Immunoprotective responses against murine sarcocystosis by ß - Irradiated sporocysts.

This study aimed to induce protective immunity against infection with Sarcocystis muris in experimental mice using ß-irradiated sporocysts. Mice were vaccinated with 50 sporocysts of S. muris which were exposed to 1.84 µSv ß-irradiation for 2, 4 and 8 h. After challenge infection, different samples were taken for evaluation. Serum and intestinal wash were assayed for IFN-γ and IgA, respectively. Mesenteric lymph nodes (MLNs) and spleen were investigated for CD4+ and CD8+ T cells using immunohistochemistry. For liver, the morphological changes in parasitic stages and the count of infiltrated CD8+ T, NK1.1+ and FasL+ cells were also investigated. Real time (RT) - PCR was used for detection of liver MHC I, CD1d, IFN-γ, perforin and FasL as well as the parasite 18S ribosomal(r) RNA in liver and muscle tissues. Alterations of liver parasitic stages as well as a decrease in the infection with the parasite in both of liver and muscle tissues were dependent on radiation exposure time. An investigation for the mechanism of immunoprotection showed an increase in liver NK1.1+ & FasL+ cells, serum IFN-γ and intestinal IgA, while CD4+ and CD8+ T showed a remarkable increase in MLNs and spleen. FasL expression increased in the liver dependently on radiation exposure time, while perforin, MHC I and CD1d were not. ß-irradiated sporocysts with 1.84 µSv for 8 h s could induce the highest protection against infection with Sarcocystis. This could be largely relied on the increased infiltration of NK cells and associated higher expression of FasL in the liver.

Influence of Moringa oleifera extract, vitamin C, and sodium bicarbonate on heat stress-induced HSP70 expression and cellular immune response in rabbits.

The current study aimed to test the effect of Moringa oleifera extract (MOE), vitamin (Vit) C, and sodium bicarbonate (NaHCO3) on heat stress (HS)-induced alterations in rabbits. Five groups of rabbits were designed as control, HS, HS + MOE, HS + Vit C, and HS + NaHCO3. HS groups were exposed to high temperatures, while treatments were given in drinking water for 6 weeks. Levels of blood cortisol, leptin, IFN-γ, TNF-α, and IL-10 were assayed using ELISA, while adrenaline was assayed calorimetrically. Expression of HSP70, FOXP3, T cell receptor (TCR) γ, and δ mRNA was tested using real-time (RT)-PCR, while HSP70 protein expression was tested using western blotting in liver and kidney tissues. Infiltration of regulatory T cells (Treg; CD25+) and NK (CD56+) cells were tested using immunohistochemistry (IHC). The levels of liver enzymes (ALT & AST), urea, and creatinine were assayed calorimetrically, while body weight gain (BWG) and feed conversion ratio (FCR) were calculated. The results showed increased levels of cortisol, adrenaline, leptin, IFN-γ, TNF-α, ALT, AST, urea, and creatinine but decreased IL-10 in the HS group. Increased expression of HSP70 on both mRNA and protein levels was associated with increased NK and γδ T cells versus decreased Treg cell infiltration in liver and kidney tissues of the HS group. In the same group, BWG was decreased, while FCR was increased with respect to the control group. All treatments used in this study reversed the effects of HS significantly. In conclusion, MOE, Vit C, and NaHCO3 can be added to rabbit diets for the amelioration of HS-induced symptoms.

Gene expression, oxidative stress and apoptotic changes in rabbit ileum experimentally infected with Eimeria intestinalis.

Coccidiosis is a parasitic disease caused by protists (apicomplexans) of the genus Eimeria Schneider, 1875 and is considered to be the most important disease faced by rabbit breeders due to its high morbidity. In the present study, the antioxidant status and changes in apoptosis and in the expression of some genes were quantified in rabbits' ilea following infection with Eimeria intestinalis Cheissin, 1948. Rabbits, orally infected with 1 × 105 sporulated oocysts of E. intestinalis, started to shed oocysts in their faeces on 8 days post infection (dpi) and reached maximum excretion on 10 dpi, with approximately 5 million oocysts. This was accompanied by a significant decrease in the live body weight of infected rabbits. Also, malondialdehyde and nitric oxide were significantly increased while catalase and glutathione were significantly decreased in the ileum tissues of the infected rabbits. In addition, a significant increase was observed in the percentages of apoptotic cells in the ilea of the infected rabbits. Furthermore, interleukin-1ß and interleukin-2 mRNA levels were significantly down-regulated and mRNA levels of interleukin-6, interferon gamma and inducible nitric oxide synthase were significantly up-regulated, while those of C-reactive protein remained unchanged. We conclude that infection with E. intestinalis induces oxidative stress, a significant increase in the percentage of apoptotic cells and a diverse and robust Th1 and Th1-related cytokine response in the ileum tissues.

