Ultrastructural changes of Drosophila larval and prepupal salivary glands cultured in vitro with ecdysone.

Alterations in the ultrastructure of in vitro cultured larval salivary glands of Drosophila melanogaster in response to the steroid hormone ecdysone were studied in relation to complex changes in puffing patterns. We found that the changes in the fine structure of cultured glands reflected progression of the puffing pattern, and they paralleled those seen in vivo. We observed that glue secretion by exocytosis, the main function of salivary glands, took place between puff stage 5 (PS5) and PS7. Glue could not be expectorated under culture conditions but was slowly released from the lumen through a duct into the medium. After the cultured glands reached PS13/PS14, further progress of puffing and fine structural alterations required that the ecdysteroid titer be transiently extremely low or absent. Under in vitro conditions we did not observe the putative new secretory program(s) described for glands in vivo after PS12. However, ultrastructural changes which unambiguously indicated that an autohistolytic process had begun in vitro started to appear after PS17. Many salivary gland cells developed numerous features of progressive self-degradation between PS18 and PS21. Actual degradation of salivary glands in vivo seemed to be rapid, but in vitro degradation was never completed, probably due to a lack of exogenous factors from the hemolymph. Manipulations of ecdysone titer in vitro in the culture medium, known during the larval puffing cycle to cause premature induction of developmentally specific puffing patterns, did not affect the normal development of ultrastructural features of the cytoplasm and nucleus.

Use of partially engorged female ticks as laboratory animals in microbiological research.

Partially engorged female ticks were used as laboratory animals in microbiological research. The ticks, which were inoculated intracoelomally, became a convenient substrate for the detection of viruses, rickettsiae and protozoal parasites. This research concerned the isolation of newly recovered micro-organisms, the study of development, structure and distribution of microbial agents in ticks, and the study of their interaction with other pathogens or symbionts during mixed infection in a tick body. The isolation and maintenance of Rickettsiella phytoseiuli, the organism not of tick-borne origin, was achieved. For use in Central Europe the tick Dermacentor reticulatus is recommended for the above investigations.

Phenomenon of Rickettsiella phytoseiuli in Phytoseiulus persimilis mite.

An unknown microorganism occurring in a predaceous mite Phytoseiulus persimilis was described by the author in 1977 as a new species Rickettsiella phytoseiuli. Some new results on the relation between this agent and its hosts are presented in this paper.

Coxiella and Rickettsiella: comparison of ultrastructure with special reference to their envelope.

Ultrastructure of Rickettsiella phytoseiuli (R.p.) multiplying in female ticks Dermacentor reticulatus was compared with that of Coxiella burnetii (C.b.) in the same ticks and in mice. C.b. in ticks and mice were always represented by 2 main cell types: small dense round or rod-like cells (DC) and larger bacteria-like cells (BC). DC were surrounded with a characteristic five-layered 20 nm thick envelope. Under the envelope DC had a stack of parallel intracytoplasmic membranes with a periodicity 5-6 nm. R.p. in tick fat body and synganglion were also inside phagosomes and formed 6 sequentially developing cell forms: dense (elementary), intermediate, bacterial, giant, and crystal-forming in which small dark bodies (initial particles) condensed. Two of them--dense and bacterial--corresponded to DC and BC of C.b. The DC envelope structure of R.p. was strikingly similar to that of some C.b. DC in mouse. We confirmed the general morphologic similarities in the structure of C.b. and R.p. DC and that of C.b. BC and intermediate cells of R.p. The envelope structure of DC type was found in other gracilicute bacteria and is supposed to have no taxonomic value but to be a reflection of population heterogeneity.

Endocytobionts in Dermacentor reticulatus ticks (Ixodidae): an electron microscope study.

Endocytobionts (ECBs) were detected in the ovaries of Dermacentor reticulatus. Their developmental cycle is directly related to the developmental stages of tick oocytes. Two basic forms of ECBs occur in tick cells, i.e. dense forms, occurring singly or in aggregates situated free in the cytoplasm of host cells, and the light forms which are larger, pleomorphic and always situated inside vacuoles of host cells. The light forms occur together with dense forms in all oocytes. The dense forms occur freely and independently in funicular cells of the oocyte, the epithelial cells of oviducts, and additionally in the cells of the Malpighian tubules. A probable function of ECBs in the tick host is discussed.

