ABSTRACT
Ticks are rich sources of serine protease inhibitors, particularly those that prevent blood clotting and inflammatory responses during blood feeding. The tick Rhipicephalus (Boophlus) annulatus is an important ectoparasite of cattle. The aims of this study were to characterize and purify the serine protease inhibitors present in R. (B.) annulatus larval extract. The inhibitors were characterized by means of one and two-dimensional reverse zymography, and purified using affinity chromatography on a trypsin-Sepharose column. The analysis on one and two-dimensional reverse zymography of the larval extract showed trypsin inhibitory activity at between 13 and 40 kDa. Through non-reducing SDS-PAGE and reverse zymography for proteins purified by trypsin-Sepharose affinity chromatography, some protein bands with molecular weights between 13 and 34 kDa were detected. Western blotting showed that five protein bands at 48, 70, 110, 130 and 250 kDa reacted positively with immune serum, whereas there was no positive reaction in the range of 13-40 kDa. Serine protease inhibitors from R. (B.) annulatus have anti-trypsin activity similar to inhibitors belonging to several other hard tick species, thus suggesting that these proteins may be useful as targets in anti-tick vaccines.
Carrapatos são uma rica fonte de inibidores da serina protease, particularmente aqueles que previnem coagulação e respostas inflamatórias durante a alimentação com sangue. O carrapato Rhipicephalus (B.) annulatus é um ectoparasita importante de bovinos. O objetivo deste estudo foi caracterizar e purificar os inibidores da serina protease presentes no extrato de larva do R. (B.) annulatus. Os inibidores foram caracterizados através de zimografia reversa uni e bidimensional e purificados com cromatografia de afinidade em uma coluna de sepharose-tripsina. A análise do extrato de larva pela zimografia reversa uni e bidimensional mostrou atividade inibitória de tripsina entre 13 e 40 kDa. Através de SDS-PAGE e zimografia reversa para proteínas purificadas pela cromatografia por sepharose-tripsina, algumas bandas de proteínas com pesos moleculares entre 13 e 34 kDa foram detectadas. Western blotting mostrou que cinco bandas de proteínas a 48, 70, 110, 130 e 250 kDa reagiram positivamente com o soro imune, enquanto não houve reação positiva nas bandas 13-40 kDa. Inibidores da serina protease do R. (B.) annulatus têm atividade antitripsina semelhante àquelas dos inibidores de outras espécies de carrapatos duros, sugerindo, assim, que essas proteínas podem ser úteis como alvo de vacinas contra carrapatos.
Subject(s)
Animals , Rhipicephalus/chemistry , Serine Proteinase Inhibitors/isolation & purification , Larva/chemistry , ProteinsABSTRACT
Staphylococcus aureus [SA] may cause infection in all body organs. Many personnel and patients in hospitals may become carriers of this species. The aim of this study was to investigate relative frequency of SA and its resistance to Methicillin. A couple of swabs were taken from nose and nails of 151 personnels working in operation room of Shahid Sadoughi Hospital in Yazd. Samples were inoculated into Mannitol salt agar medium and S. aureus colonies were antibiogramed to determine the rate of resistance to methicillin. Statistical analyses were performed by chi-square and Fishers' exact test. Overall prevalence of SA was 35.09%[19.2% and 15.9% in noses and nails, respectively]. Prevalence of methicillin-resistant SA [MRSA] was 11.92%[5.3% and 6.6% in noses and nails, respectively]. Surgeons had the highest, and workers the lowest prevalence of nasal SA carrier rate. In contrast, the nails of servants had the highest, and surgeons the lowest SA carrier rate. No significant difference was found between carrier rate and age, sex and duration of employment. But a meaningful relation was found between type of job and MRSA in both noses and nails. All of the MRSA isolates were sensitive to vancomycin. Since employees of operation room are continuously working with patients undergone operation, screening both the personnel and patients to determine the rate of carriage, preventing transfusions of fore mentioned bacterium is necessary and highly recommended