Laboratory Interpretation: MICROBIOLOGY

Laboratory Interpretation: MICROBIOLOGY

Outpatient care in the era of MRSA Julie Gutman, MD Pediatric Infectious Disease Fellow August 29, 2007 MRSA Basic Micro History of MRSA emergence Genetic markers Resistance testing Treatment Eradication

Staphylococci Gram-positive Facultative anaerobes Grow in clusters Strep usually grow in chains Catalase test (+) distinguishes from strep spp. which are (-) S. aureus S. epi Site Nose

Skin Coagulase positive negative Yellow White S. aureus & Colony

S. epidermidis are most Color important clinically Colony Size Large Small Hemolysis? yes no Emergence of MRSA First described in 1960 One year after introduction of methicillin!

Initially most cases were hospital acquired Risk factors for Hospital Acquired: Hospitalization, Surgery, Dialysis Permanent indwelling device Trach, G-tube, foley, CVL Evidence of MRSA in the Community 3% colonized with MRSA at center X 24% colonized with MRSA at center Y JID, Volume 178 (2), August 1998 Increasing incidence of CA- MRSA in

Hospitalized Children Retrospective review: Compared AugustJuly of 1988-1990 and 1993-1995 CA-MRSA disease increased: 8 in 1988-90 to 35 in 1993-95. 10/100,000 to 259/100,000 (P<.001) Noted a difference in susceptibility of isolates from children with risk factors for MRSA vs. those with no risk factors Herold et al. JAMA. 1998;279:593-598 MMWR Four Pediatric Deaths from CA-MRSAMinnesota and North Dakota, 1997-1999 First report of fatal outcomes

associated with MRSA Type MW2 (USA400) Methicillin-resistant Staphylococcus aureus (MRSA) is an emerging community-acquired pathogen among patients without established risk factors for MRSA infection This report describes four fatal cases among children with community-acquired MRSA; the MRSA strains isolated from these patients appear to be different from typical nosocomial MRSA strains in antimicrobial susceptibility patterns and pulsed-field gel electrophoresis (PFGE) characteristics. MMWR, August 20, 1999 / 48(32);707-710 Incidence of CA-MRSA Disease Fridkin, et al. NEJM 2005. 352: 1436-44.

Sequencing of CA-MRSA (MW2) Identified SCCmec IV with mecA gene Lacked the multiple resistance genes found in HA-MRSA (SCCmec II, also I and III) Homology with HA-MRSA genome ~ 95% homology is 99.7% between HA-MRSA strains 18 new virulence factors Rapid growth rate- survival advantage doubling time 13-23.5m VS. 34.8 and 46.8m for HA-MRSA Baba, et al. 2002. Lancet. 359:1819 MRSA Genetics- SCCmecIV

Staphylococcal cassette chromosome (SCCmec) mobile genetic element Methicillin resistance mecA gene Produces altered penicillin binding protein PBP2a CA-MRSA generally SCCmec IV smaller & more mobile than other SCCmec HA-MRSA carries type II predominant circulating clones USA300 and USA400 Staphylococcal Cassette Chromosomes (SCC-mec). Okuma, JCM 2002 MRSA Strain Typing

Pulsed-field gel electrophoresis (PFGE) Uses Sma1 restriction endonuclease Clones USA100- USA1000 have been designated Clones USA300 and USA400 predominate PCR-based methods spa typing DNA-sequencing of repeat regions of the Staphylococcus protein A gene (spa): repeats are assigned a numerical code and the spa-type is deduced from the order of specific repeats multilocus sequence typing (MLST) Uses the sequences of 7 house-keeping genes and assigns an allelic type to each sequence.

Antimicrobial Susceptibility CA-MRSA vs HA-MRSA Naima, et al. JAMA 2003 Egleston - ALL ISOLATES, MYSIS Incidence report,'06) EPI GP ORG N NOTES 53% VRE

VREs E.faecium 30(pt) MRSA MRSA 646 MSSA

627 ESBLs SPICE&M MRO E.coli 503 Kl.oxytoca 43

Kl.pneumoniae 160 Pr.mirabilis 46 Providencia sp. 3 Indole + Proteus

1 Citrobacter sp. 33 Enterobacter sp. 175 Morganella sp. 11 Ps.aeruginosa

722 Acinetobacter sp. 42 Steno.maltophilia 117 51% MRSA OP-SSI: 65% MRSA Courtesy of Dr. Jerris

2006 Wounds ECH MRSA [65% incidence {all sources,locations}] S.aureus Anaerobes E.coli Streptococcus pyogenes (gpA strep) Pen 100% S, Erythro 93.1% S, Clinda 99.5% (CID.05. 41:599)

