Microbial colonization promotes model cockroach gut tissue growth and development.

BACKGROUND: Digestive tissues are essential for diet processing and nutrient accessibility, especially in omnivores, and these functions occur despite and in collaboration with dynamic microbial communities that reside within and upon these tissues. Prolonged host development and reduced digestive tissue sizes have been observed in germ-free animals, and normal host phenotypes were recovered following the re-introduction of typical gut microbiomes via coprophagy. RESULTS: High-resolution histological analyses of Periplaneta americana cockroach digestive tissues revealed that total prevention of microbial colonization of the gut had severe impacts on the growth and development of gut tissues, especially the posterior midgut and anterior hindgut subcompartments that are expected to be colonized and inhabited by the greatest number of bacteria. Juveniles that were briefly exposed to normal gut microbiota exhibited a partial gut morphological recovery, suggesting that a single inoculation was insufficient. These data highlight gut microbiota as integral to normal growth and development of tissues they are in direct contact with and, more broadly, the organism in which they reside. CONCLUSIONS: We draw on these data, host life history traits (i.e. multigenerational cohousing, molting, and filial coprophagy and exuvia feeding), and previous studies to suggest a host developmental model in which gut tissues reflect a conflict-collaboration dynamic where 1) nutrient-absorptive anterior midgut tissues are in competition with transient and resident bacteria for easily assimilable dietary nutrients and whose growth is least-affected by the presence of gut bacteria and 2) posterior midgut, anterior hindgut, and to a lesser degree, posterior hindgut tissues are significantly impacted by gut bacterial presence because they are occupied by the greatest number of bacteria and the host is relying upon, and thus collaborating with, them to assist with complex polysaccharide catabolism processing and nutrient provisioning (i.e. short-chain fatty acids).

Comparative Transcriptomics Reveals the Expression Differences Between Four Developmental Stages of American Cockroach (Periplaneta americana).

The globally distributed American cockroach (Periplaneta americana) is considered a pest, but it has been widely used in traditional Chinese medicine. In the past, the American cockroach's genome and transcriptomes were sequenced, but the differential expression transcripts between developmental stages were unavailable. We performed the de novo assembly and analysis of American cockroach transcriptomes from four developmental stages. Approximately 200 million high-quality paired-end reads were generated by using Illumina Hiseq 2000 sequencer. The assembly produced 291,250 transcripts with an average length of 714 bp. In addition, 38,052 microsatellites and 11,060,020 transposable elements were identified. Based on sequence homology, 53,262 transcripts were annotated. After calculating the expression levels of all the transcripts, we found that 13 transcripts were highly expressed in all the samples and at least two, p10 and actin-related protein 1, played important roles during development. A total of 7954 differentially expressed transcripts (DETs) were identified. The adult had the largest number of DETs when compared to other samples (4818), while the 3rd and 8th larva had the least number of DETs (1332). We performed gene enrichment analysis with the DETs, and some interesting results were detected in the different groups. For example, chitin is the major component of the insect exoskeleton, and the chitin-related genes in larvae and new molted samples had higher expression levels than in adults. In addition, the enrichment analysis detected many chitin-related pathways. Our study performed the first large-scale comparative transcriptomics between the developmental stages of American cockroach, which could provide useful gene expression data for future studies.

Complex and Diverse Drivers of Parasite Loads in a Cosmopolitan Insect.

