Date of Award
Master of Science in Microbiology
Victor J. Cabelli
Clostridium perfringens is an anaerobic, spore-forming bacterium that is consistently associated with human fecal wastes. Two extraction-separation procedures were developed for use in conjunction with the mCP membrane filtration method for the enumeration of C. perfringens spores in bottom sediments. In the simpler of the two procedures a distilled water suspension of the sediment sample is pulse sonicated for 10 seconds and allowed to settle. Portions of the supernatant are removed and assayed by the mCP membrane filtration method. This procedure is recommended for general usage. A more complicated procedure using a two-phase separation system was also developed for use in situations where the presence of "toxic" materials are suspected or where relatively low spore densities are present in fine silts. The recoveries of C. perfringens spores by the two procedures, when used in conjunction with the mCP method, were comparable to each other and significantly greater than those by the British most-probably-number (MPN) metho. It was estimated that over 85 percent of the spores were recovered by the procedures. The precision of the “sonicate-and-settle” – mCP method, as determined from the coefficients of variation for paired assays on 87 sediment samples was markedly better than that theoretically obtained by the MPN method and approached that theoretically attributed to counting an average of 85 colonies on each of two plates.
Monitoring the sediments underlying coastal and estuarine waters for the spores of C. perfringens could provide rather sensitive data on the long term trends in the deposition and movement of sewage sludge. Furthermore, the comparison of C. perfringens spore densities to ��hose of the less "hardy" indicators, the fecal streptococci and coliforms, could provide information on the nature and proximity of fecal inputs to aquatic environments. Another possible application would be it's use as a "naturally occurring" conservative tracer to examine the in situ biological decay of fecal pathogens and indicator bacteria. Some of these ideas were explored using C. perfringens density data obtained from analysis by the extraction-separation methods of sediment samples from two locations, Narragansett Bay, Rhode Island and the Nev York Bight. The C. perfringens spore densities in Narragansett Bay sediments were highest in the upper reaches of the Providence River near the major sources of municipal sewage discharges into the bay and decreased with increasing distance from these sources. The highest C. perfringens densities in the Nev York Bight were found in sediment samples collected from the deeper waters to the vest and southwest of the dump site. The limited data available suggest little movement of the sludge shoreward to the north and vest beyond the 20 meter contour line.
Emerson, David J., "The Monitoring of Clostridium perfringens Spore Density in Bottom Sediments and It's Applications" (1982). Open Access Master's Theses. Paper 2389.