Initiation and orientation of the symbiote migration in the human body louse Pediculus humanus L.

Bacterial symbiotes in the human body louse Pediculus humanus migrate from the mycetome to the lateral oviducts during the adult moult. Experimental results are presented suggesting that the symbiote migration is initiated by a humoral factor associated with the adult moult; that the factor is present in both sexes but the male mycetome is unable to respond to it; and that the symbiotes are attracted by the female reproductive tract.     

Intracellular symbiotes of the pea aphid,Acyrthosiphon pisum

Histological and ultrastructural studies on the mycetome of the pea aphid, Acyrthosiphon pisum, disclosed two types of symbiotes. The more common primary symbiotes were oval in shape and were found in large mycetocytes making up the bulk of the mycetome. The secondary symbiotes were smaller, rod-shaped, and were restricted to an apparently syncytial sheath partially enclosing the primary mycetocytes. Extensive rough endoplasmic reticulum and Golgi occurred in the sheath but not in the primary mycetocytes. Lysosomal breakdown occurred in both primary and secondary symbiotes but the two processes differed markedly. In the primary mycetocytes, a small number of symbiotes were broken down individually to form small, compact residual bodies. In the sheath, breakdown of secondary symbiotes was more extensive: large numbers were broken down within cytolysomes.     

Oviduct development inDrosophila

Summary The lateral oviducts ofDrosophila virilis were investigated in normal unoperated adult females, after unilateral and bilateral ovariectomy, and following the transplantation of genital discs. Subsequent to unilateral ovariectomy at larval stages of development, mature adult females exhibited reproductive systems with a free lateral oviduct which appeared somewhat shorter and less distended than a lateral oviduct normally attached to an ovary. Histological examination revealed that such free lateral oviducts have undergone considerable growth and differentiation in the absence of direct ovarian attachment, but exhibited a smaller lumen and more highly folded epithelium. They may be distinguished from attached lateral oviducts by conformational differences and by a possibly lesser size of the epithelial cells. Free lateral oviducts observed among bilaterally ovariectomized and sham-ovariectomized specimens were indistinguishable from those obtained after unilateral ovariectomy. The results are at variance with the previously accepted conclusion that oviduct growth inDrosophila is dependent upon inductive influences emanating from the ovary and directing the outgrowth of the oviducts. Differences in the developmental performance of the oviducts as a function of age at the time of ovariectomy were not evident in the study which included larvae ranging from the second instar to late in the third instar. Transplants of larval female genital discs to other larvae revealed a lesser development of the lateral oviducts than that exhibited by a genital disc developingin situ. A range of oviducal growth which lacked any relation to ovarian influences or to other internal organs of the hosts was obtained. In general, decreased amounts of oviducal muscle were found associated with the transplants.Presented in partial fulfillment of the requirements for the degree of Doctor of Philosophy at the University of Virginia. This investigation was supported in part by: a National Science Foundation Co-operative Fellowship, by the United States Public Health Service Predoctoral Grant 1F1-GM 19,262-1, and by a Summer Research Grant, University of Missouri-St. Louis.     

Spontaneous and neurally evoked contractions of visceral muscles in the oviduct of Locusta migratoria

The oviducts of Locusta migratoria are innervated by a pair of nerves which arise from, the seventh abdominal ganglion. A distinctive network of striated muscle fibres occurs in the oviducts. The lateral oviducts and common oviduct consist of an inner circular layer of muscle and an outer longitudinal layer of muscle. At the junction of the lateral and common oviduct an additional thin longitudinal layer is found adjacent to the basement epithelium. The oviducts contracted spontaneously when isolated from the central nervous system. These myogenic contractions took the form of peristaltic contractions in the lateral oviduct, and intermittent phasic-like contractions of the posterior regions of the lateral oviduct and the common oviduct. These phasic-like contractions were associated with individual complex potentials recorded extracellularly from the muscle fibres. In locusts that had been interrupted in the process of egg laying, there were large-amplitude action potentials, firing in a bursting pattern, in the oviducal nerves. These large action potentials were absent in locusts that had not been egg-laying. These action potentials were associated with both bioelectric potentials and mechanical events in the posterior region of the lateral oviduct and the common oviduct. Electrical stimulation of the oviducal nerve mimicked the effects of spontaneous action potentials, resulting in the appearance of monophasic potentials and contractions. The contractions were graded and dependent upon both frequency and duration of stimulation. It is concluded that the oviducts of Locusta are both myogenic and neurogenic. The role of these contractions in oviposition is discussed.     

