Colonization of Greenhouse Nematode Cultures by Nematophagous Mites and Fungi

Supplement to Journal of Nematology 25(4S):789-794. 1993. © The Society of Nematologists 1993. Colonization of Greenhouse Nematode Cultures by Nematophagous Mites and Fungi D. E. WALTER, 1 D. T. KAPLAN, 2 AND E. L. DAVIS 3 Abstract: Unproductive >7-year-old greenhouse cultures of citrus nematode (Tylenchulus semipenetrans) had a well-developed soil invertebrate fauna that included nematophagous mite species characteristic of Florida citrus groves. Nematophagous mite densities in box cultures were 285 - 42 mites/liter, 2.5 to 25 times higher than densities in citrus nematode-infested groves. Vigorous root- knot nematode (Meloidogyne incognita) cultures grown in steam-pasteurized soil had few nematophagous mites until more than 3 months after inoculation. Mite species diversity had a significant (P < 0.0001) positive linear relationship with culture age that explained about one-half the variance in species number. Nematophagous mite densities rose and then fell with culture age. In root-knot cultures >3-months-old, mite densities often exceeded 1,000 mites/liter. Twelve species of nematophagous fungi also were isolated from greenhouse nematode cultures. Key words: biological control, culture, fungus, mite, nematode, nematophagous fungus, Meloido- gyne incognita, Tylenchulus semipenetran~. Monoxenic culture techniques have Walter and Ikonen (13) found that two been developed for a relatively limited species of ProtogamaseUus could consume number of plant-parasitic nematodes, and an average of 0.82 ~g (dry weight) of these techniques are often labor-intensive nematodes per day. Long-term cultures of or do not yield large numbers of nema- the citrus nematode, Tylenchulus semipene- todes (8). Thus, it is often necessary to cul- trans (Cobb), often yield population densi- ture plant-parasitic nematodes on the ties far lower than anticipated (Kaplan, un- roots of potted plants (greenhouse cul- publ.). Because nematophagous mite spe- tures). Population densities within green- cies were associated with citrus roots in a house cultures are dependent upon suit- survey of Florida citrus groves (14), we de- able edaphic conditions and the availability cided to examine the arthropod commu- of roots; however, densities can be unex- nity in greenhouse nematode cultures. pectedly low, and occasionally cultures fail. The purpose of this paper is to report on Nematophagous mites and fungi are com- antagonists that have colonized green- mon in soils, and should not be present house cultures of the root-knot nematode, initially in cultures established in steam- Meloidogyne incognita (Kofoid & White) pasteurized potting soils, but greenhouse Chitwood, and of the citrus nematode. cultures of plant-parasitic nematodes MATERIALS AND METHODS would be rich resources for any antagonists that were able to invade them. Nematode cultures: In June 1983, five Nematophagous mites are capable of wooden slat boxes 45 cm wide, 57 cm long, suppressing populations of plant-parasitic and 25 cm deep were filled to 18-cm depth nematodes in pot experiments (3,11,12). (46,170 cm 3) with 50% Astatula sand (hy- perthermic, uncoated; typic, quartzipsam- ments), 25% peat moss, and 25% vermic- ulite infested with from a Received for publication 9 June 1993. T. semipenetrans t Lecturer, Department of Entomology and Centre for variety of sources, and planted with 4-12 Tropical Pest Management, University of Queensland, St. Lucia, Queensland 4072, Australia. rough lemon (Citrus limon L.) seedlings in a Supervisory Plant Pathologist, USDA ARS, U.S. Horti- greenhouse at the U.S. Horticultural Re- cultural Research Laboratory, 2120 Camden Road, Orlando, search Laboratory (USHRL) in Orlando, FL 32803. s Assistant Professor, Department of Plant Pathology, Florida. The cooling and heating regime North Carolina State University, Raleigh, NC 27612-7616. Mention of a trademark, warranty, proprietary product, or in the greenhouse maintained a soil tem- vendor does not constitute a guarantee by the U.S. Depart- perature of 25 C -+ 5 C with air tempera- ment of Agriculture and does not imply its approval to the exclusion of other products or vendors that may also be suit- tures of 25 -+ 5 C, and the boxes were wa- able. tered daily by an automated system. 789 790 Journal of Nematology, Volume 25, Supplement to December 1993 In September 1985, existing cultures of arthropods. Other samples were taken as M. incognita race 1 from the University of needed to obtain live animals or inoculum Florida, Gainesville (UFG), were trans- for sampling nematophagous fungi. All ferred to the USHRL. Cultures were initi- mite species except Neocunaxoides andrei ated at USHRL monthly with eggs and sec- Baker & Hoffman and Proctolaelaps sp. de- ond-stage juveniles (]2) extracted from tected in nematode cultures were placed in 2-3 month-old UFG cultures with 0.53% small arenas (13) containing 500 burrow- sodium hypochlorite (4). Eggs were placed ing nematodes, Radopholus citrophilus Huet- on Baermann pans at 25 C, from which tel, Dickson, & Kaplan, reared on carrot approximately 10,000 