Mitchell A. Yakrus

Cluster of Mycobacterium Chelonae keratitis cases following laser in-situ keratomileusis ☆

PURPOSE To describe a cluster of Mycobacterium chelonae keratitis cases involving patients who underwent laser in-situ keratomileusis (LASIK) at a single refractive surgery center. DESIGN Descriptive case series of four patients and cohort study to identify disease associations. METHODS Examination schedules, diagnostic tests, and therapy were based on best medical judgment. Isolates from three patients were compared by pulsed-field gel electrophoresis. Epidemiologic studies were performed to identify the source of infection. RESULTS Seven of eight eyes developed M. chelonae keratitis following bilateral simultaneous LASIK. Each patient was thought to have diffuse lamellar keratitis initially, but all seven eyes were noted to have opacities suggestive of infectious keratitis by 13 to 21 days after surgery. All eyes had undergone hyperopic LASIK over four days in April 2001 by one surgeon in a community-based refractive surgery center. A cohort study of all patients undergoing LASIK at the same center in April 2001 revealed that M. chelonae keratitis occurred only in persons undergoing correction of hyperopia (seven of 14 eyes vs. none of 217 eyes undergoing myopic LASIK, P <.001). The only difference identified between procedures was use of masks created from a soft contact lens in hyperopic LASIK. Three isolates (three patients) were indistinguishable by pulsed-field gel electrophoresis. Eyes were treated with a combination of antimicrobial agents, including topical azithromycin in three patients, with resolution of infection in all eyes over 6 to 14 weeks. The source of infection was not identified on environmental cultures. CONCLUSION Postoperative nontuberculous mycobacterial keratitis can occur in an epidemic fashion following LASIK. Topical amikacin, azithromycin, clarithromycin, ciprofloxacin, or a combination of these agents, appears to be effective treatment for these infections.

AN OUTBREAK OF BACTEREMIAS ASSOCIATED WITH MYCOBACTERIUM MUCOGENICUM IN A HOSPITAL WATER SUPPLY

OBJECTIVE To investigate and determine the cause of an outbreak of Mycobacterium mucogenicum bacteremias in bone marrow transplant (BMT) and oncology patients. DESIGN Case-control study and culturing of hospital water sources. Isolates were typed using molecular methods. SETTING University-affiliated, tertiary-care medical center. PATIENTS Case-patients were adult and pediatric BMT patients or hematopoietic stem cell transplant (BMT) (n = 5) and oncology (n = 1) patients who were diagnosed as having M. mucogenicum bacteremia during the study period of August through November 1998. Two control-patients were selected for each case-patient matched by age, time of hospitalization, inpatient unit, and type of patient (BMT or oncology). RESULTS There were no significant differences between case-patients and control-patients regarding intravenous products received or procedures performed, frequency of bathing, neutropenia, or steroid use. Nontuberculous mycobacteria were isolated from several water sources at the medical center including tap water from sinks and showerheads, the hospital hot water source, and the city water supply to the hospital. Analysis by multilocus enzyme electrophoresis and randomly amplified polymorphic DNA showed a match between one patients blood isolate and an isolate from shower water from that patients prior hospital room. CONCLUSIONS The cause of the outbreak seemed to be water contamination of central venous catheters (CVCs) during bathing. A recommendation in early 2001 that CVCs be protected from water during bathing was followed by no M. mucogenicum bacteremias during the second half of 2001, only one in 2002, and none at all during 2003.

Repeat Positive Cultures in Mycobacterium intracellulare Lung Disease after Macrolide Therapy Represent New Infections in Patients with Nodular Bronchiectasis

The genomic DNA patterns (genotypes) of 55 episodes of late positive sputum isolates, collected after >or=4 consecutive months of negative sputum cultures, in prospective macrolide treatment trials of Mycobacterium avium complex (MAC) lung disease were assessed by pulsed-field gel electrophoresis (PFGE). Having >or=2 cultures positive for MAC after completion of therapy was documented 23 times; of 20 episodes studied by PFGE, 17 (85%) represented new genotypes (i.e., new infections), and 87% occurred in patients with nodular bronchiectasis. With >or=2 positive cultures after therapy was stopped prematurely, 6 (86%) of 7 episodes were relapses. Single positive cultures after completion of therapy occurred 16 times; only 1 (6%) was predictive of a subsequent relapse. No late isolates were macrolide resistant. Thus, relapses of MAC lung disease with these macrolide regimens are unusual, and most infections after completing therapy resulted from new strains in patients with nodular bronchiectasis.