The goal of parasite epidemiologists is to understand the factors that determine host infection levels. Potential infection determinants exist at many scales, including spatial and temporal environmental variation, among-host differences, and interactions between symbionts infecting the same host. All of these factors can impact levels of parasitism, but frequently only a subset is considered in any host-parasite system. We examined several potential determinants of pinworm infection in wild Australian cockroaches (Periplaneta australasiae) from multiple biological scales: (1) habitat; (2) season; (3) cockroach body size, developmental stage, and sex; and (4) interactions between 2 pinworm species (Leidynema appendiculata and Thelastoma sp.). Over 1 yr, we collected 239 cockroaches from 2 separate rooms in an Illinois greenhouse. We used generalized linear mixed-effects models (GLMMs) to evaluate simultaneously the influence of these factors on pinworm abundance, and nearly all had significant effects. Overall, the abundance of L. appendiculata was greater than Thelastoma sp., but the relative abundance of the 2 species was reversed in each room (i.e., a taxon × habitat effect). Abundance varied over 4 trapping seasons and increased with cockroach size. Adult cockroaches had more pinworms than nymphs, and there was also a significant taxon × stage effect: adult cockroaches had fewer pinworms than expected for their larger size, and this reduction was greater in Thelastoma sp. than in L. appendiculata. Cockroach sex had no effect on infection. Although females had more worms than males, this difference could be explained by the larger size of females. Finally, after controlling for all other potential determinants of infection, we found a strong negative association between Thelastoma sp. and L. appendiculata; cockroaches tended to be infected with either 1 pinworm species or the other. Our work underscores the importance of measuring potential determinants of infection from as many scales as possible. Such approaches are necessary to unravel the complexities of host-parasite interactions.

Antibacterial/Antifungal Activity of Extracted Chitosan From American Cockroach (Dictyoptera: Blattidae) and German Cockroach (Blattodea: Blattellidae).

We investigated bactericidal and fungicidal properties of chitosan extracted from adults and nymphs from both German cockroach, Blattella germanica (Blattodea: Blattellidae) and American cockroach, Periplaneta americana (Dictyoptera: Blattidae). The cuticle of adults and nymphs extracted from both cockroaches were dried and ground. The powders were demineralized and deproteinized followed by deacetylation using NaOH. Finally, the chitosan yields were examined for antibacterial and antifungal activities. The degree of deacetylation (DD) was different between adults and nymph stages. The antimicrobial effect of American cockroach chitosan (ACC) and German cockroach chitosan (GCC) was tested against four bacteria and four fungi. The extracted chitosans from American cockroach, Periplaneta americana and German Cockroach, Blattella germanica suppressed the growth of Gram-negative/positive bacteria except Micrococcus luteus. The growth of Aspergillus flavus and Aspergillus niger were notability inhibited by the extracted chitosans. The antimicrobial effect of the chitosan depended on the cockroach species, with chitosan of the American cockroach showing more inhibitory effect. This difference may be due to differences in the structure of chitin between the two cockroach species.

Systemic RNAi of V-ATPase subunit B causes molting defect and developmental abnormalities in Periplaneta fuliginosa.

The vacuolar (H+ )-ATPases (V-ATPases) are ATP-driven proton pumps with multiple functions in many organisms. In this study, we performed structural and functional analysis of vha55 gene that encodes V-ATPase subunit B in the smokybrown cockroach Periplaneta fuliginosa (Blattodea). We observed a high homology score of the deduced amino acid sequences between 10 species in seven orders. RNAi of the vha55 gene in P. fuliginosa caused nymphal/nymphal molting defects with incomplete shedding of old cuticles, growth inhibition, as well as bent and wrinkled cuticles of thoraxes and abdominal segments. Since growth inhibition caused by vha55 RNAi did not interfere in the commencement of cockroach molting, molting timing and body growth might be controlled by independent mechanism. Our study suggested V-ATPases might be a good candidate molecule for evolutionary and developmental studies of insect molting.

Two types of sensory proliferation patterns underlie the formation of spatially tuned olfactory receptive fields in the cockroach Periplaneta americana.

In the cockroach Periplaneta americana, to represent pheromone source in the receptive space, axon terminals of sex pheromone-receptive olfactory sensory neurons (pSNs) are topographically organized within the primary center, the macroglomerulus, according to the peripheral locations of sex pheromone-receptive single walled (sw)-B sensilla. In this study, we sought to determine when and where pSNs emerge in the nymphal antenna. We revealed two different pSN proliferation patterns that underlie the formation of topographic organization in the macroglomerulus. In nymphal antennae, which lack sw-B sensilla, pSNs are identified in the shorter sensilla, termed sw-A sensilla. Because new sw-A sensilla emerge on the proximal antenna at every molt, topographic organization in the macroglomerulus must be formed by adding axon terminals of newly emerged pSNs to the lateral region in the macroglomerulus at each molt. At the final molt, a huge number of new sw-B sensilla appeared throughout the whole antenna. Sw-B sensilla in the proximal part of the adult antenna were newly formed during the last instar stage, whereas those located in the distal antenna were transformed from sw-A sensilla. This transformation was accompanied by an increase in the number of pSNs. Axon terminals of newborn pSNs in new sw-B sensilla were recruited to the lateral part of the macroglomerulus, whereas those of newborn pSNs in transformed sw-B sensilla were recruited to the macroglomerulus according to the sensillar location. These mechanisms enable an increase in sensitivity to sex pheromone in adulthood while retaining the topographic map formed during the postembryonic development.

