Karyolysus sp. (Haemogregarinidae, Adeleida, Apicomplexa): Host-Parasite Relationships of Persisting Stages

ABSTRACT. Two persisting stages in the life cycle of a hemogregarine Karyolysus sp. are described from the liver and blood cells of its intermediate host, the lizard Lacerta raddei nairensis. The tissue cell merozoites lie in a parasitophorous vacuole. Despite the protective role of the vacuolar membrane, the intracellular parasites are progressively destroyed and eliminated during the autumn and winter. Some of the merozoites that normally survive within the host cell even in cold seasons appear to be surrounded by another type of parasitophorous vacuole which is connected to the intercellular space by narrow channels. The intraerythrocytic gamonts that persist in the circulating blood are encapsulated and undergo progressive, obvious structural changes. The two persisting stages are compared with hypnozoites of other Sporozoa.     

Plasmodium berghei: Osmotic fragility of malaria parasites and mouse host erythrocytes

The osmotic properties of intraerythrocytic and ultrasonically liberated malaria parasites (Plasmodium berghei) were analyzed and compared with those of mouse host erythrocytes utilizing a multiple tube fragility test. Cells were incubated in phosphate buffered saline solutions of varying osmolalities ranging from 20–4000 mOsm. Changes in cell ultrastructure and parasite infectivity were used as indicators of osmotic damage. Intraerythrocytic and host cell-free plasmodia showed similar patterns of cell alteration and changes in infectivity following osmotic stress. The various developmental forms within each of the preparations responded somewhat differently to hypo-osmotic stress, however. The majority of merozoites seemed to be more sensitive than many trophozoites, schizonts, and segmenters. Small trophozoites were, on the average, more resistant than other developmental forms. Incubation of parasite populations in hypotonic salt solutions with osmolalities slightly greater than the infectivity threshold of 100 mOsm lysed the majority of the merozoites, whereas many small trophozoites were still intact. While normal erythrocytes were more resistant to hypo-osmotic stress than were either intracellular or free parasites, the majority of parasitized erythrocytes was less resistant than normal erythrocytes. The predominant alteration induced by hyperosmotic stress appears in the parasite's nuclear region with myelination of the nuclear membranes and chromatin clumping. The infectivity threshold in the hypertonic range was found to be approximately 2500 mOsm. Results indicate that these obligate intracellular parasites have a wide range of osmotic sensitivities and that they are capable of existing for short periods in various osmotic environments ranging from 100–2500 mOsm without complete loss of infectivity. This suggests that these parasites have osmotic regulatory capabilities at least comparable to those of host cells.     

Parasitophorous vacuole: morphofunctional diversity in different coccidian genera (a short insight into the problem)

We present a short insight into the problem of parasitophorous vacuole (PV) formation as a most peculiar kind of cell vacuolization occurring in the course of intracellular development of coccidian pathogens of the genera Eimeria, Isospora, Toxoplasma, Sarcocystis, Cryptosporidium, Epieimeria, and Karyolysus. The review focuses on the morpho-functional diversity of PVs in these parasites. By the present time, the PVs containing different parasite genera and species have been examined to different extent. The membrane of the PV (PVM) obviously derives from the host cell plasmalemma. But soon after parasite penetration, the morphofunctional organization and biochemical composition of the PVM drastically changes: its proteins are selectively excluded and those of the parasite are incorporated. As the result, the PV becomes not fusigenic for lysosomes or any other vacuoles or vesicles, because host cell surface markers necessary for membrane fusion are eliminated from the PVM during parasite invasion.The pattern of the PVs is parasite specific and demonstrates a broad diversity within the same genera and species and even at different stages of the endogenous development. The PV is far from being an indifferent membrane vesicle containing the parasite. Instead, it represents a dynamic system that reflects the innermost events of host-parasite relationships, thus promoting the accomplishing of the parasite life cycle, which, in its turn, is a necessary prerequisite of the parasite eventual survival as a species.     

