First record of natural infection with Borrelia in ornithodoros (Ornithodoros) savignyi. Reservoir potential and specificity of the tick to Borrelia.

Field and laboratory data provided convincing evidence implicating O. savignyi at Shelateen, Halayeb province as a reservoir and a vector of a specific Borrelia. Using direct immunofluorescence Borrelia infection was detected for the first time in a natural population of O. savignyi in Egypt. The overall infection rate (IR) was relatively high (50.63%) and the infected ticks were capable of transmitting the infection to hamsters during feeding. Infected nymphs maintained borrelial infection transstadially to adults and the transstadially infected stages transmitted the spirochetes to hamsters. However, directly infected females were more efficient than males in transmitting the spirochetes to hamster and the IR of hamsters increased gradually by increasing the number of feeding infected ticks upon each hamster. Transovarial transmission of Borrelia intrinsic to O. savignyi occurred during the first and second gonadotrophic cycle of the infected females. The Borrelia sp. detected in the field collected O. savignyi showed specificity for its own natural tick host species when compared with B. crocidurae isolated from O. erraticus. Although the two tick species acquired, transstadially transferred and transmitted the two spirochetal infections, each tick species failed to transovarially transmit Borrelia isolated from the natural population of the other tick species.

Vector-host-parasite inter-relationships in leishmaniasis. I. The effect of Leishmania parasites on rate of digestion of blood proteins from various vertebrate hosts by the sand fly Phlebotomus langeroni (Diptera: Psychodidae).

Protein digestion in the gut of Phlebotomus langeroni (Nitzulescu) was studied at four subsequent 24 hour intervals post feeding on human, dog (Canis familiaris), rat (Rattus rattus) and turkey (Melagris gallopava) bloods with and without Leishmania infantum or L. major promastigotes. Most of the proteins of the studied blood meals were digested within 96 hours. The percent of blood proteins digested in the first 48 hours was higher than in the second 48 hours in all cases of the studied blood meals except the normal blood of the turkey in which the ratio of the digested blood proteins in the two periods was 1:1. During the first 48 hours, the percent of the digested blood proteins was lower than normal in the presence of L. infantum in case of human and dog blood meals. The reverse was true in case of the rat and turkey blood meals in the presence of L. infantum and in the blood meals from each of the four vertebrate hosts in the presence of L. major. The significance of these findings in considering L. infantum as a natural parasite of P. langeroni in El Agamy focus was discussed.

Vector-host-parasite inter-relationships in leishmaniasis. II. Influence of blood meal from natural vertebrate hosts with and without Leishmania infantum and L. major on the proteolytic activity in the gut of Phlebotomus langeroni (Diptera: Psychodidae).

Proteolytic activity in the gut of Phlebotomus langeroni (Nitzulescu) was studied at four subsequent 24 hours intervals post feeding on human, dog (Canis familiaris), rat (Rattus rattus) and turkey (Melagris gallopava) bloods with and without Leishmania infantum or L. major promastigotes. The gut proteolytic activity increased gradually after feeding to reach a maximum at 48 hours post feeding on any of the 12 studied blood meals. In all cases, the activity declined after 48 hours and almost terminated by 96 hours. In case of normal bloods, the proteolytic activity, at 48 hours post feeding, was the highest in case of dog followed by human, rat and turkey respectively. At this time interval the activity was relatively lower in case of human and dog blood mixed with L. infantum promastigotes than in their respective normal blood. The reverse was true in all other blood meal combinations. Promastigotes were alive and active in fresh gut smears of P. langeroni fed on human, dog and rat bloods mixed with either L. infantum or L. major, throughout the digestion period (1-4 days). They were arrested in P. langeroni within the first day post feeding on turkey blood mixed with either Leishmania species. The results of the present study indicate that the kind of blood meal and the Leishmania species affect the proteolytic activity of P. langeroni. The decrease or increase of the proteolytic activity of P. langeroni has no effect on the survival of Leishmania parasites present in the gut and the kind of blood meal is responsible for their development.

