A multi-center outbreak of Candida tropicalis bloodstream infections associated with contaminated hemodialysis machine prime buckets.

BACKGROUND: Outbreaks of fungal bloodstream infection (BSI) are uncommon among hemodialysis patients. We investigated an outbreak of Candida tropicalis BSIs involving patients at 3 of 4 affiliated hemodialysis units. METHODS: An investigation included a review of records of patients with C tropicalis BSI, a case-control study, and cultures of medications, hands of personnel, dialysis equipment, and water samples. RESULTS: Eight patients developed C tropicalis BSIs in a 3-month period. Compared to controls, cases had a higher proportion of preceding dialyses performed on a machine with a contaminated saline prime bucket (SPB) (P= .02). Observations revealed that SPBs at units A-C were rinsed with tap water, were not routinely disinfected, and that priming tubing was allowed to contact fluid in SPBs. C tropicalis was recovered from the main compartment and hollow handle of SPBs and from other environmental samples. C tropicalis isolates from patients, SPBs and other environmental samples had indistinguishable pulsed-field gel electrophoresis patterns. Following routine disinfection of SPBs, the outbreak terminated. CONCLUSIONS: This outbreak was likely due to inadequate disinfection of SPBs. The findings emphasize the importance of disinfection of SPBs. Current use of identical SPBs warrants further evaluation of hollow SPB handles as a potential infection risk.

Wild-type minimum effective concentration distributions and epidemiologic cutoff values for caspofungin and Aspergillus spp. as determined by Clinical and Laboratory Standards Institute broth microdilution methods.

Antifungal susceptibility testing of Aspergillus spp. against caspofungin has been standardized by the Clinical and Laboratory Standards Institute (CLSI). Recent studies have documented breakthrough infections with Aspergillus spp. for which the minimum effective concentration (MEC) for caspofungin ranged from 0.25 to 8 microg/mL. We tested a collection of 1590 clinical isolates of Aspergillus spp. (188 Aspergillus flavus, 1187 Aspergillus fumigatus, 114 Aspergillus niger, 71 Aspergillus terreus, and 30 Aspergillus versicolor) against caspofungin using the CLSI broth microdilution method. An epidemiologic cutoff value (ECV) of 8 microg/mL [1]), and 1 A. versicolor (4 microg/mL). The establishment of the WT MEC distributions and ECVs for caspofungin and the major species of Aspergillus will be useful in resistance surveillance and is an important step toward the development of clinical breakpoints.

Comparison of results of fluconazole and voriconazole disk diffusion testing for Candida spp. with results from a central reference laboratory in the ARTEMIS DISK Global Antifungal Surveillance Program.

The accuracy of antifungal susceptibility testing is important for reliable resistance surveillance and for the clinical management of patients with serious infections due to Candida spp. Our primary objective was to compare the results of fluconazole and voriconazole disk diffusion testing of 3227 Candida spp. performed by 47 centers participating in the ARTEMIS program with disk diffusion and MIC results obtained by the central reference laboratory. The overall categoric agreement between participant disk diffusion test results and reference MIC results was 87% for fluconazole and 95.2% for voriconazole. Likewise good agreement was observed between participant disk diffusion test results and reference laboratory disk diffusion test results, with an agreement of 90.5%, 1% very major error (VME), and 3.4% major error (ME) for fluconazole and 94.2%, 1.1% VME, and 2.5% ME for voriconazole. The disk diffusion test was reliable for detecting those isolates of Candida spp. that were characterized as resistant to fluconazole and voriconazole by MIC testing. External quality assurance data obtained by surveillance programs such as the ARTEMIS Global Antifungal Surveillance Program ensure the generation of useful surveillance data and result in the continued improvement of antifungal susceptibility testing protocols.

A global evaluation of voriconazole activity tested against recent clinical isolates of Candida spp.

Voriconazole susceptibility testing was performed on 7191 Candida spp. from 78 centers worldwide between 2004 and 2007. Voriconazole was very active in vitro (MIC(50)/MIC(90), 0.008/0.25 microg/mL; 98% susceptible). In comparison to 5866 Candida spp. isolates collected during global surveillance from 1997 to 2001, there were no changes in voriconazole mean MIC or MIC distribution.

