demonstrated after the infection of four parsley varieties of Czechoslovak assortment (Dob~enick~, Hans163 KrAtks and OlomouckA E/I). The variety KrAtks losses its symptoms soon after their maximum development. Petroselinum hortense var. crisTum (MILL.): Characteristics of the disease are similar to those mentioned above. Out of 60 inoculated individuals three became infected only. Coriandrum sativum L., coriander: Systemic leaf symptoms which appear after seven days are formed by minute grey spots which coalesce forming enlarged necrotic spots. In an advanced stage of the disease severe top necrosis occurs. Plants which survive grow slowly and are severely stunted. On reaching approximately 10 em they start to flower. Transmissibility of the virus to Coriander is difficult especially to the variety coriander HrubSick~ (3 out of 30 inoculated plants were infected). Pastinaca sativa L., parsnip: Seven to nine days after infection systemic curling and puckering of the youngest leaves followed by the formation of slight diffuse yellowish green areas occur. These symptoms disappear after a fortnight. The virus was transmitted in a very low percentage (5 out of 45 plants were infected). Anethum graveolens L., dill: Systemic transverse yellowish strips in leaves appear 10 days after inoculation. Chenopodium quinoa WILLD. and C. giganteum Do~.: These species develop 76 z. POLAK local lesions on inoculated leaves 7--10 days after inoculation. The lesions appear as minute spots, later showing a light-brown dead centre (Fig. 4 and 5). They enlarge with age, but show no tendency to coalesce. Old lesions are 2--3 mm in diameter. Virus was not recovered from uninoculated leaves. C. ~urale L., sowbane: Sowbane plants form minute, necrotic local lesions. These lesions took longer to form than did the lesions on previous Cheno- Todium species. Systemic symptoms were not noted. Attempts to transmit the virus by the dodder, Cuscuta campestris YU~CXE~ Dodder was established on diseased parsleys and stems of the parasite were trained to healthy parsleys and celeries. Twenty healthy plants of each species were parazitized, but none developed symptoms of the disease. To test the ability of the dodder to acquire virus from infected host plant, juice was extracted from stems growing on an infected and parsley tested on C. quinoa leaves. Results were compared with those obtained with infectious sap from the parsley on which doder was growing. Lesions were not produced in the test with extract from the dodder. Check test was positive. Preliminary tests with an aphid vector Green peach aphids, Myzus persicae SULZ., fed on infectious young parsley seedlings did not transmit the virus to healthy parsley and celery plants. Further detailed studies will be carried out on virus-vector interrelations using green peach aphids and some species specific for umbeliferous plants under various lengths of acquisition and test feeding periods. Thermal inactivation point Thermal inactivation point was determined in crude infectious extract from young parsleys. T~o ml of the sap was placed in injection ampullas and subjected to the desired temperature in a water bath for 10 minutes (45 ~ 50 ~ 55 ~ 60 ~ 65 ~ and 70 ~ C). Contents of ampu]las were immediately after cooling tested on sets of 10 leaves of C. quinoa. The test indicated that the thermal inactiva- tion point lay between 51 ~ and 55 ~ C. (The experiment was twice repeated.) Longevity in vitro Longevity in vitro was studied with the same crude extract used for thermal inactivation. Infectious sap was stored in a stoppered 25 ml flask under room temperature. At 24 hours' intervals 2 ml of sap was removed and tested on 10 C.