Biological and biochemical studies for characterization of some Egyptian Trichomonas vaginalis isolates

Trichomoniasis has important medical, social and economical implications regarding its serious potentially associated complications, with the possibility of HIV acquisition and transmission. T. vaginalis is a very complex organism. Studying the variation in some biological and biochemical properties of the parasite can be used for characterization of the parasites. For the characterization of T. vaginalis infecting Egyptian female patients, the growth kinetics of 20 isolates, their pathogenicity, metronidazole susceptibility and electrophoretic protein patterns, were correlated with the recorded clinical manifestations associated with these isolates. Positive samples for T. vaginalis were cultured on modified Diamond`s medium. For growth pattern study, trophozoites were counted for each isolate every 24 hours for seven days. The pathogenicity assay was performed using intra-peritoneal inoculation in mice. The isolates susceptibility to different concentrations of metronidazole was recorded by determining the minimal lethal concentration [MLC] and trophozoites viability. The biochemical variability of the studied isolates was performed using 2-dimensional electrophoresis. A broad experimental variability was recorded among the 20 T. vaginalis isolates. There was a clear relationship between 3 isolates obtained from patients with severe vaginitis and the different parameters studied. These isolates had the highest number [20-25 organisms/HPF] in wet mount of vaginal discharge, and the shortest generation time [6:34-7:31 hours]; they were also highly pathogenic to mice. Only one isolate [no. 5] proved to be metronidazole resistant. The use of the first dimensional native-polyacrylamide gel electrophoresis [Native-PAGE] demonstrated the presence of some differences. The isolates were classified into two groups according to their proteins net charge. All samples in each group were considered as one isolate. However, when the 2-dimensional electrophoresis [SDS-PAGE] was applied, five different groups could be identified according to proteins molecular weights. There is a broad experimental variability among the studied Egyptian T. vaginalis isolates regarding growth kinetics, metronidazole drug susceptibility, degree of pathogenicity, as well as the electrophoretic protein patterns

Congenital trichinellosis in experimentally infected mice

The present study aimed at allocating the time during pregnancy at which transpiacental transmission of Trichinella spiralis [T. spiralis] larvae took place. Swiss albino mice were infected at different durations of pregnancy; five days before mating, on gestation day zero and five days after mating. Furthermore, to study the effect of immunosuppression on transplacental transmission, half of the experimental mice were immunosuppressed using cyclophosphamide drug [Endoxan]. The percentage of infected uteri, embryos and placentas and the mean larval count were calculated on day 8 post infection [PT]. Moreover, the percentage of infected offspring and the mean larval count in their muscles were estimated on day 30 P1. The results of the present study revealed that, transpiacental transmission of T. spiralis could occur in offspring of mice when their mothers are infected before or after pregnancy. This was documented by the presence of larvae in the muscles of offspring. However, the rate of this transmission increased when the mothers are infected at late pregnancy whether they were immunosuppressed or not. The administration of the immunosuppressive drug whether before or after pregnancy increased the rate of infection and the mean larval count in both uteri of mothers and muscles of their offspring. A higher percentage of abortion was demonstrated in females infected before mating in different studied groups

Vaccination trial against experimental trichinellosis using autoclaved trichinella spiralis larvae vaccine [ATSLV]

The autoclaved Trichinella spiralis larvae vaccine [ATSLV] was tested and showed unpredictable effect on the immune system of mice experimentally infected with T. spiralis. The vaccine was given with Bacillus Calmette-Guerin [BCG] as an adjuvant at different durations and by different routes of administration. The best result was achieved by given the vaccine twice intradermally with two-week interval as evidenced by a significant reduction in adult and larval count, as well as reproductive capacity index. Histopathologically, there was a significant reduction in the number of the encysted larvae, which showed degeneration and hyalinization of the cyst wall accompanied by early pericystic fibrosis

Detection of trichinella pseudospiralis antigen in the parenteral phase in experimentally infected mice

Antigen distribution of T. pseudospiralis in muscles was studied at 10 days and 2, 3 and 5 weeks post-infection. Immunocytolocalization of antigen in the muscle was performed by peroxidase antiperoxidase [PAP] and immunogold silver staining techniques. Weak positive reaction of the stichosome and the cuticle was observed at 10 days post- infection which increased to attain its highest level in the fully infective stage larvae, 3 and 5 weeks post-infection. The intensity of staining reaction in the cytoplasm and nucleoplasm of infected host cell and surrounding tissue was the highest at 5 weeks post- infection. The small nuclei of the host cells and the inflammatory cells were unstained

Detection of trichinella pseudospiralis antigen in the enteral phase in experimentally infected mice