Biochemical efficacy, molecular docking and inhibitory effect of 2, 3-dimethylmaleic anhydride on insect acetylcholinesterase.

Evolution of resistance among insects to action of pesticides has led to the discovery of several insecticides (neonicotinoids and organophosphates) with new targets in insect nervous system. Present study evaluates the mode of inhibition of acetylchlonesterase (AChE), biochemical efficacy, and molecular docking of 2,3-dimethylmaleic anhydride, against Periplaneta americana and Sitophilus oryzae. The knockdown activity of 2,3-dimethylmaleic anhydride was associated with in vivo inhibition of AChE. At KD99 dosage, the 2,3-dimethylmaleic anhydride showed more than 90% inhibition of AChE activity in test insects. A significant impairment in antioxidant system was observed, characterized by alteration in superoxide dismutase and catalase activities along with increase in reduced glutathione levels. Computational docking programs provided insights in to the possible interaction between 2,3-dimethylmaleic anhydride and AChE of P. americana. Our study reveals that 2,3-dimethylmaeic anhydride elicits toxicity in S. oryzae and P. americana primarily by AChE inhibition along with oxidative stress.

Cockroach Oothecal Parasitoid, Evania appendigaster (Hymenoptera: Evaniidae) Exhibits Oviposition Preference Towards Oothecal Age Most Vulnerable to Host Cannibalism.

Many female parasitoid wasps optimize host selection to balance the benefits of high-quality hosts and the costs of predator- or hyperparasitoid-induced mortality risks to maximize their fitness. Cannibalism exists in many insect species and affects survival of parasitoid larvae developing in or on parasitized hosts. However, little is known about how parasitoid wasps resolve the fitness consequence of host cannibalism-induced mortality risk during host selection. We examined the effect of oothecal age on cannibalism in the American cockroach Periplaneta americana (L.) (Dictyoptera: Blattidae) and its effect on host age selection and fitness of its oothecal parasitoid Evania appendigaster (L.) (Hymenoptera: Evaniidae). P. americana differentially cannibalized 1-d-old (30‒60%) versus 10- to 40-d-old oothecae (<9%). However, parasitoid females did not avoid but still preferred to parasitize 1-d-old (45%) over 10- to 40-d-old oothecae (1.6‒20%). The parasitism rate was greater and the handling time was shorter on 1-d-old compared to older oothecae. For parasitoid progeny emerging from different-aged oothecae, regression analysis showed that development time increased and body size (measured as hind tibia length) and longevity decreased with oothecal age. These results demonstrate that reduced parasitoid progeny survival due to host cannibalism did not change the parasitoid's oviposition preference for newly laid oothecae, and that E. appendigaster females traded progeny survival for fitness gains for themselves and their progeny.

Mesoionic insecticides: a novel class of insecticides that modulate nicotinic acetylcholine receptors.

BACKGROUND: As the world population grows towards 9 billion by 2050, it is projected that food production will need to increase by 60%. A critical part of this growth includes the safe and effective use of insecticides to reduce the estimated 20-49% loss of global crop yields owing to pests. The development of new insecticides will help to sustain this protection and overcome insecticide resistance. RESULTS: A novel class of mesoionic compounds has been discovered, with exceptional insecticidal activity on a range of Hemiptera and Lepidoptera. These compounds bind to the orthosteric site of the nicotinic acetylcholine receptor and result in a highly potent inhibitory action at the receptor with minimal agonism. The synthesis, biological activity, optimization and mode of action will be discussed. CONCLUSION: Triflumezopyrim insect control will provide a powerful tool for control of hopper species in rice throughout Asia. Dicloromezotiaz can provide a useful control tool for lepidopteran pests, with an underexploited mode of action among these pests. © 2016 Society of Chemical Industry.

