Humoral and cellular immune response to a crude exo-antigen and purified keratinase of Microsporum canis in experimentally infected guinea pigs

Abstract

In order to understand better the host-parasite relationship and to compare with previous observations in Microsporum canis naturally infected cats, the humoral and cellular immune responses to both a crude exo-antigen and a 31.5 kDa purified keratinase were evaluated in 12 M. canis experimentally infected guinea pigs. Humoral and cellular responses were assessed by ELISA from days 0 to 56 postinfection (PI) and by measurement of delayed-type hypersensitivity (DTH) responses on days 14 and 57 PI, respectively. Additionally, immunohistochemical staining was performed and demonstrated that the keratinase was produced in infected guinea pig skin, as previously reported in cats. Despite a marked interindividual variation, all the guinea pigs produced specific IgG to the crude exo-antigen from day 21 PI onwards, but no anti-keratinase IgG was detected. Strongly positive DTH responses to the exo-antigen were observed on both dates, whereas the keratinase elicited no and weak DTH on days 14 and 57 PI, respectively. These results are in agreement with those previously described for naturally infected cats, and indicate that the 31.5 kDa keratinase is not a major antigen in M. canis infection.

Assessment of immunogenicity and protective efficacy of Microsporum canis secreted components coupled to monophosphoryl lipid-A adjuvant in a vaccine study using guinea pigs

“…Ten µg of SC (100 µL) were injected intradermally at two sites on the flanks of animals. Both a negative (100 µL of liquid Sab medium) and a positive control consisting of 10 µg/100µL of a M. canis antigen known to induce DTH in immune guinea pigs (Mignon et al, 1999) were also performed. The skin thickness was measured before and 24 h after injection with a micrometre gauge (Kulche Coppieters, Brussels, Belgium) and the mean relative increases in skin thickness were determined.…”

“…In groups 1 and 2, blood samples (250 µL) were collected from the saphenous vein on days (Mignon et al, 1999) and the positive reference was serum from the same guinea pig collected 14 days after infection. Optical density was defined as the difference between the mean absorbance for each triplicate serum sample tested and the control wells.…”

Differential gene expression in the pathogenic dermatophyte Arthroderma benhamiae in vitro versus during infection

“…Pathogen-free, two-month-old female guinea pigs (cross-bred white albinos, Dunkin Hartley strain, Charles River Laboratories) were infected with A. benhamiae by adapting a standard infection method that has been previously described for the dermatophyte M. canis (Mignon et al, 1999). Briefly, A. benhamiae mycelium scraped from freshly grown 18-day-old Sabouraud plates and suspended in a honey/PBS (1 : 2, v/v) mixture was applied on a 16 cm 2 back skin surface that had been previously clipped and scarified.…”

Secreted Metalloprotease Gene Family of Microsporum canis

“…In fact, most authors have only reported enzymatic activities of dermatophyte proteases on different macromolecular substrates and related their in vitro keratinolytic activity to the ability of dermatophytes to invade keratinized structures in vivo. In order to better understand the role of keratinases in M. canis pathogenicity, the assessment of their in vivo production is required.Despite the fact that the M. canis 31.5-kDa subtilisin-like protease was produced in vivo in naturally infected cats (28) and in experimentally infected guinea pigs (27), this protease does not appear as a major antigen in M. canis infection in these natural and experimental hosts (26,27). However, cellular and humoral immune responses to a crude M. canis exoantigen containing mainly the 31.5-and 43.5-kDa keratinases were demonstrated in both animals, suggesting that the 43.5-kDa metalloprotease could be a valuable candidate for further immunological studies, including vaccination trials.…”

“…Although keratinases are supposed to be involved in dermatophyte pathogenicity, very few authors have assessed their in vivo production (21,27,28). Moreover, only one protease with keratinolytic activity, a recombinant T. rubrum protease, was characterized at the gene level (49).…”