Isolation of intracellular symbiotes by immune lysis of flagellate protozoa and characterization of their DNA

A new method dependent on immune lysis is described for the isolation of intracellular symbiotes from two species of flagellate protozoa Blastocrithidia culicis and Crithidia oncopelti. The symbiote- containing flagellates are exposed to complement and antisera prepared in rabbits against symbiote-free organisms. The immune lysis seems to weaken the plasma membranes of the flagellates so that subsequent application of gentle shearing force liberates the intracellular entities in an undamaged condition. The symbiotes are then separated from other cellular components by DNAse digestion and differential centrifugation. The average recovery of symbiotes isolated by this method is 20%. Light and electron microscopy establishes the structural integrity and numerical abundance of isolated symbiotes in the final fractions. Integrity of symbiotes is further indicated by the high activity of a marker enzyme, uroporphyrinogen I synthetase. The DNA's of symbiote-containing and symbiote-free flagellates, and of isolated symbiotes were purified and compared after isopycnic centrifugation. The comparison establishes the presence of DNA's in symbiotes of both species. The guanine-cytosine (G-C) content of symbiote DNA differs from that of host DNA's in C. oncopelti, but resembles that of kinetoplast DNA in B. culicis. The latter observation was further shown by heat denaturation study. Renaturation kinetics indicate that the genome complexity of symbiote DNA in B. culicis is similar to that of bacteria.     

Larval-adult oviduct transformation in an insect (Oncopeltus fasciatus): cytological study

The lateral oviducts of the milkweed bug undergo a drastic larval-adult transformation during the last larval instar. This transformation is hormonally regulated and is connected with considerable morphological and cytological reorganization. Prior to transformation, the ducts are long and thin. The epithelial cells are fusiform with their long axes running cranial-caudal. They are tilted and slightly helically arranged. During transformation, the lateral oviducts shorten dramatically, the cells become columnar, and their long axes shift into the plane of cross sections. Whereas the cells appear cytologically uniform in the larval state, after transformation, the lateral oviducts are divided into a cranial secretory part, and a caudal part which exhibits a cuticular lining. Secretory processes and deposition of the cuticle start in the larva, immediately after transformation. Cuticle synthesis is first observed next to the common oviduct and progresses craniad. The cuticle-lined luminal surface reveals circular ridges and an intricate pattern of cteniform spines which are directed caudally. The novel mode of cell rearrangement during oviducal transformation and the functional significance of the two parts of the adult lateral oviducts are discussed.     

Trichomycete symbiotes of Crozetia seguyi, a primitive black fly

Crozetia is a genus of black flies endemic to the Crozet Islands in the Indian Ocean. No internal symbiotes were previously known from Crozetia species. We report two species of trichomycete symbiotes Stachylina litoralis and Smittium culicisoides from Crozetia seguyi. Larvae of C. seguyi were examined from three sites. The infection rates for St. litoralis was 10.0-33.3% (n=47) of the larvae and Sm. culicisoides was 46.1-85.7% (n=47). No other symbiotes were discovered.     

Ultrastructure and resistance to water loss in eggs of Acanthoscelides obtectus Say (Coleoptera: Bruchidae)

The mature oöcyte of Acanthoscelides obtectus is surrounded by three envelopes: an external layer, a chorion and a vitelline membrane. The external layer is secreted by the walls of the lateral oviducts. The chorion and vitelline membrane are secreted by the follicular cells. The vitelline membrane becomes very compact during the hour following fertilization and laying. The chorion is composed of three layers, one of which has a paracrystalline ultrastructure.Mature, unfertilized, chorion-containing oöcytes, whose vitelline membranes are loose, dehydrate rapidly in a dry atmosphere after laying or after removal from the lateral oviducts. Fertilized eggs are quite resistant to desiccation: after 12 days at 25°C and 5% relative humidity, viable larvae are obtained.The compact vitelline membrane is the most effective protection against dehydration. The chorion and the external layer are much less effective in preventing water loss from the egg.The retention of eggs in the lateral oviducts does not seem to lead to any modification of the structure of their envelopes.     

