Journal of Nematology 32(4):362–369. 2000. © The Society of Nematologists 2000. Bacterial-Feeding Nematode Growth and Preference for Biocontrol Isolates of the Bacterium Burkholderia cepacia Lynn K. Carta1 Abstract: The potential of different bacterial-feeding Rhabditida to consume isolates of Burkholderia cepacia with known agricultural biocontrol ability was examined. Caenorhabditis elegans, Diploscapter sp., Oscheius myriophila, Pelodera strongyloides, Pristionchus pacificus, Zeldia punctata, Panagrellus redivivus, and Distolabrellus veechi were tested for growth on and preference for Escherichia coli OP50 or B. cepacia maize soil isolates J82, BcF, M36, Bc2, and PHQM100. Considerable growth and preference variations occurred between nematode taxa on individual bacterial isolates, and between different bacterial isolates on a given nematode. Populations of Diploscapter sp. and P. redivivus were most strongly suppressed. Only Z. punctata and P. pacificus grew well on all isolates, though Z. punctata preferentially accumulated on all isolates and P. pacificus had no preference. Oscheius myriophila preferentially accumulated on growth- supportive Bc2 and M36, and avoided less supportive J82 and PHQM100. Isolates with plant-parasitic nematicidal properties and poor fungicidal properties supported the best growth of three members of the Rhabditidae, C. elegans, O. myriophila, and P. strongyloides. Distolabrellus veechi avoided commercial nematicide M36 more strongly than fungicide J82. Key words: accumulation, attraction Caenorhabditis elegans, Diploscapter sp., Distolabrellus veechi, ecology, Escherichia coli OP50, nutrition, Oscheius myriophila, Panagrellus redivivus, Pelodera strongyloides, phylogeny, Pristionchus pacificus, repellence, Rhabditida, toxicity, Zeldia punctata. Bacterial-feeding nematodes are impor- pacia (ex Burkholder) Palleroni and Holmes, tant for soil-nutrient cycling in agricultural 1981). Current taxonomic opinion suggests systems (Freckman and Caswell, 1985). this is a polyphyletic species (Govan et al., Plant productivity might be affected if ap- 1996) where some isolates are plant or op- plied biocontrol bacteria were toxic to these portunistic-human pathogens (Hales et al., common non-target organisms. Alterna- 1998). Isolates of this gram-negative bacte- tively, some soil bacterial-feeding nematodes rium are commonly found in soil and water. may consume applied biocontrol bacteria. A number of biocontrol strains with excel- Acrobeloides nanus is known to feed on and lent rhizosphere-colonizing ability have reduce the field efficacy of the introduced been isolated from maize rhizospheres bacterial-biocontrol agent, Pseudomonas cor- (Hebbar et al., 1992a). rugata, against Gaumannomyces graminis, the Burkholderia cepacia isolate M36 is regis- take-all fungus of wheat (Ryder and Bird, tered with the U.S. Environmental Protec- 1983). tion Agency as a nematicide (Stine Micro- Another agriculturally important bacte- bial Products, marketed by Market VI, L. L. rium applied as a biocontrol agent is Burk- C., Shawnee, KS) against plant-parasitic holderia cepacia (ex Burkholder) Yabuuchi, nematodes except for cyst nematodes (Noel, Kosako, Oyaizu, Yano, Hotta, Hashimoto, 1990) but has shown variable nematicidal ac- Ezaki and Arakawa, 1993 (= Pseudomonas ce- tivity (J. O. Becker, pers. comm.). Burkhold- eria cepacia isolates Bc2 (Meyer et al., 2000) Received for publication 1 November 1999. and BcF (unpub.) reduced populations of 1 Research Plant Pathologist, USDA, ARS, Nematology Labo- root-knot nematodes. A soil isolate of B. ce- ratory, Bldg. 011A, Rm. 165B, BARC-West, 10300 Baltimore Ave., Beltsville, MD 20705-2350. pacia that was not associated with maize sup- Mention of trade names or commercial products in this pub- ported good population growth of the bac- lication is solely for the purpose of providing specific informa- tion and does not imply recommendation or endorsement by terial-feeders Acrobeloides sp. (Cephalobidae) the U. S. Department of Agriculture. and Pristionchus lheritieri (Neodiplogastri- The author thanks S. Ochs and K. Campbell for lab help; P. Martin, D. P. Roberts, and K. P. Hebbar for advice; V. Illum, D. dae) (Anderson and Coleman, 1981; Ander- P. Roberts, K. P. Hebbar, W. Mao, D. H. A. Fitch, and the Caenorhabditis Genetics Center for cultures. The author also son et al., 1981). However, nothing is known thanks S. L. F. Meyer, K. P. Hebbar, L. Levy, and E. Bernard for about the effects of maize-associated isolates manuscript review. E-mail: [email protected] with pesticidal properties on non-target bac- This paper was edited by P. Grewal. terial-feeding nematode survival or bacterial 362 Growth and Attraction for Burkholderia cepacia: Carta 363 consumption. Part of the