Morphological study of the heart innervation of bats Myotis daubentoni and Eptesicus serotinus (Microchiroptera: Vespertilionidae) during hibernation

The capability of bats to have heart rates fewer than 10 beats/min during hibernation and greater than 700 beats/min during flight surprises biologists and cardiologists. Cardioacceleration of hibernating bats is considered to be a function of their intracardiac nervous system. In the present study we investigated the morphology of the heart innervation of ten M. daubentoni and four E. serotinus bats during their natural hibernation in order to determine which intracardiac structures may be involved in cardioacceleration during their short-term (in av. 15-30 min) arousal from hibernation. The primary conclusions were as follows: (1) The innervation pattern of bats differs from many mammals in that bats have: (a) a subepicardiac nerve plexus which is vastly developed and contains a large number of intrinsic ganglia on both atria and ventricles, and (b) very small diameter axons within the unmyelinated nerve fibres, from 0.15 to 0.7 microm. (2) During hibernation an intercellular space of the sinoatrial node of M. daubentoni bats was in part filled with a cottony substance which can presumably be considered to be a temporary barrier between the conductive cardiomyocytes and nerve fibres. (3) In the hibernating bats, the acetylcholine vesicles were aggregated in the synaptic bulbs away from the presynaptic membrane. Possibly, the aggregation of the acetylcholine vesicles is capable of modifying cholinergic influences on the heart activity of hibernating bats. (4) The dense cores of catecholamine synaptic vesicles within, adrenergic axon terminals were seldomly observed in hibernating bats. Therefore, catecholamines probably do not play a crucial role in the cardioacceleration of hibernating bats.     

Functional consequences of ultrastructural geometry in "backwards" fluid-transporting epithelia

Many fluid-transporting epithelia possess dead-end, long, and narrow channels opening in the direction to which fluid is being transported (basal infoldings, lateral intercellular spaces, etc.). These channels have been thought to possess geometrical significance as standing-gradient flow systems, in which active solute transport into the channel makes the channel contents hypertonic and permits water-to-solute coupling. However, some secretory epithelia (choroid plexus, Malpighian tubule, rectal gland, etc.) have "backwards" channels opening in the direction from which fluid is being transported. It is shown that these backwards channels can function as standing-gradient flow systems in which solute transport out of the channel makes the channel contents hypotonic and results in coupled water flow into the channel mouth. The dependence of the transported osmolarity (isotonic or hypertonic) on channel radius, length, and other parameters is calculated for backwards channels for values of these parameters in the physiological range. In addition to backwards channels' being hypotonic rather than hypertonic, they are predicted to differ from "forwards" channels in that some restrictions are imposed by the problem of solute exhaustion, and in the presence of a sweeping-in effect on other solutes which limits the solutes that may be transported.     

Neonicotinoids override a parasite exposure impact on hibernation success of a key bumblebee pollinator

1. Seasonal adaptations enabling the bridging of periodic challenging abiotic conditions are taxonomically widespread. However, sensitivity to other environmental stresses can be heightened during these periods. 2. Several temperate insects with over‐wintering strategies play key ecosystem and economic roles, including wild bee pollinators. For example, hibernation survival in temperate bumblebees is crucial, as only new queens of future social colonies over‐winter. These bees are also faced with other abiotic and biotic stressors, some of which have been linked to recent pollinator declines, such as exposure to pesticides and parasites. 3. Using a fully crossed experiment, the influence of dietary exposure to neonicotinoid insecticides (thiamethoxam and clothianidin) and/or the prevalent bumblebee parasite Crithidia bombi on hibernation survival and hibernation weight change of Bombus terrestris bumblebee queens was investigated. 4. Both neonicotinoid and C. bombi exposures reduced hibernation success individually, but no additive or synergistic effects between the stressors were found. Further, effects were asynchronous, with early neonicotinoid effects on hibernation mortality overriding later parasite effects under combined exposures. Neonicotinoid exposure also increased hibernation weight loss of surviving queens. 5. Diapause periods, employed by numerous temperate organisms, are likely to be especially vulnerable to environmental stresses, besides the seasonal challenge for which these periods are an adaptation. Thus, diapause requires inclusion during the consideration of the impacts of such stresses. Accordingly, it is demonstrated here that naturally relevant exposures of pesticides and parasites have important detrimental effects on bumblebees during a critical hibernation period, with potential consequences for populations of these key wild pollinators.     

