Eleanor Shoshany Anderson with Nichole Broderick and Jo Handelsman

Departments of Plant Physiology and Industrial Engineering, UW-Madison

Bacillus thuringiensis (Bt) is a naturally occurring bacterial insecticide, widely used to control pests of economic importance.  One of these pests, Gypsy moth (Lymantria dispar),defoliates more than one million acres of forest annually in this country alone.  Bt is an important tool with which to stop Gypsy moth, because it is very effective and has minimal impact on the surrounding environment.

Bt kills larvae by binding to their gut's epithelial cells, thus causing holes to form in the gut wall.  It is speculated that normal gut bacteria then enter the hemocoel through these holes, causing sepsis and death.  If these bacteria are not present, the Bt is ineffective.  This speculation was arrived at by rearing larvae on antibiotics that eliminated their gut bacteria.

To further understand the effects of antibiotics on Gypsy moth gut bacteria, we assessed the bacterial community’s ability to regrow following antibiotic pressure.  To do this, caterpillars were raised on artificial diet both with and without antibiotics.  Upon reaching the mid-second instar, the larvae raised on antibiotics were taken off the treated diet, thus removing the antibiotic pressure and allowing bacteria to grow in their guts.  To monitor the culturable bacteria, the guts of five larvae were extracted and plated daily for seven days.  These plates were then compared against the guts of larvae that had been raised on the untreated diet.  Molecular methods were then used to assess the total bacterial community in these guts over time.
          Both methods show that early gut samples contain no bacteria following the elimination of antibiotic pressure.  By the second day, bacteria were present (albeit in smaller populations than in the untreated guts), and by the seventh day, the caterpillars had developed communities comparable in size to those of untreated caterpillars.  Based on culturable morphologies, it seems that the bacterial communities that grew in these previously empty guts were different than the bacteria in untreated guts.  Thus it seems that Gypsy moth larval bacterial midgut communities are able to grow in caterpillars reared on, and then taken off of, antibiotics.