INVESTIGATION OF THE IMMUNE RESPONSE PROCESS AFTER THE INFECTION OF GALLERIA MELLONELLA LARVAE WITH SALMONELLA ENTERICA IN RELATION TO THE MECHANISMS OF PROGRAMMED CELL DEATH

Project no. 31910 · Status: Kapanmış (Yayın bekleniyor) · 2023

Summary

<p class="ql-align-justify"><span style="color: rgb(0, 0, 0);">In&nbsp;this&nbsp;thesis,&nbsp;it&nbsp;was&nbsp;aimed&nbsp;to&nbsp;investigate&nbsp;the&nbsp;changes&nbsp;occurring&nbsp;in&nbsp;the&nbsp;insect&nbsp;physiology&nbsp;of&nbsp;Galleria&nbsp;mellonella&nbsp;larvae,&nbsp;known&nbsp;as&nbsp;the&nbsp;greater&nbsp;wax&nbsp;moth&nbsp;and&nbsp;causing&nbsp;economic&nbsp;losses&nbsp;in&nbsp;honeycombs,&nbsp;after&nbsp;being&nbsp;infected&nbsp;with&nbsp;Salmonella&nbsp;enterica,&nbsp;a&nbsp;foodborne&nbsp;pathogen,&nbsp;in&nbsp;connection&nbsp;with&nbsp;apoptosis,&nbsp;pyroptosis,&nbsp;and&nbsp;autophagic&nbsp;cell&nbsp;death,&nbsp;which&nbsp;are&nbsp;among&nbsp;the&nbsp;programmed&nbsp;cell&nbsp;death&nbsp;mechanisms.Within&nbsp;this&nbsp;scope,&nbsp;an&nbsp;infection&nbsp;model&nbsp;was&nbsp;established&nbsp;using&nbsp;the&nbsp;oral&nbsp;gavage&nbsp;method,&nbsp;which&nbsp;mimics&nbsp;the&nbsp;natural&nbsp;transmission&nbsp;route&nbsp;of&nbsp;the&nbsp;pathogen.&nbsp;Hemolymph&nbsp;protein&nbsp;profiles&nbsp;for&nbsp;the&nbsp;humoral&nbsp;responses&nbsp;of&nbsp;the&nbsp;insect&nbsp;immune&nbsp;system&nbsp;were&nbsp;examined&nbsp;by&nbsp;SDS-PAGE,&nbsp;and&nbsp;changes&nbsp;at&nbsp;the&nbsp;cellular&nbsp;level&nbsp;were&nbsp;evaluated&nbsp;by&nbsp;histological&nbsp;analyses.&nbsp;The&nbsp;conventional&nbsp;PCR&nbsp;method&nbsp;was&nbsp;applied&nbsp;to&nbsp;investigate&nbsp;the&nbsp;invasion&nbsp;of&nbsp;the&nbsp;bacteria&nbsp;into&nbsp;epithelial&nbsp;cells.&nbsp;To&nbsp;determine&nbsp;changes&nbsp;at&nbsp;the&nbsp;tissue&nbsp;level,&nbsp;light&nbsp;microscopy&nbsp;examination&nbsp;of&nbsp;the&nbsp;midgut&nbsp;was&nbsp;performed,&nbsp;alongside&nbsp;transmission&nbsp;electron&nbsp;microscopy&nbsp;analysis&nbsp;at&nbsp;the&nbsp;ultrastructural&nbsp;level.&nbsp;For&nbsp;the&nbsp;alterations&nbsp;occurring&nbsp;in&nbsp;cell&nbsp;death&nbsp;processes,&nbsp;specific&nbsp;acid&nbsp;phosphatase&nbsp;enzyme&nbsp;activity&nbsp;and&nbsp;Western&nbsp;blot&nbsp;detection&nbsp;of&nbsp;the&nbsp;Atg8&nbsp;protein&nbsp;were&nbsp;carried&nbsp;out&nbsp;as&nbsp;autophagy&nbsp;markers,&nbsp;DNA&nbsp;fragmentation&nbsp;was&nbsp;determined&nbsp;as&nbsp;an&nbsp;apoptosis&nbsp;marker,&nbsp;and&nbsp;Caspase&nbsp;3&nbsp;enzyme&nbsp;activity&nbsp;was&nbsp;measured&nbsp;colorimetrically.&nbsp;For&nbsp;the&nbsp;pyroptosis&nbsp;process,&nbsp;Caspase&nbsp;1&nbsp;expression&nbsp;levels&nbsp;were&nbsp;detected&nbsp;by&nbsp;qRT-PCR.</span></p><p class="ql-align-justify"><span style="color: rgb(0, 0, 0);">When&nbsp;the&nbsp;results&nbsp;are&nbsp;evaluated&nbsp;together&nbsp;with&nbsp;the&nbsp;literature,&nbsp;they&nbsp;indicate&nbsp;that&nbsp;among&nbsp;the&nbsp;programmed&nbsp;cell&nbsp;death&nbsp;mechanisms&nbsp;during&nbsp;infection&nbsp;processes,&nbsp;apoptosis&nbsp;plays&nbsp;a&nbsp;role&nbsp;in&nbsp;pathogen-related&nbsp;degenerative&nbsp;processes&nbsp;in&nbsp;the&nbsp;gut,&nbsp;whereas&nbsp;autophagy&nbsp;shows&nbsp;an&nbsp;increase&nbsp;as&nbsp;a&nbsp;response&nbsp;to&nbsp;cellular&nbsp;stress&nbsp;depending&nbsp;on&nbsp;the&nbsp;presence&nbsp;of&nbsp;the&nbsp;pathogen,&nbsp;though&nbsp;this&nbsp;increase&nbsp;does&nbsp;not&nbsp;occur&nbsp;at&nbsp;a&nbsp;level&nbsp;to&nbsp;cause&nbsp;death.&nbsp;Considering&nbsp;the&nbsp;findings&nbsp;of&nbsp;caspase&nbsp;I&nbsp;expression&nbsp;analyzed&nbsp;as&nbsp;a&nbsp;pyroptosis&nbsp;marker,&nbsp;strong&nbsp;evidence&nbsp;was&nbsp;obtained&nbsp;indicating&nbsp;that&nbsp;caspase&nbsp;I&nbsp;is&nbsp;not&nbsp;a&nbsp;valid&nbsp;indicator&nbsp;for&nbsp;pyroptosis&nbsp;analysis&nbsp;in&nbsp;insects.&nbsp;Furthermore,&nbsp;it&nbsp;has&nbsp;been&nbsp;demonstrated&nbsp;that&nbsp;even&nbsp;if&nbsp;the&nbsp;pathogen&nbsp;cannot&nbsp;invade&nbsp;the&nbsp;larval&nbsp;midgut&nbsp;epithelium,&nbsp;it&nbsp;can&nbsp;persist&nbsp;and&nbsp;proliferate&nbsp;in&nbsp;the&nbsp;lumen&nbsp;and&nbsp;can&nbsp;activate&nbsp;different&nbsp;processes&nbsp;in&nbsp;the&nbsp;host-pathogen&nbsp;conflict,&nbsp;likely&nbsp;through&nbsp;the&nbsp;virulence&nbsp;factors&nbsp;it&nbsp;secretes.</span></p><p></p>

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