Biomedical subjects

M A Pfaller

Publications and source records attributed to M A Pfaller.

At least 271 records · Page 15Linked to original sources

PubMed1996

DNA macrorestriction profiles and antifungal susceptibility of Candida (Torulopsis) glabrata.

Candida (Torulopsis) glabrata is an emerging nosocomial pathogen that may be relatively resistant to fluconazole. A series of 75 isolates (blood, urine, tissue, and other sites) from 16 patients (1 to 12 isolates per patient) at a single university medical center were analyzed by pulsed field gel electrophoresis (PFGE) of restriction endonuclease digests of chromosomal DNA. The MICs of the isolates for amphotericin B, flucytosine, fluconazole, and itraconazole, were determined by a microdilution broth method. A preliminary study of seven restriction enzymes, three producing small fragments (Hinf I, Hind III, Eco RI) and four producing large fragments (Eag I, BssH II, Sfi I, Not I) identified Not I as giving interpretable banding patterns. Isolates were considered of different types if they differed by two or more bands. Sixteen distinct DNA types (A to P) were identified. Karyotyping was used an an additional technique to compare strains with a common PFGE type. Most patients were colonized or infected with a single type at multiple body sites and over time. One PFGE type was shared by four patients housed in different areas of the institution at different times. For two of these four patients, the karyotype was also indistintuishable. Five patients were each colonized with two distinct types. The MIC of the strains studied were amphotericin B 0.5-1.0 microgram/ml (MIC90 = 1.0 microgram/ ml), 5-fluorocytosine 0.25-->256 micrograms/ml (MIC90 = 2 micrograms/ml), fluconazole 0.25-->128 micrograms/ml (MIC90 = 32 micrograms/ml), and itraconazole 0.06-8.0 micrograms/ml). Molecular typing by PFGE using Not I digestion is a useful technique for epidemiological investigation as epidemiologically related isolates are generally identical and epidemiologically unrelated isolates are different by this method.

Candida↗

PubMed1996

Clinical evaluation of a dried commercially-prepared microdilution panel for antifungal susceptibility testing.

A commercially-prepared dried broth microdilution panel (Sensititre) was compared with a reference microdilution method for antifungal susceptibility testing of two reference yeast strains and 98 clinical isolates of Candida spp. The antifungal agents tested include 5-fluorocytosine (5FC), fluconazole, itraconazole, and D0870. Microdilution testing was performed according to National Committee for Clinical Laboratory Standards (NCCLS) recommendations. Minimum inhibitory concentration (MIC) endpoints were read visually after 48 hours of incubation and were assessed independently for each microdilution panel. The MICs for the reference strains were within published control limits for both reference and Sensititre microdilution panels. Discrepancies among MIC endpoints of no more than two dilutions (two wells) were used to calculate the percent agreement. An acceptable level of agreement between Sensititre and reference panels was observed for all antifungal agents when tested against the 98 clinical isolates. Agreement ranged from 83% for itraconazole to 93% for 5FC. The Sensititre dried microdilution panel appears to be a viable alternative to inhouse prepared microdilution panels and to the NCCLS microdilution reference method.

Antifungal Agents↗

PubMed1996

Phenotypic detection of mec A-positive staphylococcal blood stream isolates: high accuracy of simple disk diffusion tests.

