Synthesis and biological activity of 2-thiono-[1H]-5-spirocyclohexylimidazo[4,3-b]quinazolone and 8-azaquinazolone derivatives.

The reaction of 5-spirocyclohexylimidazole-2,4-dithion with one and two mole equivalent of b-aminocarboxylic acid derivatives (2a-b and 5) to give new heterocyclic systems of synthetic and potential biological interest viz. compounds (3a-b, 4a-b, 6 and 7). The structures of the products have established by chemical and spectroscopic evidence. The antimicrobial activity of the synthesized compounds was tested against 10 bacterial and yeast strains.

Metabolic changes concerning the effect of castration on some blood constituents in male rabbits.

This experiment was conducted to determine the effects of castration on some metabolic alterations concerning proteins, lipids and minerals metabolism in both immature and mature rabbits. Twenty male New Zealand White rabbits (10 immature and 10 mature) were used. Blood samples were collected from the immature and mature intact animals (control non castrated). Moreover, the castrated animal groups (immature and mature) were sampled at 2, 4, 6 and 8 weeks after castration. The obtained results revealed that, castration caused a significant decrease in serum total protein, albumin, alpha 1-globulins and alpha 2-globulins concentrations. Meanwhile, gamma-globulins level was decreased temporarily at 2 weeks after castration in mature rabbits. beta-globulins level revealed non significant decrease at 2 weeks after castration in immature rabbits followed by a non significant increase. In mature rabbits beta-globulins level non significantly increased. A significant decrease in serum total nucleic acid concentrations was observed after castration throughout the experimental periods, whereas the value of serum uric acid level markedly increased after castration. Serum lipids concentrations (total lipids, total cholesterol, phospholipids and non esterified fatty acids) were increased after castration. The value of serum trace elements including copper, iron, zinc and manganese concentrations showed a significant decrease after castration. The decrease in serum copper and zinc levels were markedly observed in mature castrated rabbits. Also, serum sodium and potassium concentrations were decreased after castration. Testosterone propionate administration in mature castrated rabbits restored most of the investigated serum blood parameters to the levels of intact non castrated mature rabbit. It could be concluded that castration (deficiency of testosterone) induced marked decrease in protein synthesis and increased protein degradation, but serum lipids markedly elevated after castration in both immature and mature animals. Also, castration caused a marked decrease in some serum minerals and trace elements.

Evaluation of FAST-ELISA for the diagnosis of experimental trichniosis.

In order to compare between FAST-ELISA and ELISA for the diagnosis of experimental trichinosis and study the kinetics of antibody and eosinophilic responses, six New Zeland rabbits were infected orally by Trichinella spiralis larvae. Blood was collected every other day for the first 2 weeks, then weekly for eleven weeks post infection. T. spiralis crude larval antigen was prepared for coating of ELISA plates and FAST-ELISA beads. Blood was examined for eosinophilic count and for serum antibody level by ELISA and FAST-ELISA techniques. The burden of infection was assessed by counting encysted larvae in muscle samples of the infected rabbits. By FAST-ELISA antibodies were detected seven days post infection (P.I.), while with ELISA technique antibodies were detected after 10 days. Both tests detected maximum antibody levels on the 4th week. The eosinophilic count reached its peak by the 2nd week. There was a significant inverse correlation between the mean eosinophilic count and the mean larval count. FAST-ELISA proved to be more sensitive than ELISA in early detection of infection, besides being a simple, fast and sensitive assay for antibody detection against T. spiralis.

Study of factors affecting growth of Leishmania in cultures.

The rate of growth of Leishmania major and L. infantum in El-On's culture media supplemented with human, dog, rat and avian blood was studied in vitro. Rabbit blood was used as a control. The effect of culture with these types of blood on the infectivity of both Leishmania strains to albino mice was also studied. The results showed that a good yield of both L. major and L. infantum parasites can be obtained in culture by using avian blood as substitute for rabbit blood in El-ON's medium. In addition, rat blood gave good results with L. infantum. The morphological forms of L. major and L. infantum on all types of blood supplemented media: elongated promastigotes, spindle promastigotes, paramastigotes and amastigoes were present all through the culture period with variable percentages. The infectivity to experimental animals was not affected by culture of both Leishmania strains on rabbit, human, rat, dog as well as avian blood supplemented media.

Plasma concentrations of lipids and lipoproteins in newborn kids and female Baladi goats during late pregnancy and onset of lactation.