Mixed infection of Rickettsiella phytoseiuli and Coxiella burnetii in Dermacentor reticulatus female ticks: electron microscope study.

Mixed infection of Rickettsiella phytoseiuli and Coxiella burnetii was investigated in hemolymph and organs of experimentally infected females of Dermacentor reticulatus ticks. Following intracoelomic infection, both agents, with the exception of Gene's organ, multiplied well in the cells of the tick host's organs. Two out of six developmental stages of R. phytoseiuli, i.e., crystal-forming and small dark particles, in dual infection with C. burnetii revealed marked morphological alterations. C. burnetii in the presence of R. phytoseiuli penetrated into the cortical layer of the synganglion and into the alveoli of the second and third type of salivary glands, but did not occur in the single infection.

Virus-like particles in Dermacentor reticulatus ticks.

Virus-like particles were found in the brain and salivary glands of Dermacentor reticulatus ticks double infected with Coxiella burnetii and Rickettsiella phytoseiuli. The particles were spherical (diameter of about 70 nm) and consisted of an electron-dense inner core and a dense external coat.

Interaction between Dermacentor reticulatus cells and Coxiella burnetii in vivo.

By electron microscopy the distribution of Coxiella burnetii was followed in females of Dermacentor reticulatus injected intracoelomally in a dose of about 10(3) EID50 per tick. The heaviest infestation with coxiellae was noticed in the cells of haemolymph, fat body, Malpighian tubulus and tracheal complex. No rickettsiae were found in Gene's organ. Unexpected was the propagation of rickettsiae in muscle fibres. C. burnetii multiplied in all organs affected. Heavy infection resulted at the marked damage of cell components. Coxiellae were evident in the haemocytes; in organs they formed small and large cell variants; endospore formations were observed free in the haemolymph.

Experimental infection with Rickettsiella phytoseiuli in adult female Dermacentor reticulatus (Ixodidae): an electron microscopy study.

Rickettsiella phytoseiuli naturally occurring in Phytoseiulus persimilis mites was cultivated in adult female Dermacentor reticulatus ticks. It demonstrates all six known developmental stages: dense, intermediate, bacterial, giant, crystal-forming and small dark particles. These stages of rickettsiae were found in salivary glands, Malpighian tubules, synganglion, ovaries, tracheal complex, haemolymph, fat body and alimentary tract. Rickettsiella phytoseiuli did not infect the Gené's organ. It multiplied in female ticks in a manner similar to that in the typical host mite, P. persimilis.

Electron microscopic study of developmental stages of Rickettsiella phytoseiuli in Phytoseiulus persimilis Athias-Henriot (Gamasoidea:Phytoseiidae) mites.

Rickettsiella phytoseiuli was found in great amounts in all tissues except of the nervous system of adult Phytoseiulus persimilis mites. Six morphologically different stages (dense, intermediate, bacterial, giant, crystal-forming and small dark particles) of R. phytoseiuli were detected. No rickettsiae were seen in the larvae and in phase 1 and 2 nymphae of these mites.

Attempt to cultivate Rickettsiella phytoseiuli in Dermacentor reticulatus ticks: an electron microscopic study.

Rickettsiella phytoseiuli occurring in Phytoseiulus persimilis mites was cultivated in Dermacentor reticulatus ticks. Rickettsiella multiplied similarly as in mites exerting all six developmental stages: dense, intermediate, bacterial, giant, crystal-forming and small dark particles.

Rickettsiella phytoseiuli and virus-like particles in Phytosfiulus persimilis (Gamasoidea: Phytoseiidae) mites.

Electron microscope investigation of tissue of Phytoseiulus persimilis mites revealed its double infection with Rickettsiella phytoseiuli and virus-like particles. The presence of rickettsiae in tissues of the mite seemed to be decisive for establishment of virus infection.