Intercellular channels in the pars tuberalis of the rat hypophysis and their relationship to the subarachnoid space

Summary A system of intercellular channels is described in the pars tuberalis (PT) of the female rat. These spaces are lined by all types of cells found in the PT and are not sealed off by tight junctions. Ventrally and dorsally, the intercellular spaces open toward the basement membranes separating the PT from (i) the subarachnoid space, and (ii) the perivascular space of the portal capillaries, respectively. These intercellular channels differ from the follicles, which are also found in the PT, being lined by a particular type of cell.In a second group of female rats an epoxy mixture was injected into the third ventricle; 10 min thereafter horseradish peroxidase was infused into the cisterna magna. After processing the brain for the demonstration of exogenous peroxidase, it was found that the tracer had reached the subarachnoid space adjacent to the hypothalamus and entered into all ventricular cavities with the exception of the infundibular recess. Under these experimental conditions it was found that the tracer fills all intercellular channels of the PT, thus indicating that there is no barrier between the subarachnoid space and the PT. It is suggested that the subarachnoid space should be regarded as a probable route for the transport of trophic factor(s) and/or secretory product(s) of the PT.Supported by Grant S-80-13 from Directión de Investigaciones, Universidad Austral de Chile     

Etude des premiers stades de développement de la Coccidie Coelotropha durchoni

Summary This study considers the earlier growth stages of Coccidium Coelotropha durchoni in its host, Nereis diversicolor. Before evolving into free trophozoites and gamontes in coeliac fluid, the parasites remain in muscular and coeliac cells in microscopic intracellular form. Electron microscope reveals that these stages show an intermediary fine structure between that of a sporozoite — from which they keep some typical characteristics such as the conoid, the fibers and the involuted tubuli — and that of the future free trophozoites. The wall consisting in two clear membrans is provided with one or several micropores. The classical cytoplasmic organites clearly stand out: dictyosomes show constant relationship with ergastoplasm, the mitochondria contain short inner tubuli. Besides the paraglycogen granules and lipoid vacuoles, at least three types of vacuoles may be observed. Peculiar topographic relationship connects mitochondria and paraglycogen granules probably in formation. In the nucleus with classical membrane and heterogeneous structure, a rather voluminous nucleolus may be seen.     

Avian epidermal differentiation: role of lipids in permeability barrier formation

Though avian skin is known to possess a highly lipogenic epidermis, little is known about its permeability barrier function. We correlated epidermal barrier function, fine structure and lipid biochemistry in the pigeon, Columbia livia, and compared these features with terrestrial mammalian systems. Whereas barrier function, as assessed by transepidermal water loss was not as efficient as in mammals, both groups shared certain morphological features including substantial compartmentalization of lipids in stratum corneum intercellular domains. Avian intercellular lipids derive from extrusion of intracellular non-membrane-bound droplets from lowermost corneocytes, rather than by secretion of lamellar discs from multigranular bodies, as previously reported in some avians, and in mammals. Instead, both the internal lamellae and the limiting membranes of multigranular bodies appear to degenerate, leading to the formation of non-membrane-bound droplets. The lipid content of avian epidermis and stratum corneum demonstrates important similarities to terrestrial mammals, i.e. abundant sphingolipids, a paucity of phospholipids, and abundant neutral lipids, but also certain striking differences, i.e. persistence of glycosphingolipids and triglycerides into the stratum corneum. Thus, avian stratum corneum forms a two-compartment system of lipid-depleted cells embedded in non-polar-lipid enriched intercellular domains, analogous to mammals. But, in contrast to mammals, the highly attenuated corneocytes of avians, which results from a paucity of keratin filaments, produce a 'straws-and-mortar' tissue, rather than the 'bricks-and-mortar' tissue of mammals.     

