Hyalesthes obsoletus is an active vector of Wood blackening in the Republic of Moldova

Hyalesthes obsoletus (Signoret) is an important vector of Wood Blackening in the Republic of Moldova. To identify the causes of the epidemiology of this disease in vineyards, the period of flight and the preference of the host plant H. obsoletus in the field were studied. Monitoring of vineyards showed that this species was found on the Field Bindweed (Convolvulus arvensis L.), and during the period of mass flight, the leafhopper was observed on Xanthium strumarium and Gorets bindweed (Fallopia convolvulus L.). The captured leafhoppers were diagnosed with the presence of Bois Noir stolbur phytoplasm (STOL). The adults of H. obsoletus were also caught from other herbaceous plants: Povoy fence (Calystegia sepium L.), Garden quinoa (Ariplex hortensis L.), Sow thistle (Sonchus oleraceus L.), Medicinal dandelion (Taraxacum officinale L.), as well as c Stinging nettle (Urtica dioica L.). Changes in climatic conditions in the region, causing premature drying of the grass cover, forces cicadas vectors to switch to actively growing plants during this period, including grapes, which contributes to the widespread of the disease.


Introduction
In the vineyards of Moldova, a disease of phytoplasmic etiology such as "jaundice" (yellows) has become widespread. The most famous representatives of this type of disease are Golden yellowing (Flavescence doree, FD -from French) and Blackening of wood (Bois noir, BNfrom French) showing identical symptoms on the affected vine. A characteristic sign of phytoplasmic damage to the vine is yellowing of the leaves and curling their edges downwards in whites and reddening in red grapes. The leaves become firm and brittle to the touch. The next distinguishing feature is the delay in the ripening of the vine. Affected shoots do not ripen and remain green until late autumn. Affected shoots, as a rule, do not yield because the inflorescences dry out and fall off. If the infection occurred at a later period, then the set berries wrinkle due to partial or complete dehydration of the bunches.
Under the natural conditions, leafhoppers are carriers of phytoplasmic diseases of grapes. Thus, the vector of FD grapes is the leafhopper Scaphoideus titanus (Ball). The insect lives and eats only on grapes, without causing any harm to the grape plant. Its harmfulness lies in the transfer of a dangerous disease. It is a highly mobile and efficient carrier that is largely responsible for the spread of Golden Yellowing. Therefore, FD, in the presence of a leafhopper Sc. titanus, can become epidemic.
Vine disease transfer BN belonging to the species Ca. Phytoplasma solani -stolbur phytoplasma (STOL) and belongs to the ribosomal group -16SrXII -A, is associated with the leafhopper H. obsoletus [1,2]. The grape plant is a random feeding place for the leafhopper [3][4][5]. The leafhopper transfers the stolbur phytoplasma to various weeds and cultivated herbaceous plants [6][7][8]. The transfer of phytoplasma by the vector is carried out as follows: the leafhopper, feeding on an infected plant, absorbs phytoplasma with plant juice, entering the food tract, and then into the body of the insect. Phytoplasma in the body of an insect acquires a latent form and incubates for 1 to 4 weeks. During this time, it multiplies and parasitizes most of the tissues and organs of the insect, then reaches the salivary glands. Starting from this period, the leafhopper, each time feeding on a healthy plant, infects it [9]. Phytoplasmas belonging to the 16SrXII -A group infects many cultivated plants [10]. In Europe, BN disease is transmitted to the grape plant from bindweed [6] and nettle [11,12], however, with the growth of other weeds on plantations, cases of the spread of infection have been recorded [13,14].
The high harmfulness of diseases of grapes of phytoplasma etiology necessitates a comprehensive study in order to develop effective control measures. To solve the problem, it is necessary: to identify the pathogen, to establish the vector of the disease and the factors contributing to the spread on the vineyards of the Republic of Moldova.

Materials and methods
The work was carried out in the Laboratory of Virology and Phytosanitary Control at the Institute of Horticulture, Viticulture and Food Technologies (IŞPHTA).
Monitoring of cicadas was carried out by taking samples from plants located in vineyards and adjacent territories with a total area of more than 10 hectares in the Ialoveni and Kaushany districts, using a trapping net and an entomological aspirator, as well as using yellow sticky traps. Monitoring was carried out annually from May to September. The traps were changed and the entomological net (30 cm in diameter) was caught every 10 days. Traps were installed both along the edges of the vineyards and in the center of the cages. Insects were caught, both from grape plants and from the grass cover in the vineyard and adjacent territories (height above the soil level 0-50 cm), one catch included 50 strokes.
The leafhoppers were photographed using a digital microscope camera. The leafhoppers were identified using the guides Anufriev G.

Results and discussion
In the period from 2018-2020, expeditionary surveys of vine plantations were carried out on an area of about 800 hectares in all zones of its cultivation. The aim was to study the geography of the spread of phytoplasmic disease in vineyards and to take samples for testing and identification of the pathogen. Surveys were carried out from September to October. In white grape varieties (Chardonnay, Riesling, Sauvignon, Aligote, etc.) during this period, the leaves were golden-yellow in color, hard to the touch and curled downward. The leaves of the Chardonnay variety often showed necrotic spots, and the leaves themselves were twisted to such an extent that they had a triangular shape (Fig. 1).
Symptoms of the disease in varieties with red berries (Cabernet-Sauvignon, Merlot, Pinot black, Feteasca Neagra, Rara Neagra, Moldova, etc.) were observed in the form of reddening of the leaf blades. On the affected shoots, leaves were often found, on which reddening occupied only one sector, limited by two or three veins (Fig. 1). Diseased shoots were usually green, with no signs of ripening and no harvest. On shoots with symptoms, an unconditioned crop was observed, the berries were shriveled due to partial or complete dehydration.  (Fig. 2). To establish a causal relationship between grapes, vectors and weeds, leafhoppers were caught from plants growing on vineyards and nearby areas. The captured leafhoppers were tested by the Nested PCR method for the presence of phytoplasmic pathogens FD and BN (Table 1).  Phytoplasma solani, 16SrXII), which causes BN disease on the vine, was previously found in herbaceous weeds, such as Field Bindweed, Highlander Bindweed, Mary's whole-leaved, as well as in Rose and Elm, which allowed them to be attributed to reserve plants phytoplasma [15]. The life cycle of the insect H. obsoletus is mainly associated with weeds. The female lays eggs in August in the soil at the base of the stem of the host plant (Fig.3). The hatched larvae feed on the roots of the host plant. If the plant is a reserve of phytoplasma, then the larvae, feeding on the roots of this plant, acquire phytoplasma and become viroform. Imagoes formed from viroform larvae are also viroform. It should be noted that in the Republic of Moldova, over the past 10-15 years, there has been significant climate change. According to meteorological observations over the past four years (2017-2020), the average annual air temperature has increased by 2.2 °C compared to the average annual data