Rima Hanna-Wakim

Flow cytometry diagnosis of dedicator of cytokinesis 8 (DOCK8) deficiency

To the Editor: Biallelic mutations in the dedicator of cytokinesis 8 (DOCK8) gene cause autosomal-recessive hyper-IgE syndrome, a combined immunodeficiency characterized by sinopulmonary infections, skin and systemic viral infections, eczema, and food allergy. DOCK8 deficiency can lead to early death from infection and malignancy. The disease is curable by hematopoietic cell transplantation (HCT). Thus, it is important to ascertain the diagnosis of DOCK8 deficiency to institute early treatment. The vast majority of DOCK8-deficient patients lack DOCK8 expression and many have deletions in the DOCK8 gene. Confirmation of the diagnosis has relied on immunoblotting of blood cell lysates and/or gene sequencing, techniques that are not routinely available. We present here a flow cytometry assay that could facilitate the diagnosis of DOCK8 deficiency, detection of carrier status, and investigation of lineage-specific DOCK8 expression following HCT.

GENETIC DIVERSITY AND ANTIVIRAL DRUG RESISTANCE OF PANDEMIC H1N1 2009 IN LEBANON

BACKGROUND In June 2009, the World Health Organization announced the 21st centurys first influenza pandemic caused by pandemic influenza H1N1 2009 (H1N1 pdm). OBJECTIVES Our goal was to analyze antiviral drug resistance and the phylogenetic relationships among hemagglutinin (HA) and neuraminidase (NA) genes of H1N1 pdm samples in Lebanon. STUDY DESIGN Nasopharyngeal swabs were collected from 197 patients with influenza-like illness from May 2009 through January 2010. Of the 50 influenza A-positive samples, 30 were analyzed for antiviral drug resistance by using in vitro susceptibility assays and cycling-probe real-time PCR. The HA and NA genes were also analyzed. RESULTS The results of hemagglutination-inhibition assays confirmed that all 30 analyzed samples were H1N1 pdm. In July 2009, community transmission of H1N1 pdm was detected in Lebanon, and an outbreak occurred in October 2009. The outbreak cases were caused by a strain with 4 mutations in the NA gene (i.e., V42I, N68T, N248D, and E462K) and 2 mutations in the HA gene: 1 in the Ca1 antigenic site (i.e., S206T) and 1 in the Ca2 antigenic site (i.e., D225E). This strain was closely related to a major H1N1 pdm cluster that was isolated worldwide. All 30 samples were amantadine-resistant, and none were zanamivir-resistant. The 1 oseltamivir-resistant sample appeared to be from a community-transmitted case in an otherwise healthy 2-year-old child. CONCLUSION Continuous monitoring of oseltamivir susceptibility among H1N1 pdm is essential to guide the effective use of this drug.

Epidemiologic characteristics, serotypes, and antimicrobial susceptibilities of invasive Streptococcus pneumoniae isolates in a nationwide surveillance study in Lebanon

Invasive pneumococcal disease (IPD) associated with Streptococcus pneumonia is a major public health problem worldwide for all age groups, including in Lebanon. Prevention through vaccination remains the most valuable tool to decrease the burden of disease. Pneumococcal conjugate vaccine 7 (PCV7), marketed internationally including in the Middle East and North Africa region for the prevention of IPD, was introduced in Lebanon in 2006, followed by PCV10 and PCV13 in 2010. However, none of these is currently part of the Extended Program of Immunization schedule and published data on IPD incidence, pneumococcal serotypes and vaccine coverage in the region are lacking. The Lebanese Inter-Hospital Pneumococcal Surveillance Program is a surveillance system set up to determine the burden of IPD and the prevalent serotypes responsible. The aim of this prospective 6-year study carried out in 78 hospitals throughout Lebanon was to obtain such data to help health authorities make informed decisions on the implementation of pneumococcal vaccination at the national level. A total of 257 isolates of culture-confirmed Streptococcus pneumoniae were evaluated. Considering all age groups, vaccine coverage was 41.4%, 53.9%, and 67.2% for PCV7, PCV10, and PCV13 serotypes, respectively; for patients <2, 2-5, and >60 years of age, PCV7 coverage was 50%, 51%, and 35%, respectively; PCV10 coverage was 53%, 74%, 45%, respectively; and PCV13 coverage was 63%, 80%, and 68%, respectively. Overall, 17.4% of these isolates were penicillin-G non-susceptible using the latest established breakpoints and mortality occurred in 23.5% of the patients with non-susceptible isolates. In addition, 10.9% of isolates were multi-drug-resistant. The highest mortality rates were observed in the eldest (>60 years of age) and youngest (<2 years of age) patients. The most prevalent invasive serotypes identified were those found in currently available pneumococcal conjugate vaccines, emphasizing the importance of implementing the vaccine in the routine immunization schedule at the national level. Continuation of current surveillance practices will help assess the impact of vaccine implementation on IPD epidemiology, serotype distribution and antibiotic resistance patterns.