J2 and eggs were disk culture (7). Voucher specimens of used to infest pots containing 1-month-old mites were deposited at the Florida De- pepper (Capsicum annuum L.) cv. California partment of Agriculture and Consumer Wonder or eggplant (Sotanum melongena Services, Division of Plant Industry, L.) cv. Black Beauty. Plants were potted in Gainesville, Florida, and with the Cana- 15-cm-d clay pots filled with steam- dian National Collection, Biosystematics pasteurized (60 minutes at 66 C) Astatula Canada, Ottawa, Ontario. sand. Young plants and established cul- Root-knot nematode populations in pot tures were maintained in a greenhouse cultures were estimated on 19 July 1989 during the colder months. During the from 3 equidistant 2.1-cm-d cores taken to summer, cultures were maintained in a 7-cm depth. The three cores from each pot screen house and watered daily. Cultures were bulked, roots were removed with a were identified by the month in which they 6-mm-pore sieve, and a 30-cm 3 subsample were infested, i.e., cultures infested in was processed with a sieving-centrifuga- June were subsequently referred to as tion technique (6). Approximately 30 cm "June series" cultures. of root segments from each sample were Arthropod and nematode sampling: Begin- inspected for galls at x40 magnification ning in July 1989, nematode cultures were for an index of root-knot nematode infec- sampled for arthropods with a 2.1-cm-d tion (galls per cm root). Statistical analyses cork borer. Six evenly spaced cores were for mite and nematode data were per- taken to 8.0-cm depth from box cultures, formed using the GLM procedure of SAS and three equidistant cores to 7.0-cm (9). depth from pot cultures. The holes were Nematophagous fungi: Cores containing refilled with the appropriate potting soil. soil and roots from box and pot cultures The cores were bulked, and arthropods were taken for extraction of nematopha- were extracted with Tultgren funnels (14). gous fungi by a sprinkle-bait plate method Box cultures were sampled for arthropods (14) from 0.15 g of soil shaken from roots in July, October, and December 1989 and or small nematode-infected root segments January, February, and March 1990. On placed on a thin layer of water agar (WA) 19 July, three uninfected pepper plants, (4 ml of 1% agar) in 9-cm-d petri dishes. and equal numbers ofM. incognita cultures Following the addition of 5,000 burrowing on pepper (2 or 3 cultures) and eggplant (2 nematodes derived from carrot disk cul- or 3 cultures) were arbitrarily chosen at 41 ture (7) to each dish, they were incubated (June series), 72 (May series), 103 (April at 25 C. In addition, four samples of 5,000 series), and 134 (March series) days after root-knot nematode eggs and J2 prepared being infested with root-knot nematodes. as for infestation of new nematode cul- Six samples (n = 6) were collected from tures were plated on WA dishes and incu- each nematode culture for arthropod anal- bated at 25 C for 5-7 days. ysis, and four samples (n = 4) were collected from each nematode culture to esti- RESULTS mate nematode population densities. Over the next 20 weeks, the May and Nematophagous fungi: The root-knot June series were periodically sampled for nematode pot cultures contained 12 spe- Colonization of Nematode Cultures: Walter et al. 791 cies of nematophagous fungi, including In contrast, both series of root-knot Arthrobotrys dactyloides Drechsler, A. musi- nematode pot cultures were dominated by formis Drechsler, A. oligospora Fresenius, the mites Lasioseius subterraneus Chant, Ge- Catenaria anguillulae Sorokine, DactyleUa olaelaps sp., Coleoscirus simplex (Ewing), cionapaga Drechsler, D. eUipsospora Grove, Gamasellodes bicolor (Berlese), G. rectiventris Dactylaria eudermata Drechsler, Haptoglossa Lindquist, and Protogamasellus mica (Ath- heterospora Drechsler, Myzocytium sp., Paeci- ias-Henriot), whose combined totals repre- lomyces lilacinus (Thom.) Samson, Rhizophi- sented 97.3 and 79.2% of the nematopha- dium sp., and Verticillium chlamydosporium gous mites in the May and June series, re- Goddard. Five species of nematode-trap- spectively. Only P. mica and Neoscirula sp. ping fungi (all of the above except D. were relatively abundant in both ckrus and eudermata) and the egg parasite V. chlamy- root-knot nematode cultures. dosporium were recovered from the four In July 1989, nematophagous mite den- samples of root-knot nematode inoculum. sities in root-knot nematode pot cultures Arthrobotrys dactyloides, C. anguiluUae, D. cio- were variable (Table 2). Uninfested potted napaga, D. ellipsospora, and an unidentified plants had nearly undetectable densities of Oomycete were isolated from citrus nema- mites, and even 41 days after nematode tode box cultures. infestation, few pots had been colonized by Nematophagous mites: Twelve species of mites. Numbers of root-knot nematode J2 nematophagous mites were isolated from and root-knot galls per cm root were not greenhouse nematode cultures (Table 1), significantly correlated with nematopha- and all except two very rare species, Neo- gous mite density.

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