Waterborne viruses associated with repeated abortion.

In this study we tried to find the role of some waterborne viruses in repeated abortion of women. The study includes maternal blood serum and fetal tissue. The serum of full-term delivered women was taken as a control. All collected samples were inoculated on BGM and Hep2G cells to detect entero and Hepatitis E viruses. Enzyme-linked immunosorbent assay was also carried out for IgM and IgG antibodies against HEV in all serum samples. HEV-Ag was determined by dot-ELISA, which used also for enterovirus typing. Reverse transcriptase polymerase chain reaction was used for detection of entero and HE virus RNAs in the collected serum samples. To follow up the source of virus transmission, the wastewater treatment plant which serves the area of samples population was studied at the intake and the final effluent for the presence of hepatitis E virus and enteroviruses with special reference to coxsackieviruses. Wastewater samples were collected for 1 year and for enterovirus concentration the adsorption-elution on nitrocellulose membrane was used and for HEV, two methods of virus concentration were used, urea arginine phosphate buffer (U-APB) and PEG8000. The results of HEV investigation of aborted women sera was 22% for IgG, 3% for IgM, 20% HEV-Ag, and 16% of HEV-RNA by RT-PCR. For fetal tissue, HEV-Ag was detected in 5% of the collected samples. The detected enteroviruses were coxsackieviruses types 2, 3,4 and 5 in all serum samples and wastewater samples. The results showed also, that virus concentration by U-APB is much better than PEG-8000 but not highly efficient.

Chronic liver disease and hepatitis C virus in Egyptian patients.

BACKGROUND/AIMS: This study was designed to evaluate anti-HCV and anti-GOR in chronic liver disease (CLD) caused by HCV alone or with bilharzia. METHODOLOGY: The parameters of hepatitis C virus (HCV) infection among 45 chronic liver disease (CLD) patients are the subject of this study. The samples that were collected included serum, saliva and liver biopsy. For comparison, 44 serum, saliva and liver biopsies were also collected from non liver disease (NLD) patients undergoing surgery at the Gastroenterology Surgical Center, Mansoura University. RESULTS: Screening of antihepatitis C (anti-HCV) with a second generation ELISA test showed that 37/45 (82.2%) sera and 17/45 (37.7%) saliva samples from CLD patients were positive for the presence of anti-HCV (IgG), while, anti-HCV (IgG) was detected among 32/44 (72.7%) sera and 6/44 (13.6%) saliva samples from NLD patients. HCV antigen was detected by immunostaining in the liver biopsy sections of 11/45 (24.4%) CLD and in 6/44 (13.6%) NLD patients. HCV antigen was detected in hepatocyte cytoplasm and nuclei, in some endothelial cells lining the hepatic cell cords, and in some bile duct cells. The serum and saliva samples from both CLD and NLD patients were also tested by ELISA for the presence of anti-GOR to determine the prevalence of autoantibody in HCV infected and non-infected patients. Anti-GOR was detected in 19/45 (42.2%) sera and in 1/45 (2.21%) saliva samples from CLD patients, while in the case of NLD patients, anti-GOR antibodies were found in 7/44 (15.9%) sera and in 4/44 (9%) saliva samples. GOR antigen was detected by an indirect immunoperoxidase stain of liver biopsies. Positive GOR antigen signals were found in hepatocytes but granular cytoplasmic, and extrahepatic localization was also noticed. A correlation between the detection of anti-GOR and anti-HCV revealed that, out of 37 anti-HCV positive CLD patients, there were 19 (51.3%) positive for anti-GOR, while 7/32 (21.8%) NLD patients were positive for anti-HCV and for anti-GOR. CONCLUSIONS: The results of the present study confirm the published anti-HCV high seropositivity among Egyptian CLD patients and point to an autoimmune processes in CLD. The liver biopsy findings stress the presence of HCV antigen in extra hepatic cells as well as in hepatocytes in CLD. Our data confirm that anti-GOR is commonly present in sera from CLD patients and show that anti-GOR are secreted in saliva. Our results showed that saliva can not be used reliably, instead of serum, for the diagnosis of HCV infection or auto-antibodies related to HCV infection, but can be used as a parameter for the evaluation of CLD activity, when repeated sampling is necessary.