Vector-host-parasite inter-relationships in leishmaniasis. III. Impact of blood meal from natural vertebrate hosts on the survival and the development of Leishmania infantum and L. major in Phlebotomus langeroni (Diptera: Psychodidae).

Phlebotomus langeroni collected from a leishmaniasis endemic focus at Et Agamy, Alexandria, Egypt, were found to have fed on blood from man, dogs (Canis familiaris) and rats (Rattus rattus). The effect of the kind of blood meal on the development and the life-cycle of L. infantum and L. major in laboratory reared P. langeroni was therefore investigated. A membrane feeding technique was used to infect sand flies. Gut smears of infected females were examined immediately after feeding and daily for 16 days. Nectomonads and short promastigote forms of L. infantum or L. major were detected in females fed on human, dog and rat bloods at all intervals. Paramastigotes (infective stage) were present only in females fed on dog blood containing L. infantum or L. major and in those fed on rat blood containing L. major. It is concluded that among the factors influencing the Leishmania-phlebotomus relationship is the natural medium in which the parasite is present in vivo. The blood of the natural reservoir host(s) is the key factor for the development of the infective parasite form in the sand fly and P. langeroni could be considered a potential vector for transmitting L. infantum from dogs and L. major from rats and dogs but not from man. This investigation offers a new concept for the study of interactions among vector, host and parasites in Leishmania transmission.

Vector-host-parasite inter-relationships in leishmaniasis. IV. Electrophoretic studies on proteins of four vertebrate bloods with and without Leishmania infantum or L. major.

Fifty five protein bands with relative mobilities of 8,954 to 245,471 kilo Daltons (kD) were electrophoretically separated from 12 feeding media of blood from 4 natural vertebrate hosts of Phlebotomus langeroni. The feeding media included human, dog (Canis familiaris), rat (Rattus rattus) and turkey (Melagris gallopava) bloods without or with Leishmania infantum or L. major promastigotes. Protein bands were identical among the feeding media of one host's blood but varied in number (24-28 bands) and relative mobilities among the various hosts' blood. Some protein fractions were common among the various hosts blood, others were only present in two or three hosts' blood and some were restricted to one host blood and were unique for each host. This study provides data which may help in understanding why blood from different natural hosts may variably influence the life cycle of Leishmania parasite in the sand fly gut.

Transfection of Na,K-ATPase alpha-subunit: regulation of enzyme abundance.

DNA-mediated gene transfer into mammalian cells was used as a model for investigating the regulation of Na,K-ATPase abundance. Complementary DNA encoding the catalytic alpha 1-submit from rat was introduced into ouabain-sensitive monkey kidney cells, and transfectants were selected by their ability to survive in normally cytotoxic concentrations of ouabain. The overall specific activity of Na,K-ATPase in the membranes of transfectants was not significantly different from that in control cells, suggesting that there was a partial replacement, rather than an addition, of introduced alpha 1 for the endogenous subunit in the functional enzyme. Immunoblotting with specific antibodies confirmed the similarities in overall alpha abundance between control and transfected cells. Hybridization analysis of total RNA, however, revealed a higher abundance of the mRNA encoding total alpha 1 in transfected cells. The results suggest that endogenous and introduced alpha-submit compete for a limited amount of beta, with rapid degradation of unassembled subunits.

Mammalian alpha 1-subunit of Na(+)-K(+)-ATPase does not need its amino terminus to maintain cell viability.

The amino terminus of the catalytic alpha-subunit from Na(+)-K(+)-adenosinetriphosphatase (ATPase) is not conserved among the various isoforms and species sequenced to date, yet it always includes a lysine-rich motif. To investigate the functional role played by this highly charged region, we altered the amino terminus of rat alpha 1 and evaluated the ability of the mutant to sustain cell viability. Nucleotide sequence encoding a 10-amino acid epitope from the human c-myc-oncogene product was substituted for the wild-type sequence encoding the first 31 amino acids of alpha 1. The chimeric cDNA containing the myc substitution was then introduced into ouabain-sensitive monkey kidney cells. Selection in ouabain produced viable colonies, suggesting that the introduced mutant was functional and conferred the ouabain-resistant phenotype of rats, despite the removal of the highly charged region from its amino terminus. Subsequent enzymatic analysis confirmed the presence of low-affinity binding sites for ouabain in the recipient colonies, and immunoblotting revealed the myc epitope on the expressed polypeptides in a membrane fraction. These results suggest that the first 31 amino acids are not required for function of alpha 1 and that the posttranslational cleavage associated with the amino terminus is unnecessary.

