Widad Al-Nakib

Risk factors for the development of diabetes mellitus in chronic hepatitis C virus genotype 4 infection.

Background and Aim:  A high occurrence of type 2 diabetes (T2D) in patients with chronic hepatitis C virus (HCV) infection has been reported in Kuwait and other countries. However, HCV genotype 4 has been underrepresented in all previous studies. Our aim was to investigate the viral and host risk factors associated with the development of T2D in patients with chronic hepatitis C genotype 4 infection in the absence of liver fibrosis and steatosis.

Echoviruses are a major cause of aseptic meningitis in infants and young children in Kuwait.

BackgroundThe etiologic agents of aseptic meningitis (AM) often include human enteroviruses. The role of enteroviruses causing AM in young children was investigated during a 3-year period in Kuwait.ResultsEnteroviral RNA was detected in cerebrospinal fluid (CSF) by reverse transcription-PCR and specific genotypes of enteroviruses were identified by direct DNA sequencing of VP4-VP2 region. Enteroviral RNA was detected in 92 of 387 (24%) suspected AM cases and the results were confirmed by hybridization of amplicons with an internal, enterovirus-specific probe. The CSF samples from 75 of 281 (27%) children < 2 years old but only from 3 of 38 (8%) 4-12 year-old children were positive for enteroviral RNA (p = 0.011). Majority of infections in children < 2 years old (49 of 75, 65%) were due to three echoviruses; echovirus type 9 (E9), E11 and E30. Only three other enteroviruses, namely coxsackievirus type B4, coxsackievirus type B5 and enterovirus 71 were detected among AM cases in Kuwait.ConclusionsOur data show that three types of echoviruses (E9, E11 and E30) are associated with the majority of AM cases in Kuwait. To the best of our knowledge, this is the first report to characterize different enterovirus genotypes associated with AM in the Arabian Gulf region.

Hepatitis C virus infection in a population with high incidence of type 2 diabetes: Impact on diabetes complications

A growing number of reports suggest a connection between hepatitis C virus (HCV) infection and type 2 diabetes (T2D). However, the association of HCV infection with diabetes-related complications has not yet been clarified. The aim of this study was to determine the prevalence of HCV infection in T2D-patients in Kuwait which has a high incidence of type 2 diabetes, and to investigate the association between HCV viremia and diabetes-related complications. A total of 438 patients with T2D (325 Kuwaitis and 113 Egyptians), and 440 control subjects, were enrolled for this study. HCV infection was assessed by testing for serum HCV-specific antibodies, and by detection of HCV RNA. HCV viral load and hemoglobin A1c (HbA1c) levels were assessed in patients with and without diabetes complications. Thirty one (7%) out of 438 T2D-patients had evidence of HCV infection compared to 4 (1%) out of 440 control adults (p<0.0001). The prevalence of HCV infection in Kuwaiti and Egyptian T2D-patients was 3% and 18%, respectively. Most of the HCV sequences detected in T2D patients and control subjects were of genotype 4. The HbA1c levels in T2D-patients with HCV viremia were significantly higher than those in HCV-negative patients. HCV viremia, female sex, age, family history of diabetes were found to be independent risk factors for diabetes complications. The results suggest that T2D-patients in Kuwait have higher prevalence of HCV infection than controls, and that HCV viremia is associated with diabetes-related complications.

Comparative evaluation of INNO-LiPA HBV assay, direct DNA sequencing and subtractive PCR-RFLP for genotyping of clinical HBV isolates.

Genotypes (A to H) of hepatitis B virus (HBV) influence liver disease progression and response to antiviral therapy in HBV-infected patients. Several methods have been developed for rapid genotyping of HBV strains. However, some of these methods may not be suitable for developing countries. The performance of INNO-LiPA HBV Genotyping assay (LiPA), direct DNA sequencing and subtractive PCR-RFLP of genotype-specific HBV genome regions were evaluated for accurately determining the HBV genotypes by analyzing sera (n = 80) samples from chronic HBV patients. Both, LiPA and DNA sequencing identified 63, 4 and 13 HBV strains as belonging to genotype D, genotype A and mixed genotype A and D, respectively. On the contrary, the PCR-RFLP-based method correctly identified all 4 genotype A but only 56 of 63 genotype D strains. Seven genotype D strains yielded indeterminate results. DNA sequence comparisons showed that a single nucleotide change in the target region generated an additional restriction site for Nla IV that compromised the accuracy of this method. Furthermore, all the mixed genotype A and D strains were identified only as genotype A strains. The data show that the PCR-RFLP-based method incorrectly identified some genotype D strains and failed to identify mixed genotype infections while LiPA and DNA sequencing yielded accurate results.

