Comparative characterization of proteolytic enzymes fromTrichophyton gallinaeandTrichophyton verrucosum

Grzywnowicz, G.; L̷obarzewski, J.; Wawrzkiewicz, K; Wolski, T.

doi:10.1080/02681218980000431

Cited by 35 publications

(20 citation statements)

References 28 publications

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“…The level of anti-T. verrucosum antibodies was higher (2.30) after challenge with the virulent strain of the organism compared with the control group (1.57). These results go hand in hand with those obtained by Grzywnowicz et al (1989) and Lee et al (1988). The efficacy of the prepared vaccines was tested through challenge three groups of rabbits (vaccinated by culture filtrate, vaccinated by formalin-inactivated vaccine and control +ve group) were challenged at 10 days post second vaccination by virulent strain of T. verrucosun The result revealed that control +ve group showed alopecia with erythema and scales formation in site of infection and by mycological examination were positive for fungal elements (septated hyphae and arthrospores), as shown in Fig.…”

Section: Discussioncontrasting

confidence: 56%

EVALUATION OF THE IMMUNE RESPONSE TO T. VERRUCOSUM VACCINES

Zahran¹,

Abdeen²

2013

American Journal of Immunology

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Dermatophytosis is considered as one of the most important fungal skin disease affecting both humans and animals otherwise its importance as zoonotic disease. Treatment of ringworm in cattle is expensive and time consuming especially on herd level. Vaccination can be a giant step forward to prevent the occurrence of mycozoonosis between the affected animals with ringworm and susceptible man. Fifty skins scraping samples were collected from animals (cattle, sheep and calves) suffering from skin lesions suspected to be ringworm. Then these collected samples were subjected to mycological examination and identification. T. verrucosum is the main causative agent of collected skin scrapings. Two types of vaccines were prepared from this strain (Culture filtrate and formalin inactivated vaccines). Rabbit model was used for challenge testing of the vaccines. The rabbits were separated into four groups as follow: Each group contained 3 rabbits each one was injected subcutaneously with 1ml of culture filtrate with adjuvant twice for seven days intervals. Group No. 2 rabbits each one was injected intramuscularly with 1 mL formalin-inactivated with adjuvant. The 3rd group is control positive and 4th one is control negative. Vaccination programme is done via injection twice with one week interval and the challenge by scarification of the skin with 0.2 mL of 5×10 7 fungal elements/mL. For 3 days. Humeral immune response was assessed by measuring anti-T. verrucosum antibodies using ELISA technique (direct method). By using statistical analysis of ANOVA, the mean optical density of anti-T. verrucosum specific IgG level in rabbits artificially immunized with culture filtrate with adjuvant vaccine significantly increase from (1.97) post first vaccination to reach (2.43) post second vaccination and still high after challenge with the virulent T. verrucosum strain (2.32) when compared with control positive (1.57). The results revealed that both type of vaccines induce good humeral immune response in vaccinated animals. Culture filtrate vaccine used as a treatment in infected farm animals by two doses within 2 weeks and noted that healing occurred.

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Section: Discussioncontrasting

confidence: 56%

EVALUATION OF THE IMMUNE RESPONSE TO T. VERRUCOSUM VACCINES

Zahran¹,

Abdeen²

2013

American Journal of Immunology

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“…Contrarily, collagen was degraded by subtilisin and collagenase. The inactivity upon collagen is a remarkable difference compared to all other known keratin-degrading enzymes, which hydrolyze collagen (3,7,11,14,15,18). Indeed, damage to collagen gives unacceptable physical properties to the finished leather (9,16,23,26).…”

Section: Dehairing Assaymentioning

confidence: 99%

Novel Keratinase from Bacillus subtilis S14 Exhibiting Remarkable Dehairing Capabilities

Macêdo¹,

Beys‐da‐Silva²,

Gava³

et al. 2005

Appl Environ Microbiol

128

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We report the isolation of a keratinolytic-producing Bacillus subtilis strain and the characterization of the exceptional dehairing properties of its subtilisin-like keratinase. This enzyme can be an alternative to sodium sulfide, the major pollutant from tanneries, and may completely replace it. Its unique nonactivity upon collagen enhances its industrial potential.Enzymatic dehairing in tanneries has been envisaged as an alternative to sulfides (4,6,9,22,23). Tanneries are constantly concerned about the obnoxious odor and pollution caused by the extremely toxic sodium sulfide used in the dehairing process step (24). Deaths due to this toxic chemical process have even been reported (2,8). Worldwide, it is estimated that 315 million bovine leathers are produced per year. Considering a waste treatment cost of $0.30 per m 2 of leather produced (A. Klein, personal communication), more than $1 million is spent per day to treat the waste from tanneries around the world. We report here a novel keratinase from Bacillus subtilis that has the potential to replace sodium sulfide in the dehairing process.Microorganism isolation. Bovine hair, skins wastes, and soil samples were suspended and cultivated in a feather-broth medium (composition in grams per liter: delipidated feather meal [the sole carbon and nitrogen source], 10.0; NaCl, 0.5; K 2 HPO 4 , 0.3; and KH 2 PO 4 , 0.4 [pH 7.5]).The best keratinase-producing organism was identified as a B. subtilis strain (named strain S14) after classification based on homology (99%) of its 16S fragment with sequences from the NCBI databank by use of BLASTN 2.2.6 (1) (accession number AY345856).Keratinase production. The microorganism was cultivated in a 14-liter bioreactor in a culture medium composed of whey milk (a dairy byproduct containing 94.0% water, 5% lactose, 0.9% protein, and 0.1% fat), pH 8.5, and bovine hair (40 g/liter). A crude extract (supernatant) was obtained after centrifugation of the culture. Keratinolytic, subtilisin, and collagenase activities were assayed by using azokeratin, as described elsewhere (15). One unit of enzyme activity was defined as the amount of enzyme that increases absorbance by 0.1 per hour in the conditions described above (15).Dehairing assay. A fresh fleshed bovine hide was washed with a commercial detergent solution and cut into 15-by 5-cm pieces. Two hundred grams of skin (usually two pieces) was processed in a drum flask at 4 rpm with crude extract or water (control) in a proportion of 1.0 ml of liquid per g of skin. When necessary, pH was adjusted with lime. At the end of the process, the skin pieces were gently scraped with fingers to remove loose hairs. This procedure was necessary because rubbing in this laboratory-scale process was not as vigorous as in industrial drums. Total skin depilation was observed in the pH range from 7 to 10, with 4.8 U/g of skin. A complete depilation was reached in 9 h at pH 9.0, 24°C, with 4.8 U/g of skin.Samples of bovine skin were kept in contact with the crude extract, and the skin fragments were fi...

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“…After a further 30 min at 4°C, the tubes were centrifuged for 15 min at 2500 g in a refrigerated centrifuge at 4°C and absorbance was measured in the supernatant at 280 nm. One unit of keratinolytic activity was the amount of enzyme that caused a change of absorbance of 0.01 at 280 nm within 1 h at 37°C (23,24). …”

Section: Quantitative Keratinase Assaymentioning

confidence: 99%

“…Three mutant strains produced larger degradation halos than the wild-type strain and were identified as H36, I7 and J5 (Figure 1). The enzymatic assay of Grzywnowicz et al (23) was used to quantify and compare differences in the enzyme activities. The keratinolytic activity of strain J5 was a serine peptidase 70% higher than that of the wild-type strain (Figure 2).…”

Section: Mutagenesismentioning

confidence: 99%