Concentrations of blood lipids and some lipoproteins were investigated in normal female Baladi goats during late pregnancy, parturition and onset of lactation as well as in their newborn kids during the first two weeks of life. A total number of 60 herparinized blood samples was collected from does at 4, 3, 2 and 1 weeks pre-partum, day of parturition and at 1, 2, 3 and 4 weeks postpartum. In addition, blood samples were also collected from their newborn kids during the first two weeks of life (day of birth, 1 and 2 weeks of age). Plasma was separated and analyzed for concentration of total lipid, total cholesterol, triacylglycerols, phospholipids, non esterified fatty acids (NEFA) and some lipoproteins as high density lipoprotein cholesterol (HDL-C) and low density lipoprotein cholesterol (LDL-C). The obtained results revealed that there was a significant decrease in plasma level of total lipids at one week after parturition. Plasma level of triaclyglycerols was significantly higher at 4, 3 and 2 weeks before parturition. This increase became very highly significant at one week before parturition. Meanwhile, plasma phospholipid concentrations showed a significant decrease at 3 weeks before parturition, followed by an significant increase at 2 and 3 weeks after parturition and highly significant increase at 4 weeks after parturition. The concentration of plasma NEFA showed a significant increase at 4 weeks before parturition followed by a very highly significant increase at 2 and 1 week before parturition. On the other hand plasma NEFA was non detected at 2, 3 and 4 weeks post-partum when compared with the value reported at day of parturition. Regarding plasma lipoprotein concentrations the obtained results showed that there was a significant increase in plasma HDL-C level at 2 and 3 weeks after parturition, followed by a very highly significant decrease at the fourth week post-partum. However, plasma LDL-C level showed a significant decrease at 3, 2 and 1 weeks before parturition, followed by a further highly significant decrease at 1 and 2 weeks post-partum. In newborn kids plasma concentrations of total lipids, total cholesterol, phospholipids, HDL-C and LDL-C were very markedly increased at 1 and 2 weeks of age. However, plasma triacylglycerol concentrations showed a highly significant decrease at 1 and 2 weeks of age. The concentration of plasma NEFA showed a very highly significant decrease at 2 weeks of age, whereas, at one week of age plasma NEFA were not detected in comparison with first day of life.

Evaluation of specific immunoglobulin G avidity enzyme linked immunosorbent assay (IgG avidity ELISA) in diagnosis of early and chronic Schistosomiasis mansoni.

This work was performed to study variations of Schistosoma specific immunoglobulins between early and chronic schistosomiasis mansoni in both children and adults. A modified enzyme linked immunosorbent assay (ELISA), based on dissociation of antigen antibody complexes with 8 mol/L urea (8 M urea) solution, was used to measure levels of low avidity immunoglobulin G antibodies (IgG Abs) against Schistosoma mansoni soluble egg antigen (SEA). The study included eighty (80) patients with active mansonian schistosomiasis. They were classified according to age, history, clinical symptomatology and examination and direct parasitological methods of diagnosis into early and chronically infected children and adults. Sera of all patients were subjected to: ELISA measuring Schistosoma specific IgM and IgG., immunoglobulin G (IgG) avidity ELISA and indirect haemagglutination test (IHA). Schistosoma specific IgG avidity ELISA detected higher levels of both urea IgG inhibition percentage and low avidity IgG Abs in early cases of schistosomiasis than chronic ones in both children and adults. Levels of urea IgG inhibition percentage were higher in children than adults. Schistosoma specific IgM/IgG ratio was more than one (> 1) in early cases in both children and adults and less than one (< 1) in chronic cases in children and 70% of adults. IHA titres were statistically higher in chronic cases than early ones in children only. So, it can be concluded that IgG avidity ELISA is a valuable method that helps to differentiate early from chronic schistosomiasis mansoni infection in both children and adults.

Application of neural networks to the real-time diagnosis of acute toxoplasmic infection in immunocompetent patients.

Neural networks constitute a relatively new, radically different approach to the interpretation and recognition of subtle diagnostic patterns in multivariate data. In this study the use of neural networks with a single serum sample for rapid real-time recognition of recent toxoplasmic infection was investigated. A neural-network model was implemented on the basis of data obtained by four serological methods--dye test, indirect fluorescence assay, indirect hemagglutination assay, and IgM immunosorbent agglutination assay--and was "trained" to extract features of acute infection by application to an analysis of 65 immunocompetent patients, 10 of whom were in fact acutely infected. The trained model correctly classified all 10 cases of acute infection. On its application to 61 additional infected patients, this method correctly identified seven cases as potentially acute. Our study shows that neural networks can discern diagnostic patterns from variables that individually have limited utility in the diagnosis of acute toxoplasmosis.

Evaluation of two cultural media (CPLM & TYM) for isolation and maintenance of Trichomonas vaginalis stocks in the laboratory.

