High prevalence of three prothrombotic polymorphisms among Palestinians: factor V G1691A, factor II G20210A and methylenetetrahydrofolate reductase C677T.

Factor V leiden G1691A/R506Q (FVL), prothrombin G20210A (FII) and methylenetetrahydrofolate reductase (MTHFR) C677T are related genetic risk factors for venous thromboembolism. Analysis for those mutations is increasingly being performed on patients exhibiting hypercoagulability. The objective of this study was to determine the prevalence of FVL, FII-G20210A and MTHFR-C677T polymorphisms and their coexistence among apparently healthy Palestinians. After institutional approval, 303 apparently healthy students from An-Najah University representative to North and South regions of West Bank with no previous history of cardiovascular diseases participated in this study. A uniform questionnaire was used to collect relevant information through personal interview with the subjects. The collected information included gender, age, smoking habits, weight and height, diseases such as diabetes, cardiovascular and family history of CVD. The frequencies of allelic distribution of the three prothrombotic polymorphisms factor V G1691A/R506Q), prothrombin G2010A, and MTHFR-C677T were 0.114, 0.050 and 0.071, respectively. The prevalence of the three thrombotic polymorphisms (FVL, FII G20210A and MTHFR-C677T) were 20.1, 9.1 and 13.8 %, respectively. Statistical analysis for factor V leiden showed no significant association between place of residence (P value = 0.953) and gender (P value >0.082). The data presented in this study showed the highest prevalence of FVL among healthy Palestinians compared to other populations and this important finding should be followed in terms of clinical significance.

Prevalence of intestinal parasites among school children in northern districts of West Bank-Palestine.

OBJECTIVES: To assess the prevalence of intestinal parasite infections in northern districts of West Bank, Palestine and to determine associated sociodemographic factors. METHODS: Random sampling of schoolchildren from rural and urban areas was carried out. Participants provided faecal samples and answered a questionnaire about their demographics and hygiene habits. Faecal samples underwent microscopy and PCR to screen for protozoan and helminths. RESULTS: Seven hundred and thirty-five samples were collected from children aged 9.5 years on average. The overall prevalence of parasitic infection was 22.2%. The rates of infections with amoeba, Giardia intestinalis, Entrobius vermicularis and Ascaris lumbricoides were 9.7%, 4.1%, 1.6% and 3.8%, respectively. Real-time PCR was performed to differentiate between Entamoeba histolytica and Entamoeba dispar. Results showed that 14% of samples positive with microscopy for amoeba were positive for E. histolytica. There was no significant association between sex and rates of infections (P-value > 0.05). There were, however, significant association between parasite infections and parents' education, place of residence, washing hands habits (P-value > 0.05). No significant association was found with number of family members or eating in school canteens (P-value > 0.05). CONCLUSIONS: Intestinal parasite infections are endemic in West Bank. Interventions such as health education and sanitation are needed.

Cryptosporidium parvum causes gastroenteritis epidemics in the Nablus region of Palestine.

A total of 30 faecal samples collected from individuals admitted to a local hospital in Nablus city in Palestine with gastroenteritis symptoms, plus five faecal samples from healthy individuals living in the same area were screened for the presence of Cryptosporidium spp. by microscopic analysis using malachite green negative staining. Molecular techniques were used to confirm the microscopic identification. All 30 samples from individuals with gastroenteritis symptoms were positive by both techniques. No other parasites were found in the faecal material of patients or healthy individuals. To explore the source of the outbreak, water was collected from various reservoirs and springs that supply the city with drinking water. Al-Qaryoon water spring was found to be contaminated with Cryptosporidium using both microscopic and molecular analysis. No other water resources were found to be contaminated. Genotyping analysis of Cryptosporidium oocysts using PCR-RFLP technique identified the parasite as C. parvum.

Nematode acetylcholinesterases are encoded by multiple genes and perform non-overlapping functions.

Nematodes are unusual in that diverse molecular forms of acetylcholinesterase are the product of distinct genes. This is best characterised in the free living organism Caenorhabditis elegans, in which 3 genes are known to give rise to distinct enzymes, with a fourth likely to be non-functional. ACE-1 is an amphiphilic tetramer associated with a hydrophobic non-catalytic subunit, analogous to vertebrate T enzymes, whereas ACE-2 and ACE-3 are glycosylphosphatidylinositol-linked amphiphilic dimers. The different ace genes show distinct anatomical patterns of expression in muscles, sensory neurons and motor neurons, with only a few examples of coordinated expression. Clear homologues of ace-1 and ace-2 have now been isolated from a variety of parasitic nematodes, and the predicted proteins have very similar C-terminal amino acid sequences, implying an analogous means of anchorage to membranes. In addition to these membrane-bound enzymes, many parasitic nematodes which colonise mucosal surfaces secrete acetylcholinesterases to the external (host) environment. These hydrophilic enzymes are separately encoded in the genome, so that some parasites may thus have a total complement of six ace genes. The secretory enzymes have been characterised from the intestinal nematode Nippostrongylus brasiliensis and the lungworm Dictyocaulus viviparus. These show a number of common features, including a truncated C-terminus and an insertion at the molecular surface, when compared to other nematode acetylcholinesterases. Although the function of these enzymes has not been determined, they most likely alter host physiological responses to promote survival of the parasite.