Pulsed-Field Gel Electrophoresis Study of Mycobacterium abscessus Isolates Previously Affected by DNA Degradation

ABSTRACT DNA degradation (which results in a smear pattern) occurs with almost 50% of Mycobacterium abscessus strains during pulsed-field gel electrophoresis (PFGE). We assessed the potential benefit of using thiourea-containing buffer with M. abscessus by studying 69 isolates not previously typeable by PFGE (i.e., those with a smear pattern). Random (epidemiologically unrelated) isolates that were typeable (no smear pattern) were included as controls. Genomic DNA was digested with DraI, XbaI, and AseI. PFGE gels were run in regular gel buffer with and without 100 μM thiourea. All 69 isolates that generated smear patterns had clear band profiles when the thiourea buffer was used. These isolates were divided into only 30 patterns with DraI, 20 patterns with XbaI, and 20 patterns with AseI. The molecular profiles were all closely or possibly related, and the differences between the isolates ranged from zero to six bands. By multilocus enzyme electrophoresis (MEE), 45 of 53 smear isolates (85%) belonged to two closely related electrophoretic types. These isolates contained at least one enzyme allele seen almost exclusively in this group. Isolates without smear patterns were unaffected by thiourea and produced unrelated PFGE profiles, as well as multiple MEE types. The hsp65 and 16S rRNA gene sequences of the isolates with smear patterns were identical to those of M. abscessus type strain ATCC 19977, which had a nonsmear pattern, suggesting that this clone is a subgroup within M. abscessus. This demonstrates that the inability to type M. abscessus by PFGE is associated with a single clone of organisms.

Comparison of Methods for Identification of Mycobacterium abscessus and M. chelonae Isolates

ABSTRACT Mycobacterium abscessus and Mycobacterium chelonae are two closely related species that are often not distinguished by clinical laboratories despite the fact they cause diseases requiring different treatment regimens. Multilocus enzyme electrophoresis, PCR-restriction fragment length polymorphism analysis of the 65-kDa heat shock protein gene, biochemical tests, and high-performance liquid chromatography of mycolic acids were used to identify 75 isolates as either M. abscessus or M. chelonae that were originally submitted for drug susceptibility testing. Only 36 of these isolates were submitted with an identification at the species level. Using the above methods, 46 of the isolates were found to be M. abscessus and 29 were identified as M. chelonae. Eight isolates originally submitted as M. chelonae were identified as M. abscessus, and one isolate submitted as M. abscessuswas found to be M. chelonae. The four identification methods were in agreement in identifying 74 of the 75 isolates. In drug susceptibility testing, all isolates of M. abscessusexhibited resistance to tobramycin (MIC of 8 to ≥16 μg/ml), while all isolates of M. chelonae were susceptible to this drug (MIC of ≤4 μg/ml). The results suggest that once an identification method is selected, clinical laboratories should be able to easily identify isolates of M. abscessus and M. chelonae.

Furunculosis Due to Mycobacterium mageritense Associated with Footbaths at a Nail Salon

ABSTRACT We report two cases of lower-extremity furunculosis caused by Mycobacterium mageritense. Both patients were patrons of the same nail salon, where they received footbaths prior to pedicures. M. mageritense bacteria isolated from two whirlpool footbaths were determined to be closely related to the patient isolates by pulsed-field gel electrophoresis.

Structural analysis of biofilm formation by rapidly and slowly growing nontuberculous mycobacteria.

