Evaluation of protective immunity of 28KD-GST as a vaccine against experimental Schistosoma Mansoni

Schistosoma mansoni GST was purified from adult worm homogenates by affinity chromatography. SDS-PAGE analysis of the purified antigen revealed that SmGST has molecular weight around 28 KD was used as a vaccine in a dose of 25 and 35 micro g. Eleven groups of BALB/c mice of 10 mice each were vaccinated by GST. Complete Freund's adjuvant [CFA] and BCG vaccine were used as adjuvant. Booster doses were given after 2 and 4 weeks. Two weeks after the last dose of vaccine, mice were challenged by S. mansoni cercariae. Blood samples were taken 7 weeks post infection for detection of IgG1, IgE and circulating antigens. Then mice were sacrificed for histopathological study of the liver. Highly significant [p > 0.001] increase in the mean optical density of IgG1 and IgE in groups vaccinated by 35 micro g GST and CFA was demonstrated. On the other hand, highly significant [P < 0.001] decrease in circulating antigen, grnuloma number and size in the same group

Detection of altered secondary lymphoid tissue chemkine responsiveness in balb/c mice infected with schistosoma mansoni

To determine the extent to which Balb/c mice splenic T cells were affected by S. mansoni infection, this study aims to investigate the ability of the T cells to produce interferon [IFN]-alpha, and their chemotactic ability at 7 weeks postinfection. The splenic T cells were capable of producing levels of IFN-alpha comparable with splenic T cells from naive mice. However, the T cells exhibited altered chemotactic activity, as evidenced by an inability to respond to seconddary lymphoid-tissue chemokine [SLC/CCL21]. Although no difference in chemokine expression was found between the spleens of infected versus control mice, chemokine production was greater in the livers of infected versus control mice. Collectively, these data indicate that Balb/c mice with 7-wk S. mansoni infection possess splenic T cells with altered chemotactic activity and that the alterations may be a consequence of granulomatous response in the liver

Enhancement of fracture healing in crossbred dogs using subcutaneous injection of salmon calcitonin

The impaired fracture healing results in pseudoarthrosis or skeletal deformity that causes functional disability bone is composed of cells and organic matrix [30%], and minerals [70%]. Fracture healing consists of three interrelated phases: inflammatory, repair, and remodeling, to return to original tissue structure. To asses the effect of salmon calcitonin on fracture healing during the mineralization process. Twenty crossbred dogs were used in this study. After a fracture was created at the midshaft of the radius and ulna of the animals and stabilized with an external splint. Ten dogs of them were injected subcutaneously with salmon calcitonin. Blood samples were taken from the cephalic or recurrent tarsal vein before the operation and at 2,4, and 6 weeks after the operation. For histological assessment bone biopsy was taken from the fractured sites at 2,4,6 weeks post-operative from both groups. The radiographic analysis revealed complete union at 6 weeks post fracture fixation in the salmon calcitonin treated group [2]. In the other group the fracture remained unhealed till the 10[th] week post-operative. The serum biochemical analysis showed highly significant increase in alkaline phosphatase in group [2] and non significant decrease in calcium level in the group [1]. The histopathological examination. Declared that the periostium was activated, the osteoblasts appeared early and the formation of osteoid callus was noticed at the 4[th] week. At the 6[th] week complete ossification and formation of the boney specules were observed in the group [2]. While in the group [1] all these observations were delayed 4-6 weeks. Salmon calcitonin accelerates fracture healing and therefore proposes that it is a potent bone anabolic agent for clinical use. It also gives us simple and easier interferences by using of external fixation rather than using of the most massive, complex, and expensive internal fixations for treating the simple, single long bone fractures in dogs. Fracture healing, salmon calcitonin, dogs

Detection of anti-fasciola istypes among poatients with fascioliasis before and after treatment with mirazid

Stool examination using modified Kato thick smear method was performed to detect Fasciola eggs and other parasites. Abdominal pain was the major presenting symptom [87.7%], followed by pallor [83.3%] and fever [16.7%]. Anemia and hepatomegaly were recorded in 77.7% of the patients compared with 27.7% with splenomegaly. Abdominal ultrasonography revealed hepatomegaly and common bile duct dilatation in 77.7% of the patients. Moreover, five cases showed diagnostic olympic game rings. All patients had positive IgG4 levels, 55 cases were positive for specific total IgG and IgG1; whereas only 24 cases had positive IgG2 levels. All negative controls showed no cross reactions. On the other hand, ELISA detecting IgG4 showed the highest specificity [95%], followed by IgG2 [85%] and the least specific test was obtained with the detection of IgG [70%] and IgG1 [65%]. One month after treatment, 91.1% of the patients were completely cured and even after another two-month follow up. In completely cured patients, none of anti- Fasciola isotypes was significantly changed. So, the detection of anti- Fasciola isotypes, especially IgG4, is very specific for the accurate diagnosis of human fascioliasis