Formation and diversity of parasitophorous vacuoles in parasitic protozoa. The Coccidia (Sporozoa, Apicomplexa)

Data on parasitophorous vacuole (PV) formation in host cells (HC) harbouring different intracellular protozoan parasites have been reviewed and critically analysed, with special reference to the main representatives of the Coccidia. The vacuole membrane (PVM) is the interface between host and parasite, playing a role in nutrient acquisition by the parasite from the HC. The PV phenomenon is regarded as a generalized HC response to the introduction of alien bodies (microorganisms), which eventually reflects the evolutionary established host-parasite relationships at cellular, subcellular and molecular levels. Special attention has been paid to the existing morpho-functional diversity of the PVs within the same genera and species of parasites, and even at different stages of the parasite life cycle. The PVM is generally considered to derive from the HC plasmalemma, whose biochemical composition undergoes significant changes as the intravacuolar parasite grows. The original HC proteins are selectively excluded from the PVM, while those of the parasite are incorporated. As the result, the changed PVM becomes not fusigenic for HC lysosomes. For Toxoplasma gondii and other cyst-forming coccidia (Isospora, Sarcocystis), a definite correlation has been noticed between the extent of rhoptry and dense granule secrets released by a zoite during HC internalization, on the one hand, and the pattern of the PV that forms, on the other one. In T. gondii, tachyzoites, known to discharge abundant secrets, commonly force the development of PVs limited with a single unit membrane and equipped with a tubulovesicular network in the lumen. Unlike, bradyzoites known to be deficient in secretory materials trigger the formation of PVs with a three-membrane lining composed of the changed invaginated plasmalemma in addition to two membranes of endoplasmic reticulum. The two different types of PV harbour, respectively, exoenteric and enteric stages of T. gondii, the latter being confined to the cat intestine only. Unlike, all endogenous stages of the classic intestinal coccidia (Eimeria spp.) develop within PVs limited with a single membrane, with some invaginations extending into the PV lumen. Unusual PV patterns are characteristic of the extracytoplasmic eimerian coccidia (Cryptosporidium, Epieimeria) and adeleid haemogreagarines (Karyolysus). In cyst-forming coccidia, the PVM is actively involved in tissue cyst wall formation, thus protecting the encysted parasites from recognition by the host immune system. All this strongly suggests that the PV is far from being an indifferent membraneous vesicle containing a parasite, but represents a metabolically active compartment in infected cells. Since all the coccidia are obligate intracellular parasites, the mode of their intimate interaction with the HC, largely accomplished via the PV and its membrane, is vital for their survival as biological species.     

Binding of fluorescein-labelled lectins on trophozoites and cysts of 3 strains of Toxoplasma gondii

The presence of carbohydrates on the surface of trophozoites and cysts of Toxoplasma gondii was investigated using 26 fluorescein labelled lectins. Three strains of parasites at different phases were tested; intra-cellular, intra-cystic and free trophozoites did not stain. However Soja hispida lectin intensly bind on cyst wall. The fluorescence could be eliminated by preincubation with certain carbohydrates, but not with trophozoite soluble antigen. These results suggest to the authors that cyst wall may be of host rather than of parasite origin.     

Evolutionary complication of life cycles in Coccidea (Sporozoa: Coccidea)

Similar strategies to preserve a species were evolved independently in different groups of Coccidea. Polyenergid oocysts and tissue cysts are found in representatives of the orders Protococcidiida and Eimeriida. Hypnozoits are found in Karyolysus lacerate and Plasmodium vivax; transovarial transmission of parasites occurs in life cycles of Coccidea of the genera Karyolysis and Babesia. Formation of heteroxenity in groups of Coccidea apparently was developed by different ways and in different periods. In some groups (Cystoisospora, Toxoplasma, Aggregata, Atoxoplasma, Schelackia, Lankesterella, and Calyptospora), recent definitive hosts were initial hosts; in other groups (Sarcocystis, Karyolysus, Haemogregarina, Hepatozoon, Plasmodium, Haemoproteus, Leucytozoon, Babesiosoma, Theileria, and Babesia), intermediate hosts were initial hosts.     

