The peritrophic membrane is a semi-permeable chitinous matrix lining the gut of most insects and is thought to have important roles in the maintenance of insect gut structure, facilitation of digestion, and protection from invasion by microrganisms and parasites. Proteins are integral components of this matrix, although the structures and functions of these proteins have not been characterized in any detail. The peritrophic membrane from the larvae of the fly Lucilia cuprina, the primary agent of cutaneous myiasis in sheep, was shown to contain six major integral peritrophic membrane proteins. Two of these proteins, a 44-kDa glycoprotein (peritrophin-44) and a 48-kDa protein (peritrophin-48) together represent >70% of the total mass of the integral peritrophic membrane proteins. Peritrophin-44 was purified and its complete amino acid sequence was determined by cloning and sequencing the DNA complementary to its mRNA. The deduced amino acid sequence codes for a protein of 356 amino acids containing an amino-terminal signal sequence followed by five similar but nonidentical domains, each of approximately 70 amino acids and characterized by a specific register of 6 cysteines. One of these domains was also present in the noncatalytic regions of chitinases from Brugia malayi, Manduca sexta, and Chelonus. Peritrophin-44 has a uniform distribution throughout the larval peritrophic membrane. Reverse transcriptase-polymerase chain reaction detected the expression of peritrophin-44 in all three larval instars but only trace levels in adult L. cuprina. The protein binds specifically to tri-N-acetyl chitotriose and reacetylated chitosan in vitro. It is concluded that the multiple cysteine-rich domains in peritrophin-44 are responsible for binding to chitin, the major constituent of peritrophic membrane. Peritrophin-44 probably has roles in the maintenance of peritrophic membrane structure and in the determination of the porosity of the peritrophic membrane. This report represents the first characterization of an insect peritrophic membrane protein.

Larvae of the blowfly Lucilia cuprina (Wied.) (Diptera: Calliphoridae) were grown in vitro on a serum-free medium in the presence of a number of lectins. Lectins with specificities for/3-(1,4)-N-acetylglucosamine (wheat germ lectin) and c~-D-mannopyranosyl and (c~-D-glucopyranosyl residues (lentil lectin and Con A) caused strong concentration-dependent inhibition of the growth of the larvae and substantial mortality. Wheat germ lectin had the strongest effects, showing 50% inhibition of larval growth at a concentration of 2 #M and 100% mortality at 25 #M. Other lectins with different sugar specificities had much less effect. The mechanism of the larvicidal action(s) of wheat germ lectin, lentil lectin and Con A was investigated, There were at least three effects of these lectins on L. cuprina larvae. First, these lectins bound to and reduced the permeability of the peritrophic membrane of the larvae. Second, they reduced ingestion of diet medium by larvae. Third, the lectins also bound to the apical membranes of larval gut epithelial ceils although there were no obvious signs of damage to these cells. It is concluded that the combination of these effects probably results in the starvation of the larvae. The implications of these results in terms of possible control strategies for L. cuprina are discussed.

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Many insects feed on blood or tissue from mammalian hosts. One potential strategy for the control of these insects is to vaccinate the host with antigens derived from the insect. The larvae of the f ly Lucilia cuprina feed on ovine tissue and tissue f luids causing a cutaneous myiasis associated with considerable host morbidity and mortality. A candidate vaccine antigen, peritrophin 95, was purified from the peritrophic membrane, which lines the gut of these larvae. Serum from sheep vaccinated with peritrophin 95 inhibited growth of first-instar L. cuprina larvae that fed on this serum. Growth inhibition was probably caused by antibody-mediated blockage of the normally semipermeable peritrophic membrane and the subsequent development of an impervious layer of undefined composition on the gut lumen side of the peritrophic membrane that restricted access of nutrients to the larvae. The amino acid sequence of peritrophin 95 was determined by cloning the DNA complementary to its mRNA. The deduced amino acid sequence codes for a secreted protein containing a distinct Cys-rich domain of 317 amino acids followed by a mucin-like domain of 139 amino acids. The Cys-rich domain may be involved in binding chitin. This report describes a novel immunological strategy for the potential control of L. cuprina larvae that may have general application to the control of other insect pests.Many insects feed on blood, tissue, or tissue fluids of mammals. The insect feeding process itself is often detrimental to the host and in addition many of these insects are vectors for the transmission of viral, protozoal, and helminth parasites that cause considerable host morbidity and mortality, particularly in humans and livestock animals. Current strategies for control of insect pests in livestock industries are becoming inadequate because of rapid and widespread development of insecticide resistance and concerns relating to the presence of chemical residues derived from these insecticides in livestock products and the immediate environment. Thus, alternative means of controlling these insects are required.One possible approach is to vaccinate the host against the offending insect life stage by using an insect gut antigen (1, 2). The gut of most insects is lined with a peritrophic membrane (PM or peritrophic matrix) at least at some stage in the life cycle (3). This semipermeable membrane is composed of chitin, proteoglycans, and proteins. The functions of the PM involve facilitating the digestive process and protecting the gut epithelial cells from invasion by microrganisms and parasites (3). The permeability of the PM is such that it either excludes or severely restricts access of ingested host antibodies to the underlying digestive epithelial cells of the gut (3, 4). The PM, however, is bathed in ingested host antibodies and, therefore, PM proteins may be candidate vaccine antigens. Preliminary studies have shown that sheep vaccinated with crude PM protein extracts from the larvae of the fly Lucilia cuprina induce an anti-larval i...