Phase contrast and electron microscopy of the mycetocytes and symbiotes of the pea aphid, Acyrthosiphon pisum

Phase contrast, scanning electron, and transmission electron microscopy of the symbiotes of Acyrthosiphon pisum was undertaken. Some staining properties of the symbiotes were also studied.The symbiotes of the pea aphid were found to be coccoid bodies 2 to 5 μ in diameter, gram negative, stained slightly by Fuelgen's, and stained blue by Machiavello's. The symbiotes appear to be surrounded by three membranes. Ribosomes may occur within the cytoplasm of the symbiotes. The cytoplasm of the mycetocytes contains large numbers of mitochondria, endoplasmic reticulum, ribosomes, and a large nucleus, and nucleolus.A discussion of the classification of the symbiotes is also presented.     

Ultrastructure of Symbiotic Bacteria in Normal and Antibiotic-Treated Blastocrithidia culicis and Crithidia oncopelti*

SYNOPSIS. An electron microscope study of diplosomes in Blastocrithidia culicis and bipolar bodies in Crithidia oncopelti has shown that both entities appear to be intracellular symbiotes and have a similar fine structure. They are enclosed by 2 unit membranes which are separated by a large space of very low density. The outer membrane is derived probably from the host cell. The matrix of the symbiotes is composed of dense ribosome-like particles and of areas of low density containing fine fibrillae. The particles are of the same size as ribosomes in bacteria and the fibrils have the characteristics of bacterial DNA. Thus, the lucid areas with fibrillae correspond to the nucleoids in bacteria. These observations suggest that the symbiotes are bacteria. The effect of chloramphenicol (CAP) and penicillin G (PCL) on these symbiotic bacteria was studied by culturing the host flagellates in media containing the antibiotics. The effect was analyzed at different intervals after the treatment by electron microscopy. After single treatment in the blood broth containing 0.08% (w/v) CAP, symbiotes appeared to have enlarged nucleoids, became deformed and eventually degenerated. In Grace's medium (supplemented with 10% fetal bovine serum) containing 0.6 or 2.4% (w/v) PCL, symbiotes of C. oncopelti remained unaltered, whereas some symbiotes of B. culicis became pleomorphic. Symbiotes of both species persisted after repeated transfers in PCL media and reverted to normal forms when transferred to PCL-free media. Sensitivity of symbiotes to CAP provides further evidence of their bacterial nature. The effect of PCL on the symbiotes of B. culicis suggests the presence in their cell envelopes of mucopeptide, which probably provides rigidity for maintaining the bacterial shape of the symbiotes.     

New aspect of the “mycetome” of a thrips,Bactrothrips brevitubus Takahashi (Insecta: Thysanoptera)

Morphology and cytochemical properties of “mycetomes” are described in the developing oocytes and eggs of an idolothripine thrips, Bactrothrips brevitubus (Thysanoptera). The “mycetome” is an aggregation of numerous granules of various sizes. We found no membrane encapsulating the aggregation of granules. Two types of granules are distinguishable: the smaller ones filled with electron-dense material and the larger ones with inclusion of myelin-like structures. Each of the granules has a limiting membrane. The limiting membrane is a simple unit membrane but shows no characteristics of cell walls. No nucleoid or nucleoplasm is detected in the granules. The “mycetome” takes up dyes whose specific incorporation into lysosomes has been demonstrated. In addition, a high activity of acid phosphatase is demonstrated in the “mycetome.” These characteristics apparently indicated that the “mycetome” of Bactrothrips brevitubus is an aggregation of lysosomes but not a clump of microorganisms. Thus we propose that the structure being regarded as the mycetome should be renamed the “lysosomal aggregation.” © 1994 Wiley-Liss, Inc.     

Inhibition of the onset of vitellogenesis by abnormal retention of mature eggs in the stick insect,Carausius morosus

Mature eggs stored in the lateral oviducts are unusual in Carausius morosus. When egg laying is experimentally prevented, ovarian production is inhibited by the accumulation of mature eggs within the lateral oviducts. Such storage affects only the very small terminal oocytes and not the vitellogenic follicles, which attain to term and are ovulated. The fact that unilateral retention of eggs affects only the corresponding ovary strongly suggests that the mature eggs themselves inhibit ovarian production in the stick insect without the involvement of an endocrine organ. It is likely that this mode of action, different from that suggested for the housefly or Rhodnius prolixus, is correlated with the independence of ovarian development from the corpus allatum hormone in the stick insect.     

