Molecular characterization of human T-lymphotropic virus, type 1 (HTLV-1) found in Kuwait: close similarity with HTLV-1 isolates originating from Mashhad, Iran.

Human T-lymphotropic virus, type 1 (HTLV-1) infection was detected in two unrelated Kuwaiti patients with tropical spastic paraparesis (HAM/TSP) and in the asymptomatic mother of one of them. The family roots of these patients were traced to the Najaf region of Iraq. The DNA sequence of three PCR-amplified fragments (env, 512 bp; pol, 140 bp; LTR, 704 bp) was determined for each of Kuwaiti HTLV-1 isolates (KUW-1,2,3). All three Kuwaiti HTLV-1 were identical in env and pol fragments and virtually identical in LTR. Two rare substitutions were found in the env and pol fragments. They were shared only with two isolates from Reunion Island (substitution in env), and two isolates from India and the Caribbean (substitution in pol). The sequences of env and pol fragments of the Middle Eastern HTLV-1 isolates were not available. However, the comparison of Kuwaiti isolates with representative Middle Eastern HTLV-1 was possible for the LTR fragment. The phylogenetic analysis of LTR sequences of KUW and 34 other HTLV-1 isolates has shown that Kuwaiti HTLV-1 belongs to a cosmopolitan "a" subtype of HTLV-1 and tends to cluster together with HTLV-1 originating from the Mashhad region of Iran. These results suggest that common origin of Mashhadi and Kuwaiti (Najafi) HTLV-1 and the possibility of another pocket of HTLV-1 infection in the Middle East, located in the Najaf region of Iraq.

HIV type 1 subtypes B and C from new regions of India and Indian and Ethiopian expatriates in Kuwait.

PIP: HIV-1 subtype B isolates have previously been described in India only in the state of Andhra Pradesh, while subtype C isolates have been reported as widespread in the Bombay and Goa regions of India. Gag subtype was determined in HIV-1 isolates from six Indians and one Ethiopian. One Indian was a native of Goa residing in Kuwait, and the others were natives of Bihar, Haryana, West Bengal, and New Delhi states. Five subjects were males aged 20-26 years. The remaining two subjects were females aged 34 and 40. Four of the men acquired HIV through sexual transmission; the other man was presumably infected through contaminated blood. Six isolates were identified as subtype C and one as subtype B. These preliminary findings obtained by arranging the HIV-1 gag sequences according to their similarity score were confirmed by cladogram and nested analysis. HIV-1 subtype C isolates are therefore present in Bombay and Goa as well as in other regions of northern and eastern India. The subtype has also infected Indian and Ethiopian expatriates living in Kuwait.^ieng

Polymerase chain reaction assay for detection of human cytomegalovirus.

Direct detection of human cytomegalovirus (HCMV) from clinical specimens was examined by using the polymerase chain reaction (PCR) for amplifying HCMV DNA. The efficiency of the amplification reaction was examined by using three different buffers and concentrations of deoxynucleotide triphosphates. The PCR assay was most efficient with a reaction mixture containing 17 mM ammonium sulfate, 67 mM Tris hydrochloride (pH 8.5), 7 mM MgCl2, 10 mM 2-mercaptoethanol, 170 micrograms of bovine serum albumin per ml, and each deoxynucleotide triphosphate at a final concentration of 1.5 mM. After 35 cycles of amplification, 0.15 fg of a plasmid containing the cloned target gene (corresponding to approximately six gene copies) was detected. The PCR assay correctly identified all of 24 clinical isolates of HCMV. Virus in urine specimens could be disrupted by heating at 93 degrees C for 30 min. The viral DNA was amplified directly from 5 microliters of preheated urine, with no further treatment before amplification. We tested the PCR assay on urine specimens from patients who had undergone renal transplantation that had been screened for the presence of HCMV by enzyme-linked immunosorbent assay, hybridization assay, and direct virus isolation. Specimens that were positive by one or more of these assays were screened by PCR. HCMV was consistently detected by PCR in all specimens that were positive by at least one other test. No cross-reactivity to other herpesviruses or MRC-5 cellular DNA was observed.

Hepatitis C virus infection in patients on maintenance dialysis in kuwait: epidemiological profile and efficacy of prophylaxis.