New insights into the infection of the American cockroach Periplaneta americana nymphs with Metarhizium anisopliae s.l. (Ascomycota: Hypocreales).

AIMS: To study the marked resistance of Periplaneta americana to entomopathogenic Metarhizium anisopliae. METHODS AND RESULTS: The low susceptibility of 4th instar nymphs applied topically with conidia seemed to be related to an active removal of conidia by the cockroach and to a disabled or retarded germination and subsequent development of conidia on the cuticle (up to 80% germination in the next 7 days after application). Inhibitions or delays of germination were related to the composition of the epicuticular fatty acids (30·1% w/w oleic, 28·3% w/w linoleic, 24·5% w/w palmitic and 11·7% w/w stearic acid) reported here. Propagules invading the nymphs through the cuticle took at least 3 days to reach the haemocoel, and no propagules were found after day 8 post-treatment. Strain IP 46 infected >50% of nymphs treated with doses ≥2 × 104  hyphal bodies (HB) nymph-1 and reduced the survival of nymphs ≤50%. Most nymphs (>70%) survived after injection of 6 × 103 and 2 × 103  HB nymph-1 . CONCLUSIONS: Findings emphasize a distinct resistance of nymphs of the American cockroach to infections by M. anisopliae. SIGNIFICANCE AND IMPACT OF THE STUDY: Our findings provide support for the development of biological control of this synanthropic cockroach pest.

CHARACTERIZATION OF NICOTINE ACETYLCHOLINE RECEPTOR SUBUNITS IN THE COCKROACH Periplaneta americana MUSHROOM BODIES REVEALS A STRONG EXPRESSION OF ß1 SUBUNIT: INVOLVEMENT IN NICOTINE-INDUCED CURRENTS.

Nicotinic acetylcholine receptors are ligand-gated ion channels expressed in many insect structures, such as mushroom bodies, in which they play a central role. We have recently demonstrated using electrophysiological recordings that different native nicotinic receptors are expressed in cockroach mushroom bodies Kenyon cells. In the present study, we demonstrated that eight genes coding for cockroach nicotinic acetylcholine receptor subunits are expressed in the mushroom bodies. Quantitative real-time polymerase chain reaction (PCR) experiments demonstrated that ß1 subunit was the most expressed in the mushroom bodies. Moreover, antisense oligonucleotides performed against ß1 subunit revealed that inhibition of ß1 expression strongly decreases nicotine-induced currents amplitudes. Moreover, co-application with 0.5 µM α-bungarotoxin completely inhibited nicotine currents whereas 10 µM d-tubocurarine had a partial effect demonstrating that ß1-containing neuronal nicotinic acetylcholine receptor subtypes could be sensitive to the nicotinic acetylcholine receptor antagonist α-bungarotoxin.

The contribution of the melanin pathway to overall body pigmentation during ontogenesis of Periplaneta americana.

The most prominent colors observed in insects are black or brown, whose production is attributed to the melanin pathway. At present, though, the contribution of this pathway to overall body pigmentation throughout ontogenesis is still lacking. To address this question we examined the roles of 2 key melanin genes (TH and DDC), in embryonic and postembryonic development of the American cockroach, Periplaneta americana. Our results show that the melanin pathway does not contribute to the light brown coloration observed in the first nymphs. However, the dark brown coloration in mid nymphs and adults is produced solely from the melanin pathway. In addition, the DDC RNAi results reveal that it is dopamine melanin, not DOPA melanin, acts as the main contributor in this process. Overall, present study provides a new insight into insect pigmentation suggesting that genetic mechanisms of coloration can change during ontogenesis. Future studies of additional basal insect lineages will be required to assess in more details the generality of this phenomenon.

EVIDENCE THAT ECDYSIS IN THE LARVAL COCKROACH, Periplaneta americana L. IS TRIGGERED BY AN INCREASE IN THE CONCENTRATION OF HEMOLYMPH SUGAR.

