Cellular organization and peritrophic membrane formation in the cardia (Proventriculus) of Drosophila melanogaster

The peritrophic membrane of Drosophila melanogaster consists of four layers, each associated with a specific region of the folded epithelial lining of the cardia. The epithelium is adapted to produce this multilaminar peritrophic membrane by bringing together several regions of foregut and midgut, each characterized by a distinctively differentiated cell type. The very thin, electron-dense inner layer of the peritrophic membrane originates adjacent to the cuticular surface of the stomadeal valve and so appears to require some contribution by the underlying foregut cells. These foregut cells are characterized by dense concentrations of glycogen, extensive arrays of smooth endoplasmic reticulum, and pleated apical plasma membranes. The second and thickest layer of the peritrophic membrane coalesces from amorphous, periodic acid-Schiff-positive material between the microvilli of midgut cells in the neck of the valve. The third layer of the peritrophic membrane is composed of fine electron-dense granules associated with the tall midgut cells of the outer cardia wall. These columnar cells are characterized by cytoplasm filled with extensive rough endoplasmic reticulum and numerous Golgi bodies and by an apical projection filled with secretory vesicles and covered by microvilli. The fourth, outer layer of the peritrophic membrane originates over the brush border of the cuboidal midgut cells, which connect the cardia with the ventriculus.     

Permeability of the peritrophic membranes of some Diptera to labelled dextrans

The permeability of the continuous, tube-like peritrophic membrane of some Diptera was investigated. Dyes, haemoglobin, cytochrome c, horseradish peroxidase, and especially dextran fractions labelled with the fluorescent dye fluorescein isothiocyanate were used as markers. The peritrophic membrane of larvae of Aedes aegypti was permeable only to dextran with a molecular weight of less than 2,400 Daltons; Evans Blue (960.8 Daltons) permeated only slowly through this peritrophic membrane. Labelled dextran with a mol. wt of 32,000 Daltons did not penetrate the peritrophic membrane of larvae of Anopheles stephensi. Dextrans larger than 32,000 Daltons did not permeate through the peritrophic membrane of Culex pipiens, Odagmia ornata, Anisopus (Phryne) cinctus, Sarcophaga barbata and Calliphora erythrocephala. Labelled dextran with mol. wt of 4,000–6,000 Daltons penetrated only slowly, and dextran of 6,200 Daltons did not penetrate the peritrophic membranes of adults of Sarcophaga barbata. The peritrophic membranes of the blowfly, Calliphora erythrocephala, were permeated slowly by dextran of 6,200 Daltons but not by dextran of 17,200 Daltons. Dextrans are readily soluble in water where the long chains form coils of round or oval shape. Charged protein molecules are more compact with smaller radii when compared to a dextran fraction of the same molecular weight. Therefore the results of investigations on permeability have to be compared in terms of the effective radii and not of the molecular weight.     

Peritrophic matrix in the midgut of tick females of the genus Ixodes (Acari: Ixodidae)

The formation of the peritrophic matrix in the midgut of females of 5 ixodid tick species (Ixodes pacificus, I. pavlovskyi, I. persulcatus, I. ricinus and I. scapularis) was studied by means of light and electron microscopy in different periods of the feeding and after detachment. The formation of the peritrophic matrix started when the first food portions came into the gut lumen, 9-12 hours after the attachment. Renovation of the peritrophic matrix took place during the whole feeding period; every new generation of midgut cells synthesized their own matrix. It was deposited on the apical surface of every midgut cell in the beginning of differentiation, and was functioning during the life of the cell. The peritrophic matrix separates spaces of the cavitary and cytozoic digestions.     

Chitin in the peritrophic membrane of Acarus siro (Acari: Acaridae) as a target for novel acaricides

