title: Chitin is a necessary component to maintain the barrier function of the peritrophic matrix in the insect midgut.
abstract: In most insects, the peritrophic matrix (PM) partitions the midgut into different digestive compartments, and functions as a protective barrier against abrasive particles and microbial infections. In a previous study we demonstrated that certain PM proteins are essential in maintaining the PM's barrier function and establishing a gradient of PM permeability from the anterior to the posterior part of the midgut which facilitates digestion (Agrawal et al., 2014). In this study, we focused on the effects of a reduction in chitin content on PM permeability in larvae of the red flour beetle, Tribolium castaneum. Oral administration of the chitin synthesis inhibitor diflubenzuron (DFB) only partially reduced chitin content of the larval PM even at high concentrations. We observed no nutritional effects, as larval growth was unaffected and neutral lipids were not depleted from the fat body. However, the metamorphic molt was disrupted and the insects died at the pharate pupal stage, presumably due to DFB's effect on cuticle formation. RNAi to knock-down expression of the gene encoding chitin synthase 2 in T. castaneum (TcCHS-2) caused a complete loss of chitin in the PM. Larval growth was significantly reduced, and the fat body was depleted of neutral lipids. In situ PM permeability assays monitoring the distribution of FITC dextrans after DFB exposure or RNAi for TcCHS-2 revealed that PM permeability was increased in both cases. RNAi for TcCHS-2, however, led to a higher permeation of the PM by FITC dextrans than DFB treatment even at high doses. Similar effects were observed when the chitin content was reduced by feeding DFB to adult yellow fever mosquitos, Aedes aegypti. We demonstrate that the presence of chitin is necessary for maintaining the PM's barrier function in insects. It seems that the insecticidal effects of DFB are mediated by the disruption of cuticle synthesis during the metamorphic molt rather than by interfering with larval nutrition. However, as DFB clearly affects PM permeability, it may be suitable to increase the efficiency of pesticides targeting the midgut.
authors.affiliation: Department of Biology, Chemistry, University of Osnabrück, Osnabrück 49069, Germany.
authors.forename: Marco
authors.initials: M
authors.lastname: Kelkenberg
authors.affiliation: Department of Biology, Chemistry, University of Osnabrück, Osnabrück 49069, Germany.
authors.forename: Jothini
authors.initials: J
authors.lastname: Odman-Naresh
authors.affiliation: Department of Biochemistry and Molecular Biophysics, Kansas State University, 141 Chalmers Hall, Manhattan, KS 66506, USA.
authors.forename: Subbaratnam
authors.initials: S
authors.lastname: Muthukrishnan
authors.affiliation: Department of Biology, Chemistry, University of Osnabrück, Osnabrück 49069, Germany. Electronic address: merzendorfer@biologie.uni-osnabrueck.de.
authors.forename: Hans
authors.initials: H
authors.lastname: Merzendorfer
mesh_terms: D000330:Aedes / Q000378:metabolism; D000818:Animals; D002686:Chitin / Q000378:metabolism*; D002687:Chitin Synthase / Q000378:metabolism; D004132:Diflubenzuron / Q000494:pharmacology*; D004064:Digestive System / Q000378:metabolism; D005216:Fat Body / Q000378:metabolism; D007814:Larva / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D018978:Molting / Q000187:drug effects; D034622:RNA Interference; D014231:Tribolium / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*
chemical_list: D002686:Chitin; D002687:Chitin Synthase; C400821:chitin synthase 2; D004132:Diflubenzuron
keywords: Aedes aegypti; Chitin; Diflubenzuron; Peritrophic matrix; Permeability; RNA interference; Red flour beetle; Tribolium castaneum