Detection of E. histolytica, G. lamblia and Cryptosporidium copro-antigens in stool samples

A double antibody sandwich ELISA technique using a chromatography purified anti sera against E. histolytica, G. lamblia and Cryptosporidium antigens was applied to detect coproantigens of the corresponding parasites in 90 patients. All positive cases were diagnosed by a parasitological examination and proved to have the infection. In addition to the 90 positive cases, 30 age-matched controls were included in the study, of which 20 individuals were infected with other parasites, but not Cryptosporidium, E. histolytica or G. lamblia [acted as an infected control group] and the other 20 individuals with no intestinal parasites [normal control group]. The assay could detect 100% of those infected with both of G. Lamblia and E. Histolytica and 96.6% of patients with Cryptosporidium infection. False positive reactions were detected in three cases using G. lamblia antisera [92.5%], five cases using E. histolytica antisera [87.5%] and two cases using Cryptosporidium antisera [95%]. A direct increase in the mean antigen level was observed with the increasing intensity of infection in the three parasites; so, higher mean OD reading was observed in the heavily infected cases than the moderately infected ones than lighter intensity of infection. Only those in the elder age group [>20 years] infected with E. Histolytica had statistically higher OD readings of the antigen than the middle age group [10-20 years]. On the other hand, no statistically significant difference was observed between the different age groups and antigen level in cases with either G. lamblia or Cryptosporidium

Anti-fasciola IgG isotypes among patients with fascioliasis before and after treatment

Stool examination using modified Kato thick smear method was performed to detect Fasciola eggs and other parasites. Abdominal pain was the major presenting symptom [83.3%] followed by pallor [71.6%] and fever [16.7%]. Anemia and hepatomegaly were recorded in 70% of patients compared to 25% with splenomegaly. Abdominal ultrasonography revealed hepatomegaly and common bile duct dilatation in 70% of patients. Moreover, six cases showed Olympic game rings which are diagnostic. All of patients had positive IgG4 levels, 58 cases were found positive for specific total IgG and IgG1, whereas, only 36 cases had positive IgG2 levels [60%]. All negative control groups showed no cross- reactions. On the other hand, ELISA detecting IgG4 showed the highest specificity [95%], followed by IgG2 [85%] and the least specific test was obtained with detection of IgG [70%] and IgG2 [65%]. One week after treatment, 90% of patients were completely cured. One and three months after treatment, the cure rate was 83.3%. In completely cured patients, none of anti-Fasciola isotypes was significantly changed

Specific circulating immune complexes in sera of children infected with S. haematobium

Schistosomiasis is a major health problem in our country. This work is carried out for detection of specific circulating immune complexes [CICs] in S. haematobium infected children as a trial to evaluate their potential use in immunodiagnosis of the disease and in assessment of disease intensity and morbidity. Sixty seven Egyptian children from El-Minia and Sharkia Governorates were included in this study, 50 of them were infected with S. haematobium [active cases], 5 infected with parasites other than Schistosoma [infected control] and 12 children were parasites free [normal control]. Sera of all cases were examined to detect specific schistosomal circulating immune complexes. Indirect ELISA assay using monoclonal antibody 128C3/3/21 as a coating antibody was used. Forty seven out of fifty actively infected cases had positive circulating immune complexes level yielding a test sensitivity of 94 percent. All of the normal control group had negative CIC level yielding a test specificity of 100 percent. The level of CICs was significantly higher in heavily infected children [those excreting >50 eggs/l0ml urine] when compared with those with light infection [excreting <50 eggs/l0ml urine]. ELISA using highly purified monoclonal antibody appeared to be a specific and sensitive test for detection of schistosomal CICs level in the serum and evaluating the intensity of infection

Sensitive and specific assay for detecting circulating antigens in individuals infected with schistosoma mansoni