The peritrophic membrane in Acarus siro L. (Acari: Acaridae) is produced by distinct cells located in the ventriculus. In this study, the chitin inside the peritrophic membrane was detected using wheat germ-lectin conjugated with colloidal gold (10 nm). The chitin fibrils of the peritrophic membrane were a target for chitin effectors, including 1) chitinase, which hydrolyzes chitin fibers inside the peritrophic membrane; 2) calcofluor, which binds to chitin and destroys the peritrophic membrane mesh structure; and 3) diflubenzuron, which inhibits chitin synthesis. In addition, soybean trypsin protease inhibitor (STI) and cocktails of chitinase/calcofluor, diflubenzuron/calcofluor and chitinase/STI were tested. These compounds were supplemented in diets and an increase of population initiated from 50 individuals was observed after 21 d of cultivation. Final A. siro densities on experimental and control diets were compared. The chitin in the peritrophic membrane was determined to be a suitable target for novel acaricidal compounds for suppressing the population growth of A. siro. The most effective compounds were calcofluor and diflubenzuron, whereas the suppressive effects of chitinase and STI were low. The failure of chitinase could be due to its degradation by endogenous proteases. The combination of chitinase and STI suppressed A. siro population growth more effectively than when they were tested in oral admission separately. The combinations of calcofluor/chitinase or calcofluor/difluorbenzuron showed no additive effects on final A. siro density. The presence of chitin in peritrophic membrane provides a target for novel acaricidal compounds, which disrupt peritrophic membrane structure. The suitability of chitin effectors and their practical application in the management of stored product mites is discussed.     

Secretion of the type 2 peritrophic matrix protein, peritrophin-15, from the cardia

The midgut of most insects is lined with a peritrophic matrix, which is thought to facilitate digestion and protect the midgut digestive epithelial cells from abrasive damage and invasion by ingested micro-organisms. The type 2 peritrophic matrix is synthesised by a complex and highly specialised organ called the cardia typically located at the junction of the cuticle-lined foregut and midgut. Although the complex anatomy of this small organ has been described, virtually nothing is known of the molecular processes that lead to the assembly of the type 2 peritrophic matrix in the cardia. As a step towards understanding the synthesis of the peritrophic matrix, the synthesis and secretion of the intrinsic peritrophic matrix protein, peritrophin-15 has been followed in the cardia of Lucilia cuprina larvae using immuno-gold localisations. The protein is synthesised by cardia epithelial cells, which have abundant rough endoplasmic reticulum, Golgi, and vesicles indicative of a general secretory function. Peritrophin-15 is packaged into secretory vesicles probably produced from Golgi and transported to the cytoplasmic face of the apical plasma membrane. The vesicles fuse with the plasma membrane at the base of the microvilli and release peritrophin-15 into the inter-microvilli spaces. The protein then becomes associated with the nascent peritrophic matrix, which lies along the tips of the epithelial cell microvilli. It is proposed that peritrophin-15 binds to the ends of chitin fibrils present in the nascent peritrophic matrix, thereby protecting the fibril from the action of exochitinases.     

Evidence for a sugar receptor (lectin) in the peritrophic membrane of the blowfly larva, Calliphora erythrocephala Mg. (Diptera)

For the first time a sugar receptor (lectin) has been localized by electron microscopy in an invertebrate. The peritrophic membrane of the blowfly larva, Calliphora erythrocephala, is shown here to express lectins with high specificity for mannose. The lectin is restricted to the lumen side of the peritrophic membrane. The surface of the midgut epithelium is devoid of mannose-specific lectins. It is suggested that the midgut epithelium has lost these lectins during the course of evolution in favour of the peritrophic membrane which is secreted by specialized cells only at the beginning of the midgut.Peritrophic membranes and the midgut epithelium lack lectins specific for galactose. The lumen side of the peritrophic membrane of the larvae has mannose and/or glucose residues, and it is densely packed with two species of bacteria, Proteus vulgaris and P. morganii. These also have mannose-specific lectins as well as mannose residues on their pili. The existence of mannose-specific receptors and mannose residues on both, peritrophic membranes and bacteria, leads to the assumption of mutual adherence between the two surfaces.     

Identification and characterisation of the excreted/secreted serine proteases of larvae of the old world screwworm fly, Chrysomya bezziana

Serine proteases are the major proteolytic activity excreted or secreted from Chrysomya bezziana larvae as demonstrated by gelatin gel analyses and the use of specific substrates, benzoyl-Arg-p-nitroanilide and succinyl-Ala-Ala-Pro-Phe-p-nitroanilide. Serine proteases were identified through their inhibition by 4-(2-aminoethyl)-benzene sulphonyl fluoride and classified as trypsin- and chymotrypsin-like on the basis of inhibition by tosyl-L-lysine chloromethyl ketone and tosyl-L-phenylalanine chloromethyl ketone, respectively. Like most insect serine proteases, the C. bezziana enzymes were active over broad pH range from mildly acidic to alkaline. The excreted or secreted serine proteases were purified by affinity chromatography using soybean trypsin inhibitor. A different subset of the serine proteases was isolated by salt elution from washed larval peritrophic matrices. Amino-terminal sequencing identified both trypsin and chymotrypsin-like sequences in the excreted or secreted pool with the latter being the dominant protease, whereas trypsin was the dominant species in the peritrophic matrix eluant. These results suggest that trypsin was possibly preferably adsorbed by the peritrophic matrix and may act as a final proteolytic processing stage as partially digested and ingested polypeptides pass through the peritrophic matrix. Immunoblot analysis on dissected gut tissues indicated that the anterior and posterior midguts were the main source of the serine proteases, although a novel species of 32 kDa was predominantly associated with the peritrophic matrix. Proteases are a target for a partially protective immune response and understanding the complexity of the secreted and digestive proteases is a necessary part of understanding the mechanism of the host's immunological defence against the parasite.     

