Thin-layer chromatography of lipids in adenovirus-treated human erythrocytes

Summary Lipid changes of human erythrocytes treated with adenovirus type 8, 9 and 10 have been studied with thin-layer chromatography. It has been established that each of the virus types induced several qualitative changes, which were manifested either in the disappearance of spots or in the development of new ones, as well as in the changes of Rf values and staining characteristics. These changes varied with the individual types and were different from those observed with influenza virus. The decrease of aminophosphatides and other amino acid-containing lipids characterized the effect of type 8 adenovirus. Erythrocytes treated with type 9 adenovirus displayed the highest number of spots, while the decrease of unsaturated lipids and neuraminic acid characterized the effect of type 10 adenovirus.     

Thin-layer and liquid column chromatographic analyses of the lipids of adultOnchocerca gibsoni

Lipids were extracted from adultOnchocerca gibsoni with chloroform/methanol and the total lipid content was characterized. Glycolipids were isolated from other lipid classes by Florisil column chromatography and were then fractionated by DEAE-Sephadex ionexchange chromatography. HPTLC revealed the presence of 9 neutral glycolipid bands and of 15 acidic glycolipid bands that stained for sialic acid with resorcinol. Lipids that contained no carbohydrates were analyzed by a combination of TLC and amino column chromatography. Triacylglycerols, cholesterol, cholesterol esters, and free fatty acids were found to be major components of the neutral lipid fraction, and diacylglycerols and monoacylglycerols were minor components. Phosphatidylethanolamine and phosphatidylcholine were the predominant phospholipids. Phosphatidylserine, phosphatidylinositol, sphingomyelin, lysophosphatidylcholine, and lysophosphatidylethanolamine were also present in significant amounts, whereas only traces of cardiolipin and phosphatidic acid were detected. Several minor lipids and phospholipids remained unidentified. These results indicate that adultO. gibsoni have a nonglycosylated lipid composition resembling that of other parasitic nematodes as well as a substantial repertoire of glycolipids, including many with the characteristics of gangliosides.     

Histochemical localisation of FMRFamide-gated Na+ channels in Helisoma trivolvis and Helix aspersa neurones

FMRFamide-gated Na⁺ channels of molluscan neurones belong to the ENa/Deg family of channels which have diverse functions. FMRFamide (Phe-Met-Arg-Phe-NH₂) Na⁺ channels were detected electrophysiologically in specified neurones of Helix (Helix aspersa) and Helisoma (Helisoma trivolvis), and clones (FaNaCs) subsequently identified. We have now made a study to determine the distribution of mRNA for the clones HaFaNaC (Helix) and HtFaNaC (Helisoma) in the nervous systems of these species using standard in situ hybridization techniques. Immunohistochemical experiments were also made using an HtFaNaC antibody to detect the channel protein in Helisoma neurones. Many neurones in the central ganglia, including those which exhibit the FMRFamide Na⁺ current, stained for FaNaC-mRNA, suggesting a much wider distribution of the channel than was indicated by the earlier work. An immunoreactive response to the channel antibody was also observed in some Helisoma neurones, such as the giant dopamine neurone of the left pedal ganglion, also shown to possess HtFaNaC-mRNA and to exhibit the FMRFamide Na⁺ current. Taken together, these experiments suggest that the clones HaFaNaC and HtFaNaC are major, if not the only, subunits of the FMRFamide-gated Na⁺ channel detected electrophysiologically in the identified neurones of these species. However, fewer neurones in Helisoma reacted with the HtFaNaC-antibody than those which exhibited message for the channel. This discrepancy may be due to a difference in sensitivity of the two techniques, or because not all of the channel mRNA is normally expressed as a membrane protein.     