Do gut symbiotes reflect the endemism of their host black flies (Diptera: Simuliidae) in the Caucasus of Armenia?

Aim We tested the hypothesis that endemism of black flies in the Caucasus of Armenia is reflected in their gut symbiotes, i.e. trichomycete fungi and ichthyosporean protists. Location The study area, Armenia, is a biodiversity hotspot, with high levels of plant and animal endemism. Located in the southern Caucasus, Armenia is a rugged, mountainous, landlocked country of 29,800 km², with fast‐flowing rivers and little forested land. About half of the 51 species of black flies in Armenia are endemic to the Caucasus. Methods The larvae of 22 species of black flies, including seven endemic to the Caucasus, were collected from 35 sites throughout Armenia, from 1998 to 2004, and assayed microscopically for gut‐inhabiting trichomycete fungi and ichthyosporean protists. Results Nearly 68% of larval black flies in Armenia contained at least one species of gut symbiote, with a mean of 0.9 ± 0.04 species per host larva. Eight species of trichomycete fungi and two species of ichthyosporeans were found in larval guts. Seven of these species were recorded for the first time in Asian black flies. Species accumulation curves showed no additional species of gut symbiotes after 17 host species and 22 sites had been examined. All gut symbiotes were widely distributed, indicating an absence of endemic trichomycetes in the black flies of Armenia. Main conclusions Although the Caucasus Region is noted for its high levels of endemism for plants and animals, including black flies, the gut symbiotes of black flies are all widespread species. The absence of endemic gut symbiotes in larval black flies might be explained by their lack of host specificity; an absence of host specificity would not constrain the geographical distribution of the symbiotes.     

In vivo sterol biosynthesis by pea aphid symbiotes as determined by digitonin and electron microscopic autoradiography

Summary Pea aphid primary symbiotes have previously been shown to synthesize cholesterol in vitro. Two electron microscopic techniques were used here to determine whether the symbiotes also synthesize cholesterol in vivo and whether this cholesterol is made available to the aphid. We also inquired into a possible role of secondary symbiotes in cholesterol biosynthesis. Treatment of aphids with digitonin resulted in significant alteration of ultrastructural sites in primary and secondary symbiote membranes. We concluded that these sites are areas of high cholesterol concentration in the symbiotes.Electron microscopic autoradiography with ³H-mevalonate precursor indicated that both primary and secondary symbiotes synthesize cholesterol; in both cases, the majority of grains were associated with the symbiote membranes. While the frequency of grains on the symbiotes remained constant, irrespective of incubation time in labelled media, the frequency of grains over surrounding tissues increased exponentially as the time of incubation was increased from 30 min to 8 h, indicating that symbiote cholesterol is transported to other tissues. High voltage electron microscopic autoradiography permitted thick section autoradiography, reducing the time of emulsion exposure from 54 days (thin section) to 12 days (0.5 m sections).Research supported by the College of Agricultural and Life Sciences, University of Wisconsin, and by a research grant (PCM 74-2401 A01) from The National Science FoundationThe authors wish to thank Dr. G.A. DeZoeten for his invaluable advice and assistance with the autoradiographic techniques, Mr. Gary Gaard for his help with electron microscopy, and Dr. Dale Johnston and Dr. Damien Neuberger for their generous help in the use of the high voltage EM     

Pattern of bacteriocyte formation in Periplaneta americana (L.) (Blattaria: Blattidae)

In the embryos of Periplaneta americana (L.) (Blattaria: Blattidae), bacterial symbionts, together with vitellophages, form a mycetomic structure inside the deutoplasm; this regresses between the 15th and 16th day after deposition of the ootheca. In this article we describe the migration of bacteria across the wall of the midgut from the mycetome, and the topographic distribution of pre-bacteriocyte cells. We also report that the pre-bacteriocytes are present only on the lateral surface of the internal abdominal fat bodies. We discuss the possible embryological origin and evolution of these cells, and put forward the hypothesis that pre-bacteriocytes are derived from oenocytes activated to perform phagocytosis.     

Symbiosis in the Larger Foraminiferan Sorites marginalis (with Notes on Archaias spp.)