Monoclonal antibody 128C3/3/21 in an antigen-capture ELISA was used to detect circulating antigen in individuals infected with Schistosoma mansoni. This antibody recognizes a carbohydrate epitope expressed on the major group of acidic egg glycoproteins and on glycoproteins and glycolipids in all other stages of parasite development. The overall sensitivity of the assay was 78%, with a sensitivity of 100% for patients excreting >100 egg/g feces [EGF] and 71% for those excreting <100 EGF. By increasing the degree of antibody biotinylation, sensitivities of 92.4% overall and 82% for those excreting <100 EGF. A direct increase in the mean level of circulating antigen was found with increasing egg counts. The difference between those excreting >100 EGF [53 individuals] and those excreting <100 EGF [39 cases] was statistically significant [P <0.01]. None of the control sera [23 uninfected individuals and 16 patients infected with other parasites] had circulating antigen level >80 ng/ml. Thus, the test specificity was >99% and its accuracy was 94.7%, the positive predictive value 100% and the negative predictive value 84.8%

Idiotypic regulation in schistosomiasis mansoni

The role of anti-idiotype Ab2 in the regulation of the immune response In Schistosomiasis was studied. Positive Ab2 could be detected among the newborns of infected mothers and experimentally S. mansoni-infected mice. Negative anti- idiotypic antibodies were observed among children above three years and control mice. Increasing levels of anti- idiotypic antibodies were detected among S. Mansoni infected children and adults. Higher mean anti-idiotype A2 levels were found among patients with hepatosplenomegaly than those with intestinal manifestations. The kinetics of anti-idiotype Ab2 among the S. mansoni-infected mice started from the end of the first week after infection up to eight weeks, it was continuous, but with undulating levels. Moreover, the anti-idiotype antibody was able to stimulate lymphocyte proliferation at eight weeks post infection

Evaluation of the levels of circulating carbohydrate antigen and anti-SWAP antibodies in patients with active Schistosoma haematobium infection and its relation to morbidity changes

The immune responses of schistosomiasis and its relation to morbidity changes is important to understand the pathogenesis of this disease. The aim of this study is to evaluate the levels of circulating antigen and anti-SWAP antibodies and its relation to morbidity changes in patients with active Schistosoma haemtobium infection. An antigen enzyme-linked immunosorbent assay [ELISA] employing monoclonal [mAb] 128C3/ 3/21 was used to detect circulating parasite-derived antigen in the sera of 35 of actively infected schistosoma haematobium patients [31 males and 4 females, 5 to 25 years of age] seen in the out patient clinic of Assiut University Hospital. AntiSWAP [soluble adultworm antigen preparation] immunoglobulins IgG1. IgG4 and IgE were performed for 25 of them. Patients were treated with praziquantel [PZQ] and re-evaluated after 1, 3, and 6 months. Changes in morbidity were evaluated using ultrasonographic grading of urinary bladder lesions. It was found that all patients had significantly high levels of circulating antigens in their sera i. e. above the cut-off value. The antigen level fell significantly in the follow cup visits [p<0.001]. Although the mean antigen level was still significantly reduced [p<0.001] at 6 months visit, 16 patients had high mean antigen level and 9 had rising levels of antigenaemia, reflecting reinfection in 6 patients and persistence of infection in the others. On the other hand, all patients had positive ELISA reaction for IgG1 and IgG4, while 5 patients had negative reaction for IgE through the different visits before and after treatment. The decrease in the mean levels of IgG1 and IgG4 were statistically significant only after 6 months of treatment, but the mean levels of IgE showed significant drop at 3 and 6 months of treatment. A significant correlation was found between the circulating antigen and the anti SWAP IgE during the active infection, but no significant correlation was found between the antigen level and IgG1 and IgG4. There was a significant correlation between the level of circulating carbohydrate antigens and morbidity changes of the urinary bladder. On the other hand there was no significant correlation between the anti-SWAP antibodies and morbidity changes. We conclude that ELISA assay for detection of circulating carbohydrate antigen of S.haematobium is valuable and sensitive in diagnosis of active infection, measurement of intensity of infection and detection of reinfection as well as evaluation of the efficacy of treatment. Its level correlates with anti-SWAP IgE during active infection. In addition it correlates significantly with

