A test for concordance between the multilocus genealogies of genes and microsatellites in the pathogenic fungus Coccidioides immitis.
(17/344)
Uncovering the correct phylogeny of closely related species requires analysis of multiple gene genealogies or, alternatively, genealogies inferred from the multiple alleles found at highly polymorphic loci, such as microsatellites. However, a concern in using microsatellites is that constraints on allele sizes may occur, resulting in homoplasious distributions of alleles, leading to incorrect phylogenies. Seven microsatellites from the pathogenic fungus Coccidioides immitis were sequenced for 20 clinical isolates chosen to represent the known genetic diversity of the pathogen. An organismal phylogeny for C. immitis was inferred from microsatellite-flanking sequence polymorphisms and other restriction fragment length polymorphism-containing loci. Two microsatellite genetic distances were then used to determine phylogenies for C. immitis, and the trees found by these three methods were compared. Congruence between the organismal and microsatellite phylogenies occurred when microsatellite distances were based on simple allele frequency data. However, complex mutation events at some loci made distances based on stepwise mutation models unreliable. Estimates of times of divergence for the two species of C. immitis based on microsatellites were significantly lower than those calculated from flanking sequence, most likely due to constraints on microsatellite allele sizes. Flanking-sequence insertions/deletions significantly decreased the accuracy of genealogical information inferred from microsatellite loci and caused interspecific length homoplasies at one of the seven loci. Our analysis shows that microsatellites are useful phylogenetic markers, although care should be taken to choose loci with appropriate flanking sequences when they are intended for use in evolutionary studies. (+info)
Drug interaction studies of a glucan synthase inhibitor (LY 303366) and a chitin synthase inhibitor (Nikkomycin Z) for inhibition and killing of fungal pathogens.
(18/344)
The interaction between inhibitors of components of the fungal cell wall, glucan and chitin, was studied in vitro with the respective synthase enzyme inhibitors LY 303366 and nikkomycin Z. With Aspergillus fumigatus synergy was noted for inhibition and killing, and synergistic activity was also noted for some isolates of other species presently regarded as difficult to treat. (+info)
Disruption of the gene which encodes a serodiagnostic antigen and chitinase of the human fungal pathogen Coccidioides immitis.
(19/344)
Disruption of genes in medically important fungi has proved to be a powerful tool for evaluation of putative virulence factors and identification of potential protein targets for novel antifungal drugs. Chitinase has been suggested to play a pivotal role in autolysis of the parasitic cell wall of Coccidioides immitis during the asexual reproductive cycle (endosporulation) of this systemic pathogen. Two chitinase genes (CTS1 and CTS2) of C. immitis have been cloned. Preliminary evidence has suggested that expression of CTS1 is markedly increased during endospore formation. The secreted CTS1 chitinase has also been shown to react with patient anti-Coccidioides complement-fixing (CF) antibody and is a valuable aid in the serodiagnosis of coccidioidomycosis. To examine the role of CTS1 in the morphogenesis of parasitic cells, the CTS1 gene was disrupted by a single, locus-specific crossover event. This resulted in homologous integration of a pAN7.1 plasmid construct that contained a 1.1-kb fragment of the chitinase gene into the chromosomal DNA of C. immitis. Results of Southern hybridizations, immunoblot analyses of culture filtrates using both CTS1-specific murine antiserum and serum from a patient with confirmed coccidioidal infection, an immunodiffusion test for CF antigenicity, and substrate gel electrophoresis assays of chitinase activity confirmed that the CTS1 gene was disrupted and nonfunctional. This is the first report of a successful targeted gene disruption in C. immitis. However, loss of CTS1 function had no effect on virulence or endosporulation. Comparative assays of chitinase activity in the parental and Deltacts1 strains suggested that the absence of a functional CTS1 gene can be compensated for by elevated expression of the CTS2 gene. Current investigations are focused on disruption of CTS2 in the Deltacts1 host to further evaluate the significance of chitinase activity in the parasitic cycle of C. immitis. (+info)
Detection of fungi in clinical specimens by phase-contrast microscopy.
(20/344)
During 1973 and 1974, the following fungi were detected in clinical specimens by using phase-contrast microscopy: Blastomyces dermatitidis, 5; Coccidioides immitis, 3; Cryptococcus neoformans, 11; other yeasts 918; dermatophytes, 863; Mucor species, 1; and Aspergillus fumigatus, 16. This technique allows rapid detection and, in many instances, immediate identification of fungi in clinical specimens. (+info)
Comparative efficacies of terbinafine and fluconazole in treatment of experimental coccidioidal meningitis in a rabbit model.