Maintenance of the life cycle of Echinostoma trivolvis (Trematoda) in dexamethasone-treated ICR mice and laboratory-raised Helisoma trivolvis (Gastropoda)

 Echinostoma trivolvis is a ubiquitous 37-collar-spined echinostome found in aquatic birds and mammals and in the planorbid snail Helisoma trivolvis. This echinostome has not been cycled continously in the laboratory. The present report provides details on the continuous life cycle of E. trivolvis in dexamethasone-treated ICR mice and laboratory-raised H. trivolvis snails. Previous attempts to obtain patent adult of E. trivolvis in mice hosts failed because of worm rejection within 2 weeks of infection. ICR mice infected with encysted metacercariae and injected with 2 mg/kg dexamethasone daily for 28 days yielded gravid worms that produced 250–500 eggs/worm at 21 and 28 days postinfection (p.i.). Miracidia derived from these eggs or eggs containing fully developed miracidia were capable of infecting 3- to 5-mm shell-diameter, laboratory-reared H. trivolvis snails. These snails released cercariae by 35 days p.i. Cercariae encysted in the kidney-pericardium of the snails. Encysted metacercariae could be excysted in vitro in an alkaline trypsin-bile salts medium or in vivo when fed to domestic chicks. Received: 13 May 1996 / Accepted: 25 June 1996     

Effects of various larval digeneans on the calcium carbonate content of the shells of Helisoma trivolvis, Biomphalaria glabrata, and Physa sp.

The calcium carbonate concentrations in the shells of Helisoma trivolvis and Physa sp. naturally infected with larval trematodes and Biomphalaria glabrata experimentally infected with larval trematodes were analyzed quantitatively. The larval trematode-snail relationships studied were H. trivolvis infected with larval Echinostoma trivolvis and Physa sp. infected with various larval digeneans, and B. glabrata infected with Echinostoma caproni or Schistosoma mansoni. The calcium carbonate concentrations of the shells of infected snails and uninfected cohorts and of the water in which the snails were maintained were determined by ion exchange chromatography. No significant differences in the calcium carbonate concentrations of shells of infected versus uninfected snails were found. The shells of B. glabrata infected with E. caproni contained significantly less calcium carbonate than the shells of uninfected B. glabrata. The hypercalcification hypothesis, i.e., larval trematodes induce an increase in the calcium concentrations in the shells of their snail hosts, was not upheld in any of the snail-larval digenean systems studied herein.     

Chemoattraction and penetration of Echinostoma trivolvis and E. caproni cercariae in the presence of Biomphalaria glabrata, Helisoma trivolvis, and Lymnaea elodes dialysate

A petri-dish bioassay was used to study the chemoattraction and penetration of the cercariae of Echinostoma trivolvis and E. caproni in the presence of snail dialysates from Helisoma trivolvis (Pennsylvania and Colorado strains), Biomphalaria glabrata, and Lymnaea elodes. Significant chemoattraction was seen with E. trivolvis cercariae in the presence of all snail dialysates released from nonperforated dialysis sacs with a molecular-weight exclusion of 12,000. Under the same conditions, E. caproni was significantly attracted to B.␣glabrata and H. trivolvis (CO strain) but not to L.␣elodes or H. trivolvis (PA strain). Dialysis sacs were perforated with needles to allow the release of snail substances of all molecular weights into the bioassay. Cercariae of both species were significantly attracted to all snail dialysates released from perforated sacs. Moreover, cercariae entered these sacs and penetrated the snails, and 24 h later the percentage of cysts per snail species ranged from 70% to 83% for E. trivolvis and from 73% to 93% for E. caproni. Dialysates released from intact sacs were extracted in choloroform-methanol (2:1) to obtain hydrophilic and lipophilic fractions. When these extracts were placed on agar plugs in the bioassay, the lipophilic fraction, but not the hydrophilic fraction, was mainly chemoattractive. Received: 9 July 1996 / Accepted: 9 September 1996     

High-performance thin-layer chromatographic analysis of neutral lipids and phospholipids in Biomphalaria glabrata patently infected with Echinostoma caproni

This study examined the effects of larval trematode infection on the neutral lipid and phospholipid content of Biomphalaria glabrata patently infected with the daughter rediae of Echinostoma caproni. Uninfected snails were used as matched controls. As determined by qualitative high-performance silica gel thin-layer chromatography (HPTLC), the major neutral lipids present in the whole bodies and digestive gland–gonad complexes in both snail populations were free sterols, free fatty acids, and triacylglycerols, and the major polar lipids were phosphatidylcholine and phosphatidylethanolamine. Quantitative analysis by HPTLC with visible and UV scanning reflectance densitometry showed no significant differences in the concentrations of these lipids in whole bodies of infected snails vs the controls, but the concentration of triacylglycerols in the infected digestive gland–gonad complex was significantly less than that of the uninfected. No qualitative differences in neutral lipids and phospholipids in shell or plasma samples were found between infected vs uninfected snails.     