Ecdysis in insects can be defined as shedding of the cuticle at the end of a larval stadium. This event can only occur after the peak titer of ecdysteroid in the hemolymph has returned to a low level. In the cockroach Periplaneta americana, ecdysis is strongly correlated with a rise in the concentration of trehalose and glucose in the hemolymph, leading to the idea that a causal relationship may exist between both events. The objective in this study was to determine if an increase in hemolymph sugar level would shorten the time to ecdysis in cockroach larvae with experimentally delayed ecdysis. The last larval stadium of P. americana averages 33.5 days but this increases significantly if the larva is injected with a small volume of saline. Injection of 10 µl of saline on day 20 and on four successive days lengthened the stadium by as much as 2 weeks. If, however, trehalose or glucose is incorporated into the saline, approximately 40% of the treated larvae undergo ecdysis at the same time as uninjected larvae. Injection of Peram-AKH, the hypertrehalosemic hormone, also decreases the time for ecdysis to occur. This suggests that peak levels of ecdysteroid trigger the release of Peram-AKH, which then leads to activation of trehalose synthesis. The results support the hypothesis that elevated hemolymph sugar is a contributing factor in the removal of ecdysteroid from the hemolymph.

Desenvolvimento de Periplaneta australasiae Fabricius (Blattodea, Blattidae) em temperatura constante de 30º c e em temperatura ambiente de laboratório / Development of Periplaneta australasiae Fabricius (Blattodea, Blattidae) at a constant temperature of 30° c and at ambient laboratory temperature

Objetivou-se avaliar o desenvolvimento de ovos e ninfas de Periplaneta australasiae (Fabricius) à temperatura de 30 ± 0,2° C, umidade relativa 80 ± 15% e fotofase de 12h e em condições ambientais de laboratório, sem controle de temperatura e umidade relativa; visando à subsídios para medidas de prevenção e controle. As ootecas foram individualizadas em tubos de ensaio até a eclosão. As ninfas foram transferidas para cubas de vidro e alimentadas com ração comercial para coelhos e água ad libitum até a emergência das imagos. Avaliou-se, período de incubação, número de ovos/ ooteca, viabilidade de ovos, número de ninfas/ooteca, período ninfal, viabilidade de ninfas e período ovo/adulto. A diferença do período médio de incubação à temperatura de 30° C (38 dias) e no ambiente (44,5 dias) foi significativa (p < 0,0001); eclodiram, em média 18,1 ninfas/ooteca a 30° C e 21 ninfas/ooteca em condições ambientais (p = 0,006); o período médio de ninfa a 30° C foi de 155,9 dias e no ambiente 279,7 dias (p < 0,0001); a viabilidade de ninfas foi superior a 50%, tanto a 30° C (55,1%) quanto em condições de laboratório (57,2%); no período médio de ovo-adulto de P. australasiae, houve diferença significativa (p< 0,001) entre a temperatura de 30 °C (194,1 dias) e em condições ambientais de laboratório (337,3 dias). Em condições de laboratório, os períodos de incubação, de ninfa e de ovo-adulto de P. australasiae foram aumentados em relação à temperatura de 30° C, não ocorrendo, entretanto, perda nem redução de viabilidade em nenhuma das fases.

With the objective of obtaining standards of measurement for prevention and control, this study compared the development of the eggs and nymphs of Periplaneta australasiae (Fabricius) at a temperature of 30 ± 0.2° C, relative humidity 80 ± 15% and photoperiod of 12 hours versus ambient conditions in the laboratory without controls of temperature and RH. Single ootheca were maintained in test tubes until ecolosion, and nymphs were transferred to glass cubes and fed commercial rabbit ration and water ad libitum until emergence of the imagos. The incubation period, number of eggs/ ootheca, viability of the eggs, number of nymphs/ootheca, nymphal duration, viability of nymphs and duration of egg to adult were all evaluated. The mean difference in the incubation period between the temperature of 30° C (38 days) and ambient conditions (44.5 days) was significant (p < 0.0001); a mean of 18.1 nymphs/ootheca ecloded at 30° C, while 21 nymphs/ootheca ecloded under ambient conditions (p = 0.006); the mean nymphal period at 30° C was 155.9 days while for the ambient it was 279.7 days (p < 0.0001); nymphal viability was greater than 50% for both the 30° C laboratory (55.1%) and the ambient (57.2%); and the mean period from egg to adult of P. australasiae was significantly different (p < 0.001) between the 30° C temperature (194.1 days) and the ambient conditions of the laboratory (337.3 days). Under ambient conditions, the duration of nymphal incubation and egg to adult development of P. australasiae were increased relative to the temperature of 30° C without a reduction in viability in any of the stages.