Detection of oxacillin-resistance in staphylococci by phenotypic methods remains problematic. Although standardized susceptibility test methods are adequate for Staphylococcus aureus, many are less satisfactory for the coagulase-negative staphylococci (CNS). We have studied 108 consecutive blood culture isolates of staphylococci. The mec A gene was detected by PCR in one S. aureus and 55 CNS isolates. Susceptibility testing was performed as follows: oxacillin (1-microgram), ceftizoxime (30-microgram), and cephalothin (30-microgram) by disk diffusion; oxacillin, ceftizoxime, cephalothin, methicillin, ampicillin, ampicillin/ sulbactam, penicillin, cefazolin, imipenem, and meropenem by the broth microdilution method. In addition, isolates were tested by the oxacillin agar screen plate method. The single oxacillin-resistant S. aureus strain was detected by all oxacillin susceptibility test methods and by the ceftizoxime disk and MIC methods. Two oxacillin-susceptible S. aureus were intermediate (minor error) by ceftizoxime broth microdilution (MIC, 16 micrograms/mL). The most sensitive, simple phenotypic methods for detection of oxacillin-resistant CNS (mec A positive) were as follows: oxacillin disk diffusion at 98%, oxacillin screen plate at 91%, oxacillin broth microdilution at 87%, ceftizoxime disk diffusion at 100%, ceftizoxime broth microdilution at 87%, and methicillin broth microdilution at 83%. These results indicate that oxacillin and ceftizoxime disk diffusion tests are the most accurate phenotypic methods in routine clinical use for detection of oxacillin-resistant CNS. Oxacillin broth microdilution MIC testing (2% NaCl supplement) would perform more satisfactorily (100% sensitivity) with an adjusted interpretive breakpoint at < or = 0.5 microgram/mL, in contrast to the lower accuracy of the "so-called" reference agar screen test.

Anti-Bacterial Agents↗

PubMed1996

Transmission of Citrobacter koseri from mother to infant documented by ribotyping and pulsed-field gel electrophoresis.

We describe a case in which Citrobacter koseri (formerly C. diversus) was transmitted from a pregnant mother with chorioamnionitis and bacteremia to her infant who was bacteremic at birth and in apparent septic shock. Two highly discriminating molecular methods, ribotyping and pulsed field gel electrophoresis, were used to examine restriction fragment length polymorphisms within the genomic DNA of maternal and infant isolates. Both techniques identified the maternal and infant isolates as the same strain, distinct from epidemiologically unrelated controls, thus confirming their common origin.

Adult↗

PubMed1996

A prolonged outbreak of methicillin-resistant Staphylococcus aureus in the burn unit of a tertiary medical center.

OBJECTIVE: To report an outbreak of methicillin-resistant Staphylococcus aureus (MRSA) in our burn unit and the steps we used to eradicate the organism. DESIGN AND SETTING: Outbreak investigation in the burn unit of a 900-bed tertiary-care medical center. OUTBREAK: Between March and June 1993, MRSA was isolated from 10 patients in our burn unit. All isolates had identical antibiograms and chromosomal DNA patterns. CONTROL MEASURES: Infection control personnel encouraged healthcare workers to wash their hands after each patients contact. The unit cohorted all infected or colonized patients, placed each affected patient in isolation, and, if possible, transferred the patient to another unit. Despite these measures, new cases occurred. Infection control personnel obtained nares cultures from 56 healthcare workers, 3 of whom carried the epidemic MRSA strain. One healthcare worker cared for six affected patients, and one cared for five patients. We treated the three healthcare workers with mupirocin. Subsequently, no additional patients became colonized or infected with the epidemic MRSA strain. CONCLUSIONS: The outbreak ended after we treated healthcare workers who carried the epidemic strain with mupirocin. This approach is not appropriate in all settings. However, we felt it was justified in this case because of a persistent problem after less intrusive measures.

Burn Units↗

PubMed1996

The positive predictive value of isolating coagulase-negative staphylococci from blood cultures.

We used four criteria to define true bloodstream infections after isolation of coagulase-negative staphylococci (CNS) from Isolator (Wampole Laboratories, Cranbury, NJ) blood cultures: (1) the patient's temperature was > or = 38 degrees C, (2) appropriate treatment was administered, (3) the physician diagnosed bloodstream infection or criteria for nosocomial bloodstream infection were met, and (4) at least one clinical sign or laboratory value was consistent with infection. Sixty (26.4%) of 227 episodes met these four criteria. By logistic regression, variables associated with meeting the definition of infection were admission to a service other than the surgical intensive care unit, the biotype of the Staphylococcus epidermidis isolates, the log of the weighted average of the total number of bacteria per milliliter of blood in all positive cultures, resistance to at least six antimicrobial agents, and the positivity of a BACTEC blood culture specimen that was drawn with the first positive Isolator culture specimen. In a high-risk population, 26% of Isolator blood cultures positive for CNS represented infections, a rate two to four times greater than that reported in the literature. Information regarding the species, biotype, antibiogram, and number of organisms per milliliter of blood might help physicians distinguish between CNS bloodstream infections and contamination.

