Biomedical subjects

G M Scalarone

Publications and source records attributed to G M Scalarone.

45 records · Page 3Linked to original sources

PubMedDate not supplied

In vitro comparative evaluations of the postantifungal effect: synergistic interaction between flucytosine and fluconazole against Candida albicans.

In vitro comparative evaluations were performed to study the efficacy of combinations of flucytosine and fluconazole in producing a postantifungal effect (PAFE) on Candida albicans. Initial studies were done to determine MIC, FIC (fractional inhibitory concentration) and optimal PAFE parameters. A turbidometric method was used to measure yeast cell growth following exposure to different concentrations of the two drugs for periods of 0.5, 1 or 2 h at temperatures of 30 degrees C and 37 degrees C. The PAFE was determined by the difference in time (h) required for growth of the control and test cultures to reach the 0.5 absorbance level following removal of the drug by dilution. Ten strains of C. albicans were then assayed (30 degrees C; 2 h exposure time) and a synergistic PAFE was evidenced with the two drugs at concentrations well below their individual MICs. PAFEs ranging from 3.8 to 10.5 h, which persisted for 1.2-2.5 h longer than those achieved with either agent separately, were evidenced when flucytosine and fluconazole were combined (flucytosine: fluconazole ratios of 1:16-1:32) at concentrations ranging from 0.024 to 0.098 micrograms ml-1 and 0.78 to 1.56 micrograms ml-1 respectively. The concentrations of each agent required to produce an optimal PAFE varied according to the C. albicans strain being assayed.

Candida albicans↗

PubMedDate not supplied

Comparative studies on the detection of delayed dermal hypersensitivity in experimental animals with lysate and filtrate antigens of Blastomyces dermatitidis.

Yeast cell and mycelial lysate and filtrate antigens prepared in our laboratory from two strains of Blastomyces dermatitidis (canine isolate T-58 and human isolate Le) were evaluated with respect to the detection of delayed dermal hypersensitivity in hairless guinea pigs previously immunized with killed whole yeast cells from two strains of Blastomyces dermatitidis (T-58 and Le) and Histoplasma capsulatum (strain G-217A). The optimal potential reactivity (reactivity minus cross-reactivity) with regard to eliciting a dermal response in animals sensitized with B. dermatitidis was achieved with the yeast phase lysate and filtrate antigens prepared from both T-58 and Le isolates (mean axes of induration values ranging from 14.0 to 15.9 mm). In contrast, the mycelial phase reagents exhibited lower potential reactivity (mean axes of induration values ranging from 5.3 to 10.5 mm).

Animals↗

PubMedDate not supplied

Comparative stability, sensitivity and specificity studies with different lots of Blastomyces dermatitidis yeast and mycelial lysate antigens.

Comparative studies were performed to assess the stability and lot-to-lot variation of Blastomyces dermatitidis yeast and mycelial phase lysate antigens. Four lots were prepared from each growth phase of B. dermatitidis strain T-58 (canine isolate) during a 14-month period. Serum specimens from dogs with blastomycosis, histoplasmosis, coccidioidomycosis, cryptococcosis and aspergillosis were assayed for antibody content using an alkaline phosphatase enzyme-linked immunosorbent assay (ELISA). The four lots of the yeast phase reagents were similar with respect to sensitivity and specificity, and the absorbance readings were approximately four times greater with sera from dogs with blastomycosis than with histoplasmosis or coccidioidomycosis. Even less cross-reactivity was evidenced when the sera from dogs with cryptococcosis and aspergillosis were assayed. In contrast, the four lots of the mycelial lysate reagents were considerably less reactive and more cross-reactive than the yeast phase antigens and, as above, the four reagents retained their activity after prolonged storage. Therefore the results indicated that the lysate antigens exhibited a great deal of stability and lot-to-lot variations in activity were not observed.

Animals↗

PubMedDate not supplied

Sensitivity and specificity of an isoelectric focusing fraction of Blastomyces dermatitidis yeast lysate antigen for the detection of canine blastomycosis.