Five hundreds vaginal discharge specimens were inoculated simultaneously in 2 axenic culture media (CPLM & TYM), in order to compare their ability to isolate and to maintain the growth of T. vaginalis in the laboratory. While both media were found to be equally good in detecting the organisms in vaginal discharges, yet, T. vaginalis stocks were maintained for a longer time in TYM medium (one year), than in the CPLM medium (2 weeks). The yields of the parasites with different inocula subcultured and after different incubation periods were counted in the TYM medium.

Evaluation of different serological tests in mansonian schistosomiasis and correlation to hepatitis B virus infection.

A total of 70 bilharzial mansonian patients (Group I: 20 early intestinal mansonian patients, Group II: 20 hepatosplenic mansonian patients without ascites, Group III: 30 hepatosplenic mansonian patients with ascites) and 30 normal controls were studied. Using the circumoval percipitin test (COPT), there was a statistically significant difference between sensitivities in Group I and Group III with higher sensitivity for Group I. Using either the indirect haemagglutination test (IHA) or the indirect fluorescent antibody test (IFAT), there was no statistically significant difference between the sensitivities in the different bilharzial groups. No statistically significant difference was found between the 3 tests in Group I, II respectively. A statistically significant difference was found between the sensitivities of COPT and IHA in Group III. A highly significant difference was found between the sensitivities of COPT and IFAT in Group III also. No statistically significant difference was found between the sensitivities of IFAT and IHA in Group III. There was no statistically significant difference between the percentages of positivity of hepatitis B surface antigen (HBSAg) in the different bilharzial groups while each of the 3 groups showed highly statistically significant difference with the control group. No statistically significant difference was found between the positivities of HBSAg and the different titers of either IHA of IFAT in each group.

Detection of Schistosoma mansoni antigen in the liver and antibodies in serum of patients with hepatic schistosomiasis.

Liver biopsies were taken from 28 patients with chronic hepatitic schistosomiasis for histopathological study and S. mansoni antigen detection by indirect immunofluorescence test. Circulating antibodies were estimated by indirect haemagglutination test. Thirteen cases (46.4%) revealed diffuse S. mansoni antigen in the liver blood sinusoids and in the formed granulomata. Circulating antibodies were detected in 21 cases, 11 of them showed antigen in the liver. Five cases only revealed definite granulomatous reactions around bilharzial ova, while the remaining 23 cases showed periportal fibrosis. So, for proper diagnosis of chronic hepatic schistosomiasis, it is recommended to examine the liver for S. mansoni antigen together with circulating antibodies detection.

Serodiagnosis of giardiasis by counterimmunoelectrophoresis and indirect immunofluorescence tests.

Serodiagnosis of giardiasis by counterimmunoelectrophoresis assay (CIEP) and indirect immunofluorescence test (IFAT) were evaluated versus stool examination. Giardia lamblia antibodies were detected in sera of 46.9% and 53.1% using CIEP and IFAT respectively out of 49 giardiasis-infected children (5-10 years) diagnosed microscopically. Among the control group (22 children) 13.6% and 18.2% were positive by CIEP and IFAT respectively. The findings of the study showed that we can not rely on serology to diagnose giardiasis specially in cases with low excretion rates.

Evaluation of an ELISA test in past and present schistosomiasis.

An enzyme linked immunosorbent assay (ELISA) was evaluated in relation to an indirect haemagglutination (IHA) test in schistosomiasis patients who were classified by clinical, sonographic and direct methods of diagnosis. Sensitivities of ELISA and IHA respectively proved to be 100% and 69.23% in acute simple intestinal schistosomiasis; 95.5% and 90.4% in chronic active schistosomiasis patients; 86.06% and 67.41% in patients with past history of exposure; 80% and 64.28% in patients with hepatosplenomegaly with past history of schistosomiasis, and 96% and 80% in patients with hepatic fibrosis as shown by sonar. It was apparent that ELISA is more sensitive than IHA in acute simple schistosomiasis in patients with past history of exposure, those with bilharzial hepatosplenomegaly and those with hepatic fibrosis. Both tests were nearly equally sensitive in chronic active schistosomiasis.

Comparative study of three tests (indirect haemagglutination, direct agglutination, and indirect immunofluorescence) for detection of antibodies to Toxoplasma gondii in pregnant women.

Sera of 600 asymptomatic pregnant women were tested by IHAT for Toxoplasma antibodies. The positive reactors were further tested by IFAT and direct agglutination test (DAT). The prevalence of Toxoplasma antibodies was found to be 27.3% by IHAT, and 58.5% of those found positive were also positive by IFAT, while only 51.8% of them were positive by DAT with 71.34% degree agreement between both tests. So, IFAT was found to be more sensitive than DAT in reference to IHAT and is recommended as a confirmatory test for those found positive on screening by IHAT, DAT, compared to IFAT, gave 69.7% sensitivity and 73.5% specificity. Results support previous findings that IHAT, IFAT and DAT measure different antibodies of Toxoplasma gondii.

