Summary
Bacteria-containing commercial products, sold to facilitate biodegradation of human and animal wastes, consist of complex mixtures of bacteria. These mixtures are often of undetermined composition and grown in batch cultures from diverse bacteria-rich inocula of proprietary origins. In order to provide a means of testing for the presence of small numbers of microorganisms, pathogenic to terrestrial or avian wildlife, in bacteria-containing biodegradation products, five DNA extraction protocols were tested for their ability to purify total genomic DNA from nine biodegradation products of different formulations. A diatomaceous earth and guanidine thiocyanate-based DNA extraction method was found to be the most reliable. Fourteen microorganism-specific polymerase chain reaction (PCR)-based assays were developed. Each PCR assay was demonstrated to be specific using DNA from 61 other species of microorganisms (105 different isolates). The mean detection limit for 10 assays using cultured organisms spiked into biodegradation products was assessed. The mean was 1 × 102.62 ± 1 × 101.58 c.f.u.g−1 (bacteria) and 1 × 103.88 ± 1 × 101.14 cells g−1 (protozoa). There were no target wildlife pathogens detected by the 14 PCR assays in unspiked biodegradation products. This study has demonstrated that molecular diagnostic means can be used to detect small numbers of selected wildlife pathogens in complex biodegradation products.
Similar content being viewed by others
References
F.C. Argyros M. Ghosh L. Huang N. Masubuchi D.R. Cave P. Grübel (2000) ArticleTitleEvaluation of a PCR primer based on the isocitrate dehydrogenase gene for detection of Helicobacter pylori in feces Journal of Clinical Microbiology 38 3755–3758 Occurrence Handle11015397
M.D. Braid L.M. Daniels C.L. Kitts (2003) ArticleTitleRemoval of PCR inhibitors from soil DNA by chemical flocculation Journal of Microbiological Methods 52 389–393 Occurrence Handle10.1016/S0167-7012(02)00210-5 Occurrence Handle12531508
R.T. Bryan R.W. Pinner R.L. Berkelman (1994) ArticleTitleEmerging infectious disease in the United States Annals of the New York Academy of Science 740 316–361
C. Burtscher S. Wuertz (2003) ArticleTitleEvaluation of the use of PCR and reverse-transcriptase PCR for the detection of pathogenic bacteria in biosolids from anaerobic digestors and aerobic composters Applied and Environmental Microbiology 69 4618–4627 Occurrence Handle10.1128/AEM.69.8.4618-4627.2003 Occurrence Handle12902250
N.D. Cohen L.J. Martin R.B. Simpson D.E. Wallis H.L. Neibergs (1996) ArticleTitleComparison of polymerase chain reaction and microbiological culture for detection of salmonellae in equine feces and environmental samples American Journal of Veterinary Research 57 780–786 Occurrence Handle8725799
M.A. Cotta T.R. Whitehead D. Zeman (2003) ArticleTitleIsolation, characterization and comparison of bacteria from swine faeces and manure storage pits Environmental Microbiology 5 737–745 Occurrence Handle10.1046/j.1467-2920.2003.00467.x Occurrence Handle12919409
S. Cui C.M. Schroeder D.Y. Zhang J. Meng (2003) ArticleTitleRapid sample preparation method for PCR-based detection of Escherichia coli O157:H7 in ground beef Journal of Applied Microbiology 95 129–134 Occurrence Handle10.1046/j.1365-2672.2003.01951.x Occurrence Handle12807462
J.A. Entry R.E. Sojka M. Watwood C. Ross (2002) ArticleTitlePolyacrylamide preparations for protection of water quality threatened by agricultural runoff contaminants Environmental Pollution 120 191–200 Occurrence Handle10.1016/S0269-7491(02)00160-4 Occurrence Handle12395830
L.H. Lauerman F.J. Hoerr A.R. Sharpton S.M. Shah V.L. Santen Particlevan (1993) ArticleTitleDevelopment and application of a polymerase chain reaction assay for Mycoplasma synoviae Avian Diseases 37 829–834 Occurrence Handle7504919
G. Lindergard D.V. Nydam S.E. Wade S.L. Schaaf H.O. Mohammed (2003) ArticleTitleA novel multiplex polymerase chain reaction approach for detection of four human infective Cryptosporidium isolates: Cryptosporidium parvum, types H and C, Cryptosporidium canis, and Cryptosporidium felis in fecal and soil samples Journal of Veterinary Diagnostic Investigations 15 262–267
J. Lu S. Sanchez C. Hofacre J.J. Maurer B.G. Harmon M.D. Lee (2003) ArticleTitleEvaluation of broiler litter with reference to the microbial composition as assessed by using 16S rRNA and functional gene markers Applied and Environmental Microbiology 69 901–908 Occurrence Handle10.1128/AEM.69.2.901-908.2003 Occurrence Handle12571010
J. McLauchlin S. Pedraza-Díaz C. Amar-Hoetzeneder G.L. Nichols (1999) ArticleTitleGenetic characterization of cryptosporidium strains from 218 patients with diarrhea diagnosed as having sporadic cryptosporidiosis Journal of Clinical Microbiology 37 3153–3158 Occurrence Handle10488169
Millner P., Karns J. (2003). Animal manure: bacterial pathogens and disinfection technologies. EPA Report (in press).
J. Miller A. Jenny J. Rhyan D. Saari D. Suarez (1997) ArticleTitleDetection of Mycobacterium bovis in formalin-fixed, paraffin-embedded tissues of cattle and elk by PCR amplification of an IS6110 sequence specific for Mycobacterium tuberculosis complex organisms Journal of Veterinary Diagnostic Investigations 9 244–249
D.G. Pitcher N.A. Saunders R.J. Owen (1989) ArticleTitleRapid extraction of bacterial genomic DNA with guanidinium thiocyanate Letters in Applied Microbiology 8 151–156
Sambrook J., Fritsch E.F., Maniatis T., (1989) Molecular Cloning. A Laboratory Manual. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory Press, ISBN 0 87969 309 6.
T. Trochimchuk J. Fotheringham E. Topp H. Schraft K.T. Leung (2003) ArticleTitleA comparison of DNA extraction and purification methods to detect Escherichia coli O157:H7 in cattle manure Journal of Microbiological Methods 64 165–175 Occurrence Handle10.1016/S0167-7012(03)00017-4
J.L. Williamson T.E. Rocke J.M. Aiken (1999) ArticleTitleIn situ detection of the Clostridium botulinum Type C1 toxin gene in wetland sediments with a nested pcr assay Applied and Environmental Microbiology 65 3240–3243 Occurrence Handle10388729
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Sibley, J.A., Cross, R.H., Quon, A.L. et al. Development of diagnostic test methods for detecting key wildlife pathogens in bacteria-containing commercial biodegradation products. World J Microbiol Biotechnol 21, 417–423 (2005). https://doi.org/10.1007/s11274-004-1765-8
Issue Date:
DOI: https://doi.org/10.1007/s11274-004-1765-8