Use of proteolytic enzymes as an additional tool for trypanosomatid identification

Abstract

The expression of proteolytic activities in the Trypanosomatidae family was explored as a potential marker to discriminate between the morphologically indistinguishable flagellates isolated from insects and plants. We have comparatively analysed the proteolytic profiles of 19 monoxenous trypanosomatids (Herpetomonas anglusteri,H. samuelpessoai,H. mariadeanei,H. roitmani,H. muscarum ingenoplastis,H. muscarum muscarum,H. megaseliae,H. dendoderi,Herpetomoassp.,Crithidia oncopelti,C. deanei,C. acanthocephali,C. harmosa,C. fasciculata,C. guilhermei,C. luciliae,Blastocrithidia culicis,Leptomonas samueliandLept. seymouri) and 4 heteroxenous flagellates (Phytomonas serpens,P. mcgheei,Trypanosoma cruziandLeishmania amazonensis) byin situdetection of enzyme activities on sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS–PAGE ) containing co-polymerized gelatine as substrate, in association with specific proteinase inhibitors. All 23 trypanosomatids expressed at least 1 acidic proteolytic enzyme. In addition, a characteristic and specific pattern of cell-associated metallo and/or cysteine proteinases was observed, except for the similar profiles detected in 2Herpetomonas(H. anglusteriandH. samuelpessoai) and 3Crithidia(C. fasciculata,C. guilhermeiandC. luciliae) species. However, these flagellates released distinct secretory proteinase profiles into the extracellular medium. These findings strongly suggest that the association of cellular and secretory proteinase pattern could represent a useful marker to help trypanosomatid identification.