Detection for cross-reactive proteins in filarial worm Setaria equina, MCF-7 human breast cancer, and Huh-7 hepatoma cells.

This study aimed to detect the cross-reactive proteins in filarial parasite adult worm Setaria equina and two different tumor cell lines (MCF-7 human breast cancer and Huh-7 hepatoma cells). This was performed using rabbit anti-S. equina extract (SeqE) or DEC (Diethylcarbamazine citrate) polyclonal IgG antibodies by indirect ELISA and western blotting. The results indicated cross-reactive bands at 70 and 75 kDa in all extracts by anti-DEC and SeqE antibodies, respectively. In addition, the expression of 70 kDa protein was only reduced in filarial worms and Huh-7 after in vitro DEC treatment compared to the control.

A new record of Myxobolus brachysporus and M. israelensis in the tilapia (Oreochromis niloticus) collected from the Nile River, Egypt.

The present study was carried out as part of an ongoing general survey for myxosporean parasites infecting tilapias in the River Nile, Egypt. In the present study, 77 Nile tilapia (Oreochromis niloticus) were collected from boat landing sites at Beni-Suef governorate, Egypt and examined for the myxosporean infection. The infection was encountered as a huge number of free spores in the kidney and the spleen. The infection showed a prevalence of 51.9% (40/77) for Myxobolus brachysporus while it was 25.9% (20/77) for Myxobolus israelensis. Mature spores of M. brachysporus were ellipsoidal and measured 8.6 × 13.2 µm. The polar capsules were subcircular with 5-6 filament turns and measured 4.7 × 3.6 µm. Spores of M. israelensis were ellipsoidal in the frontal view and fusiform in the lateral view. Spore measurements were 13.4 µm long and 8.7 µm wide. The polar capsules were elongated with 6-7 filament coils and measured 8.6 × 3.1 µm. The findings presented here proved that tilapia fishes in the Nile River are still suffering from infections with Myxobolus species. Therefore, further studies should be carried out to survey the Myxobolus infection among tilapias under culture conditions to clarify the pathological impacts of this parasite in tilapias aquaculture.

Re-description of Myxobolus fahmii from the gills of Barbus bynni with new data on the precise infection site, histological impacts, and seasonality.

Originally, Myxobolus fahmii was described from the gills of Barbus bynni collected from the Nile River, Egypt. The original study provided only mensural characteristics of the spores and referred to gill lamellae as the site for the infection. In this study, B. bynni were collected from almost the same locality, with a similar parasite being found. Our investigations enabled us to determine the precise infection site, which corresponded with the "filamental vascular type" and to provide, for the first time, information on pathogenicity and on the seasonal variation in the prevalence of M. fahmii in gills. The infection was recorded as whitish ellipsoidal plasmodia within the afferent artery of the gill filaments. Due to continuous growth of the plasmodia, gill lamellae were atrophied and obliterated at the plasmodium site. Spores were drop to ovoid shaped with a mean length of 11 µm and mean width of 8 µm. Polar capsules were pear shaped, measuring 7 µm in length × 3.0 µm, with 5-7 filament coils. The overall prevalence of infection was 32.9% (79/240). The highest prevalence was in autumn 63.3% (38/60) while the lowest was in winter 6.6% (4/60). The infection showed a significant difference in prevalence between all the seasons except autumn and spring and summer and winter.

Two Myxobolus spp. infecting the kidney of Nile tilapia (Oreochromis niloticus) in the River Nile at Beni-Suef governorate, Egypt, and the associated renal changes.

Two Myxobolus spp. are described from the kidney of the Nile tilapia (Oreochromis niloticus) collected from the River Nile, Egypt. The prevalence of infection was 61 % (47/77), with the infected fish in each case parasitized by the two Myxobolus species simultaneously. The infection was exhibited as free spores in Bowman capsules and renal glomeruli, which makes their original structures difficult to discern. In some cases, the infection appeared as a fibrous plasmodia-like structure containing degenerated developmental stages and spores in the interstitium. The paper identifies each species based on the morphological characteristics of its spores and identifies the histological impacts of Myxobolus infection in this species of fish.

Immunomodulatory effect of diethylcarbamazine citrate plus filarial excretory-secretory product on rat hepatocarcinogenesis.