Postembryonic development of sexually dimorphic glomeruli and related interneurons in the cockroach Periplaneta americana.

In most insects, sex pheromone is processed by an enlarged glomerular complex (macroglomerular complex, MGC) in the male antennal lobe (first-order olfactory center). The MGC of the American cockroach consists of two closely located A- and B-glomeruli which are responsible for processing the major sex pheromone components, periplanone-A and -B, respectively. Using anterograde dye injection, we investigated sexual dimorphism in sensory afferents and interneuron. The A- and B-glomeruli exist in the first larval instar of both sexes. The female MGC homolog grows at a relatively constant rate (1.2-1.8-fold growth per molt) throughout development, whereas the male MGC shows a period of accelerated growth between the fifth and ninth instars, where volume can be more than double in a single molt. These different growth patterns resulted in a 1:30 ratio in glomerular complex volumes of adult females versus males. In the female MGC homolog, afferents originating from the dorsal and ventral antennal surfaces were biased toward anterior and posterior regions, and segregation of these afferents was less clear compared to the adult male. The staining of interneurons projecting to the protocerebrum revealed that projection patterns characteristic of sex pheromone processing appear in the late eighth instar in males, while possibly homologous projections in the female were far fewer in number. These results suggest that the glomerular complexes in pre-eighth larval males, and probably females, are not differentiated for specific detection of sex pheromone. Male-specific projections for sex pheromone detection may be formed by modification of pre-existing neural circuitry.

[Effects of several exorphins and endorphins on the escape reaction of the cockroach Periplaneta americana under elevated temperature conditions].

The ability of several alimentary opioid peptides (exorphin C, rubiscolin-5, cytochrophi-4) and endorphins (met-enkephalin, dynotphin A(1-10), beta-neoendorphin) to change the escape reaction of the cockroaches Periplaneta americana at their placement into a hot chamber was studied. The ED50 values increasing twice the insect stay time in the hot chamber as well as duration and dynamics of the effects were determined. It has been shown that ED5 decreases statistically significantly with increase of the length of the peptide molecule and its affinity of duration of the effects and to an increase of their affinity to delta-receptors - to prolongation of the reaction (more than150 min). In the group of alimentary peptides (exorphins) the most active was a fragment of D-ribulose-1,5-biphosphate carboxylase/oxigenase rubiscolin-5 (ED5 = 386 nM per individual). This might indicate a specific ability of some plant proteins to regulate (decrease) the insect protective behavior.

Different growth patterns of two adjacent glomeruli responsible for sex-pheromone processing during postembryonic development of the cockroach Periplaneta americana.

In many insect species, sex pheromone is processed by specific, enlarged glomeruli in the antennal lobes of males. In the male American cockroach, two closely located glomeruli (A and B) are responsible for processing the major pheromone components (periplanone-A and -B, respectively), and these collectively form the macroglomerular complex. Afferents originating from the dorsal and ventral surfaces of the antenna tend to project to the anterior and posterior regions of the macroglomerular complex via the dorsal and ventral antennal nerves, respectively. This topographic segregation of afferents is seen only in the macroglomerular complex, and not in other glomeruli that process normal environmental odors. Using differential, anterograde dye injection into the two antennal sensory nerves, we show that the macroglomerular complex is not formed by fusion of several glomeruli, as suggested in previous studies, but that the precursors of the A- and B-glomeruli already exist in the first larval instar. The volume of afferents in the macroglomerular complex precursor increases nearly exponentially with molting times, 430-fold from the first instar to the adult. The A- and B-glomeruli both undergo continuous growth during postembryonic development, but peak growth rates occur in different larval stages. The growth rate of the B-glomerulus peaked in the mid-developmental stage then declined, while growth of A-glomerulus was maintained at low level in early- to mid-developmental stages but increased greatly in later stages. These results suggest perception of sex pheromone occurs in early instars, and that PA and PB have distinct roles in different developmental stages.