Detection and monitoring of cytomegalovirus infection in renal transplant patients by quantitative real-time PCR.

Objectives: To establish a sensitive and specific real-time PCR for quantitation of cytomegalovirus (CMV) DNA in clinical specimens.Subjects and Methods: In a prospective study, CMV DNA was quantified in blood samples of 255 kidney recipients with and without CMV-related symptoms between the years 2000 and 2005 in Kuwait. In a selected group of patients, the effect of anti-CMV chemotherapy was monitored by quantitative real-time PCR (qRT-PCR). Results: The established qRT-PCR assay had a sensitivity to detect 30 CMV DNA copies. CMV DNA was detected in 54/255 (24%) patients; of these, 17 (31.5%) were asymptomatic, and 37 patients (68.5%) had symptomatic CMV infection. Sequential blood specimens were collected from all CMV-positive patients and tested by CMV pp65 antigenemia and qRT-PCR assays. There was a moderate positive correlation between the two assays (Pearson’s correlation = 0.52). The median CMV viral load measured by qRT-PCR was higher in symptomatic (6.5 × 104 copies/ml) than in asymptomatic (185copies/ml) patients (p = 0.001). The estimated cut-off value of CMV DNA for CMV symptoms/disease was ≧800 copies/ml of blood. Testing of sequential samples from patients treated with symptomatic CMV infection showed that the viral load was significantly reduced after 3 weeks of anti-CMV chemotherapy (p = 0.001). Conclusion: The reported qRT-PCR is a sensitive method for quantitation of CMV DNA in the blood of kidney recipients and can be useful in monitoring the efficacy of anti-CMV therapy.

Frequency of enterovirus detection in blood samples of neonates admitted to hospital with sepsis‐like illness in Kuwait

This study investigated the role of enteroviruses in sepsis‐like illness among neonates in Kuwait. Serum samples from 139 consecutive neonates presenting with sepsis‐like illness during a three and a half‐year‐period whose blood cultures were negative for bacterial pathogens were tested. Enterovirus RNA was detected by single‐step reverse‐transcription PCR (RT‐PCR). Specific genotypes were identified by direct DNA sequencing of enteroviral genome. Serotype‐specific antibodies in serum samples from some selected patients were detected by virus neutralization test using coxsackievirus B types (CBVs). All 139 neonates presented with sepsis‐like illness and blood samples were uniformly negative for aerobic/anaerobic bacterial cultures. Fifty‐six (40%) neonates had further complications of sepsis including carditis (n = 34) and multi‐organ involvement (n = 22). Enterovirus RNA was detected by RT‐PCR in 34 of 139 (24%) serum samples which is among the highest frequency reported so far in non‐epidemic settings. Genotyping identified CBVs as most common enteroviruses, causing 19 of 34 (56%) enteroviral sepsis episodes in neonates. Of 34 carditis cases, 18 were positive for CBVs by serotyping including all 10 enterovirus RNA‐positive samples. Only one fatality was observed due to liver failure in a neonate with hepatitis. Our data showed that enteroviruses are responsible for 24% of neonatal sepsis cases due to non‐bacterial causes in Kuwait. The data indicate that enteroviruses should be considered in the differential diagnosis of sepsis‐like illness among neonates, particularly those with negative blood cultures for bacterial pathogens. J. Med. Virol. 85:1280–1285, 2013.

Mixed Viral Infections Circulating in Hospitalized Patients with Respiratory Tract Infections in Kuwait

The aim of this study was to determine the frequency of viral mixed detection in hospitalized patients with respiratory tract infections and to evaluate the correlation between viral mixed detection and clinical severity. Hospitalized patients with respiratory tract infections (RTI) were investigated for 15 respiratory viruses by using sensitive molecular techniques. In total, 850 hospitalized patients aged between 3 days and 80 years were screened from September 2010 to April 2014. Among the 351 (47.8%) patients diagnosed with viral infections, viral mixed detection was identified in 49 patients (14%), with human rhinovirus (HRV) being the most common virus associated with viral mixed detection (7.1%), followed by adenovirus (AdV) (4%) and human coronavirus-OC43 (HCoV-OC43) (3.7%). The highest combination of viral mixed detection was identified with HRV and AdV (2%), followed by HRV and HCoV-OC43 (1.4%). Pneumonia and bronchiolitis were the most frequent reason for hospitalization with viral mixed detection (9.1%). There were statistical significance differences between mixed and single detection in patients diagnosed with bronchiolitis (P = 0.002) and pneumonia (P = 0.019). Our findings might indicate a significant association between respiratory virus mixed detection and the possibility of developing more severe LRTI such as bronchiolitis and pneumonia when compared with single detection.

Human metapneumovirus in patients with respiratory tract infection in Kuwait.