Data on hepatitis C virus (HCV) infection in patients undergoing maintenance hemodialysis (HD) in Kuwait were collected retrospectively in December 1994. Ninety three of 232 patients (40%) studied had hepatitis C antibodies (anti-HCV) when tested by a second generation enzyme linked immuno-sorbent assay (ELISA-II). Since October 1992, all HD patients who tested positive for anti-HCV were dialysed on separate machines and blood transfusions were limited to acute life-threatening emergencies through regular use of recombinant human erythropoeitin. The prevalence of anti-HCV positivity among dialysis patients who received treatment during the "HCV-prophylaxis period" was 33/163 (20.2%), as compared to 46/55 (83.6%) of those who received HD during the 27 months prior to October 1992 (p< 0.0001), and had similar average duration on dialysis (12 + 7 and 13 + 7 months, respectively). Excluding the 15 patients who had anti-HCV on entry to HD during "HCV-prophylaxis period", the estimated incidence of positive anti-HCV seroconversion was 11.5 per 100 patients per year on HD. In the 93 anti-HCV positive patients, alanine aminotransferase (ALT) levels were elevated for more than six months in 32 (34.4%), elevated in multiple peaks in 22 (23.7%) and showed combined variation of the latter two abnormalities in 16 (17.2%). Histological evidence of chronic active hepatitis was present in five of six patients who manifested persistent ALT abnormalities. Vaccination against hepatitis B virus produced positive seroconversion in 76.1% patients, and those with positive anti-HCV were not at a disadvantage. In conclusion, HCV infection is common in patients undergoing HD in Kuwait.Improvement in screening assays, isolation of anti-HCV positivepatients during dialysis and limitation of blood transfusions may decrease the transmission of this disease in this patient population.

Anti-HCV-positive cirrhosis associated with schistosomiasis.

Hepatosplenic schistosomiasis is occasionally associated with cirrhosis and progressive hepatic decompensation. The aim of the present study was to determine the prevalence of antibody to hepatitis C virus in patients with schistosomiasis and cirrhosis. The prevalence of anti-HCV was studied in 12 consecutive cases of schistosomiasis associated with biopsy proven cirrhosis. All patients had a past history of schistosomiasis and high titers of schistosomal antibodies in serum (1:32 to 1:4096). Five of the 12 patients had hepatic catheterization and were found to have sinusoidal involvement with corrected sinusoidal pressures ranging from 19 to 23 mm Hg. Four had ascites, six had pedal edema, and eight had peripheral signs of chronic liver disease in the form of palmar erythema, spider nevi, and/or gynecomastia. Ten of the 12 cases (83%) were repeatedly positive for anti-HCV/ELISA. These results suggest that when patients with schistosomiasis develop cirrhosis, associated hepatitis C virus infection should be suspected.

Comparison of two immunization schedules with recombinant hepatitis B vaccine and natural immunity acquired by hepatitis B infection in dialysis patients.

In a prospective study over a 2-year period we compared two practical dosage schedules to vaccinate dialysis patients against hepatitis B virus (HBV) infection using a yeast-derived recombinant hepatitis B vaccine (Engerix-B). In addition, the natural history of this acquired immunity was compared with that developed through HBV infection in dialysis patients and healthy subjects. Patients on dialysis treatment (haemo or peritoneal) who were tested to be negative for hepatitis B surface antigen (HBsAg), anti-HBs and anti-HB core were allocated at random to receive HB vaccine according to one of the two schedules. The two groups receiving the vaccine were matched for age, sex, mean duration on dialysis and the form of dialysis treatment received. The group of patients who received a four-dose schedule (at 0, 1, 2 and 6 months) of 40 micrograms of HB vaccine each time (group 2) achieved a seroconversion rate of 79% 1 month after the last dose (at month 7) compared with a seroconversion rate of 55% in those who received three doses (at 0, 1 and 6 months) of 40 micrograms each (group 1). Healthy controls who received half the amount of vaccine on a three-dose schedule (group 3) attained 100% seroconversion (p < 0.05). When retested at 24 months, 30% of seroconverters in group 1 had lost their protective immunity, compared with only 6% in group 2 and 15% in group 3. The magnitude of antibody response (total and anti-(a)-specific) was assessed in the vaccinees at 24 months and compared with that of two other control groups, dialysis patients (group 4) and healthy volunteers (group 5), who had acquired immunity from HBV infection. In general, the total and anti-(a)-specific HBs titres in the dialysis patients (groups 1, 2 and 4) were lower than in their corresponding healthy controls (groups 3 and 5), irrespective of whether the protective immunity was acquired by vaccination or HBV infection. However, the anti-HBs titres in dialysis patients who received four doses were significantly higher than in those who received only three doses (p < 0.05), which indicated a better protective immunity in favour of the former regime. The magnitude of antibody response in the vaccinees of groups 2 and 3 compared well with their respective controls, groups 4 and 5, who had acquired their immunity through HBV infection. This implied that the yeast-derived vaccine was sufficiently immunogenic and provided lasting protection in patients and healthy subjects vaccinated by an appropriate dosage schedule.(ABSTRACT TRUNCATED AT 400 WORDS)