Evaluation of disk diffusion and Etest compared to broth microdilution for antifungal susceptibility testing of posaconazole against clinical isolates of filamentous fungi.

We performed Etest, disk diffusion, and broth microdilution susceptibility testing of posaconazole against 146 clinical isolates of filamentous fungi. By using provisional breakpoints for comparison purposes only, categorical agreement between the results of the agar-based methods and those of broth microdilution were 96 to 98%, with no very major errors. These agar-based methods hold promise as simple and reliable methods for determining the posaconazole susceptibilities of filamentous fungi.

Evaluation of Etest and disk diffusion methods compared with broth microdilution antifungal susceptibility testing of clinical isolates of Candida spp. against posaconazole.

We performed Etest, disk diffusion, and broth microdilution susceptibility testing of 2,171 clinical isolates of Candida spp. against posaconazole. By using provisional breakpoints for comparison purposes only, the categorical agreement between the agar-based methods and broth microdilution results ranged from 93 to 98%, with <1% very major errors. The essential agreement (within 2 well dilutions) between the Etest and broth microdilution methods was 94%. These agar-based methods hold promise as simple and reliable methods for determination of the posaconzole susceptibilities of Candida spp.

Antifungal susceptibilities of Candida species causing vulvovaginitis and epidemiology of recurrent cases.

There are limited data regarding the antifungal susceptibility of yeast causing vulvovaginal candidiasis, since cultures are rarely performed. Susceptibility testing was performed on vaginal yeast isolates collected from January 1998 to March 2001 from 429 patients with suspected vulvovaginal candidiasis. The charts of 84 patients with multiple positive cultures were reviewed. The 593 yeast isolates were Candida albicans (n = 420), Candida glabrata (n = 112), Candida parapsilosis (n = 30), Candida krusei (n = 12), Saccharomyces cerevisiae ( n = 9), Candida tropicalis (n = 8), Candida lusitaniae (n = 1), and Trichosporon sp. (n = 1). Multiple species suggesting mixed infection were isolated from 27 cultures. Resistance to fluconazole and flucytosine was observed infrequently (3.7% and 3.0%); 16.2% of isolates were resistant to itraconazole (MIC > or = 1 microg/ml). The four imidazoles (econazole, clotrimazole, miconazole, and ketoconazole) were active: 94.3 to 98.5% were susceptible at < or =1 microg/ml. Among different species, elevated fluconazole MICs (> or = 16 microg/ml) were only observed in C. glabrata (15.2% resistant [R], 51.8% susceptible-dose dependent [S-DD]), C. parapsilosis (3.3% S-DD), S. cerevisiae (11.1% S-DD), and C. krusei (50% S-DD, 41.7% R, considered intrinsically fluconazole resistant). Resistance to itraconazole was observed among C. glabrata (74.1%), C. krusei (58.3%), S. cerevisiae (55.6%), and C. parapsilosis (3.4%). Among 84 patients with recurrent episodes, non-albicans species were more common (42% versus 20%). A > or = 4-fold rise in fluconazole MIC was observed in only one patient with C. parapsilosis. These results support the use of azoles for empirical therapy of uncomplicated candidal vulvovaginitis. Recurrent episodes are more often caused by non-albicans species, for which azole agents are less likely to be effective.

Evaluation of three molecular typing techniques for nonfermentative Gram-negative bacilli.

OBJECTIVE: To evaluate three different DNA techniques for typing nonfermentative gram-negative bacilli isolated from Latin American hospitals. DESIGN: One hundred twenty-six nonfermentative gram-negative bacilli were typed. PARTICIPANTS: Pseudomonas aeruginosa (n = 64) and Acinetobacter baumannii (n = 42) samples were obtained from blood cultures of patients admitted to 10 medical centers in Latin America during 1998 and Stenotrophomonas maltophilia (n = 20) samples were obtained from patients admitted to the Hospital São Paulo between 1999 and 2001. METHODS: All samples were typed using automated ribotyping, PFGE, and ERIC-PCR. The discriminatory power for each technique was calculated using Hunter's generalized formula. RESULTS: All strains could be typed by automated ribotyping and ERIC-PCR, but two strains (1.6%) were not typeable by PFGE. All three techniques showed 100% reproducibility. The time to obtain the results was shorter for automated ribotyping and ERIC-PCR compared with PFGE. Likewise, the costs for ERIC-PCR and PFGE were lower than those for automated ribotyping. The interpretation of results was more complicated and more difficult with ERIC-PCR than with both PFGE and automated ribotyping. All techniques presented excellent discriminatory power for P. aeruginosa (0.98). PFGE presented the highest discriminatory power (0.94) for A. baumannii, and both PFGE and ERIC-PCR showed higher discriminatory power (0.90 for both) than automated ribotyping (0.82) for S. maltophilia. CONCLUSIONS: PFGE showed the highest discriminatory power for typing these nonfermentative gram-negative bacilli. However, automated ribotyping and ERIC-PCR can provide results in a shorter time period with similar discriminatory power.