variable plant- as noted. They include Oscheius myriophila parasitic nematode control may be a result DF5020 (Rhabditidae: Rhabditinae) (from of bacterial-feeding nematodes using B. ce- D. Fitch, New York University, New York, NY pacia as an energy source. Nematicidal iso- 10003); Caenorhabditis elegans N2 (Rhabditi- lates, however, might kill or repel bacterial- dae: Peloderinae) and Panagrellus redivivus feeders before the nematodes can consume PS1163 (Panagrolaimidae) (from the Cae- substantial amounts of bacteria. Therefore, norhabditis Genetics Center (CGC), Univer- one objective of this study was to determine sity of Minnesota, 1445 Gortner Ave., St. whether different B. cepacia isolates might Paul, MN 55108); Diploscapter sp. PS2123 differentially attract and support growth of (Diploscapteridae); Pelodera strongyloides different nematode species. The null hy- PS1129 (Rhabditidae: Peloderinae); Pristion- pothesis is that all isolates equally support chus pacificus PS312 (Neodiplogastridae); growth and do not repel all bacterial- Distolabrellus veechi LKC10 (Rhabditidae: Me- feeding nematodes. sorhabditinae); and Zeldia punctata PS1192 None of the B. cepacia soil isolates tested (Cephalobidae). Voucher specimens are de- so far have shown human-pathogenic poten- posited in the U.S. Department of Agricul- tial (Hales et al., 1998), but the bacterium is ture Collection, Beltsville, Maryland. known to be highly mutable and resistant to All nematode stock cultures were grown at antibiotics. Under certain conditions, nor- 20 °C on NGM agar spotted with Escherichia mally benign environmental B. cepacia coli OP50 (CGC, St. Paul, MN) grown in L- strains might be transmissible to cystic- Broth at 37 °C and transferred from a stock fibrosis patients. Because of this concern for LB agar plate (Stiernagle, 1999). Besides its B. cepacia strains in the agricultural environ- use for stock cultures, E. coli is used here ment (Holmes et al., 1998), nematodes that experimentally as a bacterial control to com- consume this bacterium may be useful pare population growth or relative prefer- agents for environmental remediation. ence with B. cepacia isolates. Better discrimination among agricultur- All B. cepacia cultures used in this study ally and medically important B. cepacia iso- were isolated from maize rhizospheres and lates and their modes of action might also have demonstrated biocontrol potential. be obtained from Caenorhabditis elegans mu- These include antifungal isolates J82 (EPA tants with pharmacologically interesting registration no. 006419) (Mao et al., 1997), genes (Rand and Johnson, 1995). There- BcF (Mao et al., 1998), PHQM100 (Hebbar fore, a second purpose of this study was to et al., 1998), and plant-parasitic nematode quantify growth support of C. elegans on B. suppressive isolates M36 (nematicide, EPA cepacia isolates as a basis for future genetic or registration no. 006464, Stine Microbials, toxicological tests. The population quantifi- Adel, IA) and Bc2 (Meyer et al., 2000). Burk- cation also serves as a check on the reliability holderia cepacia sources and alternate isolate of the growth-rating scale for this nematode. designations with American Type Culture Collection (ATCC) numbers and Agricul- Materials and Methods tural Research Service (USDA-ARS) num- bers include (i) Stine Microbials, Adel, Iowa, The bacterial-feeding nematodes used in for Wisconsin isolates J82 (alternate desig- the assays were selected to represent distinct nations = ATCC 51993 or ARS BcB) and groups within the Rhabditida based on mo- M36 (= ATCC 51995 or ARS B18379); (ii) D. lecular (Baldwin et al., 1997; Blaxter et al., P. Roberts and K. P. Hebbar, USDA, Belts- 1998) and morphological criteria (Andrassy, ville, Maryland, for Maryland isolates ARS 1984; Sudhaus, 1976). Nematodes used in Bc2 and ARS BcF; and (iii) K. P. Hebbar for the study were isolated and identified by the French isolate PHQM100. author in the current lab (LKC) or in the Bacterial stock cultures were grown Sternberg lab (PS) at the California Institute monthly on nutrient agar plates incubated of Technology, Pasadena, California, except for 24 hours at 30 °C. For experimental use, 364 Journal of Nematology, Volume 32, No. 4, December 2000 most bacteria were grown for 24 hours in as evidenced by severely reduced optical nutrient broths (Difco Laboratories, De- density. troit, MI) to achieve densities of 106 to 107 Preference plate tests: Nematodes were CFU/ml. Isolates Bc2 and M36 required an grown to adulthood on lawns of E. coli OP50 extra 12–24 hours to reach this density. on NGM agar plates and rinsed once with Burkholderia strains were grown at 30 °C and sterile distilled water from plates recently E. coli at 37 °C. depleted of bacteria to reduce interference