SYNOPSIS. It was demonstrated with the aid of light and electron microscope studies that Sorites marginalis (Lamarck) harbors zooxanthellae. The hosts were scraped from Thalassia testudinum Konig growing in Key Largo Sound (Florida, U.S.A.) and immediately preserved in appropriate fixatives. Zooxanthellae were distributed unevenly throughout all the chamberlets; only a few symbiotes were found in the embryonic chambers and the inner or outer chambers, but the intermediate chambers were packed with symbiotes. The outer chambers contained many food vacuoles in addition to symbiotes. Some zooxanthellae might have been in the process of degeneration or digestion. The symbiotes were found to have a typical dinoflagellate nucleus, a single large lobate cortical chloroplast with one stalked pyrenoid, an accumulation body, and many starch granules. The nonmotile stage of the zooxanthella was similar, but perhaps not identical, to Symbiodinium microadriaticum Freudenthal from various hosts.
The foraminiferan host is heterokaryotic with hundreds of generative (small) nuclei and scores of vegetative (large) nuclei. Most of the generative nuclei were found in the embryonic apparatus and the inner chambers. Most of the vegetative nuclei were found in the inner and outer chambers.     

Arthropod symbiotes of Laonastes aenigmamus (Rodentia:Diatomyidae)

Arthropod symbiotes of the Laotian rock-rat, Laonastes aenigmamus (Rodentia:Diatomyidae), from Laos are examined. This host is a member of Diatomyidae previously thought to have gone extinct >10 million yr ago. Permanent symbiotes are represented by 2 species, a new species of sucking louse, Polyplax sp., near rhizomydis (Phthiraptera:Polyplacidae), and a new species of fur mite, Afrolistrophorus sp., near maculatus (Acariformes:Listrophoridae). The temporary parasites are represented by 18 species, i.e., 1 mesostigmatan species, i.e., a new species of Androlaelaps near casalis (Parasitiformes:Laelapidae); immature stages of 2 tick species, Ixodes granulatus and Haemaphysalis sp. (Parasitiformes:Ixodidae); and a rich fauna of chiggers (Acariformes:Trombiculidae) comprising 8 genera and 15 species. It is hypothesized that this host completely lost its initial fauna of ectosymbiotes and that ancestors of the recorded symbiotes switched to this host from rodents of the superfamily Muroidea.     

Pharmacological profile of octopamine and 5HT receptors on the lateral oviducts of the cockroach,Periplaneta americana

The effects of the amines 5HT and octopamine on the myogenic activity of Periplaneta americana (L.) oviducts and the pharmacological profile of octopamine and 5HT receptors on the lateral oviducts have been determined. Application of 5HT to the oviducts resulted in a dose-dependent increase in basal tonus and amplitude of contractions. Antagonist studies revealed that the 5HT receptor on the cockroach oviduct most resembles the mammalian 5HT₂ receptor. Application of octopamine resulted in a decrease in basal tonus and had a biphasic effect on the amplitude of contractions, being stimulatory at low doses and inhibitory at higher ones. The inhibitory effects of octopamine appear to be mediated via cAMP and are blocked by antagonists which indicate that the octopamine receptor is of the octopamine-2 type. © 1995 Wiley-Liss, Inc.     

Ovulation and the mechanism of follicle rupture

Summary The effects of penicillin and chlortetracyline HCl on the fine structure of the intracellular symbiotes of the pea aphid were studied in an attempt to remove the symbiote population. High penicillin concentrations, 1% and 0.1%, caused symbiote breakdown but were toxic and/or repellent to the aphids; at 0.1% specific effects were observed on the symbiotes' cell walls. After the use of 0.01% penicillin in the aphid diet, the symbiotes had abnormal cell walls and were abnormally dilated; however, symbiote division and transmission from one aphid generation to the next seemed unaffected and the aphids appeared normal. Aphids fed 0.1% chlortetracycline failed to reproduce. After 7 days, their symbiotes were found to break down at a high rate but aphid mitochondria were also adversely affected at this stage. Following 0.002% chlortetracycline, the aphids produced aposymbiotic progeny with apparently normal mitochondrial populations; these larvae failed to develop.Research supported by the College of Agricultural and Life Sciences, University of Wisconsin, and by a research grant (GB 31904 X) from the National Science Foundation.