Blastomyces dermatitidis yeast lysate antigen (T-58, dog isolate) fractions prepared using the Rotofor preparative isoelectric focusing (IEF) cell (Bio-Rad) were compared with B. dermatitidis yeast lysate and filtrate reagents with respect to the detection of antibodies in sera from dogs with blastomycosis, histoplasmosis, coccidioidomycosis, cryptococcosis and aspergillosis. A horseradish peroxidase enzyme immunoassay with Turbo TMB substrate was used in the study. One particular IEF fraction (pH 4.3) was optimal in the assay, and it exhibited greater sensitivity (100%) and specificity (93%) than the lysate or filtrate preparations. The highest degree of cross-reactivity was encountered with the histoplasmosis and coccidioidomycosis specimens and considerably less with the cryptococcosis and aspergillosis sera. Studies are in progress to purify further the optimal IEF fraction.

Animals↗

PubMedDate not supplied

Induction and detection of cell-mediated reactions with different Blastomyces dermatitidis antigenic preparations.

Blastomyces dermatitidis yeast-phase antigens (killed cell, KWY; lysate, Lys; and filtrate, Fil) from canine isolate T-58 were compared with respect to the induction and detection of cellular immune responses in mice. The antigens exhibited good sensitivity and specificity when used to detect a delayed-type hypersensitivity (DTH) response in mice previously immunized with T-58 or Histoplasma capsulatum antigens. Greater reactivity was observed with the KWY and Lys antigens as DTH-inducing agents (immunogens) than with the Fil antigen. The antigens were also compared with regard to the induction and detection of a lymphoproliferative reaction using splenocytes from normal and sensitized mice, and optimal reactivity was demonstrated with the KWY and Fil antigen preparations both as immunogens (absorbance values 0.22-1.60 and 0.20-0.90, respectively) and as in vitro testing antigens (absorbance values 0.60-1.60 and 0.35-1.00 respectively) (P < 0.01). Peritoneal macrophages from mice sensitized with Fil and KWY antigens showed the greatest in vitro replication inhibition (RI) of B. dermatitidis yeast cells (RI values of 53% and 50% respectively) (P < 0.05). When mitogenic or antigenic lymphocytic supernatants were compared with respect to their ability to enhance the phagocytic activity of unelicited macrophages from normal mice to kill yeast cells, the T-cell mitogens (concanavalin A and phytohaemagglutinin) optimally activated the naive macrophages (45% and 44% RI respectively) compared with the B-cell mitogen (LPS) (23% RI) (P < 0.05). Similar results were obtained with the lymphocytic supernatants from mice immunized with KWY cells when activated using KWY or Fil antigens (46% and 40% RI respectively) (P < 0.05).

Animals↗

PubMedDate not supplied

Comparison of enzyme immunoassay and immunodiffusion for the detection of canine blastomycosis.

Two Blastomyces dermatitidis commercial immunodiffusion antigens, Meridian Diagnostics and Nolan-Scott Laboratories, and two B. dermatitidis yeast phase lysate antigens, T-58 and K-Le, prepared in our laboratory were utilized in an enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies in 61 serum specimens from dogs with blastomycosis. All antigens used in the ELISA were diluted to 50 ng of protein ml-1. Immunodiffusion (ID) tests were also performed on the sera using a Nolan-Scott ID antigen. Greater reactivity, based on the ELISA index value, was evidenced with the two yeast phase lysate antigens over the two commercial reagents in the ELISA. In addition the sensitivity of the ELISA with the T-58, K-Le and Meridian antigens was considerably greater than that of the ID test. The Nolan-Scott antigen, however, was able to detect more positive specimens when used in the ID procedure than in the ELISA. Therefore encouraging results were obtained with respect to the continued development and optimization of the ELISA using B. dermatitidis cell lysate antigens for the diagnosis of blastomycosis.

Animals↗

PubMedDate not supplied

Enzyme immunoassay detection of antibodies in canine blastomycosis using Blastomyces dermatitidis lysate antigens.