Human metapneumovirus (hMPV) has been recognized as an important cause of respiratory tract infections in all age groups and in all geographical area. The role of hMPV in causing respiratory tract infections in Kuwait was not yet investigated. The aim of this study was to determine the prevalence of hMPV infection in Kuwait among patients with respiratory tract infection with respect to other respiratory viruses. During January–December 2009, 460 respiratory samples from 388 patients with respiratory tract infection were collected from different hospitals. They were tested for hMPV RNA by real‐time PCR, and for other respiratory viruses by conventional PCR. Out of 388 patients, 110 (28%) were positive for viral respiratory infections; 21 (5.4%) were positive for hMPV, 29 (7.5%) were positive for rhinovirus, 13 (4%) were positive for respiratory syncytial virus, and 10 (3%) were positive for adenovirus. Most (n = 19, 90.5%) of hMPV‐positive patients were admitted to the intensive care unit, 76% of them were of age 2 years and below, and 24% of age 59 years and above. All hMPV‐positive elderly patients had pneumonia while 50% of hMPV‐positive infants had bronchopneumonia. Children with hMPV/rhinovirus co‐infection (n = 3, 1%) had recurrent chest infection and frequent intensive care unit admission. The hMPV infection was mostly detected between December and May, and genotype B was more prevalent than genotype A. This is the first study demonstrating the prevalence of hMPV infection in Kuwait, and suggests that hMPV infection is prevalent in infants and elderly patients with lower respiratory tract infection. J. Med. Virol. 83:1811–1817, 2011.

Low Levels of Th1-Type Cytokines and Increased Levels of Th2-Type Cytokines in Kidney Transplant Recipients With Active Cytomegalovirus Infection

BACKGROUND Cytomegalovirus (CMV) infection is a major complication after kidney transplantation. It is clear that Th1 and Th2 cell subsets are of major importance in determining the class of immunoprotective function in infectious diseases. Given the strong influence exerted by Th1- and Th2-type immunity on the outcome of infections, we felt it important to elucidate the levels of Th1- and Th2-type cytokines to CMV-related antigens in kidney recipients and to identify antigens that play an essential role in preventing the development of CMV infection and/or disease. METHODS One hundred twenty subjects were followed for CMV infection by the antigenemia assay. We investigated peripheral blood mononuclear cells (PBMCs) responses to five CMV-related peptide antigens (pp65, gB, pp150, pp28, and pp38). Stimulation index was determined by radioactive thymidine uptake, while the production of Th1-type cytokines (interferon-gamma and tumor necrosis factor-alpha) and Th2-type cytokines (interleukins-4 and -10) were measured by enzyme-linked immunosorbent assay. RESULTS The levels of Th1-type cytokine production after stimulating PBMCs with CMV-related antigens gB and pp150 resulted in significant decreases in the levels of interferon-gamma, while pp65, pp150, and pp38 produced significant decreases in the level of tumor necrosis factor-alpha between the two groups (P < .05). For Th2-type cytokines only pp28 produced a significant increase in the level of interleukin-10 between the two groups (P < .05). Regarding the Th1:Th2 ratios, a lower Th1-bias was observed among the CMV-positive patients for PBMCs stimulated with three CMV-related antigens (pp65, pp38, and pp28). CONCLUSION Low levels of Th1-type cytokines and increased levels of Th2-type cytokines upon stimulation with CMV-related peptide antigens were associated with reduced cell-mediated immunity to CMV, thus seeming to correlate with active CMV infections.

The Prevalence of Human Bocavirus, Human Coronavirus-NL63, Human Metapneumovirus, Human Polyomavirus KI and WU in Respiratory Tract Infections in Kuwait

Objective: The aim of this study was to investigate the prevalence of human coronavirus (HCoV)-NL63, human metapneumovirus (hMPV), human bocavirus (Boca), human polyomavirus KI (KIV) and human polyomavirus WU (WUV) in respiratory tract infections (RTI) in Kuwait. Materials and Methods: Respiratory samples from 735 hospitalized patients with RTI from September 2010 to April 2013 were evaluated for the presence of HCoV-NL63, hMPV, Boca, KIV and WUV using molecular assays, polymerase chain reaction (PCR) and reverse-transcription PCR. Results: Of the 735 patients, 285 (38.8%) were diagnosed with viral RTI. The distribution of respiratory viruses was hMPV: 15 (5.3%), Boca: 14 (4.9%), WUV: 10 (3.5%) and KIV: 4 (1.4%). HCoV-NL63 was not detected in any of the samples. Conclusions: These newly discovered viruses were associated with the development of RTI in Kuwait. The rapid identification of these viral infections could aid in the control of nosocomial transmission, reduce the use of antibiotics and improve treatment and management strategies.