Interferon-gamma-mediated inhibition of the development of Eimeria tenella in cultured cells

The effect of treating cultured Madin-Darby bovine kidney cells (MDBK) with recombinant bovine interferon-alpha 1 (IFN-alpha 1) or recombinant bovine interferon-gamma (IFN-gamma) on the intracellular development of Eimeria tenella was studied. Treatment of the MDBK cells with IFN alpha-1 for 24 hr before infection and for 48 hr after infection had no effect on the development of E. tenella. However, following the same treatment regime with serial dilutions of IFN-gamma induced a significant reduction in the number of total intracellular parasites (sporozoites, trophozoites, and meronts) compared to the untreated controls. Of these intracellular parasites, less than 30% had developed beyond the sporozoite stage. These results are suggestive of a role for IFN-gamma in protecting or limiting the development of E. tenella in their host cells. These results could be relevant to the control of these organisms and may be exploited for use with a coccidiosis vaccine.     

Intracellular antioxidant enzymes are not globally upregulated during hibernation in the major oxidative tissues of the 13-lined ground squirrel Spermophilus tridecemlineatus

Hibernating mammals exhibit oxidative stress resistance in brain, liver and other tissues. In many animals, cellular oxidative stress resistance is associated with enhanced expression of intracellular antioxidant enzymes. Intracellular antioxidant capacity may be upregulated during hibernation to protect against oxidative damage associated with the ischemia-reperfusion that occurs during transitions between torpor and arousal. We tested the hypothesis that the 13-lined ground squirrel (Spermophilus tridecemlineatus), upregulates intracellular antioxidant enzymes in major oxidative tissues during hibernation. The two major intracellular isoforms of superoxide dismutase (MnSOD and CuZnSOD), which catalyze the first step in superoxide detoxification, were quantified in heart, brain and liver tissue using immunodetection and an in-gel activity assay. However, no differences in SOD protein expression or activity were found between active and hibernating squirrels. Measurements of glutathione peroxidase and glutathione reductase, which catalyze hydrogen peroxide removal, were not broadly upregulated during hibernation. The activity of catalase, which catalyzes an alternative hydrogen peroxide detoxification pathway, was higher in heart and brain of torpid squirrels, but lower in liver. Taken together, these data do not support the hypothesis that hibernation is associated with enhanced oxidative stress resistance due to an upregulation of intracellular antioxidant enzymes in the major oxidative tissues.     

Site preference of myxozoans in the kidneys of Hungarian fishes

Myxozoans are common parasites of fish kidneys, with most having specific sites of development. Five specific sites of development include (1) the lumen of renal tubules, (2) the renal corpuscles followed by location in renal tubules, (3) intracellular location within the tubular epithelium followed by a stage in the lumen of the ducts, (4) haematopoietic tissue with dispersed trophozoites, and (5) haematopoietic tissue with large, localized plasmodia. A coelozoic development preceded by presporogonic multiplication characterises most Sphaerospora spp. Early plasmodial stages of Myxidium and Chloromyxum spp. are frequently found in the renal glomerules, while spores develop in the urinary channels in plasmodia released from the renal corpuscles. In Hoferellus and Myxobilatus spp., spores are formed in small plasmodia inside the lumen of the urinary ducts after several internal cleavages in the epithelium of renal tubules. The presence of dispersed trophozoites among haematopoietic tissue cells of the renal interstitium characterises the development of Sphaerospora tincae and Tetracapsuloides bryosalmonae, the causative agent of proliferative kidney disease (PKD). Spores of S. tincae are formed at the place of plasmodial development, while spore formation of PKD is in the renal tubules. A large mass of spores, often surrounded by a connective tissue capsule, can appear in the renal interstitium during infections by several Myxobolus spp.; furthermore, a large number of these spores formed in plasmodia in distant tissues can also accumulate in melano-macrophage centres.     

