RESUMO
Esca and petri diseases are important grapevine trunk diseases in South Africa and most other grape-producing countries. The causal pathogens are Phaeomoniella chlamydospora and several species of Phaeoacremonium. In total, 25 species of Phaeoacremonium have been isolated from grapevines of which seven species have been linked to Togninia teleomorphs obtained through in vitro mating studies (3). Of these species, only perithecia of T. minima, T. fraxinopennsylvanica, and T. viticola have been found on grapevines in California (1,2,4). T. minima is heterothallic, and although both mating types are present in South African vineyards, perithecia have never been observed (3). In the current study, grapevine cordons and trunks were collected from vineyards and rootstock mother vines within Western Cape Province for examination in the laboratory under a dissecting microscope. The grapevines displayed general decline symptoms, including reduced vegetative growth, dead or dying shoots and cordons, as well as internal vascular streaking and/or a red/black/brown margin next to decayed wood typically associated with esca and petri disease. Rootstock mother vines were apparently healthy, although many old, cracked pruning wounds were visible. Togninia-like perithecia with distinctive long necks were found along the wood crevices, often on old pruning wounds. The perithecia were removed and placed on microscope slides with sterile water. Structures were measured and slides were washed with 500 µl of sterile water onto potato dextrose agar amended with chloramphenicol (250 mg/liter). Ascospores were allowed to germinate overnight to obtain single ascospore colonies. Perithecia were found on cultivars Muscat d' Alexandrie and Pinotage (Vitis vinifera) at Stellenbosch in May 2011 and on Ramsey (V. champinii) rootstock mother vines at Slanghoek in June 2012. Perithecia were globose to subglobose, black, and often embedded in the wood tissue but also present on the surface of the wood. The length of the necks was 250 to 300 × 47.5 to 55 µm. The asci were hyaline and ranged from 16 to 25 × 3.5 to 5 µm. Ascospores were hyaline, ellipsoid, and ranged from 5 to 6 × 1.5 to 2 µm. These measurements were similar to those reported by Mostert et al. (3) and Rooney et al. (4). Colony growth was typical of T. minima. DNA was extracted from the colonies and the partial betatubulin gene was amplified and sequenced using the primers T1 and Bt2b. Sequences were deposited into GenBank (JX962864 to 67). Based on a megablast search of the NCBI's GenBank nucleotide database, 100% similarity was found with other T. minima sequences (JQ691670.1, HQ605018.1, HQ605014.1; identities = 647/647 [100%], gaps = 0/647 [0%]). To our knowledge, this is the first report on the occurrence of T. minima perithecia on grapevines in Western Cape Province of South Africa. The removal of dead spurs and cordons will be instrumental in lowering the inoculum originating from perithecia, especially in rootstock mother blocks where no control strategies are applied for petri disease or esca. Spore trapping studies are currently in progress to study spore release patterns in order to determine whether pruning wounds are at risk during traditional pruning periods. References: (1) A. Eskalen et al. Plant Dis. 89:528, 2005. (2) A. Eskalen et al. Plant Dis. 89:686, 2005. (3) L. Mostert et al. Stud. Mycol. 54:1, 2006. (4) S. Rooney-Latham et al. Plant Dis. 89:867, 2005.
RESUMO
Biological control is a promising non-chemical approach for the control of gastrointestinal nematodes of sheep. Use of combinations of biocontrol agents have been reported to be an effective method to increase the efficacy of biological control effects. In this study, combinations of either two Bacillus thuringiensis (Bt) or Clonostachys rosea (C. rosea) isolates and Bt+C. rosea isolates were evaluated in vitro in microtitre plates for their biocontrol activity on sheep nematodes. The Baermann technique was used to extract the surviving L3 larval stages of intestinal nematodes and counted under a dissecting microscope to determine the larval counts. Results indicate that there was a significant reduction of nematode counts due to combination of biocontrol agents (P<0.001). Combinations of Bt isolates reduced nematodes counts by 72.8%, 64% and 29.8%. The results revealed a control level of 57% when C. rosea isolates P3+P8 were combined. Combination of Bt and C. rosea isolates B10+P8 caused the greatest mortality of 76.7%. Most combinations were antagonistic, with only a few combinations showing an additive effect. None were synergistic. The isolate combinations were more effective than when isolates were used alone.