Morphological characterization and sexual dimorphism of the antennal sensilla in Bactericera gobica Loginova (Hemiptera: Psyllidae)-a scanning and transmission electron microscopic study.

Bactericera gobica is the major pest of Goji berry plants and causes severe damage. Psyllids mainly use the antennal sensilla to recognize olfactory cues necessary to find host plants and mates. However, the structure and function of the antenna and the antennal sensilla of B. gobica remains previously unexplored. Here, we identify the external and internal morphology of the antennal sensilla of B. gobica using both scanning electron microscopy (SEM) and transmission electron microscopy (TEM). We found seven types of sensilla on the filiform antennae, including apical setae (LAS, SAS), sensilla basiconica (SB1, SB2), sensilla campaniform (SCA), sensilla chaetica (ChS1, ChS2), cavity sensilla (CvS1, CvS2), antennal rhinaria (AR1, AR2), and sensilla trichodea (ST). Five of these sensilla types-apical setae, sensilla basiconica, sensilla chaetica, cavity sensilla, and antennal rhinaria-may have olfactory functions based on their porous surfaces and internal dendritic outer segments (DOS). We also found several differences between the two sexes of B. gobica in the sensilla array and internal structure. ChS and DOS in the protrusions of AR were more abundant in males than females. Altogether, we comprehensively revealed the fine structure and probable function of B. gobica antennae and identified differences in the distribution and structure between psyllid sexes. Our findings provide important insights for future studies on defining the olfactory function of psyllid antenna using electrophysiological methods.

Fungal Associates of Soft Scale Insects (Coccomorpha: Coccidae).

Ophiocordyceps fungi are commonly known as virulent, specialized entomopathogens; however, recent studies indicate that fungi belonging to the Ophiocordycypitaceae family may also reside in symbiotic interaction with their host insect. In this paper, we demonstrate that Ophiocordyceps fungi may be obligatory symbionts of sap-sucking hemipterans. We investigated the symbiotic systems of eight Polish species of scale insects of Coccidae family: Parthenolecanium corni, Parthenolecanium fletcheri, Parthenolecanium pomeranicum, Psilococcus ruber, Sphaerolecanium prunasti, Eriopeltis festucae, Lecanopsis formicarum and Eulecanium tiliae. Our histological, ultrastructural and molecular analyses showed that all these species host fungal symbionts in the fat body cells. Analyses of ITS2 and Beta-tubulin gene sequences, as well as fluorescence in situ hybridization, confirmed that they should all be classified to the genus Ophiocordyceps. The essential role of the fungal symbionts observed in the biology of the soft scale insects examined was confirmed by their transovarial transmission between generations. In this paper, the consecutive stages of fungal symbiont transmission were analyzed under TEM for the first time.

Functional foregut anatomy of the blue-green sharpshooter illustrated using a 3D model.

Sharpshooter leafhoppers (Hemiptera: Cicadellidae: Cicadellinae) are important vectors of the plant pathogenic bacterium Xylella fastidiosa Wells et al. (Xanthomonadales: Xanthomonadaceae). This pathogen causes economically significant diseases in olive, citrus, and grapes on multiple continents. Bacterial acquisition and inoculation mechanisms are linked to X. fastidiosa biofilm formation and fluid dynamics in the functional foregut of sharpshooters, which together result in egestion (expulsion) of fluids likely carrying bacteria. One key X. fastidiosa vector is the blue-green sharpshooter, Graphocephala atropunctata (Signoret, 1854). Herein, a 3D model of the blue-green sharpshooter functional foregut is derived from a meta-analysis of published microscopy images. The model is used to illustrate preexisting and newly defined anatomical terminology that is relevant for investigating fluid dynamics in the functional foregut of sharpshooters. The vivid 3D illustrations herein and supplementary interactive 3D figures are suitable resources for multidisciplinary researchers who may be unfamiliar with insect anatomy. The 3D model can also be used in future fluid dynamic simulations to better understand acquisition, retention, and inoculation of X. fastidiosa. Improved understanding of these processes could lead to new targets for preventing diseases caused by X. fastidiosa.

Three-dimensional reconstruction of a whole insect reveals its phloem sap-sucking mechanism at nano-resolution.