The distribution of T. pseudospiralis antigen in tissue sections of the intestine of infected mice was studied at different intervals [[10, 21, 35 hours and 10 days post-infection [PI]]. Immunocytolocalization of the antigen in the intestine was done by peroxidase antiperoxidase staining technique. Positive staining reaction of the enterocytes was noted in all durations examined. The mononuclear cells of the lamina propria were positively stained at 10 hours PI while patchy staining of muscularis mucosa was observed at 10 days PI. The present study revealed intense staining reaction in the stichosome and the cuticle of 10-hour old larvae L1, the intensity of the reaction decreased throughout the worm development, until it almost disappeared at 10 days PI. The gut lining and its occupying substance were positive throughout the enteral phase. Intense staining reaction of the hypodermal cells was observed in L2, L3 and adult stage worm. The genital primordium was weakly stained in the larval stages while the male and female reproductive systems were moderately stained at the adult stage

Modulation of host response during infection with trichinella pseudospiralis using immuno-stimulant drug

Infection of the mouse with Trichinella pseudospiralis is accompanied by pronounced suppression of host inflammatory response. This study examines the effect of infection with this parasite on one major component of cell mediated immunity, delayed type hypersensitivity [DTH] reaction and the effect of administration of immunostimulant drug [levamisole] on host response to T. pseudospiralis infection. Serum cortisol levels and the response of skin test to phytohemagglutinin were used to construct the time table of drug regimen. Three groups were studied; group I [infected mice], group II [received Levamisole on day 6 pi], and group II [received Levamisole on day 14 pi]. Early administration of levamisole restored DTH response to normal and histopathological examination of infected muscles revealed evident collagen deposition around the muscle larvae

Studies on T spiralis and T pseudospiralis larvae recovered from mice immunized with heterologous trichinella antigen

In this study, S3 antigen of T. spiralis and T. Pseudospiralis was given to the heterologous infection in albino mice. The recovered larvae were studied as regards their number, length, and steroscan examination. It was found that vaccination of mice with T. spiralis [S3] antigen induced reduction in T. pseudospiralis larval count by 66.43% and significant reduction in length. Using T. pseudospiralis [S3] antigen reduction of 74.77% in T. spiralis larval count was detected as well as significant reduction in length, scanning electron microscopic [SEM] examination revealed more or less similar finding in both species; loss of integrity of the cuticle, some of the larvae were attacked by granulocytes mainly eosinophils and few larvae appeared completely destructed with loss of normal architecture. A peculiar finding was noticed in case of T. pseudospiralis larvae in form of fine electron dense deposits giving the cuticular surface a mottling appearance

Scanning electron microscopic study of trichinella pseudospiralis muscle phase after modulation of host response using an immunostimulant drug

In a previous study, evident collagen deposition around T. pseudospiralis larvae was noticed after administration of Levamisole to infected mice. The present investigation was undertaken to confirm these findings. Scanning electron microscopic examination revealed multiple clear cystic formations on the surface of the diaphragm muscle. The presence of such structures was seen to be more frequent when the drug was given as early as the 6th day post infection. Membrane like structure was observed around T. pseudospiralis larvae, an interesting finding which was by far not reported before

Cyclosporine A in experimental trichinosis: histopathological study

Cyclosporin A [CsA], a relatively new drug, was previously used in a series of studies on experimental trichinosis. This work was conducted to study the histopathological changes of CsA treatment at different time intervals from T. spiralis infection in mice. The work included study of the effect of the drug on the larvae treated either in vivo or in vitro. The drug was most effective when it was administered on the same day of infection, then when given one week before. The effect was evidenced by diminution both in the number of larvae and in the cellular reactions

Heterophyids: experimental chemotherapy and immunotherapy

Heavy infections with the tiny flukes heterophyids can cause intestinal pain and mucous diarrhea, thus the study of praziquantel [Pzq] and the new drug cyclosporin A [CsA] as antiparasitic drugs, were undertaken in this work, as well as the effect of matecercarial and adult antigens as immunizing agents. To assess the result of the work, the number and length of the recovered heterophyids were studied. Description of the surface by SEM was carried out for the groups; treated by cyclosporin A and immunized by adult heterophyid antigen [third fraction] which gave the highest percentage reduction

Cyclosporine A in experimental trichinosis scanning electron microscopic study

The surface morphology of adult and larvae of Trichinella spiralis after cyclosporin A treatment was studied by scanning electron microscopy [SEM] of fixed, dried and metal coated specimens. The earliest topographic changes revealed in adults after the drug treatment were in form of disorganization of the cuticle, especially between the hypodermal pores which appeared somewhat thickened and irregular. Other changes were in the form of bullae formation covering the whole width of the adult surface. Other specimens showed wrinkling of the outer layer that was seen later to peel off. In few worms, layer outer was damaged, releasing some of the internal organs. Smoothening of the cuticle with obliteration of the grooves between the cuticular ridges were observed as the earliest surface larval changes, ulcer formation which was deeply seated was seen in patchy areas in some specimens. The interaction between the parasite surface and the drug is of significant importance, as this can throw light on the possibilities of its mode of action

attempt to induce immunity against toxocara canis infection in albino mice parasltological studies