importance of detecting circulating toxoplasma antigens in human cases

This work was performed on 100 suspected toxoplasmosis cases including 75 females with complicated obstetric histories, 15 children presented with hydrocephalus, retino-chorditis and lymphadenitis and 10 children suffering from leukemia. Also, 40 age-matched controls were included in this study. Serum samples from all patients and controls were examined to detect anti-Toxoplasma IgG and IgM using indirect ELISA. Antiserum against Toxoplasma avirulent strain was prepared in New Zealand white rabbit, then it was used after purification for detecting circulating Toxoplasma antigens in the sera of these studied groups using a double antibody sandwich ELISA technique. Positive anti-Toxoplasma IgG was detected in 45% of the female group, while 19% were IgM positive. According to IgM/IgG ratio, 9 cases were considered as acutely infected-females and 25 as chronically infected ones. The mean optical density of both IgG and IgM among acutely and chronically infected females were found significantly higher than of the control groups. Ten non-leukemic children were IgG and IgM positive cases and were considered as acutely infected patients. On the other hand, all children with leukemia were negative for anti- Toxoplasma IgG and IgM. Eleven out of 19 acutely infected cases [58%] had positive Toxoplasma antigens, whereas only one case [4%] with chronic infection had positive antigen level

Transmission of circulating schistosomal antigens from infected mothers to their newborns

This work tried to answer the question of antigen transfer possibility using a high sensitive assay in 50 mothers and their newborns at birth and 1, 3, and 6 months after delivery. The assay used in this work could detect circulating S. mansoni antigens in 45 infected mothers [90%] with active S. mansoni infection. A significant direct increase in mean antigen levels was found with the intensity of infection evaluated by egg counting [p <0.01]. The clinical stage of the diseased mothers was apparently unrelated to the ELISA test values as no significant relations were observed. Positive antigen levels were detected in 33 newborns [66%] of the 45 positive antigen mothers, then the percentage positivity was directly decreased with the advancement of age, as only 5 infants [10%] had positive antigen levels compared with 0% at 6 months of age. A positive correlation between newborn serum antigen concentration and concentration of antigen in sera of their mothers was obtained

Parasitic causes of hepatomegaly in children

Three hundred children with hepatomegaly were selected and subjected to full clinical and laboratory examinations. Also, serum samples were examined to detect IgG using ELISA against SEA, chromatography purified hydatid cyst antigen, commercially available Toxoplasma antigen, partially purified adult antigen fasciola and second- stage larvae Toxocara canis antigen. IFAT was used to detect IgG against Toxoplasma and T. canis and a commercially available IHAT kit for leishmaniasis was used. Based on immunological assays, 125 cases were suffering from various parasitic infections; 30 cases with schistosomiasis, 26 cases fascioliasis, 18 Toxocariasis, 35 toxoplasmosis, 3 cases hydatidosis and 13 cases mixed parasitic infections. No parasitic causes could be found in 175 cases. It was concluded that parasitic infections should be considered as important causes of liver enlargement in children; and serological methods using purified antigenic fractions are important tools for diagnosis

Detection of pneumocystic carinii in immuno-suppressed rats by different histological stains and immunological assays

Detection of P. carinii in lung sections of 35 immunosuppressed albino rats was evaluated using five histological stains [Toluidine blue "O", Giemsa, Gram's stain, PAS and H and E] and two immunological assays [indirect immunofluorescence [IIF] and indirect immunoperoxidase [IIP]], using polyclonal antibody raised in white New Zealand rabbits against rat P. carinii. In spite of the high sensitivity of the histological stains, they yield insufficient details for easy identification with difficult interpretation. However, Gram and PAS stains provided a better visualization with easy identification of the parasite. On the other hand, both immunological assays allowed an accurate rapid interpretation to the stained slides. The IIP technique, having 100% sensitivity in detecting the precipitated parasite antigens with absolute specificity, seems to be a good diagnostic tool for detecting P. carinii in lung sections

Evaluating the detection of circulating filarial antigen in diagnosis of bancroftian filariasis and filarial hydrocele