Using serial block-face scanning electron microscopy, we report on the internal 3D structures of the brown planthopper, Nilaparvata lugens (Hemiptera: Delphacidae) at nanometer resolution for the first time. Within the reconstructed organs and tissues, we found many novel and fascinating internal structures in the planthopper such as naturally occurring three four-way rings connecting adjacent spiracles to facilitate efficient gas exchange, and fungal endosymbionts in a single huge insect cell occupying 22% of the abdomen volume to enable the insect to live on plant sap. To understand the muscle and stylet movement during phloem sap-sucking, the cephalic skeleton and muscles were reconstructed in feeding nymphs. The results revealed an unexpected contraction of the protractors of the stylets and suggested a novel feeding model for the phloem sap-sucking.

Since the 19^th century, scientists have been investigating how the organs of insects are shaped and arranged. However, classic microscopy methods have struggled to image these small, delicate structures. Understanding how the organs of insects are configured could help to identify new methods for controlling pests, such as chemicals that target the mouthparts that some insects use to feed on plants. Most insects that feed on the sap of plants suck out the nutrient via their stylet bundle ­ a thin, straw-like structure surrounded by a sheath called the labium. As well as drying out the plant and damaging its tissues, the stylet bundle also allows the insect to transmit viruses that cause further harm. To investigate these mouthparts in more detail, Wang, Guo et al. used a method called SBF-SEM to determine the three-dimensional structure of one of the most destructive pests of rice crops, the brown planthopper. In this technique, a picture of the planthopper was taken every time a thin slice of its body was removed. This continuous slicing and re-imaging generated thousands of images that were compiled into a three-dimensional model of the brown planthopper's whole body and internal organs. Previously unknown features emerged from the reconstruction, including a huge cell in the planthopper's abdomen which is full of fungi that provide the nutrients absent in plants. Next, Wang, Guo et al. used this technique to see how the muscles in the labium and surrounding the stylet move by imaging planthoppers that were frozen at different stages of the feeding process. This revealed that when brown planthoppers bow their heads to eat, the labium compresses and pushes out the stylet, allowing it to pierce deeper into the plant. This is the first time that the body of such a small insect has been reconstructed three-dimensionally using SBF-SEM. Furthermore, these findings help explain how brown planthoppers and other sap-feeding insects insert their stylet and damage plants, potentially providing a stepping stone towards identifying new strategies to stop these pests from destroying millions of crops.

Morphological and Developmental Traits of the Binucleation of Male Accessory Gland Cells in the Benthic Water Bug, Aphelocheirus vittatus (Hemiptera: Aphelochiridae).

The male accessory glands (MAGs) in insects are pair(s) of internal reproductive organs that produce and secrete the plasma component of seminal fluid. In various insects, MAG size is important for male reproductive success because the fluid provides physiologically active substances and/or nutrients to females to control sperm as well as female reproductive behaviors. Although the MAG epithelial cells in most insect species are standard mononucleate cells, those in some insect taxa are binucleate due to incomplete cytokinesis (e.g., Drosophila [Fallén] [Diptera: Drosophilidae]) or cell fusion (e.g., Cimex [Linnaeus] [Hemiptera: Cimicidae]). In the case of Drosophila, the apicobasal position of the two nuclei relative to the epithelial plane changes from vertical to horizontal after nutrient intake, which allows the volume of the MAG cavity to expand effectively. On the other hand, in the case of Cimex, the positions of the two nuclei do not change apicobasally in response to feeding, but their position relative to the proximodistal axis varies depending on the tubular/spherical organ morphology. Here, we report that the MAG of the benthic water bug Aphelocheirus vittatus (Matsumura) (Hemiptera: Aphelochiridae) shows binucleation in all epithelial cells. Despite the phylogenetically close relationship between Aphelocheirus and Cimex, the MAG cells in Aphelocheirus showed a Drosophila-like apicobasal change in the position of the two nuclei in response to feeding. Furthermore, the cytological processes during binucleation are more similar to those in Drosophila (incomplete cytokinesis) than to those in Cimex (cell fusion). These results indicate that the physiological role and mechanism of binucleation in MAG cells changed during the evolution of Hemiptera.

Sensory armature and stylets of the mouthparts of Stephanitis nashi (Hemiptera: Cimicomorpha: Tingidae), their morphology and function.