The origin and functions of the insect peritrophic membrane and peritrophic gel

There is a a fluid (peritrophic gel) or membranous (peritrophic membrane, PM) film surrounding the food bolus in most insects. The PM is composed of chitin and proteins, of which peritrophins are the most important. It is proposed here that, during evolution, midgut cells initially synthesized chitin and peritrophins derived from mucins by acquiring chitin-binding domains, thus permitting the formation of PM. Since PM compartmentalizes the midgut, new physiological roles were added to those of the ancestral mucus (protection against abrasion and microorganism invasion). These new roles are reviewed in the light of data on PM permeability and on enzyme compartmentalization, fluid fluxes, and ultrastructure of the midgut. The importance of the new roles in relation to those of protection is evaluated from data obtained with insects having disrupted PM. Finally, there is growing evidence suggesting that a peritrophic gel occurs when a highly permeable peritrophic structure is necessary or when chitin-binding molecules or chitinase are present in food.     

A novel family of chitin-binding proteins from insect type 2 peritrophic matrix. cDNA sequences, chitin binding activity, and cellular localization

The peritrophic matrix is a prominent feature of the digestive tract of most insects, but its function, formation, and even its composition remain contentious. This matrix is a molecular sieve whose toughness and elasticity are generated by glycoproteins, proteoglycans, and chitin fibrils. We now describe a small, highly conserved protein, peritrophin-15, which is an abundant component of the larval peritrophic matrices of the Old World screwworm fly, Chrysomya bezziana, and sheep blowfly, Lucilia cuprina. Their deduced amino acid sequences code for a 8-kDa secreted protein characterized by a highly conserved and novel register of six cysteines. Two Drosophila homologues have also been identified from unannotated genomic sequences. Recombinant peritrophin-15 binds strongly and specifically to chitin; however, the stoichiometry of binding is low (1:10,000 N-acetyl glucosamine). We propose that peritrophin-15 caps the ends of the chitin polymer. Immunogold studies localized peritrophin-15 to the peritrophic matrix and specific vesicles in cells of the cardia, the small organ of the foregut responsible for peritrophic matrix synthesis. The vesicular contents are disgorged at the base of microvilli underlying the newly formed peritrophic matrix. This is the first time that the process of synthesis and integration of a peritrophic matrix protein into the nascent peritrophic matrix has been observed.     

Permeability of the Peritrophic Envelopes of Herbivorous Insects to Dextran Sulfate: a Test of the Polyanion Exclusion Hypothesis

We tested the hypothesis that the permeability of the peritrophic envelope in herbivorous insects is greatly reduced for polyanions as a result of an extensive network of anionic sites in the proteoglycans of the matrix. 14C-Dextran sulfate (polyanionic, 8000 M(w)) and fluorescein isothiocyanate-labeled (FITC) dextran (monoanionic, 9400 M(w)) were introduced together into the endoperitrophic space of the midguts of Orgyia leucostigma (Lepidoptera) larvae and Melanoplus sanguinipes (Orthoptera) adults. In all cases more of the 14C-dextran sulfate permeated the peritrophic envelope than the FITC-dextran, the opposite of the result predicted by the polyanion exclusion hypothesis. We conclude that polyanion exclusion is not a mechanism that contributes significantly to the permeability properties of the peritrophic envelopes of these two species, or that explains the failure of tannic acid to cross the peritrophic envelopes of lepidopteran larvae. Copyright 1997 Elsevier Science Ltd. All rights reserved     

Plant resistance and its effect on the peritrophic membrane of southwestern corn borer (Lepidoptera: Crambidae) larvae