Adult↗

PubMed1996

Vertical and horizontal transmission of unique Candida species to premature newborns.

The number of nosocomial bloodstream infections due to Candida species in critically ill newborns is increasing. This pathogen may be vertically transmitted from the mother or nosocomially acquired in the nursery. The goal of this study was to identify the route of transmission of unique Candida species and strains from mothers to their preterm offspring. Specimens from mothers for fungal cultures were obtained before delivery, and specimens from infants for sequential fungal cultures were obtained at defined intervals. Candida species were identified by standard methods and were typed by electrophoretic karyotyping (EK) and restriction endonuclease analysis of genomic DNA (REAG) with pulsed-field gel electrophoresis. Antifungal susceptibility testing was performed on all isolates. Fungal cultures were positive for Candida species in 12 (63%) of 19 mothers' specimens and in seven (33%) of 21 infants' specimens. EK and REAG revealed that both the mother and the infant in three (14%) of 21 mother-infant pairs were colonized with the identical strain of Candida albicans. C. albicans was most commonly transmitted vertically. Candida parapsilosis colonized other infants and could not be accounted for by a maternal reservoir.

Candida↗

PubMed1996

Nosocomial candidiasis: emerging species, reservoirs, and modes of transmission.

During the 1980s, the frequency of nosocomial candidiasis increased dramatically. This trend has continued into the 1990s, and Candida species remain a major cause of nosocomial infections. Although Candida albicans remains the most frequent cause of fungemia and hematogenously disseminated candidiasis, a number of reports have documented infections caused by other Candida species: C. tropicalis, C. glabrata, C. parapsilosis, C. krusei, and C. lusitaniae. Many of these infections arise from an endogenous source, and their frequency is influenced by the patient population, the various treatment regimens, and the antibiotics or other supportive care measures employed at specific institutions. Additional infections may be accounted for by exogenous acquisition via the hands of health care workers, contaminated infusates and biomaterials, and the inanimate environment. Ongoing investigation should help improve our understanding of the epidemiology of candidiasis and facilitate the development of rational preventive measures.

Antifungal Agents↗

PubMed1996

Use of macrorestriction analysis to demonstrate interhospital spread of multiresistant Acinetobacter baumannii in São Paulo, Brazil.

We evaluated the spread of Acinetobacter baumannii strains among three hospitals located in São Paulo, Brazil. A total of 46 isolates, which were typed by chromosomal DNA analysis with use of pulsed field gel electrophoresis (PFGE), were tested for susceptibility to the fluoroquinolones, carbapenems, aminoglycosides (amikacin), cephalosporins, polymyxin B, and ampicillin/sulbactam by means of the broth microdilution method, disk diffusion, and the E-test. Isolates with an identical PFGE pattern (pattern B) that were susceptible only to carbapenems, polymyxin B, and ampicillin/ sulbactam were recovered in all three hospitals. In addition, isolates with PFGE pattern A that were susceptible only to polymyxin B and ampicillin/sulbactam were recovered in hospitals 1 and 2. The results of our study strongly suggest the interhospital transmission of multiresistant epidemic strains of A. baumannii in São Paulo. Once in the hospital, these strains can disseminate and cause outbreaks with devastating consequences.

Acinetobacter↗

PubMed1996

Molecular subtypes and antifungal susceptibilities of serial Cryptococcus neoformans isolates in human immunodeficiency virus-associated Cryptococcosis. Cryptococcal Disease Active Surveillance Group.

