SBIR-STTR Award

DNA hybridization probes specific for common pathogenic bacteria have been developed. The long-term objective of this research is the perfection of simple protocols and a kit using DNA hybridization probes for the detection and identification of pathogens in clinical specimens. The specific aim of this project is to assess the feasibility of this approach to pathogen identification in clinical specimens by: (1) determining numbers and methods of recovery of pathogens in clinical specimens; (2) assessing the facility with which they may be lysed and their DNA prepared for hybridization; (3) determining the utility of procedures developed for rapid hybridization; and (4) assessing probe modifications to enhance delectability. The methods employed include cloning and identifying bacterial DNA fragments with unique base sequences, developing procedures for lysing bacteria and recovering DNA, developing protocols for rapid DNA-DNA hybridization, and detecting hybrid complexes. Successful completion of Phase I will demonstrate that DNA hybridization procedures can accurately and quickly detect and identify pathogens in clinical material. With feasibility demonstrated, Phase 11 development will require the cloning and testing of additional probes specific for pathogens encountered in infectious disease, modifying and adapting procedures for different types of clinical specimens, and developing alternative methods for detection of hybrid complexes. The application of recombinant DNA technology to diagnosis of infectious disease represents a fundamental departure from current microbiological identification procedures and should provide unsurpassed accuracy in organism identification. Development of this procedure into a simple, reliable kit has major commercial potential.Institute Of Allergy And Infectious Diseases

The primary goal of this research is to adapt our improved DNA hyridization technology to the identification of select disease agents compatable with current levels of sensitivity of available detection systems. Tests to be developed to commercial kits include Herpes simplex I and II, Cytomegalovirus, Mycobacterium sps., and bacterial/viral agents of infectious diarrhea. The specific aim of this proposal is to develop and clinically evaluate DNA probes and methodology for each disease entity by: 1.) identifying and producing DNA probes for those infectious agents by recombinant DNA techniques; 2.) expand and refine the sensitivity of the detection system using both radiometric and non-radiometric methodologies; 3.) improve the current test methodology. Initial tests will require primary culture. As progress in these three areas is made on improving test sensitivity, our long term objective is to incorporate these advances into a series of new infectious disease diagnostic tests as a result of increasingly greater sensitivity, thus reducing the time necessary for primary culture and ultimately allowing the test to be performed directly on the clinical specimen. Application of DNA probes to infectious disease in this manner should provide unsurpassed accuracy in organism identification. Development of this procedure into a simple, reliable kit has major commercial potential.

Thesaurus Terms:
bacteria, actinomycetales, mycobacterium, communicable diseases diagnosis, diagnosis, diagnostic tests, design, development and evaluation of diagnostic tests, microbial identification and classification (techniques), microbial identification and classification, viral typing and identification, nucleic acids, dna probe, nucleic acids, synthetic nucleic acids, hybrid nucleic acids, viruses, herpesviridae, alphaherpesvirinae, human (alpha) herpesvirus 1, viruses, herpesviridae, alphaherpesvirinae, human (alpha) herpesvirus 2, viruses, herpesviridae, betaherpesvirinae, genetic manipulation bacteria, streptococcaceae, streptococcus group a, bacterial diseases, actinomycetales infections, bacterial diseases, enterobacteraceae, blood toxicology, bacteremia-septicemia, gastrointestinal disorders, diarrhea, nucleic acids, dna bacterial, nucleic acids, dna viral, virus diseases, herpesviridae, virus diseases, herpesviridae, herpes simplex, virus diseases, herpesvirodae, herpes simplex, herpes genitalis genetics, biochemical genetics, molecular cloning, human, tissues, fluids etc. from non-related sources outside immediate project, physical separation, electrophoresis, gel, radioautography, radiotracers, tissue (cell) culture