New insights on the clinical variability of FKBP10 mutations.

To date 45 autosomal recessive disease-causing variants are reported in the FKBP10 gene. Those variant were found to be associated with Osteogenesis Imperfecta (OI) for which the hallmark phenotype is bone fractuers or Bruck Syndrome (BS) where bone fractures are accompanied with contractures. In addition, a specific homozygous FKBP10 mutation (p.Tyr293del) has been described in Yup'ik Inuit population to cause Kuskokwim syndrome (KS) in which contractures without fractures are observed. Here we present an extended Palestinian family with 10 affected individuals harboring a novel homozygous splice site mutation, c.391+4A > T in intron 2 of the FKBP10 gene, in which the three above mentioned syndromes segregate as a result of skipping of exon 2 and absence of the FKBP65 protein. At the biochemical level, Hydroxylysyl pyridinoline (HP)/lysyl pyridinoline (LP) values were inversely correlated with OI phenotypes, a trend we could confirm in our patients. Our findings illustrate that single familial FKBP10 mutations can result in a phenotypic spectrum, ranging from fractures without contractures, to fractures and contractures and even to only contractures. This broad intra-familial clinical variability within one single family is a new finding in the field of bone fragility.

Genetic analysis of osteogenesis imperfecta in the Palestinian population: molecular screening of 49 affected families.

BACKGROUND: Osteogenesis imperfecta (OI) is a heterogeneous hereditary connective tissue disorder clinically hallmarked by increased susceptibility to bone fractures. METHODS: We analyzed a cohort of 77 diagnosed OI patients from 49 unrelated Palestinian families. Next-generation sequencing technology was used to screen a panel of known OI genes. RESULTS: In 41 probands, we identified 28 different disease-causing variants of 9 different known OI genes. Eleven of the variants are novel. Ten of the 28 variants are located in COL1A1, five in COL1A2, three in BMP1, three in FKBP10, two in TMEM38B, two in P3H1, and one each in CRTAP, SERPINF1, and SERPINH1. The absence of disease-causing variants in the remaining eight probands suggests further genetic heterogeneity in OI. In general, most OI patients (90%) harbor mainly variants in type I collagen resulting in an autosomal dominant inheritance pattern. However, in our cohort almost 61% (25/41) were affected with autosomal recessive OI. Moreover, we document a 21-kb genomic deletion in the TMEM38B gene identified in 29% (12/41) of the tested probands, making it the most frequent OI-causing variant in the Palestinian population. CONCLUSION: This is the first genetic screening of an OI cohort from the Palestinian population. Our data are important for genetic counseling of OI patients and families in highly consanguineous populations.

Molecular characterization of methicillin-resistant Staphylococcus aureus isolates in three different Arab world countries.

Molecular characterization of methicillin-resistant Staphylococcus aureus (MRSA) isolates in three different Arab world countries (West Bank of Palestine, Jordan, and Iraq) was the aim of the study presented here. This is done on the basis of spa sequencing and staphylococcal cassette chromosome mec (SCCmec) typing. The majority (92%) of the spa-tested isolates belonged to spa type t932 and possessed the (SCCmec) type III. These data suggest that MRSA clone, which harbors the spa type t932 and (SCCmec) type III, had been transferred throughout the three studied countries.

Determination of Helicobacter pylori Virulence Genes in Gastric Biopsies by PCR.

Aim. The aim of this study was to identify the presence of H. pylori in biopsy specimens from symptomatic patients by PCR. In addition, the rate of cagA, vacA, iceA1, and iceA2 virulence genes was determined. Materials and Methods. One hundred antral gastric biopsy specimens were collected during endoscopy from patients suffering from gastroduodenal symptoms. The samples were collected by the gastroenterologists in their own clinics in Ramallah, Palestine. DNA was extracted from the biopsies and subsequently used for PCR identification of H. pylori and the virulence genes using specific primers. Results. The rate of positive H. pylori in the collected biopsies was 44%. The rates of the virulence genes in this sample: cagA, vacA, iceA1, and iceA2 were 65.9%, 40.9%, 63.6%, and 84.1%, respectively. Conclusion. The iceA2 gene was the most frequent in this study. Much research is necessary to determine the presence of an association of this gene with gastric pathology. Variation in the rates of the iceA gene in different countries is a strong indication of its geographical distribution. This study would provide important information regarding the prevalence of virulence genes (vacA, cagA, iceA1, and iceA2) in H. pylori strains in the sample tested in this country.

Antimicrobial resistance in non-typhi Salmonella enterica isolated from humans and poultry in Palestine.

