2. How to interpret the results of antimicrobial susceptibility testing – what to do next? Mi Suk Lee, MD. Division of infectious Diseases Kyung Hee University Medical Center 2007. Mar. 20 th

3. What is antimicrobial susceptibility test? MIC by CLSI (Clinical and Laboratory Standards Institute) * National Committee for Clinical Laboratory Standards (NCCLS)

4. Choose of appropriate antibiotics Site of infection Immune status, etc Antimicrobial susceptibility test Host BacteriaAntibiotics

5. Prerequisite for interpretation G(+) or G(-) ? From what specimen ? Consistent with clinical symptom & sign ? Which antibiotic is specific to the recovered organism? Resistant to antibiotics ?

6. Prerequisite for interpretation G(+) or G(-) ? 1. Can exclude antibiotics ineffective for G(+) or G(-) from the list of treatment. 2. For G(+) : Aminoglycoside, aztreonam, FQ (ciprofloxacin, ofloxacin) can be excluded. 3. For G(-) : Glycopeptide (vancomycin, teicoplanin) can be excluded.

7. Prerequisite for interpretation From what specimen ? 1. All that glitter is not gold! – Because we human beings are not completely clean. – normal flora…. 2. Specimens normally sterile  Blood, CSF 3. Sputum, Stool…  low reliability

8. Prerequisite for interpretation Consistent with clinical symptom & Sign? 1. True infection 2. Colonization 3. Contamination during sampling specimens

9. Prerequisite for interpretation Which antibiotic is specific to the recovered organism? 1. You don’t have to interpret every antibiotic in the culture report list. 2. Knowing a few antibiotics specific to the recovered organism  That’s enough.

10. Prerequisite for interpretation Resistant to antibiotics ? 1. The interpretation of resistance is not simple. - oxacillin R in staphylococci  mean ‘do not use  -lactams’ - imipenem R in P. aeruginosa  consider R in FQs, AGs - any 3 rd cepha. or aztreonam R in G (-)  consider ESBL 2. You don’t have to know the all mechanism of resistance, but knowing resistance is mandatory.

11. What will be discussed? A brief review of sensitivity test  not in detail The meaning of S, R, I. Interpretation of report for individual organism We are often in the dilemmatic situation. – How should we solve these dilemma? – Although there’s not always correct answer…

12. Sensitivity test Dilution method – Agar dilution – Broth dilution

13. Sensitivity test Disk diffusion method – Diameter of inhibitory zone  Inversely correlated with the log 2 MIC – Easy to perform, but cannot apply to slow, fastidious organisms or anaerobes – Limited to Staphylococci spp., Enterobacteriaceae, Pseudomonas, Acinetobacter.

14. Sensitivity test E (epsiolmeter)-test – For fastidious grower such as some streptococci spp., Hemophilus influenzae, Neisseria gonorrheae, Streptococcus pneumoniae. – For anaerobes – Convenient, but costs too much.

15. Sensitivity test Automated – Microscan – Vitek

16. Report of Sensitivity “S” Sensitive : Isolates may be appropriately treated with usual dosage “R” Resistant : Predicting possible failure of the tested antimicrobial agent “I” Intermediate – or moderately susceptible

17. Conflicting decision when you meet “I” organism Increase the dosage of the antibiotics? Discard the antibiotics and choose an alternative one? My opinion  Rather not use it unless there’s no alternative.

18. Main clinically significant bacterial species you have to know G (-) – Pseudomonas – Non-pseudomonas : enterobacteriaceae G (+) – Staphylococcus – Streptococcus

19. G (-): ESBL-producer or not ? Extended spectrum  -lactams (oxyimino-  -lactams)  High affinity to PBP, stable to  -lactamase  Ceftazidime, ceftriaxone, cefotaxime, aztreonam, cefpodoxime ESBL (extended spectrum  -lactamases)  Hydrolyze oxyimino-  -lactams  Inhibited by clavulanate  No action to carbapenem or cephamycin  TEM, SHV, OXA group

20. G (-) Escherichia coli The most common cause of UTI & Sepsis Mainly intrinsic resistance & infection – In the bowel & translocation to the systemic circulation – Plasmid mediated  -lactamase : TEM-1 in 80% resistant to ampicillin, 1 st generation cephalosporins

21. Escherichia coli

22. Escherichia coli - ESBL Screening : Cefpodoxime 4 ug/mL Ceftazidime 1 ug/mL Confirm : Double disk synergy test

23. G (-) Klebsiella species A very tough guy !  can survive in the toughest environment. SHV-1  -lactamase producing ESBL : mediated via plasmid Patients at risk – Prolonged hospitalization, prolonged use of antibiotics – Immunocompromised host Can be transmitted via hands

24. Klebsiella pneumoniae

25. G (-) E. coli & Klebsiella If ESBL-producer,  Do not use ceftazidime, ceftriaxone, cefotaxime, aztreonam.  Double disk synergy test for confirmation

26. G (-) E. coli & Klebsiella - ESBL Use another one #Cephamycin (cefotetan, cefoxitin) Risky ! #  -lactam/  -lactamase inhibitor Half & half (no effect to ampC  -lactamase) #Non-  -lactam antibiotics (quinolone, aminoglycoside) Reasonable, but few clinical supporting data #4 th generation cephalosporin Half & half #Carbapenem : recommendable

27. G (-) Pseudomonas aeruginosa Widespread Can adapt themselves very well to any environment Opportunistic Outbreak in ICU Can be found in medical device, sink, disinfectant, vaporizer, bath, suction device, jelly or ointment, lotion, etc.