Diethylcarbamazine citrate (DEC) had a significance in anti-filarial chemotherapy, while excretory-secretory product (ES) is released from adult filarial females. The target of the current study was to examine the immunomodulatory effect of DEC, Setaria equina ES or a combination of them on rat hepatocellular carcinoma (HCC) induced by diethylnitrosamine (DEN). In vitro effect of combined DEC and ES or ES alone on lipopolysaccharide (LPS)-stimulated rat peripheral blood mononuclear cells (PBMCs) was tested through IFN-γ assay in culture supernatants. In addition, single or repeated doses of DEC, ES or DEC+ES have been applied in white albino rats to test the effect on HCC. Levels of IFN-γ and anti-ES IgG antibodies in rat serum were assayed using ELISA. Hemolytic complement activity (CH50) was determined in serum while the concentration of nitric oxide (NO) was assayed in liver tissue. The infiltration of NK cells as well as the expression of MHC Iproliferating cell nuclear antigen (PCNA), inducible NO synthase (iNOS), Bcl2 and p53 were determined using immunohistochemistry. There was a dose-dependent increase in IFN-γ after in vitro exposure to DEC+ES. Repeated ES doses increased NO concentration (p<0.05) and expression of iNOS but reduced CH50 (p<0.001), while repeated DEC+ES doses could increase anti-ES IgG (p<0.01), IFN-γ level (p<0.05) and NK cell infiltration. The same treatments could also reduce the expression of MHC I expression, PCNA, Bcl2 and p53. This study has shown immunomodulatory and protective effects of DEC+ES repeated doses on rat HCC.

Effect of diethylcarbamazine citrate and Setaria equina excretory-secretory material on rat hepatocellular carcinoma.

Diethylcarbamazine citrate (DEC) has been known for its efficacy to eradicate bancroftian filariasis in Egypt and other countries in the world. One of the known effects was to decrease the level of circulating filarial antigen in the patient's serum. The target of this study was to examine the effect of DEC, excretory-secretory (ES) material from the filarial parasite Setaria equina or a combination of both on the status of oxidative stress and pathogenesis of rat hepatocellular carcinoma (HCC) induced by diethylnitrosamine and 2-acetylaminofluorene. This could be tested in vitro using nitroblue tetrazolium reduction test for measuring the level of superoxide anion (O2(•-)) released from rat peritoneal macrophages. For in vivo test, a single dose before induction of carcinogenesis or continually repeated doses with DEC, ES or DEC + ES was used. Exposure of macrophages to ES could lead to a significant decrease (p < 0.01) in O2(•-) release, while DEC (200 µM) could modulate such effect with significant increase (p < 0.05). Pathogenesis of liver cancer and treatment were evaluated using histological investigation, level of antioxidant and liver function enzymes. Repeated ES doses could increase the activity of antioxidant enzymes, especially the catalase enzyme and show a protective effect on liver architecture. DEC could modulate the later effects when combined with ES. No significant effect on the liver function enzymes after treatment was observed. Nuclear factor κB was found to be localized only in the cytoplasm after single and repeated treatments with ES. This study could indicate the effect of S. equina ES as antioxidant against rat HCC, while DEC could modulate such effect when combined with it.

Seasonal fluctuation and histopathology of Henneguya ghaffari (Myxozoa: Myxosporea) infection in the gills of the Nile perch, Lates niloticus, in the River Nile: a new locality record.

Henneguya ghaffari Ali (Dis Aquat Org 38:225-230, 1999), which was originally described in Lake Wadi El-Rayan in the western desert of Egypt, has been discovered in the gills of the Nile perch, Lates niloticus, sourced from the River Nile at Beni-Suef governorate. The species identification was based on the spore morphometry. Of 180 Nile perch, 68 were found to be naturally infected with H. ghaffari (37.7%). A significant seasonal fluctuation in the prevalence was discerned, with the maximum rate occurring in the winter (68.8%) and the minimum rate in the summer (8.8%). The plasmodia of the parasite were evident as white rods, occupying almost a third of the gill filament and with mean dimensions of 0.7 × 0.2 mm. Histological investigations revealed that the present plasmodia were potentially compatible with the intrafilamental type. Infection with H. ghaffari initiated epithelial hyperplasia and curling and atrophy of the respiratory lamellae, which underpin its deleterious effect on the host by decreasing the functional respiratory surface of the gills. The present study concluded that infection with H. ghaffari originated in the River Nile before moving to the new ecosystem of Lake Wadi El-Rayan through drainage water.

Pfeifferinella sp. (Pfeifferinellidae, Apicomplexa) infecting the fresh water snail Pirenella conica light and electron microscope studies.

