Microbiologic Diagnosis of Diabetic Foot Infections Albert T.

Microbiologic Diagnosis of Diabetic Foot Infections Albert T. Sheldon, Jr. Ph.D. Microbiology Team Leader Division of Anti-infective Drug Products

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 2 Guidance for Industry- Foot Infections in Patients with Diabetic mellitus Microbiological Considerations All patients should have pre-therapy cultures. Gram stain and cultures should be obtained from acceptable sources using acceptable methods. We prefer cultures obtained by leading edge needle aspiration, soft tissue and joint aspiration, bone biopsy and/or surgical debridement. Microorganisms isolated should be assessed as true pathogens, colonizers, or contaminants. Only microorganisms designated as true pathogens should be considered in determining microbiological evaluability of enrolled subject.

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 3 Factors that Influence Infection Rates Risk of Wound Infection varies according to the following equation:* Dose of bacterial contamination x Virulence Resistance of the Host *Altemeire, W.A., & W.R. Culbertson (1965) Surgical Infection In Moyer, C. et .al. (eds.): Surgery, Principles and Practices, 3rd ed. Philadelphia, Lippincott Co.

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 4 Host Factors that Influence Infection Rates Diversity and abundance of microorganisms present in the wound are also influenced by host factors: Wound type, depth, location, and quality Presence of nonviable exogenous contamination Peripheral blood insufficiency Immune competence of the host

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 5 Manual of Clinical Microbiology* “The use of specimens for bacteriological analysis requires that specific clinical material be collected, stabilized, and transported according to exacting specifications to insure valid results.” *Murray, P. R., E. J. Baron, J. H. Jorgensen, M. A. Pfaller, and R. H. Yolken. 2003. Manual of Clinical Microbiology, 8th Edition. ASM Press, Washington D.C.

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 6 Clinical Microbiology Implicit in this definition are two issues that are of interest to the discussion of decubitus foot infections. The methods used to collect the clinical sample and the validity of the results to assess the involvement of an organism in the etiology of the disease.

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 7 1. Methods used in Collection of Microbiological Wound Samples Deep Tissue Techniques Biopsy and/or surgically debrided tissue*+ Leading edge Needle aspirate*+ Joint fluid or synovium+ Bone specimen+ Blood+ Surface Sampling Techniques Swab* Curettage* Dermabrasion Velvet pad surface imprint *Published methods used in decubitus ulcer sampling. + Method recommended in Agency guidance document are deep tissue techniques.

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 8 Comparison of Sampling Methods Sapico F.L., et. Al. (1984) Rev Infect Dis. 6:S171-S176

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 9 Comparison of Sampling Methods Sapico F.L., et. Al. (1984) Rev Infect Dis. 6:S171-S176

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 10 Relationship Between Biopsy & Swab: Burn Wounds Thomson, P.D., (1994) Amer J Surgery 167:7S-10S

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 11 Relationship Between Quantitative Culture & Swab Breidenbach W,C., & S. Trager (1994) Plats. Reconstr. Surg. 95:860-865 Study purpose: To determine the relationship between the quantity of bacteria and infection in complex extremity wounds, and To compare the predictive value for wound infection of quantitative cultures versus other factors considered to have predictive value for wound infection. I will focus on the latter purpose of the study.

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 12 Relationship Between Quantitative Culture & Swab (cont) Breidenbach W,C., & S. Trager (1994) Plats. Reconstr. Surg. 95:860-865 Evaluated 50 patients with complex wounds-defined as a soft-tissue defect that required flap for closure. Quantitative cultures (biopsy) compared to Clinical parameters (factors considered to have predictive value in wound infection; wound position, mechanism of injury, fracture type) Laboratory test (Swab culture) 28 patients had quantitative cultures obtained after debridement and high pressure wash prior to flap closure. 16 patients had swab cultures. 2-5 samples obtained per wound.

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 13 Criteria for Positive/Negative Test, Infection, and Prevalence Breidenbach W,C., & S. Trager (1994) Plats. Reconstr. Surg. 95:860-865

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 14 Criteria for Positive/Negative Test, Infection, and Prevalence Breidenbach W,C., & S. Trager (1994) Plats. Reconstr. Surg. 95:860-865

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 15 Predictive values, Sensitivity, & Specificity of Tests Studied

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 16 Predictive values, Sensitivity, & Specificity of Tests Studied

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 17 2. Interpretation of Microbiologic Diabetic foot infection Samples (or Qualitative Microbiology) Diabetic foot ulcers are polymicrobic.* S. aureus is the predominant aerobic species followed by S. epidermidis, Streptococcus spp., P. aeruginosa, Enterococcus spp., and coliform bacteria. The predominant anaerobic species are Peptostreptococcus spp., Bacteroides spp., and Prevotella spp. Application of good microbiological techniques will allow isolation of anaerobes from up to 95% of decubitus ulcers. However, such studies are not usually performed due to the labor intensive nature of anaerobic microbiological studies. *Bowler, P.G., et. al. (2001) Clin Microbiol Rev 14:244-269.

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 18 Schools of Thought* Although microorganisms are responsible for wound infections, there is controversy regarding their role. Published literature is inconclusive! The density of microorganisms is the critical factor in determining whether a wound is likely to heal. The presence of specific pathogens is of primary importance in delayed healing. Microorganisms are of minimal importance in delayed healing. Debate whether a wound should be sampled, the value of the results and the method that should be used.

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 19 Conclusions Widespread controversy still exists Regarding the exact mechanisms by which microorganisms cause wound infection, Regarding the significance of microorganisms in nonhealing wounds that do not exhibit signs of clinical infection, and Regarding the best microbiological techniques to monitor the microbiology of wounds. ASM Manual Clinical Microbiology states “A swab is not the specimen of choice…Since a swab specimen of a decubitus ulcer provides no clinical information.”

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 20 Conclusions A regulatory agency must require microbiological methods that provide us with confidence and data necessary to assess the response of antimicrobials for their intended uses. We describe, in our guidance document, what we consider to be relevant methods. These are the deep tissue techniques discussed in a previous slide.

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 21 Louis Pasteur “ The germ is nothing. It is the terrain in which it is found that is everything.” Pasteur, L. (1880) De l’attenuation virus du cholera des poules. CR Acad. Sci. 91: 673-680.

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 22 Obtaining a Swab Culture Before taking the specimen Cleanse the wound with water or saline Remove excess necrotic debris Compress edges to elicit new drainage Use swab tip to swab the healthiest tissue Do not swab exudate, pus, eschar, or heavily fibrous tissue. Gram stain helps define quality of specimen as determined by polymorphonuclear cells, squamous epithelial cells and bacteria on gram stain. ASM Manual Clinical Microbiology states “A swab is not the specimen of choice…Since a swab specimen of a decubitus ulcer provides no clinical information.”

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 23 Microbiology of noninfected wounds

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 24 Microbiology of Infected Wounds

28-Oct-2003 Albert T. Sheldon, Jr. Ph.D. DAIDP 25 Useful Wound Definitions Contamination-Presence of bacteria on the wound surface. Colonization-Presence of and multiplication of surface microbes/bacteria contaminants without infection. Infection-Invasion and multiplication of microorganisms in body tissues resulting in local cellular injury. Distinction between definitions not so clear in the clinical setting.