Mouthparts are important appendages that are specialized for detection of food sources and feeding. The pear lace bug, Stephanitis nashi Esaki and Takeya, is a major pest of pear in China, sucking the sap and affecting plant growth. Fine structure of the mouthparts including distribution and abundance of receptor sensilla occurring of adult S. nashi was examined using scanning electron microscopy and structural details are described for the first time. The mouthparts of S. nashi are generally similar to those of other Hemiptera and consist of a pyramidal labrum, a tube-like segmented labium, and a stylet fascicle made up of two mandibular and two maxillary stylets. The four segments of the labium differ in length and have five classes of sensilla including 3 types of sensilla basiconica (I, II, III), 2 types of sensilla trichodea (I, II), 1 type of sensillum campaniformium, 1 type of flower-like sensillum and a sensillum placodeum. Sensilla trichodea II are distributed on each segment of the labium. Sensilla basiconica I occur on the base of the second and fourth segment. The labial tripartite apex composes of two sensory fields and a rostral lid. Each sensory field possesses 2 sensilla basiconica II, 9 sensilla basiconica III, 1 flower-like sensillum and 1 sensillum placodeum. The mandibular stylet tips have about 30 pairs of lateral minor teeth, which may help in penetrating leaves. Externally, the end of each maxillary stylet is smooth; internally it has five teeth. There is no obvious difference between males and females in the distribution, number and types of sensilla. The mouthparts morphology of S. nashi, is consistent in many respects to that of many other phytophagous hemipterans but appear to include some features unique to the Tingidae and others that closely resemble those of both phytophagous and predaceous Miridae.

Sensory Receptors Associated with the Labial Tip and Precibarium of Philaenus spumarius L. (Hemiptera: Aphrophoride).

The meadow spittlebug, Philaenus spumarius (Linnaeus) (Hemiptera: Aphrophoridae), is an important vector for the xylem-limited bacterium Xylella fastidiosa (Wells, Raju, Hung, Weisburg, Mandelco-Paul, and Brenner), which is associated with olive quick decline syndrome in southern Italy. The mouthparts of Hemiptera have important roles in host plant selection, feeding behavior and for vectoring pathogens that cause plant diseases. In this study, the functional morphology of the sensory structures located on the labium tip and precibarium of P. spumarius was investigated using scanning and transmission electron microscopy. The labium tip is composed of two symmetrical sensory complexes, each with five different types of sensilla: aporous sensilla trichodea type 1 and 2; uniporous sensilla chaetica type 1 and 2; and multiporous sensilla basiconica. The precibarium of P. spumarius has two kinds of sensory structures: bulbous sensilla and papillae sensilla. In particular, two groups of sensilla are located on the epipharynx: a distal group that consists of ten papillae sensilla and a proximal group composed of six papillae sensilla and two bulbous sensilla, while the hypopharynx has only two papillae sensilla. The involvement of these sensory structures in the context of feeding behavior and pathogen transmission is discussed.

Transmission Characteristics of Wheat Yellow Striate Virus by its Leafhopper Vector Psammotettix alienus.

Wheat yellow striate virus (WYSV), which is found in wheat fields of Northwest China and transmitted by leafhopper vector Psammotettix alienus, is a tentative new species in the genus Nucleorhabdovirus. Although the insect vector and host range of WYSV have been characterized, many aspects of the acquisition and transmission processes by its insect vector have not been elucidated. Here, the transmission parameters of WYSV by P. alienus were determined using wheat cv. Yangmai 12 as the indicator plant under a controlled temperature (23 ± 1°C) and photoperiod (16 h of light). The results showed that the minimum periods for acquisition were 5 min and 10 min for inoculation access. The latent period for successful transmission was most commonly 16 to 20 days (minimum, 10 days; maximum, 22 days). The quantitative reverse-transcriptase PCR results indicated that the WYSV titer increased with time after acquisition, suggesting that WYSV can replicate in P. alienus. Notably, female P. alienus transovarially transmitted the virus to next generations at relatively high efficiency. Electron microscopy of the WYSV-infected leafhopper revealed bacilliform particles aggregated in the cytoplasm of the salivary gland and midgut tissues. Our present studies suggested that acquisition and transmission of WYSV by P. alienus is consistent with a propagative, circulative, and persistent mode of transmission. Details regarding transmission competencies and distribution of WYSV in P. alienus will provide a basis for designing preventive measures.

Three-dimensional architecture of a mechanoreceptor in the brown planthopper, Nilaparvata lugens, revealed by FIB-SEM.