A distinct family of acetylcholinesterases is secreted by Nippostrongylus brasiliensis.

A third variant of acetylcholinesterase (AChE A) secreted by the parasitic nematode Nippostrongylus brasiliensis has been isolated which shows 63-64% identity to AChE B and AChE C, with a truncated carboxyl terminus and a short internal insertion relative to AChEs from other species. Three of the fourteen aromatic residues which line the active site gorge in Torpedo AChE are substituted by non-aromatic residues (Y70T, W279D and F288M). All three enzymes have 8 cysteine residues in conserved positions, including 6 which have been implicated in disulphide bonds in other AChEs. Phylogenetic analysis suggests that these enzymes form a distinct group which evolved after speciation and are most closely related to ACE-2 of Caenorhabditis elegans. Recombinant AChE A secreted by Pichia pastoris was monomeric and hydrophilic, with a substrate preference for acetylthiocholine and negligible activity against butyrylthiocholine. A model structure of AChE A built from the coordinates of the Torpedo californica AChE suggests that W345 (F331 in Torpedo) limits the docking of butyrylcholine. This model is consistent with mutational analysis of the nematode enzymes. Expression of AChE A is regulated at the transcriptional level independently of the other 2 secreted variants, with maximal expression by fourth stage larvae and young adult worms. These enzymes thus appear to represent an unusual family of AChEs with conserved structural features which operate outside the normal boundaries of known functions in regulation of endogenous neurotransmitter activity.

Cloning and expression of two secretory acetylcholinesterases from the bovine lungworm, Dictyocaulus viviparus.

We describe the molecular cloning, expression and biochemical characterisation of recombinant forms of two secreted acetylcholinesterases from adult Dictyocaulus viviparus. The two variants (designated Dv-ACE-1 and Dv-ACE-2) were 613 and 615 amino acids long and showed 94.7% identity to one another. The highest level of identity to other cholinesterases was with ACE-2 of Caenorhabditis elegans. Dv-ACE-1 and Dv-ACE-2 showed 48.0 and 47.7% identity to C. elegans ACE-2 over 577 amino acids, respectively. The primary structure of both enzymes showed conservation of the catalytic triad and of a tryptophan residue known to be critical for the choline-binding site, but differed in the number of potential glycosylation sites and at one amino acid in the peripheral anionic site. Southern blotting and PCR experiments indicated that the genes encoding these enzymes are distinct. When expressed in Pichia pastoris, the enzymes were active, but differed subtly in their biochemical characteristics. Both enzymes exhibited a preference for acetylcholine as substrate, but differed in the extent of excess substrate inhibition and in their optimal pH for activity. The lack of an obvious carboxy-terminal membrane anchor and the presence of an insertion at the molecular surface were other features which, thus far, appear to be characteristic of parasite secreted acetylcholinesterases.

Trichinella spiralis secretes a homologue of prosaposin.

Infective larvae and adult stage Trichinella spiralis secrete a protein homologous to prosaposin, the precursor of sphingolipid activator proteins (saposins) A-D originally defined in vertebrates. The protein contains four saposin domains, with the six cysteine residues which form the three intramolecular disulphide bonds in close register in each case. It differs substantially from vertebrate prosaposins in the N-terminal prodomain, the region separating saposins A and B, and completely lacks the C-terminal domain which has been demonstrated to be essential for lysosomal targetting in these organisms. The protein is secreted in unprocessed form with an estimated mass of 56 kDa, and contains a single N-linked glycan which is bound by the monoclonal antibody NIM-M1, characteristic of the TSL-1 antigens which are capped by tyvelose (3,6-dideoxy-D-arabinohexose). Immuno-electron microscopy localised the protein to membrane-bound vesicles and more complex multi-lamellar organelles in diverse tissues including the hypodermis, intestine and stichosomes, although it was absent from the dense-core secretory granules typical of the latter. Possible functions of a secreted prosaposin are discussed.

Association between factor V Leiden mutation and poor pregnancy outcomes among Palestinian women.

Pregnancy is a hypercoagulable state with increased tendency for thrombus formation, a condition that is increased when combined with acquired or inherited risk factors that lead to thrombophilia. Among the inherited risk factors is Factor V Leiden mutation, an autosomal dominant trait with reduced penetrance. The mutation seems to be associated with different poor pregnancy outcomes including recurrent miscarriages. In the present study, we performed a case-control study to investigate the association between the Leiden mutation and poor pregnancy outcome among the Palestinian population in the West bank region of Palestine. The study included 145 subjects with recurrent miscarriages and 205 matched control subjects with successful pregnancies who experienced normal delivery and no apparent complications. Leiden mutation was detected in 41 of the 145 study subjects (28.2%), and in 24 of the 205 control subjects (11.7%). Subjects homozygous with the mutant allele were identified only among the test and not the control group. Data analysis indicates a significant association between the mutant allele and recurrent miscarriages (p-value<0.05). Furthermore, this association is significant between the mutant haplotype with miscarriages compared to control group showing time effect where there is no association for miscarriages before week 10. Results here also show strong association of factor V leiden polymorphism among primary aborters compared to secondary aborters or control groups. The odds ratio for the primary aborters was 75 and p<0.0001. In conclusion, these results provide evidence for a significant correlation between recurrent miscarriages and Factor V mutation in our population.