The southwestern corn borer, Diatraea grandiosella Dyar (Lepidoptera: Crambidae), is a serious pest of corn, Zea mays L., in the southern United States. Corn germplasm lines with conventional genetic leaf-feeding resistance to this pest, the fall armyworm, Spodoptera frugiperda (J.E. Smith), and other lepidopterans have been released to the public by USDA-ARS scientists located in Mississippi. Recent studies suggest the insect resistant lines disrupt the integrity of the peritrophic membrane of the fall armyworm. The objectives of this study were to investigate any morphological differences in the structure of the peritrophic membrane of southwestern corn borer larvae feeding on resistant and susceptible corn hybrids and to quantify the damage. Larvae were reared under field and laboratory conditions on three corn hybrids (two resistant and one susceptible). Scanning electron microscopy was used to examine the peritrophic membrane for abnormalities such as holes or tears and to count the holes or tears in the membrane. Differences in the degree of damage to peritrophic membrane of larvae fed on resistant and susceptible plants were not detected. Up to five distinct layers of the membrane were observed in each larva. Variation in the amounts of damage to the peritrophic membrane observed from larvae feeding on all plant material was high. Plant resistance adversely affects growth and development of southwestern corn borer larvae, and further investigations are needed to explain the role of plant resistance and its relation to peritrophic membrane in southwestern corn borer larvae.     

The peritrophic membrane as a barrier: its penetration by Plasmodium gallinaceum and the effect of a monoclonal antibody to ookinetes

We studied the point at which a monoclonal antibody (mAb C5) to a surface protein (Pgs25) on Plasmodium gallinaceum ookinetes blocked the infection of Aedes aegypti mosquitoes. The antibody did not block the development of zygotes to ookinetes in vitro. Development of ookinetes to oocysts in the mosquito was blocked to the same extent whether zygotes grew to ookinetes in the presence of mAb C5 or the antibody was added after the ookinetes had reached full development. When ookinetes developed in vitro in the presence of mAb C5, antibody remained on the surface of the parasite for the next 50 hr and did not block attachment to the peritrophic membrane. When ookinetes were fed to mosquitoes, two subpopulations of mosquitoes were observed (high numbers of oocysts per midgut and low numbers of oocysts per midgut). mAb C5 reduced the number of oocysts per midgut in the subpopulation that had low numbers of oocysts. The subpopulation that had high numbers of oocysts was unaffected by antibody, indicating that the antibody did not block invasion of the midgut epithelium. When mAb C5 was fed with gametocytes, the parasites invaded the epithelium at the same time (between 30 and 35 hr after the blood meal) as in controls, although at a markedly reduced rate. The ultrastructural observations were consistent with a block of parasites within the peritrophic membrane and not with a block at the epithelium, as parasites were not seen to accumulate within the space between the peritrophic membrane and the epithelium. The mechanism by which mAb C5 to Pgs25 of P. gallinaceum blocks the penetration of the peritrophic membrane remains undefined. We present evidence that the parasite modifies the peritrophic membrane during penetration, an observation first made for Babesia microti during penetration of the peritrophic membrane in Ixodes ticks. Ookinetes in the absence of antibodies appeared to disrupt the layers of the peritrophic membrane, suggesting an enzymatic mechanism for penetration.     

Peritrophic matrix proteins

The peritrophic matrix (or peritrophic membrane) lines the gut of most insects at one or more stages of the life cycle. It has important roles in the facilitation of the digestive processes in the gut and the protection of the insect from invasion by microorganisms and parasites. The traditional view of the peritrophic matrix as a relatively insert sieve, composed largely of proteins and glycosaminoglycans embedded in a chitinous matrix, is under revision as more is learned about the molecular characteristics of the peritrophic matrix proteins. This review summarizes emerging knowledge of the main protein constituents of the peritrophic matrix. The availability of the first sequences of integral peritrophic matrix proteins has coincided with the explosion of information in sequence databases. It is therefore possible to examine common structural themes in this family of proteins as well as in proteins of unknown location and function from a variety of other insects, nematodes and viruses. The review concludes with speculation about the biological functions of the proteins in this matrix.     

Non-absorption of ingested lipophilic and amphiphilic allelochemicals by generalist grasshoppers: the role of extractive ultrafiltration by the peritrophic envelope.