Trichoid sensilla are the most common mechanoreceptors in insects; depending on their distribution, they can act as either exteroceptors or proprioceptors. In this study, the internal structure of the trichoid sensillum from Nilaparvata lugens was studied, using focused ion beam scanning electron microscopy (FIB-SEM). We reconstructed a three-dimensional (3D) model derived from the FIB-SEM data set. The model displayed characteristic mechanosensory sensilla components, including a hair inserted in the socket, a dendrite going through the laminated cuticle, and an electron-dense tubular body at the dendrite terminal. The detailed 3D model showed the relationship between the microtubules within the tubular body and those outside of the tubular body. We also found an autocellular junction in the tormogen cell, indicating that the tormogen cell grows around the dendrite sheath to form a hollow column shape during sensilla morphogenesis.

External morphology and developmental changes of tarsal tips and mouthparts of the invasive spotted lanternfly, Lycorma delicatula (Hemiptera: Fulgoridae).

External structures of insects contribute to the ability of herbivores to select and feed on their host plants. The invasive spotted lanternfly, Lycorma delicatula (Hemiptera: Fulgoridae) is an economically important and polyphagous insect pest in the eastern US. The lanternfly causes substantial damage to many woody plants by sucking phloem sap, reducing photosynthesis, causing weeping wounds, and creating conditions for sooty mold. Lanternfly nymphs switch host plants during their development. However, little is known about relationship between the lanternfly and its plant hosts, and particularly about morphological adaptations of the lanternfly to host plant usage at each developmental stage of the pest. In this study, we focused on assessing changes in morphology of (a) the lanternfly mouthparts (stylets and labium), and (b) the lanternfly tarsal tips (arolia and tarsal claws) at each developmental stage. Our study revealed several developmental patterns among which the presence of the indentations on mandibular stylets in late instars and adults, as well as the exponential growth of the labium and stylet length, and the tarsal claw dispersal during the lanternfly development. Our findings are critical for investigating and predicting the lanternfly host range, and the lanternfly dispersal to new host trees at each developmental stage.

Vibrational behavior of psyllids (Hemiptera: Psylloidea): Functional morphology and mechanisms.

Vibrational behavior of psyllids was first documented more than six decades ago. Over the years, workers have postulated as to what the exact signal producing mechanisms of psyllids might be but the exact mechanism has remained elusive. The aim of this study is to determine the specific signal producing structures and mechanisms of the psyllids. Here we examine six hypotheses of signal producing mechanisms from both previous and current studies that include: wingbeat, wing-wing friction, wing-thorax friction, wing-leg friction, leg-abdomen friction, and axillary sclerite-thorax friction. Through selective removal of possible signal producing structures and measuring wing beat frequency with high speed videos, six hypotheses were tested. Extensive experiments were implemented on the species Macrohomotoma gladiata Kuwayama, while other species belonging to different families, i.e., Trioza sozanica (Boselli), Mesohomotoma camphorae Kuwayama, Cacopsylla oluanpiensis (Yang), and Cacopsylla tobirae (Miyatake) were also examined to determine the potential prevalence of each signal producing mechanism within the Psylloidea. Further, scanning electron microscope (SEM) was used to examine possible rubbing structures. The result of high speed video recordings showed that wingbeat frequency did not match the dominant frequency of vibrational signals, resulting in the rejection of wingbeat hypothesis. As for the selective removal experiments, the axillary sclerite-thorax friction hypothesis is accepted and wing-thorax friction hypothesis is supported partially, while others are rejected. The SEM showed that the secondary axillary sclerite of the forewing bears many protuberances that would be suitable for stridulation. In conclusion, the signal producing mechanism of psyllids may involve two sets of morphological structures. The first is stridulation between the axillary sclerite of the forewing and the mesothorax. The second is stridulation between the axillary cord and anal area of the forewing.

Mouthparts enriched odorant binding protein AfasOBP11 plays a role in the gustatory perception of Adelphocoris fasciaticollis.

Insect odorant binding proteins (OBPs), one of the most important groups of odor carriers, are believed to play essential roles in chemoreception. In the present study, we focused on AfasOBP11 in Adelphocoris fasciaticollis. Expression profiles showed that AfasOBP11 was mainly expressed in the mouthparts of A. fasciaticollis. Additionally, two types of sensilla, sensilla trichodeum and sensilla basiconicum, were found on the mouthparts of bugs. Moreover, anti-AfasOBP11 antiserum strongly labeled the sensilla basiconica. In fluorescence binding assays, recombinant AfasOBP11 displayed much stronger binding abilities to non-volatile secondary metabolite compounds than to volatile odors, suggesting a role of AfasOBP11 in taste sensing. To further investigate the biological functions of AfasOBP11, the feeding behavior of wild-type, dsGFP-injected and dsAfasOBP11-injected bugs was evaluated by performing electrical penetration graph (EPG) tests. After RNA interference of target AfasOBP11, A. fasciaticollis bugs spent a longer time and pierced more frequently on the artificial diet containing 2.0% gossypol, indicating that RNAi treated bugs reduced sensitivity to gossypol. Our findings suggest that AfasOBP11 may play a vital role in chemoreception of A. fasciaticollis, especially in gustatory perception.