Estimation of Toxoplasma gondii antibodies in patients with cardiomyopathy.

The possible involvement of Toxoplasma gondii in the pathogenesis or as a complicating factor in patients with cardiomyopathy has been investigated by the serological detection of specific antibodies by IFAT. Fifty six serum samples were collected from patients who presented with dyspnoea on effort, pericardial pains, palpitations and oedema of lower limbs. They were investigated by Echo, ECG and epidemiological history, and diagnosed as dilated, ischaemic and restrictive cardiomyopathies and myocarditis. Forty serum samples from patients with valvular lesions were examined as a control group. Twelve out of 56 samples were positive at end titre of 1:16 (21.4%). Sera with positive titres were found to occur mostly in patients with ischaemic cardiomyopathy. All samples of the control group were negative.

Investigation of Sarcocystis as a causative agent in cardiac disease.

Sera from 56 cases with cardiomyopathy and myocarditis and 40 cases with different types of valvular diseases were tested, using the indirect fluorescent antibody test (IFAT) for the detection of antibodies to Sarcocystis. None of the cases showed specific reaction where the IFAT was negative at the cut off titre 1:8 in all the cases.

Characterization of Egyptian isolates of Trichomonas vaginalis I. Isoenzyme patterns.

Three T. vaginalis isolates from Egypt were compared for their isoenzyme electrophoretic patterns on cellulose acetate. The enzymes studied were: glucose-6-dehydrogenase (G6PD); malate dehydrogenase (MDH); phosphoglucomutase (PGM); glucose phosphate isomerase (GPI); malic enzyme (ME); hexokinase (HK) and lactate dehydrogenase (LDH). The three isolates shared the same isoenzyme banding patterns of MDH; GPI; HK and LDH. Two of these isolates were similar in their banding patterns of G6PD, PGM and different from those of the third isolate. The latter was similar to one of the two isolates and different from the other in the ME isoenzyme patterns.

Characterization of Egyptian isolates of Trichomonas vaginalis: I. Serotyping.

Antitrichomonal hyperimmune sera against T. vaginalis stocks isolated from Egyptian female patients were employed for serological differentiation of somatic and soluble antigens in the Ouchterlony gel double immunodiffusion technique. It was concluded that soluble trichomonal antigens present in association with living flagellates are stock--specific reacting with some, but not all the antitrichomonal hyperimmune sera, while those present in association with dead parasites are common antigens reacting with all the sera. Three stocks, E1, E2 and E3 could be differentiated into two strains using their stock--specific antigens. The somatic antigens of six trichomonal stocks reacted with all the hyperimmune sera denoting common antigenic make up.

Evaluation of a direct agglutination test and methylene blue dye test in diagnosis of cutaneous leishmaniasis.

The usefulness and sensitivity of a direct agglutination test (DAT) in the diagnosis of cutaneous leishmaniasis infection has been investigated. Trypsin treated, formalin fixed and coomassie blue stained Leishmania promastigotes were used as antigens: L. infantum, L. donovani, L. aethiopica. Although the titres of sera from patients with localized cutaneous leishmaniasis were low, sera from lepromatous, tuberculous and toxoplasmic patients gave high titres indicating cross reactivity. Comparable results were obtained when the same sera were tested using freshly prepared antigen or antigens stored for 5 months at 4 degrees C, and with addition of 0.78% 2-mercaptoethanol to the diluent.

Naturally occurring toxoplasma antibodies in serum and milk of lactating women.

Serum and milk of lactating women were tested for toxoplasmosis using specific-IgG IFAT. Apparently healthy 70 women were selected: 54 from rural and 16 from urban areas. Serum and milk were simultaneously collected from each one. Sera were positive in 22 (31.4%) of the total 70; including 16 (29.6%) and 6 (37.5%) of rural and urban groups respectively. No statistical significant difference was found for positivity and titre levels between the two groups (P greater than 0.05). Milk was positive in 12 (17.1%) of the 70 women; including 10 (18.5%) and 2 (12.5%) from rural and urban groups respectively, having no statistical significant difference (P greater than 0.05). Comparing serum and milk for positivity and titre levels, also there was no statistical significant difference (P greater than 0.05). It is concluded that relatively low antibody levels in serum could be excreted in milk and may be protective for suckling babies. Occurrence of antibodies in serum and milk are homogeneously distributed between rural and urban inhabitants.