The permeability barrier in mammalian epidermis

The structural basis of the permeability barrier in mammalian epidermis was examined by tracer and freeze-fracture techniques. Water-soluble tracers (horesradish peroxidase, lanthanum, ferritin) were injected into neonatal mice or into isolated upper epidermal sheets obtained with staphylococcal exfoliatin. Tracers percolated through the intercellular spaces to the upper stratum granulosum, where further egress was impeded by extruded contents of lamellar bodies. The lamellar contents initially remain segregated in pockets, then fuse to form broad sheets which fill intercellular regions of the stratum corneum, obscuring the outer leaflet of the plasma membrane. These striated intercellular regions are interrupted by periodic bulbous dilatations. When adequately preserved, the interstices of the stratum corneum are wider, by a factor of 5-10 times that previously appreciated. Freeze-fracture replicas of granular cell membranes revealed desmosomes, sparse plasma membrane particles, and accumulating intercellular lamellae, but no tight junctions. Fractured stratum corneum displayed large, smooth, multilaminated fracture faces. By freeze-substitution, proof was obtained that the fracture plane had diverted from the usual intramembranous route in the stratum granulosum to the intercellular space in the stratum corneum. We conclude that: (a) the primary barrier to water loss is formed in the stratum granulosum and is subserved by intercellular deposition of lamellar bodies, rather than occluding zonules; (b) a novel, intercellular freeze-fracture plane occurs within the stratum corneum; (c) intercellular regions of the stratum corneum comprise an expanded, structurally complex, presumably lipid-rich region which may play an important role in percutaneous transport.     

Relative number of generations of hosts and parasites does not influence parasite local adaptation in coevolving populations of bacteria and phages

A potential consequence of host-parasite coevolution in spatially structured populations is parasite local adaptation: local parasites perform better than foreign parasites on their local host populations. It has been suggested that the generally shorter generation times of parasites compared with their hosts contributes to parasites, rather than hosts, being locally adapted. We tested the hypothesis that relative generation times of hosts and parasites affect local adaptation of hosts and parasites, using the bacterium Pseudomonas fluorescens and a lytic phage as host and parasite, respectively. Generation times were not directly manipulated, but instead one of the coevolving partners was regularly removed and replaced with a population from an earlier time point. Thus, one partner underwent more generations than the other. Manipulations were carried out at both early and later periods of coevolutionary interactions. At early stages of coevolution, host and parasites that underwent relatively more generations displayed higher levels of resistance and infectivity, respectively. However, the relative number of generations that bacteria and phages underwent did not change the level of local adaptation relative to control populations. This is likely because generalist hosts and parasites are favoured during early stages of coevolution, preventing local adaptation. By contrast, at later stages manipulations had no effect on either average levels of resistance or infectivity, or alter the level of local adaptation relative to the controls, possibly because traits other than resistance and infectivity were under strong selection. Taken together, these data suggest that the relative generation times of hosts and parasites may not be an important determinant of local adaptation in this system.     

Spatial frequency analysis in early visual processing

The existence of multiple channels, or multiple receptive field sizes, in the visual system does not commit us to any particular theory of spatial encoding in vision. However, distortions of apparent spatial frequency and width in a wide variety of conditions favour the idea that each channel carries a width- or frequency-related code or 'label' rather than a 'local sign' or positional label. When distortions of spatial frequency occur without prior adaptation (e.g. at low contrast or low luminance) they are associated with lowered sensitivity, and may be due to a mismatch between the perceptual labels and the actual tuning of the channels. A low-level representation of retinal space could be constructed from the spatial information encoded by the channels, rather than being projected intact from the retina.     

Coordination of hormone-induced calcium signals in isolated rat hepatocyte couplets: demonstration with confocal microscopy.

Excitable cells often display rapid coordination of hormone-induced intracellular calcium signals. Calcium elevations that begin in a single epithelial cell also may spread to adjacent cells, but coordination of hormone-induced signals among epithelial cells has not been described. We report the use of confocal microscopy to determine the inter- and intracellular distribution of cytosolic calcium in isolated rat hepatocyte couplets, an isolated epithelial cell system in which functional polarity is maintained. Both vasopressin and phenylephrine evoked sequential coordinated calcium signals in the couplets, even during cytosolic calcium oscillations. The coupling was abolished by closure of intercellular gap junction channels by treatment with octanol. These observations demonstrate that hormone-induced intracellular calcium signals are coordinated among hepatocytes and suggest that gap junction channels mediate this intercellular integration of tissue responsiveness.