Lymphocyte responses to human cytomegalovirus in different groups of patients in Britain and in adults from west Africa and the Middle East.

Antibody prevalence and lymphocyte proliferation responses to cytomegalovirus (CMV) and herpes simplex virus (HSV) were compared in several different groups of patients: genitourinary medical (GUM) patients, hemophiliacs, men with clinical acquired immunodeficiency syndrome (AIDS) and cases of primary CMV mononucleosis, and also in adults in the general population (control subjects) comprising separate groups native to Britain, West Africa, and the Middle East. Among the British control subjects who were positive for CMV IgG, all were also positive against CMV antigen by the lymphocyte transformation test (LTT). However, among those who were CMV IgG-positive in the various groups of patients, 20-86.9% gave positive responses to CMV antigen by the LTT; moreover, 75.7% and 55.5% of the CMV IgG-positive healthy control subjects from West Africa and the Middle East, respectively, gave positive LTT responses to CMV antigen. When the same groups of patients were tested for responsiveness to HSV antigen by the LTT, there was good agreement between a positive result by this test and by serology in all except those with primary CMV mononucleosis (42.8%). Overall, lymphocyte responses to CMV were significantly impaired in healthy, CMV antibody-positive subjects from West Africa and the Middle East compared to similar subjects from Britain.

Assessment of the infectivity of corneal buttons taken from hepatitis B surface antigen seropositive donors.

Sixty one corneas taken from 33 hepatitis B surface antigen (HBsAg) seropositive donors and 20 control corneas taken from 12 HBsAg seronegative donors were tested for the presence of HBsAg using reversed passive haemagglutination (RPHA) and enzyme linked immunosorbent assay (ELISA) and for the presence of hepatitis B virus core DNA (HBVcDNA) using the hybridisation technique in their epithelium, stroma endothelium, and storage media. HBsAg was detected by ELISA in the epithelium of one cornea (1.6%), in the stroma endothelial suspensions of six corneas (9.8%), and in the storage media of five corneas (8.2%). HBVcDNA was detected for the first time in the cornea; in the epithelium of four corneas (6.6%), stroma endothelium of nine corneas (14.8%), and the storage media of five corneas (8.2%). The control corneas were negative for HBsAg, while HBVcDNA was detected in the stroma endothelium of two corneas (10%) and in the media of two corneas (10%). This study confirmed that HBV can be present in the human cornea. Preservation in corneal storage media for up to 6 days could not eliminate the virus from the cornea. The possibility of HBV transmission through corneal transplantation should not be overlooked.

Evidence of a 1985-1987 outbreak of acute and chronic hepatitis in Egypt caused by a mutant hepatitis-B virus detected by spot-DNA hybridization test.

Sera from 65 acute and 113 chronic sporadic hepatitis were screened for serological markers of hepatitis-B virus (HBV) and hepatitis delta virus (HDV) and for HBV-DNA. The enzyme linked immune sorbent assay (ELISA) and dot-DNA hybridization tests were used. Two HBV-DNA probes and their labelling systems (biotin, radiolabelling with 32P and digoxigenin) were compared for sensitivity and specificity. The 65 acute sera had serological parameters of HBV infection in 38 (58%) when all these sera were HBsAg, IgM anti HBcAg positive plus HBeAg presence in 11/38 sera. Some of the acute sera had markers of acute HBV and HDV coinfection in 14 and superinfection in 13. Thus HBV with HDV represented 27 (41.5%) of the acute hepatitis in this study. Correlation of these serological markers with dot-DNA hybridization results showed that serum HBV-DNA was present in 36/38 (94.7%) of the acute HBV infection. In the case of acute HBV+HDV positive antigenemia 4/6 had serum HBV-DNA while 10/21 of acute HBV with anti-deltaV. IgM had serum HBV-DNA. There were four cases that gave HBV-DNA positivity in sera without combination of HBV markers suggesting infection with "mutant" HBV. In the chronic hepatitis sera there were markers of HBV past infection (IgG anti HBc in 63/113 and IgG anti HBs in 36/113). Yet, among these sera there was HBV-DNA positive signals (20/63 and 17/36) respectively. Analysis of some of these HBV markers also suggested infection with "mutant" HBV.