This study aimed at preparation and fractionation of the crude second stage Toxocara canis larval antigens and the determination of the optimal dose of the crude and fraction that gave the highest percentage reduction in the larval counts. The crude antigen was prepared from the second stage larvae after hatching of the cultivated larvated eggs obtained from the uteri of adult worms collected from 250 stray puppies. The antigen doses given individually in each group with a total dose of 12.5, 25, 50 and 100 mg protein, respectively. The crude antigen was further fractionated by column chromatography using Sephadex G-200. The dose of each fraction was calculated from the crude optimal dose according to their proportion in the elution volume. All immunized mice were challenged two weeks after the last imunizing dose by 800 larvated eggs. Brain larval count was done 30 days post-challenge to determine the optimal dose giveng the highest percentage reduction. It was found that a total dose of 12.5 mg protein of the crude antigen and the fourth fraction gave the highest percentage reduction in the larval count. These two antigens were further used to study the effect of immunization on humoral antibody responses. This was done by determination of the serum antibody level by the indirect fluorescent antibody test 3, 7, 15, 30 and 60 days post-infection. The antibody responses early and higher in the immunized groups than those of the controls. The maximum antibody level was reached at 15 days post-infection in all groups

Cyclosporine A in experimental trichinosis: parasitological study

During the last few years, antiparasitic effects has been related to the immunosuppressant cyclosporin A [CsA]. This work was planned to study the effect of this drug against different aspects of experimental trichinosis. The work included three experiments. The first experiment was to study the effect of [CsA] on infection when administered to mice at different durations i.e. post infection as well as before and on the same day of infection. It proved to be most effective when given on the same day of infection as evidenced by the significant reduction in adult count, length of female, larval count and index of reproductive capacity [RCI]. However, the effect though still significant, diminished when the drug was given pre- and post- infection. The second experiment was to study the effect of [CsA] on infectivity of the T. spiralis larvae treated either in vivo or in vitro. The results showed that the drug has marked effect in that part of the study as shown by the reduction in all parameters of assessment .The third experiment demonstrated the challenge that followed [CsA] treated primary compared to that of non treated one. Results showed significant reduction in all values. These results evoke ideas for further designs of other treatment and control stratigies for the disease

Effect of cyclosporine A on trichinosis; a histochemical study

The aim of this work was to reveal the possible effect of the drug CSA treatment in experimental trichinosis on some important tissue enzymes; alkaline and acid phosphatases, succinic dehydrogenase and alpha-esterase. This study revealed that the effect of the drug was apparently evident on succinic dehydrogenase enzyme which returns back to the control. The diminished acid phosphatase activity showed the effect of the drug on the T-cells

Partial purification of antigens from heterophyid metacercariae

Heterophyid encysted metacercariae were collected from Tilapia fish. These were used for the preparation of crude heterophyid metacercarial antigen which was then subjected to gel-filtration chromatography using Ultrogen ACA[44]. Three major peaks were obtained with three distinct molecular weights. Fraction I 60,000, fraction II 48,000 and fraction III 22,000 daltons. The relevance and expected further uses of these fractions are discussed in relation to the homologous parasite as well as to other heterologous parasites such as Schistosoma mansoni

Effect of albendazole on enteral and parenteral phases of trichinella spiralis

Albendazole was administered after 24 hrs, 48 hrs, and 72 hrs post infection. The best results were obtained when the drug was given 24 hrs post infection, where no adult worms were found one week later. Larval count was performed to study the effect of the drug on the parenteral phase of infection. Albendazole was given at weekly intervals from the first to the 8th week after infection caused reduction in the larval count

Preliminary study on the role of musca domestica in spreading of toxoplasma infection

IFAT technique was performed on mice previously inoculated per OS by an emulsion of flies collected from different districts of Alexandria during summer months for detection of Toxoplasma antibodies

Histopathological and histochemical studies on experimentally infected hamsters with pygidiopsis genata

Haematoxylin eosin stained sections showed the stunted growth of the villi as well as widening of the intervillous spaces in the regions where the worms were seen lodged. A focal cellular reaction was seen in the lamina propria. The enzymatic activity was markedly reduced in the villi in succinic dehydrogenase, nonspecific esterase and alkaline phosphatase; where as marked reduction in the activity of acid phosphatase was noticed particularly in the glands. Those changes suggest a disturbance in both digestive and absorptive mechanisms