Identification of Old World Leishmania species by PCR–RFLP of the 7 spliced leader RNA gene and reverse dot blot assay

The aim of the study was to assess the 7SL RNA PCR followed by restriction fragment length polymorphism (RFLP) and reverse dot blot (RDB) assays for use in identification of Old World Leishmania species. Species‐specific RFLP patterns were obtained for Leishmania major, Leishmania tropica and the Leishmania donovani complex when the 7SL RNA PCR product was digested with the restriction enzyme BsuRI, an isoschizomer of HaeIII. For the RDB assay, biotin‐labelled 7SL RNA amplicons were hybridized to Leishmania genus‐specific and species‐specific oligonucleotide probes immobilized onto a membrane. The Old World Leishmania species could be distinguished by using five probes: one that was a genus‐specific probe and hybridized to all Leishmania species (Lc), two that were specific for L. major (Lm1 and Lm2), one that was specific for L. tropica (Lt) and one that detected both L. major and L. tropica (Lmt). The PCR–RDB was 10 times more sensitive than 7SL PCR and can detect <1 parasite. In addition, the identification of species was easier and more reliable than with 7SL PCR–RFLP. 7SL PCR–RFLP detected parasites in 50 of 57 clinical samples, whereas PCR–RDB detected 53 and 55 were detected by amplification of kinetoplast (k) DNA. The 7SL RNA PCR has proven useful for direct diagnosis of Old World leishmaniasis, especially when combined with the RBD assay for species identification.     

Methylenetetrahydrofolate reductase C677T gene polymorphism and the association with dyslipidemia in type 2 diabetic Palestinian patients

BackgroundDyslipidemia in diabetes is common and characterized by hypertriglyceridemia with decreased levels of high‐density lipoprotein. The objective of this study was to assess the prevalence of MTHFR C677T polymorphism in Palestinian T2DM patients and to investigate the association between this polymorphism and lipid profile in diabetic patients with and without dyslipidemia.MethodsA total of 208 T2DM patients including 98 with dyslipidemia and 110 without dyslipidemia were enrolled in this study. The MTHFR C677T genotyping was conducted by PCR‐RFLP followed by agarose gel electrophoresis.ResultsThere were no significant differences in either the genotype distribution or allele frequency in T2DM patients with or without dyslipidemia (37.8% CC, 54% CT, 8.2% TT vs. 48.2% CC, 41.8% CT, 11% TT; p = 0.209). However, among the dyslipidemic group, the TT carriers have a higher HDL level (46.8 ± 17.8) compared to (CC+CT) carriers (34.68 + 11.9) (p = 0.01). In the group without dyslipidemia, there was a significant elevation in diastolic blood pressure (DBP) among the CC carriers (83.6 ± 10.6) compared to those who carried at least one mutant allele (CT+TT) (78.1 ± 11.1) (p = 0.009).ConclusionsThe study shows that in our Palestinian population the MTHFR 677TT genotype lowers DBP significantly in patients without dyslipidemia and is related to increased level of HDL in diabetic dyslipidemia patients.     

Methods incorporating a polymerase chain reaction and restriction fragment length polymorphism and their use as a 'gold standard' in diagnosing Old World cutaneous leishmaniasis

Three polymerase chain reaction (PCR) assays — kinetoplast DNA (kDNA) PCR, 7SL PCR, ITS1 PCR — were compared for their ability to detect leishmanial parasites in the skin tissue aspirates of 212 Palestinian suspect cases of cutaneous leishmaniasis (CL). The kDNA PCR was the most sensitive, detecting 156/170 (91.8%) of positive samples confirmed by a set ‘gold standard’ but of poor specificity in identifying the species of Leishmania. The 7SL PCR detected 154/170 (90.5%) and the ITS1 PCR only 108/170 (63.5%) of the true positives, and both were 100% specific. The highest Kappa coefficient agreement (95% CI) was obtained with the 7SL PCR (0.792 ± 0.098). Restriction analysis of the products from the ITS1 PCR and 7SL PCR enabled identification of species of Leishmania. L. tropica was the predominant cause of CL, with L. major being less frequent.     

New clinico-epidemiologic profile of cutaneous leishmaniasis, Morocco

During the past 20 years, cutaneous leishmaniasis has emerged as a major public health threat in Morocco. We describe distribution of Leishmania major and L. tropica in Morocco and a new focus of cutaneous leishmaniasis due to L. infantum. We recommend using molecular techniques to diagnose suspected leishmaniasis cases.     

Effect of the Patient’s Knowledge on Peritonitis Rates in Peritoneal Dialysis

♦ Introduction: After a training period, patients maintained on continuous ambulatory peritoneal dialysis (CAPD) assume responsibility for their own treatment. With the aid of appropriate tools, home visits help with ongoing evaluation and training for these patients.♦ Methods: We conducted a home visit survey of 50 patients maintained on CAPD in Sudan between April 2009 and June 2010. Housing conditions, home environment, and patient’s or caregiver’s knowledge about peritoneal dialysis and the exchange procedure were evaluated using structured data collection sheets. Scores were compared with infection rates in the patients before the home visit.♦ Results: Patients were maintained on CAPD for a median duration of 11 months. Their mean age was 42 ± 23 years; 70% were male; and 14% had diabetes. Only 34% of patients had suitable housing conditions, and 56% required assisted PD. Of the autonomous patients and assisting family members, 11.6% were illiterate. The median achieved knowledge score was 11.5 of 35 points. The median achieved exchange score was 15 of 20 points. Knowledge and exchange scores were positively and significantly correlated (R = 0.5, p = 0.00). More patients in the upper quartile than in the middle and lower quartiles of knowledge scores were adherent to daily exit-site care (33.3% vs 5.3%, p = 0.02). Compared with patients in the middle and lower quartiles of knowledge score, patients in the upper quartile had lower rates of peritonitis, exit-site infection, and hospitalization.♦ Conclusions: The proposed evaluation form is a valid and reliable assessment tool for the follow-up of CAPD patients. Patients in the upper quartile of knowledge score demonstrated better adherence to the recommended treatment protocols and lower infection rates.Key words: Adherence, housing conditions, knowledge, peritonitisAfter a training period, patients maintained on con tinuous ambulatory peritoneal dialysis (CAPD) assume responsibility for their own treatment. The need for a home visiting service to manage this program of self-care is widely acknowledged (1-5). The provision of specialist community care for CAPD patients was reported to be associated with significant reductions in infections and hospital admissions (1). The development of home visit protocols has also been reported to be a valuable addition to continuous quality improvement projects in established PD centers (6).Sudan currently has 7 active peritoneal dialysis (PD) units: 6 in Khartoum state and 1 in Al-Gezira state. The most recent unit was established in December 2008 as part of the adult nephrology service at Soba University Hospital in Khartoum. In that unit, strong emphasis is placed on application of the principles of the adult learning theory in patient education. Patients are evaluated in their own homes at the end of training and at regular intervals afterwards. For that purpose, we designed a concise evaluation form that focuses on practical aspects of PD-related knowledge in compliance with the International Society for Peritoneal Dialysis recommendations (7). We also designed comprehensive checklists for the evaluation of PD-related procedures and the home environment. To ensure construct validity, those tools were reviewed and refined by a group of doctors and nurses with working experience in PD, and they were piloted in a small group of patients.In the present study, we used those same tools to evaluate a group of prevalent PD patients who had been maintained on CAPD for variable durations in Sudan. The patients received conventional training upon initiation of dialysis in the other 6 PD units and were never visited in their homes by the PD team. We also evaluated the association between scores on the evaluation tools and infection rates before the home visit.