Proteolytic activity regarding Sarconesiopsis magellanica (Diptera: Calliphoridae) larval excretions and secretions.
Sarconesiopsis magellanica (Diptera: Calliphoridae) is a medically important necrophagous fly which is used for establishing the post-mortem interval. Diptera maggots release proteolytic enzymes contained in larval excretion and secretion (ES) products playing a key role in digestion. Special interest in proteolytic enzymes has also been aroused regarding understanding their role in wound healing since they degrade necrotic tissue during larval therapy. This study was thus aimed at identifying and characterising S. magellanica proteolytic enzyme ES products for the first time. These products were obtained from first-, second- and third-instar larvae taken from a previously-established colony. ES proteins were separated by SDS-PAGE and their proteolytic activity was characterised by zymograms and inhibition assays involving BAPNA (Nα-benzoyl-dl-Arg-p-nitroanilide) and SAPNA substrates, using synthetic inhibitors. The protein profile ranged from ∼69kDa to ∼23kDa; several of them coincided with the Lucilia sericata ES protein profile. Serine-protease hydrolysis activity (measured by zymogram) was confirmed when a ∼25kDa band disappeared upon ES incubation with PMSF inhibitor at pH 7.8. Analysis of larval ES proteolytic activity on BAPNA and SAPNA substrates (determined by using TLCK and TPCK specific inhibitors) suggested a greater amount of trypsin-like protease. These results support the need for further experiments aimed at validating S. magellanica use in larval therapy.
Medical and Forensic Entomology Research Group, School of Medicine and Health Sciences, Universidad del Rosario, Bogotá, Colombia. Electronic address: tatis_espinita@hotmail.com.
Yudi T
YT
Pinilla
Darwin A
DA
Moreno-Pérez
Manuel A
MA
Patarroyo
Felio J
FJ
Bello
D000818:Animals; D001586:Benzoylarginine Nitroanilide / Q000378:metabolism; D012634:Bodily Secretions / Q000201:enzymology; D002863:Chromogenic Compounds / Q000378:metabolism; D004175:Diptera / Q000201:enzymology*; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007814:Larva / Q000201:enzymology; D008970:Molecular Weight; D010447:Peptide Hydrolases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D059748:Proteolysis
D002863:Chromogenic Compounds; D004791:Enzyme Inhibitors; D001586:Benzoylarginine Nitroanilide; D010447:Peptide Hydrolases
Larvae; Larval therapy; Necrotic wounds; Proteases; Proteolytic activity; S. magellanica