The role of the peritrophic envelope in the non-absorption of three allelochemicals ingested by generalist grasshoppers was examined. This study tested the hypothesis that the association of lipophilic and amphiphilic allelochemicals with lipid aggregates (mixed micelles) reduces their permeability through the peritrophic envelope, a process similar to extractive ultrafiltration. Each of three allelochemicals (digitoxin, ouabain, and xanthotoxin) were solubilized in a lysolecithin suspension and injected separately into the midgut lumens of adult Melanoplus sanguinipes (Orthoptera: Acrididae). The low permeability of digitoxin through the peritrophic envelope was consistent with the extractive ultrafiltration of this compound. By comparison, ouabain and xanthotoxin permeability coefficients were 7- and 12-fold higher, respectively, than those of digitoxin. The results of extractive ultrafiltration assays confirmed that digitoxin is effectively extracted in lysolecithin micelles, but that neither ouabain nor xanthotoxin aggregates efficiently with these micelles.     

棉铃虫围食膜的蛋白质组鉴定

【目的】围食膜(peritrophic membrane, PM)是昆虫抵御随食物摄入的病原微生物入侵的第一道天然屏障。本研究旨在鉴定出农业重大害虫棉铃虫Helicoverpa armigera围食膜的总蛋白成分,为进一步揭示昆虫围食膜的形成机制及研发新颖的害虫控制策略奠定基础。【方法】剥离棉铃虫5龄幼虫PM,用三氟甲磺酸(trifluoromethane sulfonic acid, TFMS)处理,采用液质联用技术(LC-MS/MS)鉴定围食膜蛋白质组,然后对鉴定结果进行生物信息学分析。【结果】本研究共鉴定出棉铃虫幼虫围食膜蛋白质169个,是目前鉴定最多的棉铃虫围食膜蛋白。通过GO分析,可以将这些鉴定的蛋白分为细胞组分、分子功能和生物学过程三大类;KEGG富集结果显示,鉴定蛋白可以富集在12条代谢通路中;蛋白互作分析(protein protein interaction, PPI)结果表明,以ACC和CG3011等蛋白为核心可以形成蛋白互作网络。【结论】本研究鉴定了169个棉铃虫幼虫围食膜蛋白质,并对其进行了GO, KEGG和PPI分析,结果有助于人们全面理解昆虫围食膜的分子结构和功能。     

Peritrophins of adult dipteran ectoparasites and their evaluation as vaccine antigens

Several peritrophins of larvae of Lucilia cuprina (sheep blowfly) have demonstrated potential as vaccine antigens, and some have been characterised and cloned. These proteins are tightly associated with the peritrophic matrix, a chitinous tube or sac lining the lumen of the gut of most insects. The peritrophins require strong denaturants for their removal from peritrophic matrix. We now report the preliminary characterisation of peritrophins of the adult stage of L. cuprina and Haematobia irritans exigua (buffalo fly). Similar SDS-PAGE profiles were obtained for proteins extracted in SDS or urea from isolated adult peritrophic matrices of both species. Radioiodination of urea-extracted peritrophins improved sensitivity, indicating numerous proteins of 15-75 kDa. Direct radioiodination of L. cuprina peritrophic matrix preferentially labelled high molecular weight complexes and proteins of 80-90 kDa. Two-dimensional gel analyses of a urea extract of adult L. cuprina peritrophic matrix revealed that most proteins were moderately acidic. Antibodies produced against SDS-extracted peritrophins, or against sonicated peritrophic matrices of these two flies were crossreactive. The sera also appeared to recognise SDS-extracted components of Triton X-100 treated and washed adult peritrophic matrix of the mosquito, Aedes vigilax (Skause). This profile altered as the peritrophic matrix matured. In concordance with extracts from the adult L. cuprina and H.i. exigua peritrophic matrices, proteins in the 50-75 kDa region were immunodominant. The vaccine potential of the peritrophins of these Diptera were examined following vaccination of cattle and rabbits with adult H.i. exigua or L. cuprina peritrophins. When the adult life stages of H.i. exigua or two mosquitoes, A. vigilax and A. aegypti (Linnaeus), were fed on the sera or blood of vaccinated hosts, there were no detrimental effects to any life cycle stages of these Diptera.     