The Anatomy and Ultrastructure of the Digestive Tract and Salivary Glands of Hishimonus lamellatus (Hemiptera: Cicadellidae).

In recent years, we found that Hishimonus lamellatus Cai et Kuoh is a potential vector of jujube witches'-broom phytoplasma. However, little is known about the anatomy and histology of this leafhopper. Here, we examined histology and ultrastructure of the digestive system of H. lamellatus, both by dissecting and by semi- and ultrathin sectioning techniques. We found that the H. lamellatus digestive tract consists of an esophagus, a filter chamber, a conical midgut and midgut loop, Malpighian tubules, an ileum, and a rectum. Furthermore, both the basal region of the filter chamber epithelium and the apical surface of the midgut epithelium have developed microvilli. We also identify the perimicrovillar membrane, which ensheaths the microvilli of midgut loop enterocyte, and the flame-like luminal membrane, which covers the microvilli of the conical midgut epithelium. In addition, H. lamellatus has the principal and accessory salivary glands. Our observations also showed that the endoplasmic reticulum, mitochondria, and secretory granules were all highly abundant in the secretory cells of the principal salivary glands, while the accessory glands consist of only one ovate or elbow-like acinus. We also briefly contrast the structure of the gut of H. lamellatus with those of other leafhopper species. These results intend to offer help for the future study on the histological and subcellular levels of phytopathogen-leafhopper relationships, including transmission barriers and the binding sites of pathogens and other microorganisms within their leafhopper vectors.

Molecular characterization, ultrastructure, and transovarial transmission of Tremblaya phenacola in six mealybugs of the Phenacoccinae subfamily (Insecta, Hemiptera, Coccomorpha).

Mealybugs (Hemiptera, Coccomorpha: Pseudococcidae) are plant sap-sucking insects which require close association with nutritional microorganisms for their proper development and reproduction. Here, we present the results of histological, ultrastructural, and molecular analyses of symbiotic systems of six mealybugs belonging to the Phenacoccinae subfamily: Phenacoccus aceris, Rhodania porifera, Coccura comari, Mirococcus clarus, Peliococcus calluneti, and Ceroputo pilosellae. Molecular analyses based on bacterial 16S rRNA genes have revealed that all the investigated species of Phenacoccinae are host to only one type of symbiotic bacteria-a large pleomorphic betaproteobacteria-Tremblaya phenacola. In all the species examined, bacteria are localized in the specialized cells of the host-insect termed bacteriocytes and are transovarially transmitted between generations. The mode of transovarial transmission is similar in all of the species investigated. Infection takes place in the neck region of the ovariole, between the tropharium and vitellarium. The co-phylogeny between mealybugs and bacteria Tremblaya has been also analyzed.

New Anatomical Evidence from the Male Asian Citrus Psyllid (Hemiptera: Liviidae) Invokes Controversy Over the Accepted Function of Some Male Reproductive Organs in Psylloidea.

Males of many Psylloidea are known to possess a characteristic structure at the functional hub of their reproductive apparatus, between afferent and efferent passage of seminal fluid. The structure is a squat, cylindrical endoskeleton consisting of two sections. Classical authors named them as 'sperm pump' and 'ejaculatory duct', based on superficial resemblance to a spring-loaded, thimble-shaped cylinder, encircled by smooth, vertical columns interpreted to be muscles which, when contracted, compress the cylinder and affect seminal fluid discharge. The discovery of numerous spherules of unknown composition and function in and around the columns of the Asian citrus psyllid male genitalia invoked rigorous scrutiny of the classical literature for evidence to support its claims, and determined that the grounds for vetting the structure as a sperm pump were fully teleological. This paper raises several objections to modern acceptance of this classical interpretation, presenting them as problematic, thought-provoking, and sometimes controversial anatomical features. The two sections are herein called 'drum' and 'spout'. As an endoskeleton, the sections are an invagination of the exoskeleton and therefore cannot receive seminal fluid into their hollow. A phallus is identified inside an aedeagal tube, indicating that it is the ejaculatory duct-the tube, drum, and spout are considered its housing. A sheath envelopes the drum and is directly continuous with the spout hypodermis, another problematical feature raising the question of whether it is detached from adherence to the drum cuticles. Also, there are four afferent tubes but only two openings in the drum to receive their seminal fluids.