Proton-pyrophosphatase and polyphosphate in acidocalcisome-like vesicles from oocytes and eggs of Periplaneta americana.

Acidocalcisomes are acidic organelles containing large amounts of polyphosphate (poly P), a number of cations, and a variety of cation pumps in their limiting membrane. The vacuolar proton-pyrophosphatase (V-H(+)-PPase), a unique electrogenic proton-pump that couples pyrophosphate (PPi) hydrolysis to the active transport of protons across membranes, is commonly present in membranes of acidocalcisomes. In the course of insect oogenesis, a large amount of yolk protein is incorporated by the oocytes and stored in organelles called yolk granules (YGs). During embryogenesis, the content of these granules is degraded by acid hydrolases. These enzymes are activated by the acidification of the YG by a mechanism that is mediated by proton-pumps present in their membranes. In this work, we describe an H(+)-PPase activity in membrane fractions of oocytes and eggs of the domestic cockroach Periplaneta americana. The enzyme activity was optimum at pH around 7.0, and was dependent on Mg(2+) and inhibited by NaF, as well as by IDP and Ca(2+). Immunolocalization of the yolk preparation using antibodies against a conserved sequence of V-H(+)-PPases showed labeling of small vesicles, which also showed the presence of high concentrations of phosphorus, calcium and other elements, as revealed by electron probe X-ray microanalysis. In addition, poly P content was detected in ovaries and eggs and localized inside the yolk granules and the small vesicles. Altogether, our results provide evidence that numerous small vesicles of the eggs of P. americana present acidocalcisome-like characteristics. In addition, the possible role of these organelles during embryogenesis of this insect is discussed.

Ancestral Notch-mediated segmentation revealed in the cockroach Periplaneta americana.

Through division into segments, animal bodies can reach higher degrees of complexity and functionality during development and evolution. The segmentation mechanisms of insects and vertebrates have been seen as fundamentally different at the anatomical and molecular levels, and consequently, independently evolved. However, this conclusion was mostly based on observations of derived insects such as Drosophila. We have cloned the Delta, Notch, and hairy genes in the cockroach Periplaneta americana, a basal insect with short germ-band development, and carried out functional assays of Notch activity during its segmentation. Our results show that, in more basal insects, segmentation involves a similar developmental mechanism to that in vertebrates, including induction of segment formation by cyclic segmental stripes of hairy and Delta expression. This result indicates that Notch-mediated segmentation is the ancestral segmentation mechanism of insects, and together with previous results in the literature [Stollewerk A, Schoppmeier M, Damen WGM (2003) Nature 423:863-865], of arthropods as well. The similarity with vertebrate segmentation might suggest that Notch-mediated segmentation is an ancient developmental mechanism inherited from a common ancestor of insects and vertebrates.

Genome size of termites (Insecta, Dictyoptera, Isoptera) and wood roaches (Insecta, Dictyoptera, Cryptocercidae).

The evolution of genome size has been discussed in relation to the evolution of various biological traits. In the present study, the genome sizes of 22 dictyopteran species were estimated by Feulgen image analysis densitometry and 6-diamidino-2-phenylindole (DAPI)-based flow cytometry. The haploid genome sizes (C-values) of termites (Isoptera) ranged from 0.58 to 1.90 pg, and those of Cryptocercus wood roaches (Cryptocercidae) were 1.16 to 1.32 pg. Compared to known values of other cockroaches (Blattaria) and mantids (Mantodea), these values are low. A relatively small genome size appears to be a (syn)apomorphy of Isoptera + Cryptocercus, together with their sociality. In some phylogenetic groups, genome size evolution is thought to be influenced by selective pressure on a particular trait, such as cell size or rate of development. The present results raise the possibility that genome size is influenced by selective pressures on traits associated with the evolution of sociality.