The intrinsic peritrophic matrix protein peritrophin-95 from larvae of Lucilia cuprina is synthesised in the cardia and regurgitated or excreted as a highly immunogenic protein

The intrinsic peritrophic matrix glycoprotein, peritrophin-95, from the midgut of larvae of Lucilia cuprina can only be solubilized from the matrix using strong denaturants. This suggests that the protein has a structural role in the matrix. Consistent with this is the finding that immuno-gold and immuno-fluorescence localizations of the protein showed a uniform distribution within the peritrophic matrix. RT-PCR demonstrated that expression of peritrophin-95 mRNA was restricted to the larval cardia, a small organ located in the anterior midgut from which the type 2 peritrophic matrix originates. Immuno-blots and ELISAs demonstrated that the sera from sheep infested naturally or artificially with these larvae recognised peritrophin-95. This indicates that peritrophin-95 stimulates the ovine immune system during larval infestation even though the protein is firmly attached to the peritrophic matrix in the larval midgut and seemingly "concealed" from the ovine immune surveillance system. Analyses of larval regurgitated or excreted material by immuno-blots, immuno-affinity purification and amino-terminal sequencing demonstrated the presence of soluble monomeric peritrophin-95. These results indicate that peritrophin-95, a candidate vaccine antigen for use in sheep is not a "concealed" antigen as previously thought. The presence of soluble peritrophin-95 in the regurgitated/excreted material from larvae suggests that this protein may be involved in a maturation phase of peritrophic matrix production, a by-product of which is the excretion or regurgitation of soluble peritrophin-95.     

Chitin is only a minor component of the peritrophic matrix from larvae of Lucilia cuprina

The gut of most insects is lined with a peritrophic matrix that facilitates the digestive process and protects insects from invasion by micro-organisms and parasites. It is widely accepted that the matrix is composed of chitin, proteins and proteoglycans. Here we critically re-examine the chitin content of the typical type 2 peritrophic matrix from the larvae of the fly Lucilia cuprina using a range of techniques. Many of the histochemical and biochemical techniques indicate the presence of chitin, although they are often adversely influenced by the presence of highly glycosylated proteins, a principal component of the matrix. The alkali-stable fraction, which is used as an indicator of the maximum chitin content in a biological sample, is only 7.2% of the weight of the matrix. Larvae fed on the potent chitin synthase inhibitor polyoxin D or the chitin-binding agent Calcofluor White, showed strong concentration-dependent inhibition of larval weight and survival but no discernible effects on the matrix structure. A bacterial endochitinase fed to larvae had no effect on larval growth and no observable effect in vitro on the structure of isolated peritrophic matrix. RT–PCR did not detect a chitin synthase mRNA in cardia, the tissue from which PM originates. It is concluded that chitin is a minor structural component of the type 2 peritrophic matrix of this insect.     

Feeding, nutrient flow, and functional gut morphology in the cricket Gryllus bimaculatus

The flow of nutrients through the digestive tract of Gryllus bimaculatus is regulated by the proventriculus, which effectively triturates the partially digested food coming from the crop and shoves the mushy nutrient mass into the space between the paired caeca. The many folds at the base of the caeca form a sieve, and only fine food particles (4-10 microm) and fluids in the mush are filtered under pressure (produced by proventricular peristalsis) into the caeca. Combined with the release of enzymes in the caeca and the influx of water, the caeca are rapidly inflated on day 1 after the terminal molt. The remaining, mostly undigested food is shoved into a tube formed by the peritrophic membrane, which is first formed at the anterior end of the ventriculus. A mucous membrane (peritrophic gel) covers the caecal epithelium, and seems to merge with the true peritrophic membrane at the beginning of the ventriculus. The Type I peritrophic membrane is dragged posteriorly through the entire ventriculus and ileum by the posterior movement of the food bolus, which is shoved posteriorly at a rate of 6 mm/h by proventricular pressure. The growth rate of the peritrophic membrane is about 3 mm/h. Peristalsis does not occur in the midgut or ileum; the muscles in these regions function solely to counteract the internal pressure produced by the proventriculus. The exo- and endoperitrophic space in newly molted animals is open and fluids can flow in both directions. The endoperitrophic space becomes filled on day 1, and leads to a great reduction of the exoperitrophic space. In the ileal pouch (exoperitrophic space) the peritrophic membrane separates the mass of bacteria from the waste bolus within the endoperitrophic space. Feathery bristles arising from the cuticular covering of the finger-like invaginations of the ileal wall hold most of the bacterial mass in place. The crop weight decreases from day 1 to day 3 as the weight of caeca, ventriculus, and ileum increases. After day 3, food uptake and the weight of the entire gut system decrease in female crickets, partly in response to space restrictions in the abdomen caused by rapid ovarial growth.