28. G (-) Pseudomonas aeruginosa Resistant to penicillin, cephalosporin, tetracycline, chloramphenicol, sulfonamide Sensitive to anti-pseudomonal cephalosporin (ceftazidime, sulperazone), aminoglycoside, qunolone, carbapenem Imipenem-resistant organism  recently emerging problem

29. G (-) P. aeruginosa Sensitive to ceftazidime, aminoglycoside, FQ?  Ignore ceftriaxone, cefotaxime, ceftizoxime  Ignore 1 st & 2 nd cephalosporins If not sensitive, how about imipenem? Avoid monotherapy ; Use antibiotics in combination (eg. Ceftazidime + AMK)

30. Pseudomonas aeruginosa

31. Multi-drug resistant P. aeruginosa

32. Ubiquitous in environment - 25% in skin of healthy outpatients - 7% in pharynx, 45% in tracheostomy sites Intrinsically resistant to many antibiotics Step 1: clinically significant? - Insignificant in most cases G (-) Acinetobacter spp.

33. Step 2 : resistant to piperacillin & ceftazidime? - Natural resistance to amp, amox, AMX/CLAV, 1 st gen. cephalosporins - Hyperproduction of chromosomal AmpC + altered porin proteins : Resistant to β-lactams except carbapenems Step 3 : resistant to imipenem? - Decrease in outer membrane protein or altered PBPs - Carbapenem-hydrolyzing enzymes : VIM-2 (Korea) Step 4 : resistant to quinolones? - Naturally less susceptible to ciprofloxacin than other genera, have MICs close to the breakpoint concentration G (-) Acinetobacter spp.

34. Pan-drug resistant A. baumannii Overall 30-day mortality 49% Portal of entry : 80% resp. tract Treatment : colistin, polymyxin B, high dose amp/sulb, ± RFP

35. G (+) Staphylococcus spp. Living on the skin MRSA = oxacillin-resistance # The meaning of oxacillin-resistance  You cannot use any  -lactams !  Only glycopeptide (vancomycin, teicoplanin) is effective. # Alteration of penicillin binding protein (PBP) # Controlled by mecA gene # Nosocomial in most cases # Person-to-person  the importance of hand washing

36. G (+) Staphylococcus spp. Step 1 : From what specimen ? Is it real ? Step 2 : Resistant to Oxacillin ? Step 3 : If not, Resistant to Penicillin ?

37. G (+) Staphylococcus spp. Step 1 : From what specimen ? Is it real ?  From blood or CSF ?  Any possibility of contamination even if isolated from blood ?  Consistent with the patient’s symptom & sign ?

38. G (+) Staphylococcus spp. Step 2 : Resistant to Oxacillin ? # The meaning of oxacillin-resistance  You cannot use any  -lactams !  Only glycopeptide (vancomycin, teicoplanin) is effective. # In case of oxacillin-sensitive, but resistant to any one of aminoglycoside, other  -lactams, macrolide, tetracycline, or clindamycin  Recheck whether hidden MRSA or MRCNS. oxacillin salt agar screening (6 ug/mL) # If not, go to step 3.

39. G (+) Staphylococcus spp. Step 3 : If not, Resistant to Penicillin ? * If yes,  -lactamase producer.   -lactamase stable penicillin (flucloxacillin, nafcillin, cloxacillin, oxacillin)   -lactam/  -lactamase inhibitor  1 st generation cephalosporins * If no, S to both penicillin & oxacillin  Penicillins or any anti-staphylococcal antibiotics

40. Methicillin susceptible S. aureus

41. Methicillin resistant S. aureus

42. G (+) Enterococcus spp. Step 1 : From what specimen? Blood? Step 2 : Resistant to penicillin or AG ? Step 3 : Resistant to Vancomycin?

43. G (+) Enterococcus spp. Step 1 : From what specimen ? Blood ? * From stool or urine  Ignore! * Recovered with other organisms (mixed)  Ignore! * From blood  Prepare for trouble! Why ?  Probability of truly virulent strain is very high.  Survivor from the bowel jungle !  Acquiring virulence during the process of invasion into blood stream from bowel.

44. G (+) Enterococcus spp. Born to be resistant to lots of antimicrobials (esp. cephalosporins) Vancomycin-resistant enterococcus (VRE) – VanA, VanB, VanC1/C2/C3, VanD – Can be transferred to other organisms & other species – What if it is transferred to MRSA ?  Nightmare !