Rapid expulsion of the intestinal trematodes Echinostoma trivolvis and E. caproni from C3H/HeN mice after infection with Nippostrongylus brasiliensis

 The number of goblet cells in the small intestines of C3H/HeN mice increased rapidly following their infection with about 500 third-stage larvae (L₃) of the intestinal nematode Nippostrongylus brasiliensis. The number of goblet cells reached its peak on day 9 postinfection (p.i.). Worm burdens in the hosts’ small intestines were determined following a challenge infection with encysted metacercariae of the intestinal trematodes Echinostoma trivolvis or E. caproni on days 8 and 16 after primary infections with N. brasiliensis. All metacercariae of E. trivolvis or E. caproni used to challenge the hosts on day 8 p.i. were expelled. Considerable numbers of E. trivolvis (48.6%) and E. caproni (67.1%) remained in the intestines of hosts challenged with these echinostomes on day 16 p.i. All the controls used for E. trivolvis and E. caproni infections without primary infections with N. brasiliensis showed recovery rates greater than 70%. An enzyme-linked immunosorbent assay (ELISA) showed that the IgM titer rose remarkably and plateaued on day 11 p.i. No marked rise in the IgG or IgA titer occurred during the experiment. These results indicate that mucins increased by hyperplastic goblet cells associated with primary infections with N. brasiliensis are responsible for a rapid expulsion of the worms of the challenge infection with E. trivolvis or E. caproni from the mouse host. Received: 10 January 1996 / Accepted: 10 April 1996     

New approach to molecular characterization of Paramphistomum cervi and Carmyerius gregarius and comparative analyses with selected trematodes

Parasitology Research - The two ruminant parasites, Paramphistomum cervi and Carmyerius gregarius, were collected from fresh-slaughtered native cattle at local abattoirs in Sadat district, Menoufia...     

Fatty acid composition ofEchinostoma trivolvis (Trematoda) rediae and adults and of the digestive gland-gonad complex ofHelisoma trivolvis (Gastropoda) infected with the intramolluscan stages of this echinostome

Gas-liquid chromatographic studies were done to determine the fatty acid composition of the digestive gland-gonad (DGG) complex ofHelisoma trivolvis snails infected with the intramolluscan stages ofEchinostoma trivolvis, of rediae freed from the DGG, of uninfected DGG, and of 41-day-old adult worms grown in golden hamsters. The DGG of infected snails showed significantly higher levels of stearic acid (180), hexatrienoic acid (163n-4), and docosahexanoic acid (226n-3) than that of uninfected snails. However, the DGG of uninfected snails showed significantly higher levels of 202 non-methylene-interrupted diene (NMID) and adrenic acid (224n-6) than that of infected snails. The profiles of other fatty acids were remarkably similar in both infected and uninfected snails. Adult worms showed significantly higher amounts of numerous saturated fatty acids and dienes as compared with the rediae. However, the rediae showed significantly higher amounts of certain monoenes and trienes as compared with the adults. Fatty acid differences between rediae and adults probably reflect differences in either the available lipid pools in the immediate host sites or the metabolic activity of each stage of this echinostome.     

Light microscopic study of four plagiorchiid trematodes infecting marine fish in the south-eastern Mediterranean Sea,Alexandria City,with descriptions of two new species

Parasitology Research - During the present investigation, a total of 220 fish specimens belonging to three different species, namely, little tunny Euthynnus alletteratus, African snook Lates...     

Thin-layer chromatography of trinitrophenyl derivatives of amino acids in urine and plasma

The preparation and separation by thin-layer chromatography of the trinitrophenyl derivatives of amino acids in plasma and urine is described. The semi-quantitative procedure presents a useful addition to existing methods for screening biological fluids for aminoacidopathies.     