Genotypic characterization of carbapenem-nonsusceptible Acinetobacter spp. isolated in Latin America.

The principal aim of this study was to evaluate the genomic diversity among the imipenem-nonsusceptible Acinetobacter spp. (INSA) collected from the Latin American medical centers within the SENTRY Antimicrobial Surveillance Program. The INSA isolates were collected from patients with bloodstream infections, who were hospitalized in seven Latin American countries between 1997 and 1999. For epidemiologic comparison, 20 carbapenem-susceptible Acinetobacter spp. (CSA) isolates were collected in the same period of time from the respective medical centers. A total of 23 Acinetobacter spp. isolates exhibiting imipenem MIC values of >/=8 microg/ml were typed by ribotyping, an automated molecular method. The isolates showing an identical ribogroup were also typed by pulsed-field gel electrophoresis (PFGE). The antimicrobial susceptibility to various antimicrobial agents was evaluated using a reference broth microdilution technique. Among the INSA isolates, 13 distinct ribogroups were observed, whereas 16 ribogroups were detected among the CSA. Nearly 57% of the INSA belonged to only four ribogroups. Identical ribogroups and PFGE patterns were observed among INSA and CSA isolates collected from medical centers located in different countries (Brazil and Argentina). Our results showed: (1) a higher genomic variability among the CSA; (2) presence of epidemic clones among INSA isolates encountered in Latin American medical centers; and (3) spread of INSA and CSA epidemic clones between Latin American countries.

Caspofungin activity against clinical isolates of fluconazole-resistant Candida.

A total of 7,837 clinical isolates of Candida were tested against fluconazole, and 351 resistant (fluconazole MIC >/=64 micro g/ml) isolates were identified (4% of the total tested). All fluconazole-resistant isolates were inhibited by caspofungin at concentrations that can be exceeded by standard doses (MIC at which 90% of the isolates were inhibited, 1 micro g/ml; 99% of the MICs were

Outbreak of Candida rugosa candidemia: an emerging pathogen that may be refractory to amphotericin B therapy.

Candida rugosa has been rarely reported as a human pathogen. We retrospectively evaluated a cluster of Candida rugosa candidemia cases occurring in six hospitalized patients from a tertiary care teaching hospital in São Paulo, Brazil. Genetic relatedness among the six C. rugosa outbreak isolates was characterized by RAPD assay using 3 different 10-mer primers and by pulsed field gel electrophoresis. The source of the outbreak was not identified. All patients had been subjected to invasive medical procedures, including central venous catheterization, surgery or dialysis. Two patients were undergoing amphotericin B therapy prior to the onset of candidemia. The crude mortality rate was very high, despite antifungal therapy. C. rugosa may represent an emerging pathogen associated with invasive medical procedures, able to infect immunocompetent hosts causing serious systemic infection refractory to amphotericin B therapy.

Improving typeability of multiple bacterial species using pulsed-field gel electrophoresis and thiourea.

Although pulsed-field gel electrophoresis is considered the "gold standard" technique for molecular typing, typeability may not be excellent for some bacterial species because of DNA degradation. Previous reports suggest that the addition of thiourea in the gel buffer can improved the typeability for some species. In the present study, 66 Gram-negative strains (seven species) known to be affected by DNA degradation and four control strains were evaluated by PFGE with and without the addition of 50 microg/M of thiourea to the buffer used in the electrophoresis. Macrorestriction patterns were obtained for all K. pneumoniae, S. marcescens, P. aeruginosa, and Salmonella spp., for 95.4% of E. coli, and for 50% of E. cloacae strains from the gels performed in the buffer with throurea. However, typeability was not improved for Acinetobacter spp. The range of non-typeable species for which thiourea can limit the problem of DNA degradation is considerably wider than described in previous publications.