Influence of feeding and starvation on the persistence and transmission of Quaranfil virus by Argas (Persicargas) arboreus (Acari: Argasidae).

Viral antigen was detected in the cytoplasm and in associated membranes of salivary gland acinus cells by indirect immunofluorescence and immunoperoxidase staining. Viral ribonucleoproteins (indicated histochemically by presence of pyroninophilic granules) which had accumulated in the cytoplasm of salivary gland type B (granular) acini of unfed Argas (Persicargas) arboreus Kaiser, Hoogstraal & Kohls were no longer visible 24 h after feeding. Virus in tick salivary glands increased from 300 to 500 plaque-forming units during the brief feeding interval (approximately 1 h), but virus was not detectable by 72 h. Overall salivary gland, ovarian, and synganglion tissue levels of Quaranfil virus decreased in the 96 h after feeding, except for synganglion samples in which virus titers increased during 24 h after feeding. Starvation for 105 d resulted in a sevenfold increase in salivary gland viral content compared with those starved 45 d, whereas synganglion tissue titers for Quaranfil virus became undetectable, and ovarian tissue values were similar to those starved for 45 d. Feeding had a greater effect on viral persistence in tissues for ticks starved 60 additional d (comparing 45 with 105 d) in that no Quaranfil virus was detected in any tissue after 48 h (compared with 72 h). Feeding infected ticks (with short extrinsic incubation) on chicks resulted in a peak of host mortality on days 7 and 8, whereas long extrinsic incubation resulted in sporadic mortality over 20 d of monitoring.

Viral etiology of obscure splenomegaly in Egyptian children.

Fifty seven Egyptian children aged 1.5 to 9.5 years with mild splenomegaly (less than 3 cm below the costal margin) were screened for antibodies against the three common viruses of the Herpes group: Cytomegalovirus (CMV), Epstein-Barr (EB) and Herpes type 1 virus. A group of 57 healthy children were studied similarly. All patients were subjected to a comprehensive laboratory and clinical work up to exclude any hematological, metabolic or malignant etiology for the splenomegaly. Splenic aspirates from five cases were examined histologically and by immunohistochemistry for the antigens of CMV. Only primary or reactivation of CMV might be considered a cause of splenomegaly, as there was a statistically significant increase in the prevalence of IgM antibodies to CMV in the patients compared to normal controls (63% of patients and 19.4% of controls had IgM antibodies, P less than 0.001; 68.3% of patients and 54% of controls had IgG antibodies, P is insignificant). An almost equal proportion of children with and without splenomegaly had antibodies to EB-Viral Capsid Antigen (EBVCA) both IgG and IgM. (28% of cases and 33% of controls had IgM antibodies; 26% of patients and 21% of controls had IgG antibodies). A role of Epstein-Barr viral infection could not be ruled out in these patients. There was a higher prevalence of antibodies to Herpes type 1 virus in asymptomatic controls than in children with splenomegaly. (10% of patients and 43% of controls had IgM antibodies, 10.6% of patients and 38% of controls had IgG antibodies).

Diagnosis of tuberculous meningitis by immunofluorescence and enzyme immunoassay.

Cerebrospinal fluid was collected from 29 patients with tuberculous meningitis, 21 and 7 patients with bacterial and viral meningitis and 5 normal subjects. Pressure, aspect, glucose, protein and cellular content of CSF were studied. Detection of acid fast bacilli in direct film stained by Zeil Neilsen (Z.N.) and fluorochrom (Fl.Ch.) and Culture on Lowenstein Jensen media were done. Then specific immunoglobulin G & M to Mycobacteria were assayed by Immunofluorescence (IF using BCG) and by Enzyme Linked Immunosorbant assay (ELISA) using protein-A of M. Tuberculosis. It was found that diagnosis of M. Tuberculosis by CSF culture was more sensitive than by direct CSF film stained with Z.N. or Fl.Ch. stain (positive in 44.8%, 10.3% and 17.2% of cases respectively). It was noticed that the detection of CSF IgG antibodies was more sensitive than IgM antibodies either by IF or ELISA. By comparing ELISA and IF tests for detection of specific anti-mycobacterial immunoglobulin in CSF, it was clear that the sensitivity and specificity of ELISA was more than IF test. A positive result for antimycobacteria IgG antibodies was obtained in 79.3% and 58.6% of cases respectively (p less than 0.05). None of the CSF of normal controls, bacterial and viral meningitis cases gave positive antimycobacteria IgG by ELISA while 9.5% of the CSF of bacterial and 14.3% of aseptic meningitis cases gave positive results with IF. The sensitivity, specificity and predictive value of the described ELISA test, make it useful for early diagnosis of tuberculous meningitis.