Serial isolates of Cryptococcus neoformans from 33 human immunodeficiency virus-infected patients with cryptococcosis were analyzed to determine whether persistence might result from reinfection with a new cryptococcal strain or acquisition of antifungal resistance. Isolates were subtyped by multilocus enzyme electrophoresis (MEE), electrophoretic karyotyping (EK), random-amplified polymorphic DNA (RAPD), and the CNRE-1 DNA probe, MICs of amphotericin B, fluconazole, and 5-fluorocytosine were determined. No changes in MEE or RAPD subtypes were detected in serial isolates from any patient. Isolates from 8 patients (24%) showed alterations in EK only (mobility change in two or more bands) but not with any other subtyping method. MICs did not change significantly in isolates from 30 patients. In 1 case, the fluconazole MIC increased stepwise over 18 months, suggesting development of resistance. These overall invariant subtyping and MIC results confirm previous studies suggesting that persistent cryptococcal infection is due to relapse rather than reinfection or antifungal drug resistance.

AIDS-Related Opportunistic Infections↗

PubMed1996

Standardization of antifungal susceptibility testing.

The application of in-vitro antifungal susceptibility testing to clinical research and to the guidance of antifungal therapy has been limited by a lack of reproducibility and uncertain clinical relevance. As a result of studies of the identified variables impacting on in-vitro susceptibility results, the National Committee for Clinical Laboratory Standards have proposed a standardized antifungal susceptibility test method M27-P. The degree of intra- and inter-laboratory reproducibility which can be achieved with this method have been defined in multi-laboratory collaborative studies. More convenient methods (microdilution broth and stable gradient technology) have been evaluated relative to the proposed standard method and the potential for a similar process with a disc diffusion method is apparent. A modification of this standard method for susceptibility testing of filamentous fungi appears promising. The existence of a standardized method and of alternative methods with a defined relationship to the proposed standard, facilitates meaningful analysis of published studies addressing the issue of clinical relevance of antifungal susceptibility testing. As a result of this process, correlation of MICs determined in vitro with clinical response to therapy is beginning to emerge, most notably in relation to fluconazole therapy for oropharyngeal candidosis associated with infection with the human immunodeficiency virus.

Antifungal Agents↗

PubMed1996

Microdilution antifungal susceptibility testing of Candida albicans and Cryptococcus neoformans with and without agitation: an eight-center collaborative study.

The growth patterns observed in the trailing wells when fluconazole is being tested may give rise to readings that suggest resistance or increased MICs for known susceptible strains. We conducted a multicenter study to evaluate the intralaboratory and interlaboratory reproducibilities of a method that uses agitation to disperse these types of growth. Ten strains of Candida albicans and five strains of Cryptococcus neoformans were tested against fluconazole, flucytosine, and amphotericin B by using a microdilution adaptation of the proposed reference method of the National Committee for Clinical Laboratory Standards for yeasts (M27-T). The endpoint criterion used before agitation was consistent with the M27-T recommendation, while a criterion of 50% or more reduction of growth compared with the control was used after agitation. The results of this study showed that use of agitation and the modified endpoint criterion both improved intralaboratory and inter-laboratory agreement and increased the frequency of interpretable MICs. The MICs obtained by this method were comparable to those obtained by the broth macrodilution M27-T method. Like M27-T, this method was not able to definitely distinguish amphotericin B-susceptible from -resistant strains, although the MICs for the resistant strains were consistently higher than those for the susceptible ones. The findings imply that agitation should be seriously considered when antifungal agents, particularly fluconazole, are tested in a microdilution format.

Amphotericin B↗

PubMed1996

Stenotrophomonas maltophilia: emergence of multidrug-resistant strains during therapy and in an in vitro pharmacodynamic chamber model.