Determination and seasonal abundance of mite predators of the house fly eggs in Cairo and Giza, Egypt.

Mite predators on the house fly eggs were collected from manure in two slaughterhouses in Cairo and Giza for 2 successive years. Macrochelidae, Parasitidae and Uropodidae were generally collected in that order of abundance. Macrochelidae was represented by 3 species: M. mascaedomesticae (Scopoli), M. merdarius (Berlese) and Glyptholaspis confusa (Foa). Parasitidae and Uropodidae were represented by P. consanguineus (Oudemans & Voigts) and U. (Fuscuropoda) Marginata (C.L. Koch), respectively. The macrochelid and parasitid species generally attained their highest populations in winter and/or spring from December to May. However, the highest populations of the uropodid species were reached mainly in late spring, summer and autumn from May to October.

Abnormal aggregation accompanies abnormal platelet Ca2+ handling in arterial thrombosis.

The resting levels of cytoplasmic Ca2+ (measured by Quin 2 fluorescence) and dense tubular Ca2+ (measured by chlorotetracycline, CTC, fluorescence) are shown to be higher in platelets from patients with arterial thrombosis than from normal donors. Turbidimetric studies of aggregation of diluted platelet-rich plasma (PRP) at 135 microM Ca2+ showed increased rates of aggregation for patients relative to normal controls. For ADP-stimulated aggregation, increased maximal rates (Vmax) and decreased doses for half-maximal rates were observed. With collagen-stimulated aggregation, patient samples showed only decreased ED50 values relative to normal controls. The changes in these values are linearly correlated with the elevation of resting dense tubular Ca2+ level determined by the calcium-CTC test carried out at 2 mM external Ca2+. For ADP-stimulated aggregation this relationship can be mimicked by pre-incubating normal platelets with subcritical concentrations of the Ca2+ ionophore A23187. These results suggest that elevated cytoplasmic and dense tubular Ca2+ in the "resting state" is a major factor in arterial thrombosis, rendering the platelet more sensitive to the stimulation by physiologic agents.

Membrane-associated proteins of adriamycin sensitive and resistant murine leukemic P388 cells.

We have isolated an 84-fold adriamycin resistant subline, P388/R84, from mouse leukemia P388 cells by serial cultivation in methylcellulose in the presence of increasing drug concentrations. Electrophoresis of detergent soluble fractions of radiolabeled sensitive and resistant cells suggested marked alterations in the protein fractions of 160, 100, 60, 45, and 30 kd. In resistant clones labeled with 125I an increase in 160 and 100 kd proteins was accompanied by concomitant reduction in the 60, 45, and 30 kd proteins. In 35S methionine-labeled resistant cells, similar increases in the 160 and 100 kd components were observed but in contrast to 125I-labeled cells the 30 kd component was also higher. Alterations in surface proteins were confirmed in experiments where the cell extracts were adsorbed to concanavalin A polymers and extracted with 0.26 M methyl-alpha-D-mannopyranoside. Our data confirm earlier reported observations on cell-surface protein changes in cells resistant to anthracyclines and alkaloids.

Synthesis and identification of [76As]arsenic trichloride.

Radiosynthesis of arsenic trichloride (AsCl3) was achieved using 76As as the radionuclide of choice for preliminary investigations. The synthesis required neutron activation of arsenic trioxide (As2O3) followed by reaction with sulfur monochloride (S2Cl2) under a dry inert atmosphere. The synthesized 76AsCl3 was purified by distillation or partially purified by treatment with dry acetonitrile followed by filtration. Identification was accomplished by thin layer chromatography (TLC) and reversed phase high performance liquid chromatography (HPLC). 76AsCl3 was used for synthesis of a dihydrophenarsazine derivative.