A Blastomyces dermatitidis yeast phase lysate antigen (T-58) prepared in our laboratory was used in an enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies in 201 serum specimens from dogs with blastomycosis. In addition 36 sera from non-infected dogs from an endemic area for blastomycosis and 36 sera from non-infected dogs residing in a blastomycosis non-endemic area were assayed. Sensitivity values ranged from 96.2% (53 pre-treatment sera) to 88.5% (148 post-treatment sera) with the specimens from dogs with confirmed blastomycosis. Minimal reactivity was experienced with the sera from non-infected dogs. Positive reactions were obtained with 8.3% of the endemic area sera and with 5.6% of the sera from the non-endemic area. Mean ELISA index values ranged from 7.66 with the pretreatment sera to 1.11 and 1.03 with the endemic area sera and non-endemic area specimens respectively.

Animals↗

PubMedDate not supplied

Induction and detection of delayed dermal hypersensitivity in guinea-pig immunized with Blastomyces dermatitidis lysate and filtrate antigens.

Guinea-pigs were immunized using yeast phase antigens (lysate and filtrate preparations) from two strains of Blastomyces dermatitidis (T-58 and Le). Following a sensitization period, the animals were skin tested on days 40 and 216 using T-58 and Le yeast and mycelial phase lysate and filtrate antigen preparations for the detection of delayed dermal hypersensitivity (DTH). Using the Friedman's analysis of variance by rank test, significant differences were found in the efficacy of the immunogens to induce DTH in the animals when skin tested on both occasions (P < 0.05). Optimal reactivity was observed in guinea-pigs immunized with Le yeast lysate (mean axes of induration ranging from 12.0 to 18.8 mm and 7.0 to 18.5 mm on day 40 and 216, respectively) and T-58 yeast filtrate (mean axes of induration ranging from 7.5 to 18.0 mm and 8.0 to 17.0 mm on day 40 and 216, respectively) when the immunogens were administered with adjuvant. When the same data was analysed using the Friedman's test with regard to evaluating the efficacy of the skin test antigens to detect DTH, significant differences were found between them (P < 0.05) with optimal results using the Le mycelial filtrate and yeast lysate antigens (mean axes of induration ranging from 7.0 to 16.5 mm and 7.5 to 14.5 mm, respectively) when skin testing was done on day 40. When skin testing was done on day 216, the T-58 and Le mycelial lysate antigens gave optimal results (mean axes of induration ranging from 9.0 to 18.5 mm and 7.5 to 15.0 mm, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)

Adjuvants, Immunologic↗

PubMedDate not supplied

Detection of antibodies and delayed hypersensitivity with Rotofor preparative IEF fractions of Blastomyces dermatitidis yeast phase lysate antigen.

Blastomyces dermatitidis (dog isolate T-58) yeast phase lysate antigen was concentrated and separated by Rotofor preparative isoelectric focusing cell (Bio-Rad). The pH values of the fractions were determined and equilibrated to pH 7.2 and then analysed by enzyme-linked immunosorbent assay using horseradish peroxidase enzyme system against serum specimens from dogs with blastomycosis, histoplasmosis, aspergillosis, and coccidioidomycosis. The results showed a peak absorbance at pH 3.89-4.31 (fractions 4 and 5) with the blastomycosis serum specimens. This was a single sharp peak while the rest of the fractions were lower. In contrast the sera from dogs with histoplasmosis showed a peak absorbance at pH 5.54-5.97 (fractions 9 and 10), while the other mycoses showed patterns that did not resemble the blastomycosis or histoplasmosis specimens. Serum specimens from dogs with blastomycosis being treated with itraconazole were also assayed (pre-treatment and 1, 2, 3, and 12 months post-treatment sera). The characteristic peak for blastomycosis was observed and a decrease in the peak was seen as the treatment progressed. Fractions 3-12 were also used to detect delayed dermal hypersensitivity in hyperimmunized hairless guinea-pigs. Fraction 5 (pH 4.31) elicited the optimal response in B. dermatitidis-immunized animals, while no cross-reactivity was observed in guinea-pigs sensitized with Histoplasma capsulatum killed cells.

Animals↗