Rapid quantitative assay of rabies post-vaccination antibody by ELISA.

An improved method of ELISA for rabies post-vaccination antibody determination has been developed comparing adsorption properties of polystyrene beads and microtitre plates which were coated with different concentrations of rabies virus antigen. All 106 human post-vaccination serum samples tested were found repeatedly positive within the range of mean values (MV) from 1.4 to 43.0 international units (IU)/0.1 ml. The plates displayed much higher coefficient of variation (CV) when testing lower serum dilutions in different wells of the same plate. In higher dilutions, the sera with high and low antibody levels could easily be distinguished, the results corresponding well to those of the indirect fluorescent antibody test (IFAT), with the exception of some negative sera also to those obtained by ELISA at the Institut Pasteur (Paris). The slightest CV occurred with a 1:400 serum dilution on beads, indicating that ELISA using beads as solid phase can be recommended as a rapid, sensitive and reliable technique for quantitative assay of rabies post-vaccination antibody.

Urinary schistosomiasis associated with hepatitis C virus infection.

We studied 26 adult patients referred for cystoscopy: 13 consecutive patients with schistosome ova on bladder biopsy and antibodies to Schistosoma species in serum were classified as having urinary schistosomiasis, while 13 consecutive patients without schistosome ova on bladder biopsy and who were negative for antibodies to Schistosoma species in serum served as controls. Nine of 13 patients (70%) and none of 13 controls (p < 0.0005) had antibodies to hepatitis C virus in serum (anti-hepatitis C virus). All controls and patients who were negative for anti-hepatitis C virus had normal serum alanine aminotransferase levels, while 2 of 9 (22%) positive for anti-hepatitis C virus had elevated levels. Our study shows that patients with urinary schistosomiasis are at high risk for anti-hepatitis C virus positivity and that some of them may have active liver disease. Therefore, it is imperative to screen patients with urinary schistosomiasis for associated hepatitis C virus infection and liver disease.

Aflatoxin M1 contamination in commercial samples of milk and dairy products in Kuwait.

As part of the programme on monitoring of environmental contaminants in food stuff in Kuwait, 54 samples of fresh full cream and skimmed milk, powdered milk, yoghurt, and infant formula were analysed for aflatoxin M1 (AFM1) by HPLC following sample clean up using immuno-affinity columns. Of samples, 28% were contaminated with AFM1 with 6% being above the maximum permissible limit of 0.2 microgl(-1). Three fresh cow milk samples collected from a private local producer showed the highest level of 0.21 microg l(-1) AFM1. There was no contamination with AFM1 in powdered milk and infant formulas. These results show the necessity of a survey involving a larger number of milk and its products and suggest that presently the contamination of milk and milk products with AFM1 does not appear to be a serious health problem in Kuwait. Nevertheless, a continuous surveillance programme may be warranted to monitor regularly the occurrence of aflatoxins in the animal feeds responsible for current limited contamination and to note rapidly and worsening in the situation that may depend on market changes or on unfavourable climatic developments.

Acute sporadic hepatitis E in Kuwait.