Emergence of Stenotrophomonas maltophilia as a nosocomial pathogen is becoming increasingly apparent. Pleiotropic resistance characterizes S. maltophilia. Furthermore, a slow growth rate and an increased mutation rate generate discordance between in vitro susceptibility testing and clinical outcome. Despite original susceptibility, drug-resistant strains of S. maltophilia are often recovered from patients receiving beta-lactams, quinolones, or aminoglycosides. Given the disparity among various in vitro susceptibility methods, this study incorporated a unique pharmacodynamic model to more accurately characterize the bacterial time-kill curves and mutation rates of four clinical isolates of S. maltophilia following exposure to simulated multidose regimens of ceftazidime, ciprofloxacin, gentamicin, and ticarcillin-clavulanate. Time-kill data demonstrated regrowth of S. maltophilia with all four agents. With the exception of ticarcillin-clavulanate, viable bacterial counts at the end of 24 h exceeded the starting inoculum. Ciprofloxacin only reduced bacterial counts by less than 1.0 log prior to rapid bacterial regrowth. Resistant mutant strains, identical to their parent strain by pulsed-field gel electrophoresis, were observed following exposure to each class of antibiotic. Mutant strains also had distinct susceptibility patterns. These data are consistent with previous reports which suggest that S. maltophilia, despite susceptibility data that imply that the organism is sensitive, develops multiple forms of resistance quickly and against several classes of antimicrobial agents. Standard in vitro susceptibility methods are not completely reliable for detecting resistant S. maltophilia strains; and therefore, interpretation of these results should be done with caution. In vivo studies are needed to determine optimal therapy against S. maltophilia infections.

Anti-Bacterial Agents↗

PubMed1996

Application of CHROMagar Candida for rapid screening of clinical specimens for Candida albicans, Candida tropicalis, Candida krusei, and Candida (Torulopsis) glabrata.

CHROMagar Candida is a new differential culture medium that allows selective isolation of yeasts and simultaneously identifies colonies of Candida albicans, C. tropicalis, and C. krusei. We evaluated the use of this medium with 316 yeast isolates including 247 isolated directly on CHROMagar from clinical material. Over 95% of stock and clinical isolates of C. albicans, C. tropicalis, and C. krusei were correctly identified on the basis of colony morphology and pigmentation on CHROMagar. Additionally, CHROMagar also allowed the identification of C. (Torulopsis) glabrata at a similar level of accuracy. The overall agreement between two observers in reading the CHROMagar plates was 95%. Growth of Candida sp. isolates on CHROMagar had no adverse effect on antifungal MICs or Vitek identification results. In parallel, cultures of 548 stool and rectal swab specimens set up on CHROMagar and Sabouraud glucose agar (SGA) were positive in 234 instances. CHROMagar was positive and SGA was negative for 11 specimens, and CHROMagar was negative and SGA was positive for 18 specimens. A single yeast species was isolated on both media from 162 specimens, and in 146 (90%) of these specimens the same species was detected on both CHROMagar and SGA. A total of 43 of the 234 positive cultures contained mixtures of yeast species. Twenty (47%) of these mixed cultures were detected only on CHROMagar. CHROMagar is extremely useful in making a rapid presumptive identification of common yeast species. This capability plus the ability to detect mixed cultures of Candida spp. promises to improve and streamline the work flow in the mycology and clinical microbiology laboratory.

Adult↗

PubMed1996

In vitro susceptibility testing and DNA typing of Saccharomyces cerevisiae clinical isolates.

Saccharomyces spp. are widely distributed in nature and may colonize the normal human gastrointestinal tract. Although Saccharomyces cerevisiae isolates have been previously considered nonpathogenic, they appear to be increasingly associated with infections in immunocompromised or otherwise debilitated patients. The antifungal susceptibility and epidemiology of S. cerevisiae are poorly defined at present. A series of 76 isolates (mostly stool surveillance and throat swab isolates) from 70 bone marrow transplant patients hospitalized at two different medical centers were characterized by antifungal susceptibility testing and restriction endonuclease analysis of chromosomal DNA. For DNA typing, digestion with NotI followed by pulsed-field gel electrophoresis was applied. Typing results revealed 62 distinct DNA types among the 76 clinical isolates. Despite this genomic diversity, clusters of identical isolates were identified among different patients hospitalized concurrently in the same unit, indicating possible nosocomial transmission. The MICs of amphotericin B, 5-fluorocytosine, fluconazole, and itraconazole were determined by a broth microdilution method, as recommended by the National Committee for Clinical Laboratory Standards. The MICs at which 90% of the strains were inhibited were as follows: amphotericin B, 1.0 micrograms/ml; 5-fluorocytosine, 0.25 micrograms/ml; fluconazole, 8.0 micrograms/ml; and itraconazole, 1.0 micrograms/ml. The relative resistance of S. cerevisiae to fluconazole and itraconazole may promote the emergence of this species as a pathogen among immunosuppressed patients.