Enterococcus faecalis Cephs are ineffective SPICE and M Serratia, Providencia, Indole + Proteus, Citrobacter, Enterobacter and Morganella Source: Misys Incidence query Courtesy of Dr. Jerris 2006 Antibiogram Egleston Clinda

Single isolate per patient per year TMP/SMX Cipro Inducible Clinda Resistance Mediated through erm gene MLSB Alteration in the 23S ribosomal RNA Part of 50S ribosomal subunit Occurs through methylation of adenine Confers resistance to: erythromycin and most other macrolides (M)

lincosamides (L, lincomycin, and clindamycin) streptogramin type B (SB). Consider in all Erythro Resistant isolates!!! D- test for inducible clindamycin resistance NO induction msrA-mediated erythromycin resistance Erythro R Clinda S Negative reaction

Courtesy of Dr. Jerris D- test for inducible clindamycin resistance Inducible clindamycin resistance (erm-mediated) Erythro R Clinda R 15 - 26 mm Positive reaction

Photos courtesy of J. Jorgensen and K. Fiebelkorn. Courtesy of Dr. Jerris Egleston Inducible Clinda Resistance 06 *Note that the rates vary dramatically depending on the site and patient population Courtesy of Dr. Jerris Skin and Soft Tissue Infections CHOA - 2006 WOUNDS: 288 isolates with MRSA (1 isolate per patient per year) Erythro Resistant and Clinda Susceptible 6/288 were D-zone test positive (2.1%)

- Outpt. 2/186 (1%) - Inpat. 4/102 (4%) @30% of population carry this organism-CDC Courtesy of Dr. Jerris Syringes FOR QUALITY RESULTS, SEND TISSUE, FLUIDS ,OR ASPIRATES (SAME SPECIMEN FOR PATHOLOGY BUT NOT IN FORMALIN)

Courtesy of Dr. Jerris Vancomycin Intermediate (resistant) S. aureus (VISA) Japan, 1996: first documented case of VISA MIC=8 ug/mL US, July 1997, VISA-associated peritonitis isolate was susceptible to rifampin, chloramphenicol, bactrim, and tetracycline. National Committee for Clinical Laboratory Standards breakpoints for susceptibility: susceptible 4 ug/mL

Intermediate = 8-16 ug/mL resistant 32 ug/mL MMWR, August 22, 1997; 46(33);765-766 Spectrum of Disease more virulent than HA-MRSA. Infection of skin or soft tissue Osteomyelitis Pyomyositis Necrotizing pneumonia Severe sepsis purpura fulminans, DIC, and bilateral adrenal hemorrhage

Necrotizing fasciitis Thromboemboli Panton Valentine Leukocidin (PVL) Bicomponent, pore-forming leukotoxin LukS-PV and LukF-PV Produces lysis of PMNs Carried on an integrated bacteriophage (phiSLT) Causes dermonecrosis if injected into skin of animals Associated with severe disease in CA-MSSA and CA-MRSA More frequently associated with sepsis, high

fever, leukopenia, hemoptysis, pleural effusion and death than PVL (-) Panton Valentine Leukocidin (PVL) BUT PVL-negative (lukS/F-PV knockout) strains of USA300 and USA400 were as lethal as wild-type strains in a mouse sepsis model, and they caused comparable skin disease Staph virulence factors

Outpatient Management of CA-MRSA + Assumes >90% D test negative, erythro resistant CAMRSA Baker, AAP News September 2004 For life threatening disease Nafcillin + Vancomycin + Clinda (effect on toxin) + Gent or Rifampin

+/- Ceftriaxone for gram negatives NO good data to support this combo! A Note about Wounds For abscesses, DRAINAGE is key! Abscesses<5 cm I&D alone sufficient! Antibiotic Choices for Staph Nafcillin- preferred treatment for MSSA! More rapid clearance of blood cultures than Vanc Bactrim- some proven efficacy for SSI Does NOT cover strep!

