Techniques for Controlling Baterial Flora of Pristionchus lheritieri.

Since 1903, when Metcalf (5) described a method for obtaining and rearing bacteriafree Rhabditis brevispina (Claus) Biitschli, many procedures have been published. Glaser and Stoll (6) used sodium hypochlorite for surface-sterilization of nematodes. Dougherty and Calhoun (2, 3) eliminated the microflora of R. pellio (Schneider) Btitschli with antibiotics and then substituted Escherichia coli. The described techniques are a combination of the above and innovations adapted to our work with Pristionchus lheritieri (Maupas) Paramonov. Surface sterilization of P. lheritieri was tested by exposing them to different concentrations of chlorine prepared from commercial sodium hypochlorite. Free chlorine in these solutions was determined by combined iodometric (1) and colorimetric tests. The nematodes were immersed in various chlorine dilutions for scheduled time periods. The treatments were terminated by adding sterile sodium thiosulphate to the chlorine solutions containing the nematodes to neutralize the chlorine. The nematodes were removed from this mixture and observed for motility, a measurement of survival. Control and exchange of bacterial flora in nematodes was accomplished by transferring surface-sterilized nematodes individually to Petri dishes with nutrient agar (3 g of beef extract, 5 g of peptone and 15 g of agar in one liter of distilled water) that had