(21/344)
A rabbit model of coccidioidal meningitis was used to compare the therapeutic efficacies of terbinafine (TBF) and fluconazole (FCZ). Hydrocortisone acetate-treated New Zealand White male rabbits were infected intracisternally with either 2.2 x 10(4) or 6.4 x 10(4) Coccidioides immitis arthroconidia. Oral treatment with polyethylene glycol 200 (PEG) twice daily (n = 8), TBF twice daily (n = 9; 200 mg/kg of body weight/day), or FCZ once daily (n = 8; 80 mg/kg/day) began on day 5 and continued for 21 days. Mean survival times were 20, 24, and 32 days for rabbits treated with PEG, TBF, and FCZ, respectively. All of the FCZ-treated animals (100%; P = 0.003), 56% of the TBF-treated animals (P = 0.4), and 25% of the PEG-treated animals survived the length of the study. Both FCZ and TBF were effective at reducing the incidence of paresis. Only FCZ was effective at reducing most neurological and systemic signs. FCZ treatments resulted in lower cerebrospinal fluid (CSF) protein concentrations and leukocyte counts and faster clearing of CSF fungal cultures compared with those for PEG-treated controls, but TBF treatments had no significant effect on these parameters. Neither drug affected CSF glucose levels. Mean serum TBF levels by bioassay were within the range of 3.5 to 6.2 microgram/ml at 1, 2, and 4 h postdosing and 0.35 to 7.0 microgram/ml at 14 h postdosing. No TBF was detected in CSF. Mean FCZ levels (24 to 25.5 h postdosing) by bioassay were 16.4 to 19.2 and 13.5 to 19.2 microgram/ml in serum and CSF, respectively. The reduction in the numbers of CFU in the spinal cord and brain was over 100-fold (P = 0.0005) in FCZ-treated animals and 2-fold (P +info)
Skin test and blastogenic responses to Sporotrichun schenckii.
(22/344)
In vivo skin testing and in vitro lymphocyte blastogenesis were evaluated in a young adult population as methods for detecting cellular immunity to Sporotrichum schenckii. Similar procedures for Candida albicans and Coccidioides immitis were also investigated. 5 of 143 subjects had positive skin tests and 14 had positive blastogenic responses to S. schenckii. These 14 subjects also exhibited unusually high responses to C. albicans in vitro and 11 of the 14 were female. Data demonstrated a correlation coefficient of 0.89 when comparing the blastogenic assays for S. schenckii and C. albicans, suggesting cross antigenicity. Intact cellular immune mechanisms in combination with exposure to C. albicans may protect the host from systemic infection with S. schenckii. Although a limited number of subjects were studied, as a group, females had more vigorous cellular immune responses to C. albicans than males. The rare occurence of sporothrix infection in females as compared to males may be the result of antigenic stimulation from commonly observed vaginal colonization with C. albicans. The present data indirectly support this hypothesis. (+info)
Coccidioidin and merthiolate in previously sensitized animals.
(23/344)
The effect of merthiolate, which is used as a preservative in skin test materials, on skin test reactions was determined in guinea pigs. In four groups of animals, merthiolate in basal medium produced skin tests at 24 and 48 h characterized by erythema and/or induration in an intermediate region, i.e., 5 plus or minus 2.2 mm. One of the four groups of animals was a nonsensitized control group. The other three groups were subcutaneously sensitized with (i) merthiolate and saline, (ii) killed Coccidioides immitis arthrospores, and (iii) merthiolate with killed C. immitis arthrospores. Coccidioidin only and merthiolate in coccidioidin produced positive delayed results in groups 3 and 4, which were sensitized with arthrospores. A synergistic effect of merthiolate and coccidioidin was observed in animals of group 4 sensitized by merthiolate with killed C. immitis arthrospores. This effect was observed at 24 h when positive reactions of coccidioidin with merthiolate were significantly greater than skin tests with plain coccidioidin. (+info)
Risk factors for severe pulmonary and disseminated coccidioidomycosis: Kern County, California, 1995-1996.
(24/344)
Surveillance for coccidioidomycosis (CM) and a case-control study for risk factors among adults were conducted in Kern County, California. From January 1995 through December 1996, 905 cases of CM were identified, for an annual incidence of 86 cases per 100,000 population. A total of 380 adults were enrolled in the case-control study: 77 had severe pulmonary disease, 33 had disseminated disease, and 270 control patients had mild disease. Independent risk factors for severe pulmonary disease included diabetes, recent history of cigarette smoking, income of < $15,000 per year, and older age. Oral antifungal therapy before hospitalization was associated with a reduced risk of CM pneumonia. Risk factors for disseminated disease were black race, income of < $15,000 per year, and pregnancy. Early treatment of CM with oral antifungal agents may prevent severe pulmonary disease in groups considered to be at high risk, such as elderly individuals, persons with diabetes, and smokers. Persons at risk for severe CM may benefit from vaccination once an effective CM vaccine is available. (+info)