45. G (+) Enterococcus spp. Contact isolation & Hand washing !!

46. G (+) Enterococcus spp. Step 2 : Resistant to penicillin or AG ? * Ignore any cephalosporins * If penicillin-sensitive  not producing  -lactamase - Ampicillin, amoxicillin, unasyn, augmentin, piperacillin, etc - Combination with AG if not highly resistant to AG * If penicillin-resistant - Vancomycin + AG if not highly resistant to AG

47. G (+) Enterococcus spp. Step 3 : Resistant to vancomycin ? – If not from blood, ignore !  infection control – If you’re sure that that guy is a truly virulent strain, Almost nothing to give. That’s real… Quinupristin/dalfopristin (synercid)?, Oxazolidinone (linezolid, eperzolid), Tigecycline, daptomycin, etc Teicoplanin, if VanB phenotype (a lucky case)

48. Penicillin susceptible E. faecium

49. VSE faecium

50. VRE faecium * Synercid : only effect to E. faecium

51. G (+) Streptococcus pneumoniae Step 1 : Resistant to penicillin or oxacillin? Step 2 : If yes, also resistant to 3 rd generation cephalosporins? Step 3 : From what specimen?

52. G (+) S. pneumoniae Step 1 : Resistant to penicillin or oxacillin? * See for resistance to oxacillin * If sensitive to oxacillin  Happy end. S to penicillin & 3 rd generation cephalosporins * If resistant to oxacillin  Go to step 2. R to penicillin, but ?? to 3 rd gen. cephalosporins Check MIC of penicillin & 3 rd gen. cephalosporins

53. G (+) S. pneumoniae Step 2 : Resistant to 3 rd gen. cephalosporin? – If no,  Use 3 rd generation cephalosporin – If yes,  Add Vancomycin to the 3 rd generation cephalosporin.

54. Streptococcus pneumoniae - PSSP * Penicillin MIC : S  0.06, I 0.1-1, R  2

55. Streptococcus pneumoniae - PRSP * Penicillin MIC : S  0.06, I 0.1-1, R  2

56. * Ceftriaxone/cefotaxime MIC in S. pneumoniae (by E-test) SIR CNS  0.5 1  2 2 Non-CNS  1 2  4 4 * Not available in KHMC

57. G (+) S. pneumoniae Step 3 : From what specimen? * From blood (with pneumonia) * From CSF  consider BBB penetration

58. Pneumococcal pneumonia when in vitro susceptibilities are known PSSP (PCN MIC  0.06) – Primary : penicillin-G 1 mU q 4 h or ampicillin 1 g q 6 h – Alternative : cefuroxime, cefotaxime/ceftriaxone, macrolide PRSP (PCN MIC 0.1-2) – Primary : penicillin-G 2 mU q 4 h or ampicillin 2 g q 6 h – Alternative : cefotaxime/ceftriaxone, macrolide PRSP (PCN MIC  4) – Primary : continue with initial empiric therapy if clinically satisfactory – Alternative : imipenem, macrolide, vancomycin, new FQ – Vancomycin not routinely recommended for CAP caused by PRSP

59. Pneumococcal meningitis when in vitro susceptibilities are known Kaplan SL, et al. Clin Microbio Rev 1998;11:628 *  2

60. G (+) Streptococcus spp. viridans streptococci  Penicillin-resistance or not ?  in fact, rare Streptococcus pyogenes  No sensitivity test at all because there’s been no report of  -lactam resistance Streptococcus agalactiae  Penicillin-resistance or not?

61. Viridans Streptococci Usually contaminant If persistent bacteremia  consider infective endocarditis If PCN MIC  4 ug/mL  use vancomycin (very rare)

62. Streptococcus pyogenes

63. Dilemma, Dilemma.. ?

64. Dilemma 1. Positive blood culture. Is That Real? Even in the febrile patient. Especially coagulase (-) staphylococci (CNS) The possibility of contamination : 2~3% Any guideline for discriminating the true from false ?

65. Dilemma 1. Positive blood culture. Is That Real? Tentative guideline for discrimination  The culprit organism - CNS, Bacillus spp., Corynebacterium spp., Micrococcus spp., P. acnes, viridans streptococci, S. maltophilia, C. perfringens  How long did it take to grow ? - If longer than 72-h, probably contamination  More than 1/4 or 1/6 bottles? - (exception) immunocompromised pt, prosthetic device.  Repeated isolation of the same organism from a sterile specimen other than blood.

66. Dilemma 2. Positive sputum culture. Is That Real? The sputum is the most unreliable specimen. Dilemma in the Dx. of nosocomial pneumonia. Protected BAL

67. Dilemma 3. Mixed culture. Is That Real? Commonly seen in the urine specimen. In most cases  not true. Exception : from abscess

68. Dilemma 4. What if resistant to all antimicrobials? Frankly speaking … Nobody Knows !

69. My opinion – somewhat subjective  Check if the specimen is a significant one : - normally sterile or not ?  Recheck sensitivity - If the result is sensitive  Thank God ! - If intermediate  increase the dosage.  Simultaneously, try another new antibiotics. (state-of-the-art) Dilemma 4. What if resistant to all antimicrobials?

70. Knowing is not enough ; we must apply. Willing is not enough ; we must do. by Goethe said