Clindamycin- covers strep and staph! 10-20% resistance, should NOT use as monotx for severe infections! Vancomyin- DOC for MRSA Antibiotic Choices for Staph Gentamicin- used a an adjuntive therapysynergistic activity with Vanc Rifampin- proven efficacy in setting of prosthetic valve endocarditis, otherwise, little data Should never be used as monotherapy for treatment! New Antibiotic Choices for Staph

Linezolid Skin and soft tissue infections, pneumonia Have been reports of failure when used for endocarditis Can be given PO SE: thrombocytopenia, optic neuritis, peripheral neuropathy Daptomycin Skin and soft tissue infections, bacteremia, endocarditis with MRSA NOT for pneumonia! SE- musculoskeletal. Monitor CPK. Others you may hear about

Tigecycline- approved for MRSA skin ond soft tissue infections Extremely broad spectrum!! Covers VRE, complicated intra-abdominal infections SE: GI- Nausea, Vomiting Quinupristin-Dalfopristin- approved for VRE, MSSA, not MRSA Requires central line Severe myalgias, arthralgias Dalbavancin- once weekly dosing Not yet FDA approved! Prevention Strategies, MMWR Feb 7,2003 / 52 (05);88

Community setting: Thoroughly clean towels, bed linens, personal clothing Thoroughly wash cuts and abrasions with soap and water Disinfect athletic equipment, benches, personal equipment with commercial disinfectant or bleach ( 1 tablespoon bleach in 1 quart water) diluted Common sense personal hygiene Individual patients: Local treatment I & D (antimicrobics)

Courtesy of Dr. Jerris Decolonization- Cochrane Review 6 trials (384 participants) Trial Intervention Outcome- Eradication Harbarth 1999 IN mupirocin compared to

placebo 25% mupirocin vs. 18% placebo; RR 1.39; Not statistically significant! Parras 1995 IN mupirocin vs. IN fusidic acid + PO Bactrim Eradication: 78% mupirocin vs. 71% bactrim+ fusidic acid @ 90d Chang

2000 Oral fusidic acid vs. no treatment RR 0.67 (0.18-2.42) Not statistically significant! Walsh 1993 Novobiocin + rifampin (N+R) vs. Bactrim+ rifampin (B+R)

67% N+R vs. 53% B+R (P = 0.18) NS! Resistance to rifampin- more in the bactrim vs. novobiocin group (14% vs. 2%, P = 0.04). Peterson 1990 Cipro + rifampin vs. Bactrim + rifampin 6-month eradication in 3/11 cipro + rifampin and 4/10 Bactrim + rifampin recipients. **terminated due to cipro resistance !!!! Muder

1994 PO rifampin, minocycline, or rifampin +minocycline vs. no treatment Not statistically significant! Rifampicin better than minocycline at day 30 but not at day 90 Single agent vs. placebo RR=8 (0.53-122) Combined Tx vs. placebo RR=9.45 (0.62-145) Decolonization- Cochrane Review SE in up to 20% of systemic agent use. All trials reported development of resistance

Authors' conclusions Insufficient evidence to support use of topical or systemic antimicrobial therapy for eradicating MRSA. Need for large RCT! Decolonization 2% chlorhexidine gluconate wash + 2% mupirocin intranasally + Oral Rifampin + Doxy for 7 days 74% of treated vs. 32% untreated had negative cultures at 3 months Simor et al., CID 2007. 178:44.

Our Protocol Clinda or Bactrim + Rifampin PO Mupirocin daily x 7 d to nares of ENTIRE FAMILY Chlorhexidine gluconate washes daily for a week, then 1-2 times weekly (can substitute bleach baths) Cross your fingers and pray. Blood Culture VOLUME Volume of blood drawn for culture is the MOST IMPORTANT variable in detecting bacteremia or fungemia MAX volume per BC System For adult patients, the yield of pathogens increases in direct proportion

to the volume of blood that is cultured from 2 to 30 mL. The yield still increases when 40 mL (or even higher) volumes of blood are cultured, although the increase may no longer be in direct proportion to the volume of blood cultured. For pediatric patients, the limited data that has been published also indicate that the yield of pathogens increases in direct proportion to the volume of blood that is cultured [Li J, Plorde J, and Carlson L. 1994. Effects of volume and periodicity on blood cultures. J Clin Microbiol. 32:2829-2831] Courtesy of Dr. Jerris Questions? 2006 Antibiogram Egleston

4 pandemic MRSA clones derive from 2 distinct backgrounds Oliveira, Lancet 2002

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