Replication of Crimean-Congo hemorrhagic fever virus in four species of ixodid ticks (Acari) infected experimentally

The vector potentials of Hyalomma dromedarii Koch, H. impeltatum Schulze & Schlottke, H. truncatum Koch, and Rhipicephalus appendiculatus Neumann for Crimean-Congo hemorrhagic fever (CCHF) virus (IbAr 10200) were evaluated by intracoelomic inoculation. All three Hyalomma species became infected; infection rates ranged between 80 and 100% at 7-14 d after inoculation, and viral titers increased in unfed specimens almost 100 times above inoculation levels within the first week following infection. Only 40% of the R. appendiculatus became infected, and viral titers of unfed specimens increased less than 10 times above inoculation levels. The virus persisted to 153 d in unfed H. impeltatum. Viral titers were significantly higher in female than in male H. dromedarii, H. impeltatum, H. truncatum, and R. appendiculatus after blood feeding. Blood feeding had little effect on the viral titers of male Hyalomma species. However, the percentage of female and male ticks from which virus was recovered was significantly higher from fed ticks compared with unfed ticks. No virological evidence of transovarial transmission was found in greater than 78,000 first-generation progeny (larvae, nymphs, and adults) of inoculated female H. dromedarii, H. impeltatum, H. truncatum, and R. appendiculatus. All species transmitted CCHF virus to guinea pigs when allowed to feed at both 6 and 21 d after inoculation.     

Effects of Echinostoma caproni miracidia dose on the neutral and polar lipids of Biomphalaria glabrata as determined by high-performance thin-layer chromatography

The effects of a 5 versus 25 miracidia exposure of Echinostoma caproni on the lipid composition of Biomphalaria glabrata was studied using high performance thin layer chromatography (HPTLC)-densitometry. A 50 miracidia dose was not used because such a high level of exposure caused severe snail mortality by 3 weeks post-exposure (PE). Lipids were determined in the digestive-gland gonad complex (DGG) of the exposed snails and in the uninfected matched controls at 2 and 4 weeks PE. Extraction of lipids from DGGs was carried out by the Folch method with chloroform-methanol (2:1), and extracts were analyzed on Analtech HPTLC-HLF pre-adsorbent silica gel plates with measurement of separated bands using a CAMAG Scanner 3. For neutral lipids the mobile phase was petroleum ether-diethyl ether-glacial acetic acid (80:20:1) and the detection reagent was 5% ethanolic phosphoric acid, and for polar lipids chloroform-methanol-deionized water (65:25:4) mobile phase and 10% cupric sulfate in 8% phosphoric acid detection reagent were used. No significant differences in the concentrations of free sterols, free fatty acids, triacylglycerols, phosphatidylcholine, and phosphatidylethanolamine were seen at 2 weeks PE in any of the groups. At 4 weeks PE, the free fatty acid concentration increased significantly in the snails exposed to 25 miracidia compared to that of the 5 miracidia/snail group or the controls. Elevation of the free fatty acid fraction in the high dose snail group suggested that some changes occurred in the lipid metabolism of the snails in that group as a function of miracidia dose.     

Levels of liver enzymes and urea in rats naturally infected with larval forms of Taenia taeniformis

The aim of this study was to investigate the changes in the levels of liver enzymes and urea associated with an outbreak of cysticercosis (Taenia taeniformis) in rat liver. At the end of a previous trial, the animals were euthanized and necropsied when cysts of T. taeniformis were found. The number of cysts ranged from ten to 30 per rat liver. Blood samples were collected from ten rats with cysticercoids (from 12 to 22 cysts) and from ten non-affected rats that were kept in another animal house. Alanine aminotransferase (ALT), aspartate aminotransferase (AST), and urea values were reduced when compared with non-parasitized animals; alkaline phosphatase (ALP) and gamma-glutamyltransferase (GGT) values were increased. Since the current experiment had to be repeated due to hepatic impairment evidenced by reduced ALT, AST, and urea values and increased ALP and GGT values, this study aims to alert the scientific community to the importance of sanitary barriers in animal housing.