Antifungal Agents↗

PubMed1996

Comparison of three methods for recovery of yeasts from hands of health-care workers.

This study compared three methods for the detection of yeasts on the hands of 30 nurses: (i) direct finger impressions on inhibitory mold agar plates, (ii) bag washes in brain heart infusion broth, and (iii) bag washes in brain heart infusion broth supplemented with gentamicin and vancomycin. The antimicrobial agent-supplemented bag wash method identified the greatest number of yeast carriers and yielded the most yeast isolates, especially non-C. albicans Candida spp.

Candida↗

PubMed1996

Quality control guidelines for National Committee for Clinical Laboratory Standards--recommended broth macrodilution testing of ketoconazole and itraconazole.

Ketoconazole and itraconazole were tested in a multilaboratory study to establish quality control (QC) guidelines for yeast antifungal susceptibility testing. Two isolates that had been previously identified as QC isolates for amphotericin B, fluconazole, and flucytosine (Candida parapsilosis ATCC 22019 and Candida krusei ATCC 6258) were tested in accordance with the National Committee for Clinical Laboratory Standards M27-P guidelines. Each isolate was tested 20 times with the two antifungal agents in the five laboratories by using a lot of RPMI 1640 unique to each laboratory as well as a lot common to all five laboratories, thus generating 200 MICs per drug per organism. Overall, 96 to 99% of the MICs for each drug fell within the desired 3-log2 dilution range (mode +/- 1 log2 dilution). By using these data, 3-log2 dilution QC ranges encompassing 98% of the observed MICs for three of the organism-drug combinations and 94% of the observed MICs for the fourth combination were established. These QC ranges are 0.064 to 0.25 micrograms/ml for both ketoconazole and itraconazole against C. parapsilosis ATCC 22019 and 0.125 to 0.5 micrograms/ml for both ketoconazole and itraconazole against C. krusei ATCC 6258.

Antifungal Agents↗

PubMed1996

Multisite reproducibility of the Etest MIC method for antifungal susceptibility testing of yeast isolates.

A multicenter study was performed to establish the interlaboratory reproducibility of Etest, to provide an additional comparison of Etest MICs with reference broth macrodilution MICs, and to develop some tentative quality control (QC) guidelines for using Etest for antifungal susceptibility testing of Candida spp. Two QC strains, Candida parapsilosis ATCC 22019 and Candida krusei ATCC 6258, were tested by Etest against amphotericin B, fluconazole, flucytosine, itraconazole, and ketoconazole in each of four laboratories. The QC strains were tested 20 times each against the five antifungal agents by using a common lot of RPMI agar. A total of 80 MICs per drug per strain were generated during the study. Overall, 98 to 100% of the MICs fell within a 3 log2 dilution range for the respective yeast-antifungal agent combinations. The level of agreement of Etest MICs with broth macrodilution MICs was 86 to 100% with amphotericin B (C. krusei and C. parapsilosis), itraconazole (C. krusei and C. parapsilosis), flucytosine (C. parapsilosis), and fluconazole (C. parapsilosis). A lower level of agreement was observed with ketoconazole (C. krusei and C. parapsilosis). Although all participants reported identical Etest MICs, the MICs of flucytosine and fluconazole when tested against C. krusei fell well above the upper limits of the reference range for this strain. The tentative QC limits for the two QC strains and five antifungal agents when tested by the Etest methodology are the same as the QC limits when tested by the reference broth macrodilution method for amphotericin B and C. krusei, itraconazole and C. krusei, flucytosine and C. parapsilosis, fluconazole and C. parapsilosis, and itraconazole and C. parapsilosis. The Etest QC ranges are 1 dilution broader (4-dilution range) than the reference macrodilution method QC ranges for ketoconazole and C. krusei, amphotericin B and C. parapsilosis, and ketoconazole and C. parapsilosis.

Amphotericin B↗