Four groups of patients were selected: 16 patients with clinical evidence of obstructive filarial lymphangiopathy without microfilaremia, 12 patients with clinical evidence of obstructive filarial lymphangiopathy with microfilaremia and 9 patients with microfilaremia. Two control groups were also included. Blood films, sera and hydrocele fluid samples were collected from all subjects. Polyclonal antibody against Dirofilaria immitis worm homogenate was prepared, fractionated and conjugated with HRP. Both polyclonal antibody and monoclonal antibody [AD12] were used in a sandwich ELISA. Using polyclonal antibody, both microfilaremic groups [groups 2 and 3] had significantly higher mean OD readings than that of control groups [P <0.05], whereas, the mean OD readings of patients with symptomatic amicrofilaremia had no significant difference than control groups. Symptomatic microfilaremic group had the highest percentage of antigen positivity 7/12 [58.3%] among all groups, while symptomatic amicrofilaremic group had the least antigen positivity 2/16 [12.5%]. Patients presented with elephantiasis only or with hydrocele had no antigen positive levels 0/12 [0%] in their serum or hydrocele fluid samples

Detection of giardia antigen in stool samples before and after treatment

A double antibody sandwich ELISA technique, using a chromatography purified Giardia antiserum, was applied to detect fecal antigen in patients infected with Giardia lamblia before and after treatment. The assay could detect antigens in 98% of infected cases with false positive reactions in 3 cases infected with E. histolytica. There was a significant direct relation between the antigen level in stool samples and the number of Giardia cysts. The mean level of copro- antigen was slightly lower in children, below 10 years, than in older patients, without significant difference. On the other hand, the lowest cyst count was noticed in elder patients, over 20 years. The level of fecal antigens decreased significantly after successful treatment in patients with giardiasis. It was concluded that detection of Giardia antigens by ELISA technique in the stool samples was a highly sensitive [98%] and specific [91%] diagnostic method. It is also, considered as a good monitor for treatment success

Prevalence of fasciola infection among school children in Sharkia governorate, Egypt

This study was performed on 1350 school children from 9 different villages in Sharqia Governorate to investigate the real situation of endemicity of fascioliasis in the area. Stool examination using modified Kato thick smear method was performed to detect Fasciola infection and other parasites. Those with negative stool samples were examined serologically by ELISA test to detect anti-Fasciola IgG. All cases with positive anti-Fasciola IgG were further examined by circum- oval precipitin test [COPT] against viable S. mansoni eggs to exclude the crossly reacted Schistosoma infection. 69 cases were found to pass Fasciola eggs in their stool samples [5.1%]. Anti-Fasciola IgG was detected in the sera of 231 children [17.1%] using ELISA test. 84 out of 231 children were found positive by COPT and were considered as schistosomal cases. The remaining 147 cases who gave negative COPT were considered as Fasciola infections. All of 69 Fasciola positive stool cases were found positive by ELISA test and negative by COPT test. The sensitivity of stool analysis was 47% versus 100% sensitivity of ELISA, whereas the specificity of ELISA was 63%. The total number of Fasciola positive cases by ELISA and stool analysis were 147 cases among 1350 children indicating prevalence of 10.9% among school children in Sharqia Governorate. The results highlighting the importance of health education and snail control in decreasing the high prevalence

Low avidity antibodies in diagnosis of recent experimental schistosomal infection

ELISA test was used to measure both of low- and high-avidity specific IgG against soluble S. mansoni egg antigen, to differentiate between S. mansoni experimentally infected mice with recent and chronic infection. Mice were infected with 100 S. mansoni cercariae and serum samples were obtained and tested 4, 6, 8, 10, 12, 14 and 16 weeks after infection. The results showed that there was a statistically significant difference between low- and high-avidity specific IgG in experimentally infected mice at 4, 6, 8 and 10 weeks after infection [recent infection]. On the other hand, no difference was found in those with chronic infection [12, 14 and 16 weeks after infection]. From the results obtained, it was concluded that the comparison of low- and high-avidity specific IgG levels was able to differentiate between recent and chronic experimentally infected mice with S. mansoni

Humoral and cellular immune responses in experimental trichinosis

This work was designed to study the relation between cellular and humoral immune responses in experimentally infected albino rats with T. spiralis larvae over a period of four weeks. Cell-mediated immune response [measured by lymphoblast transformation test] revealed a marked lymphoblast response during the intestinal phase, but the reaction became irregular with larval encystation in muscles. Humoral immune response was estimated by both of micro- and dot-ELISA techniques. It was obvious that significant higher OD readings and dot color intensities were observed in the muscular phase than in intestinal phase. The results proved that micro-ELISA had the least accuracy [66.6%] and poorest specificity [81.25%], while dot-ELISA showed the best sensitivity [100%] and highest specificity [87.5%]. Lymphoblast transformation was as specific as dot-ELISA