Approach to Non-Neutropenic Fever in Pediatric Oncology Patients-A Single Institution Study.

Pediatric oncology patients with fever, even when not neutropenic, are known to be at an increased risk of bloodstream infections. However, there are no standard guidelines for management of fever in non‐neutropenic patients, resulting in variability in practice across institutions.

Dietary Zinc Intake and Plasma Zinc Concentrations in Children with Short Stature and Failure to Thrive.

Background: The burden of zinc deficiency on children includes an increased incidence of diarrhea, failure to thrive (FTT) and short stature. The aim of this study was to assess whether children with FTT and/or short stature have lower dietary zinc intake and plasma zinc concentrations compared to controls. Methods: A case-control study conducted at the American University of Beirut Medical Center included 161 subjects from 1 to 10 years of age. Results: Cases had a statistically significant lower energy intake (960.9 vs. 1,135.2 kcal for controls, p = 0.010), lower level of fat (30.3 vs. 36.5 g/day, p = 0.0043) and iron intake (7.4 vs. 9.1 mg/day, p = 0.034). There was no difference in zinc, copper, carbohydrate and protein intake between the 2 groups. The plasma zinc concentration did not differ between the cases and controls (97.4 vs. 98.2 μg/dl, p = 0.882). More cases had mild-to-moderate zinc deficiency when compared to controls with 10.3 vs. 3.6%, p = 0.095. Conclusion: Our study did not show statistically significant difference in dietary zinc intake and plasma zinc concentrations between children with FTT and/or short stature compared to healthy controls. A prospective study is planned to assess the effect of zinc supplementation on growth parameters in FTT children.

Intestinal Inflammation and Dysregulated Immunity in Patients with Inherited Caspase-8 Deficiency

Caspase-8 (CASP8) is a protease that initiates apoptosis and regulates inflammation and immune responses. We identified germline mutations in CASP8 in 3 unrelated patients with infant-onset inflammatory bowel disease: 2 patients were homozygous for the mutation 710A>G, p.Q237R, which resulted in reduced protein expression, and 1 patient carried the mutation 793C>T, p.R265W. We isolated peripheral blood mononuclear cells from our index patient and observed defects in T- and B-cell maturation, proliferation, and/or activation. Macrophages from 1 patient with CASP8 deficiency and monocytic BLaER1 cells with knockout of CASP8 or overexpression of CASP8 with the 710A>G mutation had altered inflammasome activity on stimulation with lipopolysaccharide. Patient-derived intestinal organoids and colon carcinoma cells with knockout of CASP8 had defects in cell death processes that involved loss of TRAIL signaling and increased necroptosis. These findings indicate that CASP8 controls inflammation, innate and adaptive immunity, and intestinal barrier integrity in humans.

Questionnaire-based survey in a developing country showing noncompliance with paediatric gastro-oesophageal reflux practice guidelines.

This 2015 study investigated whether Lebanese paediatricians diagnosed and managed gastro‐oesophageal reflux disease (GERD) in infants and children in accordance with the 2009 guidelines from the North American and European Societies for Paediatric Gastroenterology, Hepatology and Nutrition.

Mutations in pyrin masquerading as a primary immunodeficiency

Whole exome sequencing is increasingly used in the diagnosis of primary immunodeficiencies due to the overlapping and atypical presentations of these disorders. We report two patients who presented with recurrent infections and early onset colitis. They were investigated by whole exome sequencing due to suspicion of primary immunodeficiency and found to have mutations in pyrin known to cause familial Mediterranean fever.