Novel calcium channels used for lanthanide ion-induced adrenal catecholamine release.

Catecholamine release by La3+, Ce3+, Pr3+, or Nd3+ from isolated bovine adrenals requires the presence of calcium in the perfusing medium. Calcium dependent, La3+-induced adrenal catecholamine release is enhanced at low pH and only slightly inhibited by pretreatment with the calcium channel blocker, verapamil. Since calcium channels in adrenal medulla used by lanthanides are not effectively blocked by hydrogen ions or by verapamil, they must be different from the calcium channels normally opened by the physiological neuromediator, acetylcholine.

Effect of hydrogen ions on influx and efflux of Ba2+ and Cd2+ in adrenal medulla.

To examine the possibility that enhanced uptake of Ba2+ and Cd2+ might explain the increased secretion induced by these metals at high pH, Ba2+ and Cd2+ were perfused through isolated bovine adrenals at high pH 7.4 and 8.3. Both metals were found in greater concentrations in the tissue after exposure at the high pH. Continued perfusion of adrenals with Locke's solution at pH 7.4 or 8.3 after exposure to the metals showed that Cd remaining in the tissue was not affected by the pH of the washout but residual Ba was greater after washout at the high pH. Catecholamine release was correlated with tissue Ba content regardless of the pH of the washout, but residual Cd was not correlated with amine secretion. It appears that low H+ concentrations increase influx of Ba2+ and Cd2+ into adrenomedullary cells and decrease efflux of Ba from this tissue. Elevated intracellular levels of Ba and Cd probably mediate the enhanced adrenomedullary secretion noted after exposure to the metals at high pH. Barium in the tissue is proportional to secretion, but it appears that only part of the Cd2+ entering the cells is involved in initiating adrenal catecholamine release.

Further studies of hormone-sensitive sodium and potassium transport in red cells from developing chick embryos.

Sodium and potassium transport in the definitive series of chick embryo red cells changes significantly, both qualitatively and quantitatively, during maturation. Sodium efflux and potassium influx consist of three parts: a ouabain-sensitive, a furosemide-sensitive, and a ouabain-furosemide-insensitive component. In chick red cells of most ages, the ouabain-sensitive and furosemide-sensitive parts of the cation fluxes do not overlap. Cation transport in the more mature red cells is increased significantly by epinephrine, whereas cation transport in red cells from younger embryos is stimulated much less. This is a beta-adrenergic effect of epinephrine and is mediated by cyclic AMP. The relative lack of response in younger embryos is not due to the absence of beta-adrenergic receptor or the lack of production of cyclic AMP. Ouabain has no effect on the hormone-sensitive sodium or potassium transport. On the other hand, furosemide nearly completely abolishes the effect of epinephrine. In addition, there is a good correlation between furosemide-sensitive components of both sodium and potassium transport and the epinephrine-sensitive component. Furosemide has no effect on cyclic AMP levels in the presence or absence of epinephrine. This suggests that furosemide may act directly on the cation transport system. In the red cells from younger embryos, furosemide-sensitive units are present but cannot be fully activated by epinephrine. Therefore, the lack of the hormone effect on cation movements in these cells is consistent with the view that the appropriate units are present, but do not respond fully to intracellular cyclic AMP levels.

The subgenus Persicargas (Ixodoidea: Argasidae: Argas). 33. Effect of gamma radiation on first nymphal instar A. (P.) arboreus and adult fertility.

Gamma radiation doses higher than 1,000 rads are lethal to first nymphal instar Argas (Persicargas) arboreus. Only 50% of first nymphal instars receiving 1,000 rads reach adulthood but those receiving 100-500 rads survive normally. Males resulting from irradiated first nymphal instars are almost normally fertile. Female germinal cells tolerate higher radiation doses received in the first nymphal instar stage than in the adult stage. Females resulting from first nymphal instars receiving does higher than 100 rads are less fertile (egg number and percent hatch) than normal and produce F1 larvae of lower than normal viability. Progenies of females irradiated as nymphs apparently inherit lethal genes, which may be useful, if irradiated at critical stages, in tick control.