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A high-volume screen for anthelmintic microbial metabolites with an avermectinlike mode of action was developed. The primary screen used the free-living nematode Caenorhabditis elegans in a whole-organism assay. The specificity for avermectinlike compounds resides in the secondary screen, which takes advantage of the chloride channel-opening properties of the avermectins. By using standard microelectrode techniques, membrane conductance changes following exposure to extracts of microbial cultures were measured in the walking leg stretcher muscle fibers of the lined shore crab Pachygrapsus crassipes. The avermectins and related milbemycins give a characteristic response of rapid loss of membrane resistance coupled with a slight hyperpolarization of the membrane. This is partially (near 50%) reversible with the chloride channel blocker picrotoxinin. Four morphologically similar cultures that produced avermectinlike activities were identified by this screen. Isolation of the active components from one of these cultures (strain UC 8984) followed by nuclear magnetic resonance spectroscopy resulted in the identification of milbemycins alpha 1 and alpha 3. These metabolites are members of a large family of milbemycins produced by Streptomyces hygroscopicus subsp. aureolacrimosus NRRL 5739. Systematic studies revealed that strain UC 8984 is also a S. hygroscopicus strain, but which is taxonomically distinct from NRRL 5739.

antimicrobial agents and chemotherapy

Development of a mechanism of action-based screen for anthelmintic microbial metabolites with avermectinlike activity and isolation of milbemycin-producing Streptomyces strains