Donor-to-host transmission of Candida albicans after corneal transplantation.

PURPOSE: To report donor-to-host transmission of Candida albicans after penetrating keratoplasty. DESIGN: Interventional case report. METHODS: A 15-year-old boy who underwent penetrating keratoplasty for keratoconus with donor tissue from a drowning victim developed keratitis and a lenticular abscess 26 days postoperatively. RESULTS: Candida albicans was cultured from the donor rim and the recipient cornea. Antifungal sensitivity profiles were identical for the two isolates. DNA profiles were identical for both isolates, confirming the donor as the source of the infection. CONCLUSION: This case demonstrates the value of routine culture of corneal donor rims and the advisability of close follow-up and possible antifungal prophylaxis when donor rims are positive for fungus.

Comparison of an automated ribotyping system to restriction endonuclease analysis and pulsed-field gel electrophoresis for differentiating vancomycin-resistant Enterococcus faecium isolates.

The RiboPrinter Microbial Characterization System was compared with pulsed-field gel electrophoresis (PFGE), restriction endonuclease analysis (REA), and epidemiological data for typing 45 vancomycin-resistant Enterococcus faecium (VRE) isolates. In 21 clinically related isolates, 90 to 100% were similar by PFGE and REA, but only 57% were similar by the RiboPrinter. In another eight clinically related isolates, three isolates similar by PFGE and REA were all unique by the RiboPrinter. In contrast, in 16 clinically unrelated isolates, the predominant RiboPrinter ribotype represented 50% of the strains, while the largest PFGE and REA clones represented less than 19% of the strains. These data suggest that the RiboPrinter is not reliable for VRE investigation.

Change in colony morphology of Candida lusitaniae in association with development of amphotericin B resistance.

It is not uncommon to see amphotericin B treatment failure in patients with systemic infection caused by Candida lusitaniae. We report a patient with stage IV ovarian carcinoma and C. lusitaniae sepsis whose treatment with amphotericin B failed. The initial blood isolate was susceptible to amphotericin B in vitro; however, the MIC for a blood isolate recovered 7 weeks after treatment began showed a fourfold increase. Direct subculture of two positive blood samples obtained within a week of the patient's death showed the coexistence of two distinct colony color variants on CHROMagar Candida (CAC). One variant was susceptible to amphotericin B, and one was resistant. These results emphasize the importance of repeat amphotericin B susceptibility testing for patients with persistent C. lusitaniae infection. The presence of colony variants on CAC may signal the emergence of amphotericin B resistance in C. lusitaniae and should be investigated.

Evaluation of the discriminatory power of pulsed-field gel electrophoresis and PCR fingerprinting for epidemiologic typing of Candida species.

OBJECTIVE: To evaluate the discriminatory power of genotyping methods (PCR fingerprinting and pulsed-field gel electrophoresis) validated for Candida albicans in other Candida species. METHODS: Molecular typing methods are increasingly being applied for studies where the interpretation of data essentially relies on the typing results rather than epidemiologic data. In this situation, the discriminatory power (ability to identify differences among epidemiologically unrelated strains) of the typing method is important in allowing one to draw valid conclusions. By applying PCR fingerprinting, electrophoretic karyotyping, and restriction fragment endonuclease analysis using standard restriction enzymes and primers proven to be useful in previous studies, we evaluated whether the use of multiple genotyping methods is sufficient to delineate known unrelated strains among seven Candida species. RESULTS: All three methods identified individual genotypes for each of the seven Candida species studied. However, optimal strain delineation required the combined use of all three typing methods and was observed only within the small number of C. albicans and C. tropicalis isolates tested in this study. CONCLUSION: Typing assays that are able to delineate a certain Candida species may not be used blindly for other species of that genus. Regarding the limited number of strains tested, further validation of the discriminative power of genotyping methods (including in C. tropicalis) should be done.