Fifty-seven adult patients with acute hepatitis and 34 comparison patients without liver disease were evaluated using a newly developed Western blot assay for IgM antibody to hepatitis E virus. The mean age of patients with hepatitis was 32 years (range, 18-55 years); 88% were male. Among patients with acute hepatitis, hepatitis A (anti-HAV IgM positive) was diagnosed in two (4%), hepatitis B (anti-HBc IgM positive) in three (5%), and hepatitis E (anti-HEV IgM positive) in 34 (60%). One hepatitis patient had CMV IgM, another had EBV IgM, and 16 others (28%) were negative for all serologic markers of acute viral hepatitis. No patient with acute hepatitis A or B and none of the comparison patients without acute hepatitis had anti-HEV IgM. All but one case of acute hepatitis E were found among expatriates of Asian origin, and acute hepatitis E was associated significantly with recent travel to the Indian subcontinent. These data suggest that acute hepatitis E is common among foreign workers in Kuwait but that little HEV transmission is occurring directly in Kuwait.

Detection and differentiation of picornaviruses in clinical samples following genomic amplification.

A polymerase chain reaction (PCR) assay was used to detect and differentiate picornaviruses (PVs), using primers homologous to the 5' non-coding and VP2 regions of the PV genome. The PCR resulted in a 530 bp PCR product for human rhinoviruses (HRVs) and a 650 bp product for polioviruses, coxsackieviruses (CV) or echoviruses. The PCR assay could detect as little as 1 p.f.u. of virus in either cerebrospinal fluid (CSF) or stool, using ethidium bromide-stained gels. Standard strains of poliovirus, CV, echovirus and HRV were detected, with the exception of echovirus type 22. In contrast, heterologous viruses, such as herpes simplex virus, human cytomegalovirus, adenovirus, influenza virus and rotavirus, as well as human and monkey cell DNA, were not amplified. In nasal swabs taken from patients with respiratory infections, the PCR detected 27 of 28 HRV isolation-positive specimens. All specimens from which viruses other than HRVs were isolated were negative by PCR. The PCR definitively identified poliovirus and CVs from the CSF or stool of patients with aseptic meningitis, as well as CV in the pericardial fluid of a patient who had suffered a myocardial infarction. Specimens taken from patients with similar pathologies, and from which heterologous viruses were isolated, were uniformly negative by PCR.

Short-term IgM and IgG antibody responses to hepatitis E virus infection.

53 adult patients with acute hepatitis caused by hepatitis E virus were identified by the presence of IgM antibody to hepatitis E virus, and followed for 12 months to evaluate the kinetics of anti-HEV antibodies. All but 1 female Kuwaiti patient were expatriate workers from the Indian subcontinent, temporarily working in Kuwait. Follow-up samples obtained at 1, 3, 6 and 12 months were evaluated for IgM and IgG antibodies to hepatitis E virus. IgM-class antibodies to hepatitis E virus were detectable in 12/27 (44%) patients at 1 months, in 0/26 at 3 months, in 0/8 at 6 months and 0/6 at 12 months. IgG antibodies to hepatitis E virus were detectable in 46/47 (98%) at onset, 26/27 (96%) at 1 month, in 26/29 (90%) at 3 months, 16/16 (100%) at 6 months and 8/8 (100%) at 12 months of follow-up. This study suggests that IgM antibodies to hepatitis E virus decline rapidly after an acute infection but IgG antibodies to hepatitis E virus persists for at least 1 year in many patients.

Direct detection of human cytomegalovirus in urine specimens from renal transplant patients following polymerase chain reaction amplification.

A polymerase chain reaction (PCR) assay was used to amplify human cytomegalovirus (HCMV) directly from urine specimens taken from renal transplant patients. In serial urine samples from patients who had at least one specimen positive for HCMV; the PCR assay consistently detected the presence of HCMV DNA sequences, whereas virus detection by other tests such as enzyme-linked immunosorbent assay (ELISA), nonradioactive DNA hybridization assay, and virus isolation were variable. Of 37 specimens positive by PCR, 36 were positive by either ELISA, hybridization assay, or virus isolation. Infectious virus was detected in 13 of the 37 PCR-positive urines. HCMV DNA was detected by PCR in all samples that were positive for HCMV by either hybridization assay or virus isolation. The viral genome copy number was determined by PCR assay for several urine samples that were positive by virus isolation but negative for HCMV by ELISA or hybridization assay. Viral genome copy number estimates indicated the presence of HCMV at very low levels in these urines verifying the fidelity of the virus isolation procedures. The consistency of the PCR assay makes it an ideal method for detection of infection and monitoring antiviral drug therapy in patients infected with HCMV.