INTRODUCTION: The efficacy of chemotherapy can be compromised by drug resistance. This study was undertaken to describe the resistance profiles and fluoroquinolone resistance mechanism of non-typhoidal Salmonella (NTS) isolated from humans and poultry in West Bank, Palestine. METHODOLOGY: One hundred and fifty-one isolates of NTS, obtained from humans (71) and poultry (80), collected between September 2005 and January 2007, were tested for susceptibility to ampicillin, gentamicin, tetracycline ceftriaxone, nalidixic acid and ciprofloxacin. Mutation patterns within gyrA were determined by direct sequencing or by digestion of PCR-amplified DNA fragments with the restriction enzyme HinfI. RESULTS: Resistance rates among human and poultry isolates were respectively 59% and 51% for ampicillin, 31% and 10% for gentamicin, 59% and 80% for tetracycline, 59% and 45% for nalidixic acid, and 30% and 15% for ciprofloxacin. All the isolates were susceptible to ceftriaxone. Mutations at positions 83 and/or 87 were detected in gyrA of isolates with resistance to nalidixic acid. Isolates which were resistant to nalidixic acid but susceptible to ciprofloxacin had a single gyr A gene mutation at point 87. This gene mutation was sufficient to induce a new phenotype (6 isolates) with decreased susceptibility to ciprofloxacin. CONCLUSION: Mutations in gyrA at positions 83 or 87 were the most prevalent mutation pattern of fluoroquinolone resistant NTS isolates but other unknown mechanisms are also present. Continued surveillance of antimicrobial resistance among NTS isolates is needed to mitigate the increasing prevalence of quinolone resistance.

Detection of Trichomonas vaginalis in Vaginal Swab Clinical Samples from Palestinian Women by Culture.

Background. Trichomoniasis is a common sexually transmitted disease caused by Trichomonas vaginalis. It is a major health problem worldwide. The World Health Organization (WHO) has estimated that 180 million infections are acquired annually worldwide. Methodology. Vaginal swabs (1207) were cultured for T. vaginalis on Trichomonas Medium no. 2 (Oxoid) soon after specimen collection. The cultures were examined daily using a light microscope to detect the presence of T. vaginalis. Results. The prevalence of T. vaginalis was 13.6% (164/1207). The infection rate was the highest during pregnancy, 28.1%, and the lowest among women whose spouses use condoms, 8.6%. Conclusions. The culture method was used in this study to accurately determine the prevalence of this parasite in the West Bank, Palestine. The results of the study will eliminate ambiguities concerning trichomoniasis in this country and will contribute to better management and proper treatment.

Community-acquired meticillin-resistant Staphylococcus aureus in Palestine.

Community-acquired meticillin-resistant Staphylococcus aureus (CA-MRSA) is becoming an important public-health problem. This study attempted to investigate S. aureus and MRSA colonization in nasal swabs obtained from 843 patients without a history of hospitalization at the time of hospital admission and from 72 health-care workers chosen for comparison. Of the patients, S. aureus was detected in 218/843 (25.9%) and MRSA in 17/843 (2.0%). Of the health-care workers, S. aureus was detected in 15/72 (20.8%) and MRSA in 10/72 (13.9%). The majority of the 27 MRSA isolates exhibited a sensitivity pattern expected for CA-MRSA. Multilocus restriction fragment typing resolved the isolates into eight restriction fragment types. The predominant restriction fragment types were AAACCAA and AAAAAAA, accounting for 51.9% (14/27) of the MRSA isolates and included CC5 and CC1 groups, respectively. This study thus demonstrated the transmission of CA-MRSA strain types into a health-care setting, emphasizing the need for implementation of a revised set of control measures in both hospital and community settings.

Prevalence of Cryptosporidium spp. in children with diarrhoea in the West Bank, Palestine.

This study was conducted to investigate the prevalence of Cryptosporidium spp. in children (n=760) with diarrhea aged 1 month to 13 years, living in urban areas (n=234), rural areas (n=394) and refugee camps (n=132). Samples were collected, stained by modified acid fast stain, and examined microscopically for oocysts. The overall prevalence was 11.6% (88/760). The prevalence was higher in refugee camps at 12.9% (17/132) and in rural areas at 12.2% (48/394) as compared to 9.8% (23/234) in urban areas. According to age, the prevalence in age group I (<5 years) was significantly high (P<0.05) at 14.4% (67/464) as compared to 7.7% (15/195) in age group II (5-10 years) and 5.9% (6/101) in age group III (10-15 years). Our findings indicate that the prevalence of Cryptosporidium spp. is high when compared to that in developed countries.

Helicobacter pylori, a causative agent of vitamin B12 deficiency.