ABSTRACT Mycobacterium avium complex (MAC) and rapidly growing mycobacteria (RGM) such as M. abscessus, M. mucogenicum, M. chelonae, and M. fortuitum, implicated in health care-associated infections, are often isolated from potable water supplies as part of the microbial flora. To understand factors that influence growth in their environmental source, clinical RGM and slowly growing MAC isolates were grown as biofilm in a laboratory batch system. High and low nutrient levels were compared, as well as stainless steel and polycarbonate surfaces. Biofilm growth was measured after 72 h of incubation by enumeration of bacteria from disrupted biofilms and by direct quantitative image analysis of biofilm microcolony structure. RGM biofilm development was influenced more by nutrient level than by substrate material, though both affected biofilm growth for most of the isolates tested. Microcolony structure revealed that RGM develop several different biofilm structures under high-nutrient growth conditions, including pillars of various shapes (M. abscessus and M. fortuitum) and extensive cording (M. abscessus and M. chelonae). Although it is a slowly growing species in the laboratory, a clinical isolate of M. avium developed more culturable biofilm in potable water in 72 h than any of the 10 RGM examined. This indicates that M. avium is better adapted for growth in potable water systems than in laboratory incubation conditions and suggests some advantage that MAC has over RGM in low-nutrient environments.

Persistence of Nontuberculous Mycobacteria in a Drinking Water System after Addition of Filtration Treatment

ABSTRACT There is evidence that drinking water may be a source of infections with pathogenic nontuberculous mycobacteria (NTM) in humans. One method by which NTM are believed to enter drinking water distribution systems is by their intracellular colonization of protozoa. Our goal was to determine whether we could detect a reduction in the prevalence of NTM recovered from an unfiltered surface drinking water system after the addition of ozonation and filtration treatment and to characterize NTM isolates by using molecular methods. We sampled water from two initially unfiltered surface drinking water treatment plants over a 29-month period. One plant received the addition of filtration and ozonation after 6 months of sampling. Sample sites included those at treatment plant effluents, distributed water, and cold water taps (point-of-use [POU] sites) in public or commercial buildings located within each distribution system. NTM were recovered from 27% of the sites. POU sites yielded the majority of NTM, with >50% recovery despite the addition of ozonation and filtration. Closely related electrophoretic groups of Mycobacterium avium were found to persist at POU sites for up to 26 months. Water collected from POU cold water outlets was persistently colonized with NTM despite the addition of ozonation and filtration to a drinking water system. This suggests that cold water POU outlets need to be considered as a potential source of chronic human exposure to NTM.

Multiphasic Approach Reveals Genetic Diversity of Environmental and Patient Isolates of Mycobacterium mucogenicum and Mycobacterium phocaicum Associated with an Outbreak of Bacteremias at a Texas Hospital

ABSTRACT Between March and May 2006, a Texas hospital identified five Mycobacterium mucogenicum bloodstream infections among hospitalized oncology patients using fluorescence high-performance liquid chromatography analysis of mycolic acids. Isolates from blood cultures were compared to 16 isolates from environmental sites or water associated with this ward. These isolates were further characterized by hsp65, 16S rRNA, and rpoB gene sequencing, hsp65 PCR restriction analysis, and molecular typing methods, including repetitive element PCR, random amplified polymorphic DNA PCR, and pulsed-field gel electrophoresis (PFGE) of large restriction fragments. Three of five patient isolates were confirmed as M. mucogenicum and were in a single cluster as determined by all identification and typing methods. The remaining two patient isolates were identified as different strains of Mycobacterium phocaicum by rpoB sequence analysis. One of these matched an environmental isolate from a swab of a hand shower in the patients room, while none of the clinical isolates of M. mucogenicum matched environmental strains. Among the other 15 environmental isolates, 11 were identified as M. mucogenicum and 4 as M. phocaicum strains, all of which were unrelated by typing methods. Although the 16S rRNA gene sequences matched for all 14 M. mucogenicum isolates, there were two each of the hsp65 and rpoB sequevars, seven PCR typing patterns, and 12 PFGE patterns. Among the seven M. phocaicum isolates were three 16S rRNA sequevars, two hsp65 sequevars, two rpoB sequevars, six PCR typing patterns, and six PFGE patterns. This outbreak represents the first case of catheter-associated bacteremia caused by M. phocaicum and the first report of clinical isolates from a U.S. hospital. The investigation highlights important differences in the available typing methods for mycobacteria and demonstrates the genetic diversity of these organisms even within narrow confines of time and space.

First Isolations of Segniliparus rugosus from Patients with Cystic Fibrosis

ABSTRACT We report three cases of the new genus Segniliparus isolated from patients with cystic fibrosis. All isolates were unambiguously identified by 16S rRNA gene sequencing as Segniliparus rugosus (GenBank accession no. AY 60892). Drug susceptibility results that may enhance treatment for cystic fibrosis patients with this opportunistic pathogen are presented.