1. Clinical data to support the interpretation of susceptibility testing of anaerobes Robin Howe

2. 'Mixed anaerobes sensitive to metronidazole' (common practice in UK) Pus from cerebral abscess Primary plate after 5 days AnO 2 incubation Courtesy Val Hall

3. ? But would we report:- 'Mixed aerobes sensitive to a cephalosporin' 'Fungus present, try Athlete's Foot powder' 'Virus detected, have a hot whisky and lemon' And would we test them like this?….. Courtesy Val Hall

4. 'S.O.P.' for susceptibility testing of anaerobes Agar: Any (selective or non-selective) Inoculum: Direct sample, ?mixed, not standardised Antimicrobial agents: metronidazole Incubation: On bench till 5pm, then AnO 2 18-72hrs Controls: None Interpretation: –Any sized zone = mixed anaerobes, sens to MZ –No zone = no anaerobes isolated –Colonies within zone = aerobes Courtesy Val Hall

5. Why is susceptibility testing of anaerobes not generally used in clinical decision- making? Technical issues with testing –Slow growth –Lack of consensus regarding agar/method Anaerobic susceptibility patterns –Predictable –Unchanging over years Limited data to correlate in vitro results with outcome –Infections often polymicrobial –Outcome affected by multiple factors (eg surgery)

6. Are resistance rates predictable? Does in vitro resistance correlate with an identifiable resistance mechanism? Hecht (2004) CID39: 92

7. Are resistance rates predictable? Does in vitro resistance correlate with an identifiable resistance mechanism? Hecht (2004) CID39: 92

8. Baquero (1992) –10% C. perfringens in Spain resistant to pen (MIC >0.5 mg/L) BSAC BP 0.12 mg/L

9. Are resistance rates predictable? Does in vitro resistance correlate with an identifiable resistance mechanism? B. fragilis Class 2e cephalosporinase V. common Inhibited by clavulanate etc Cephamycinases Uncommon cepA, cfxA Zinc metallo-  -lactamase cfiA, ccrA Present in ~4% - not usually expressed Altered PBPs Rare Porin loss Reported

10. Are resistance rates predictable? Does in vitro resistance correlate with an identifiable resistance mechanism? Reduced susceptibility to metronidazole –Common in Propionibacteria & actinomycoses –Rare in B. fragilis –nim genes C.difficile REFERRALS TO ARU Bacteroides spp. n=78 (5% of all Bacteroides) Clostridium paraputrificum n=5 (4%) Clostridium ramosum n=3 (1%)

11. nim genes Nim = nitro-imidazole reductase Types A – G found in Bacteroides spp. Detected by PCR-RFLP Chromosomal / plasmid-borne Absent from some MZ resistant orgs Probable alternative mechanisms High level MZ resistance can be induced in some nim-containing strains Courtesy Val Hall

12. nim genes identified in 2% of 1,502 B. fragilis from 19 European countries

14. Animal studies Rat model of secondary peritonitis –Pooled faecal emulsion intraperitoneally –initially E. coli predominates with often fatal bacteraemia –If survive  abscesses with B. fragilis –Early gentamicin  no bacteraemia but late abscesses –Early clindamycin  no effect on bacteraemic mortality but reduced late abscesses in survivors Onderdonk et al (1974) Infect Immun 10:1256 The role of anti-anaerobic therapy

15. Animal studies

16. ?synergistic infection IP injection of mixtures of three orgs Brook (1994) JAC 34: 791

18. Reports of clinical failure associated with resistance Penicillin vs C. perfringens –NIL Metronidazole vs Bacteroides spp –YES

19. Rotimi et al (1999)CMI 5: 166 3 case reports –75 yrs female post op Hartmann’s treated with CAZ/MTZ Readmitted with paracolic abscess B. frag isolated (MTZ MIC >32 mg/L) Cured with drainage + IMI –40 yrs male Gangrenous appendix (mixed B. frag/E. coli/Pseudomonas) CXM/MTZ started Day 5 - Wound infection –B. frag + B. ovatus (MTZ MIC >32mg/L) –Cured with co-amox –37 yrs male Post renal transplant cholecystitis  cholecystectomy  necrotising pancreatitis Multiple Abs – CTX/MTZ/AMIK – MEM/AMIK/CPM Mixed isolates from lap inc B. distasonis (MIC MTZ >32 mg/L, MEM >32 mg/L) Pt died

20. Reports of clinical failure associated with resistance Penicillin vs C. perfringens –NIL Metronidazole vs Bacteroides spp –YES Penicillin vs Bacteroides spp. –YES Brook (1984) Arch Otolaryngol 110: 228 Gudiol (1990) Arch Intern Med 150: 2525

21. Reports of clinical failure associated with resistance Penicillin vs C. perfringens –NIL Metronidazole vs Bacteroides spp –YES Penicillin vs Bacteroides spp. –YES Β-lactam/β-lactamase inhibitors vs Bacteroides spp. –NIL Carbapenems vs Bacteroides spp. –YES

22. 38 year old female –Elective laparotomy for adhesions –Post-op IA collection treated with co-amox –Day 13 - surgical drainage & change to CTX + MTZ –BC grew B. fragilis (isolate 1) –Changed to imipenem –2 weeks later persistent empyema drained (isolate 2) –Cured with drainage/clindamycin/gentamicin Turner et al (1995) Lancet 345: 1275

23. Prospective evaluation of 128 patients with Bacteroides bacteraemia CID (2000) 30: 870

25. Any data relating level of resistance to outcome? NO

26. Conclusions Antimicrobial resistance is variably predictable Resistance rates are increasing –CLD – becoming common –MTZ + carbapenems – emerging Inducibility is a concern Correlation between in vitro resistance and outcome has not been established for many anaerobic infections –The role of surgery should not be forgotten

27. Finegold 1989 Susceptibility testing of anaerobes should be done in 4 settings: –Determine patterns of susceptibility to new agents –Monitor susceptibility patterns Nationally –Monitor susceptibility patterns locally –Assist in the management of individual patients Persistence of infection/ failure of usual regimes/ difficulty making decisions based on precedent Brain abscess/ endocarditis/ osteomyelitis/ prosthetic device infection/ septic arthritis