BACKGROUND: Helicobacter pylori is one of the most common causes of peptic ulcer disease worldwide and a major cause of chronic superficial gastritis leading to atrophy of gastric glands. METHODOLOGY: A total of 60 patients suffering from gastric disease due to H. pylori infection were evaluated. Endoscopy was performed and gastric biopsies were obtained for histopathology and urease test. Blood was simultaneously collected for the determination of the levels of vitamin B12 and the MCV. Vitamin B12 levels were determined by chemiluminescent assay. RESULTS: Our results indicate that the mean vitamin B12 level +/- SEM for the total population, the H.pylori infected and non-infected patients were 264.5+/-22.9, 207.7+/-21.9 and 419.7+/-39.8 respectively. H. pylori was found in 71.7% (43/60) of the patients tested. The level of vitamin B12 was lower than 200pg/ml (deficient) in 67.4% (29/43) of patients tested positive for H. pylori. CONCLUSION: H. pylori appears to be implicated in causing vitamin B12 deficiency.

Screening of indigenous Palestinian medicinal plants for potential anti-inflammatory and cytotoxic activity.

Organic extracts of 24 selected plant species, used by Palestinian traditional healers to treat different illnesses and diseases, were tested for their anti-inflammatory and anti-tumoral activities. The plant selection was based on existing ethnobotanic information and interviews with local healers. The extracts of the plants under investigation were tested for their potential anti-tumor (cytotoxic) effect on the murine fibrosarcoma L929sA cells, and on the human breast cancer cells MDA-MB231 and MCF7. Cytotoxicity screening models provide important preliminary data to select plant extracts with potential antineoplastic properties. MTT (Tetrazolium blue) colorimetric assay was used to evaluate the reduction of viability of cell cultures in the presence or absence of the extracts. The extract from Withania somnifera, L. Dunal (Solanaceae) presented an IC(50) value at 24h of 150 and 60 microg/ml, on L929sA and MCF7 cells, respectively, while the extract from Psidium guajava L. (Myrtaceae) presented an IC(50) value at 24h of 55 microg/ml on MCF7 cells. Other extracts examined, like Laurus nobilis L. (Lauraceae) and Salvia fruticosa M. (Labiatae), also displayed a remarkable activity. Additionally, as the nuclear transcription factor NFkappaB regulates the expression of various genes that play critical roles in apoptosis and immunomodulation, we further investigated the effect of nine promising plant extracts, withheld from the first cell viability screening on NFkappaB activation. The extracts showed variable degrees of NFkappaB-inhibitory activity. Whereas Withania somnifera extract demonstrated the strongest NFkappaB-inhibitory activity, other extracts derived from Laurus nobilis, Psidium guajava and Foeniculum vulgare M. (Umbiliferrae) also revealed immunomodulatory NFkappaB activities. These species are good candidates for further activity-monitored fractionation to identify active constituents.

Withaferin a strongly elicits IkappaB kinase beta hyperphosphorylation concomitant with potent inhibition of its kinase activity.

The transcription factor NFkappaB plays a critical role in normal and pathophysiological immune responses. Therefore, NFkappaB and the signaling pathways that regulate its activation have become a major focus of drug development programs. Withania somnifera (WS) is a medicinal plant that is widely used in Palestine for the treatment of various inflammatory disorders. In this study we show that the leave extract of WS, as well as its major constituent withaferin A (WA), potently inhibits NFkappaB activation by preventing the tumor necrosis factor-induced activation of IkappaB kinase beta via a thioalkylation-sensitive redox mechanism, whereas other WS-derived steroidal lactones, such as withanolide A and 12-deoxywithastramonolide, are far less effective. To our knowledge, this is the first communication of IkappaB kinase beta inhibition by a plant-derived inhibitor, coinciding with MEK1/ERK-dependent Ser-181 hyperphosphorylation. This prevents IkappaB phosphorylation and degradation, which subsequently blocks NFkappaB translocation, NFkappaB/DNA binding, and gene transcription. Taken together, our results indicate that pure WA or WA-enriched WS extracts can be considered as a novel class of NFkappaB inhibitors, which hold promise as novel anti-inflammatory agents for treatment of various inflammatory disorders and/or cancer.

Germ line transcription in mice bearing neor gene downstream of Igamma3 exon in the Ig heavy chain locus.

Class switch recombination (CSR) is preceded by germ line transcription that initiates from promoters upstream of switch (S) sequences and terminates downstream of associated constant genes. Previous work showed that germ line transcripts and their processing are required for CSR and that germ line transcription is regulated in a major part by a regulatory region located downstream of the Ig heavy chain locus. This long-range, polarized effect can be disturbed by inserting an expressed neomycine resistance (neo(r)) gene. To contribute to a better understanding of the mechanism of such a long-distance regulation, we generated knock-in mice in which a neo(r) gene was inserted downstream of Igamma3 exon leaving intact all the necessary elements for germ line transcription and splicing. We show that the expressed neo(r) gene interferes with transcription initiation from Igamma3, and that it impairs but does not block S recombination to Cgamma3. Moreover, we show for the first time that the neo(r) gene provides through chimeric neo(r)-Cgamma3 transcripts the necessary elements for splicing of germ line transcripts by activating two novel cryptic splice sites, one in the coding region of the intronless neo(r) gene and the other in the Igamma3-Cgamma3 intron.