Coccidian oocysts were proved to be found in 70 of 100 collected Pirenella conica snails, with a natural infection of 70%. It was observed that, Pfeifferinella sp. was transferred between hepatopancreas and small intestine of snail. The prepatent period of Pfeifferinella sp. infecting P. conica snails ranged from 14-18 days and the patent period was reached 50 days (P.I.). Merogony stages were the early stages observed in this study. These stages were observed in the hepatopancreas and in a large clear parasiteophorous vacuole (PV). In snails killed 4 days P.I. immature meronts were measured 12 x 10 µm containing 8 nuclei. Meanwhile, mature meronts with about 6 differentiated merozoites were detected as early as 6 days P.I., and measured 3.1 x 1.4 µm. The earliest gametogonic stages were seen in the intestine of Pirenella conica snails killed 12 days P.I. Microgamonts contained about 4 nuclei and measured 7.9 x6.7 µm. The macrogamonts measured 7.3 x 5.6 µm. Macrogametes were characterized by the presence of the vaginal tube, this tube measured 4.3 x 1.1 µm. Fertilization was occurred in the intestine of the infected snails at 12 days P.I. Zygotes developed into young oocysts after fertilization. Sporogony occurred in the intestine. In the earliest stage, the nucleus of young oocyst was occupied the central position that were observed through the examination of the intestine of infected Pirenella snails at 14 day P.I. These oocysts were found to be colorless and ellipsoid or spherical in shape measured 9.5 x 8.5 µm. The oocyst wall consists of two layers, micropyle and micropyle cap were not observed in these oocysts, and wall forming bodies were arranged at the periphery of oocyst directly under the developed oocyst wall. Sporulated oocyst contains 8 sporozoites filling the entire cavity of the oocyst without sporocyst formation. Fully sporulated oocysts were excreted in the faces of infected snails from 14-18 day P.I., these oocysts measured 9.5 x 8.5 µm, Micropyle was absent and while a residual body was observed.

Evaluation of a 14.5 kDa-Fasciola gigantica fatty acid binding protein as a diagnostic antigen for human fascioliasis.

The aim of the present study was to evaluate the efficiency of 14.5 kDa-Fasciola gigantica fatty acid binding protein (FABP) as a diagnostic antigen for human fascioliasis. 14.5 kDa FABP was isolated from the crude extract of adult F. gigantica worms by ion exchange chromatography followed by gel filtration chromatography and then analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis under reducing condition. Anti-FABP IgG polyclonal antibody (pAb) was generated in rabbits and purified by using sequential use of ammonium sulfate, caprylic acid, and then ion exchange chromatography. Conjugation of purified rabbit anti-FABP IgG with horse reddish peroxidase (HRP) was conducted and used in detecting the coproantigen in the stool and the circulating Fasciola antigen (CA) in the sera of Fasciola-infected patients using sandwich enzyme-linked immunosorbent assay (ELISA). The sensitivities of sandwich ELISA test were 96.43% and 94.74%, while the test specificities were 94.87% and 84.62% for the detection of coproantigen and CA, respectively. The parasitological diagnosis using the Kato-Katz technique revealed 64.29% sensitivity with 100% specificity. The diagnostic efficacy of sandwich ELISA was 95.52% for coproantigen and 87.93% for CA detection. In contrast, the diagnostic efficacy of Kato-Katz technique was 85.07%. It was concluded that 14.5 kDa FABP represented a valuable antigen for the immunodiagnosis of human fascioliasis using sandwich ELISA.

Lambs infected with UV-attenuated sporocysts of Sarcocystis ovicanis produced abnormal sarcocysts and induced protective immunity against a challenge infection.

The present study surveyed the prevalence of natural infection of the sheep esophagus muscle with sarcocysts of Sarcocystis ovicanis and examined induction of protective immunity using UV-attenuated sporocysts. The overall prevalence of natural infection of the sheep was 95%. Infectivity of the collected sarcocysts was confirmed by shedding of sporulated oocysts after feeding infected esophageal tissues to dogs. To induce protective immunity, lambs were immunized 3 times (once a week) with 1.5 x 10(4) sporocysts exposed to UV-light for 30 min (UV-30 group) or 60 (UV-60 group) min and then challenged with 1.5 x 10(4) normal sporocysts at the 3rd week post the 1st vaccination. These lambs showed high survival and less clinical signs of sarcocystosis than normal infected lambs. The attenuated sporocysts produced abnormal cysts; small in size and detached from the muscle fiber. These abnormalities were more obvious in UV-60 group than UV-30 group. Also, the IFN-gamma level and lymphocyte percentage were increased while the total leukocyte count was decreased in the UV-60 group compared with other groups. The high level of IFN-gamma may be an evidence for the induction of Th1 responses which may have protective effect against a challenge infection.