An outbreak of acute haemorrhagic conjunctivitis caused by enterovirus 70 in Jeddah during 1985.

Seventy-five cases of acute haemorrhagic conjunctivitis (A.H.C.) were subjected to full ophthalmic, bacteriological, virological, serological as well as cytological examinations. The majority of cases presented with bilateral conjunctivitis (70 out of 75). Although follicular reaction was the earliest sign, yet subconjunctival haemorrhages were the most constant findings in all the cases. Enterovirus 70 was isolated in 57 cases from conjunctival swabs and in 43 cases from serum samples. Complement flaxation test was positive for enterovirus 70 in 42 cases but negative for coxsackie and adenoviruses. No primary role has been found for bacteria in the pathogenesis of A.H.C. in this outbreak. Cytological examination of conjunctival scrappings showed the characteristics cytopathic effect (CPE) of enteroviruses.

Experimental infection and protection of lambs with a minute plaque variant of Rift Valley fever virus.

The injection of a single dose containing 5 X 10(3) plaque forming units (PFU) of a minute plaque variant of Rift Valley fever virus (RVFV) into two susceptible lambs resulted in no detectable viremia, pyrexia or clinical signs of disease. Immunization with the minute plaque variant induced neutralizing antibody as early as seven days postinoculation; however, no complement fixing antibodies were detected. Lambs immunized in this manner were protected when challenged with an infectious dose containing 1 X 10(3) PFU of wild type RVFV. These data indicate that the minute plaque variant may hold promise as a candidate live virus animal vaccine.

Experimental immunosuppression and infection of chickens with Quaranfil virus.

In ovo hormonal bursectomy of chick embryos was carried out by chorioallantoic membrane inoculation with testosterone propionate at the 18th day of embryonation. At hatching immunosuppression was complemented with sublethal X-irradiation. At 40 days of age the immunomanipulated and control chickens were inoculated subcutaneously with a single dose of Quaranfil virus. It was observed that bursectomy and bursectomy with sublethal X-irradiation prolonged viraemia up to the 20th day following virus infection with a significant reduction of natural haemagglutinins and of virus induced specific antibody response.

Reduced pathogenicity associated with a small plaque variant of the Egyptian strain of Rift Valley fever virus (ZH501).

Variants of Rift Valley fever virus producing plaques in CER cells of four different sizes are described. A plaque-forming unit (PFU) variant forming minute plaques was isolated and purified. Virus derived from this variant was not pathogenic to adult Swiss albino mice by the intraperitoneal (i.p.) route and was less pathogenic than the parent strain (ZH501) to adult Sprague Dawley rats by i.p. route, but produced typical severe liver necrosis in adult Syrian hamsters with intranuclear and intracytoplasmic eosinophilic inclusions. Antigen and antiserum to the minute variant prepared in mice reciprocally cross-reacted with antisera and antigens of the original strain (ZH501) in the complement fixation test. Plaque size of the minute variant remained constant after serial passages in cell culture and in suckling mouse brain. When the minute plaque variant was passaged i.p. in hamsters, virus which formed large plaques in CER cells was recovered from the hamster sera.

Rift Valley fever virus: some ultrastructural observations on material from the outbreak in Egypt 1977.

Rift Valley fever virus isolates from the 1977 outbreak in Egypt were studied at an ultrastructural level. The particles measured 90 to 110 nm in diam. using negative staining and sectioning techniques, with a core component of 80 to 85 nm. The surface of the virions was calculated to be covered by approx. 160 sub-units. The particles were found in smooth endoplasmic reticular systems, which were made up of either multi-tubular complexes, or of a single large vacuole. The majority of these membrane systems were found to be unassociated with Golgi apparatus. Inclusion bodies were found within the host cell nuclei (made up of rods and fine granules) and in the cytoplasm (aggregates of fine or coarse granules). The possible relationship of these structures to virus replication is discussed.