Functional anatomy of the precibarial valve in Philaenus spumarius (L.).

In phytophagous sap-sucking insects, the precibarial valve plays an important role in sap ingestion. We used light and electron microspcopy to study the morphology and the ultrastructure of the precibarial valve of the meadow spittlebug, Philaenus spumarius (Hemiptera, Aphrophoridae), in order to better understand the operative mechanism of this structure. The precibarial valve revealed to be a complex structure with a bell-like invagination in the middle of the precibarium (on the epipharynx). Unlike the current hypothesis, we propose that the valve opens by dilator muscles and closes through cuticular and fluid tensions, the latter leading to morphological changes to the plane of the valve based on sap flow. Moreover, the presence of a precibarial secretory structure is described for the first time for auchenorrhynchan insects. In light of these observations, functions are hypothesized and discussed for this secretory structure.

Sexual Dimorphism in Wax Secretion Offers Ecological Adaptability During Ericerus pela (Hemiptera: Coccidae) Evolution.

The scale insect, Ericerus pela Chavannes, shows a typical sexual dimorphism. Males and females are different not only in morphology, but also in their ability to secrete wax and ecological adaptability. Here we report the morphological and structural characteristics of wax glands on E. pela females and males. The differences in wax glands and wax secretion between females and males reflect their different needs for living habitats and different ecological strategies. Sciophilous male nymphs are with five types of wax glands, and the wax glands on the dorsum secrete a layer of wax filaments plausibly for protection against direct light irradiation. On the other hand, five types of wax glands were found on the abdomen of females. Heliophilous female nymphs hardly secrete any wax, but the wax glands located along the spiracle on the abdomen may help this insect to breathe. Female adults secrete wax filaments on eggs to protect them from predators and prevent themselves from sticking to each other. In summary, males appear to secreted wax for creating a shaded niche that fits their sciophilous life style, whereas females are likely to adopt an ecological strategy with thickened epidermis for heliophilous acclimatization and overwintering.

A study of the role of vision in the foraging behaviour of the pyrrhocorid bug Antilochus conquebertii (Insecta; Hemiptera; Pyrrhocoridae).

Our study aims to describe (1) external morphology of the compound eye of Antilochus conquebertii, (2) postembryonic changes involving the eye's shape and size and (3) behaviour of the animal with respect to the organization of the compound eye. With each moult of the insect, the structural units of the compound eye increase in size as well as the number, resulting in an overall increase in eye size. The resolution of the adult eye is better than the young one. The adult possesses UV and polarization sensitivity in its eye. Parallel to the changes of the eye the behaviour of the adult animal changes, rendering it increasingly nocturnal and less active in under illuminated conditions. The current study describes the eye and its functional relationship with the behaviour of the animal at the nymphal and adult developmental stage.

Obligate development of Blastocrithidia papi (Trypanosomatidae) in the Malpighian tubules of Pyrrhocoris apterus (Hemiptera) and coordination of host-parasite life cycles.

Blastocrithidia papi is a unique trypanosomatid in that its life cycle is synchronized with that of its host, and includes an obligate stage of development in Malpighian tubules (MTs). This occurs in firebugs, which exited the winter diapause. In the short period, preceding the mating of overwintered insects, the flagellates penetrate MTs of the host, multiply attached to the epithelial surface with their flagella, and start forming cyst-like amastigotes (CLAs) in large agglomerates. By the moment of oviposition, a large number of CLAs are already available in the rectum. They are discharged on the eggs' surface with feces, used for transmission of bugs' symbiotic bacteria, which are compulsorily engulfed by the newly hatched nymphs along with the CLAs. The obligate development of B. papi in MTs is definitely linked to the life cycle synchronization. The absence of peristalsis allow the trypanosomatids to accumulate and form dense CLA-forming subpopulations, whereas the lack of peritrophic structures facilitates the extensive discharge of CLAs directly into the hindgut lumen. The massive release of CLAs associated with oviposition is indispensable for maximization of the infection efficiency at the most favorable time point.