This study primarily aimed to identify the causative species of cutaneous leishmaniasis (CL) in the Khyber Pakhtunkhwa Province of Pakistan and to distinguish any species-specific variation in clinical manifestation of CL. Diagnostic performance of different techniques for identifying CL was assessed. Isolates of Leishmania spp. were detected by in vitro culture, polymerase chain reaction (PCR) on DNA extracted from dried filter papers and microscopic examination of direct lesion smears from patients visiting three major primary care hospitals in Peshawar. A total of 125 CL patients were evaluated. Many acquired the disease from Peshawar and the neighboring tribal area of Khyber Agency. Military personnel acquired CL while deployed in north and south Waziristan. Leishmania tropica was identified as the predominant infecting organism in this study (89.2%) followed by Leishmania major (6.8%) and, unexpectedly, Leishmania infantum (4.1%). These were the first reported cases of CL caused by L. infantum in Pakistan. PCR diagnosis targeting kinetoplast DNA was the most sensitive diagnostic method, identifying 86.5% of all samples found positive by any other method. Other methods were as follows: ribosomal DNA PCR (78.4%), internal transcribed spacer 2 region PCR (70.3%), culture (67.1%), and microscopy (60.5%). Clinical examination reported 14 atypical forms of CL. Atypical lesions were not significantly associated with the infecting Leishmania species, nor with “dry” or “wet” appearance of lesions. Findings from this study provide a platform for species typing of CL patients in Pakistan, utilizing a combination of in vitro culture and molecular diagnostics. Moreover, the clinical diversity described herein can benefit clinicians in devising differential diagnosis of the disease.

Despite their medical importance, natural breeding sites of sand flies have been poorly explored. Detecting such sites is imperative in vector control strategies. This study aimed to identify potential breeding and resting microhabitats of sand flies and household risk factors associated with cutaneous leishmaniasis transmission in Dir districts, Khyber Pakhtunkhwa, Pakistan. Survey of indoor and outdoor habitats in four tehsils (subdistricts) of upper and lower Dir districts was carried out using sticky traps, flit method, and aspirator. To assess risk factors associated with cutaneous leishmaniasis (CL), questionnaires were administered to household heads. Mud houses and plants belonging to Cucurbitaceae family sheltered highest number of sand flies. Excessive number of flies were collected from indoor sites (common rooms used for both people and animals) compared to outdoor habitats (cattle dungs). Phlebotomus salangensis Artemiev, 1978 and Phlebotomus sergenti were the most widely distributed species, while Sergentomyia bailyi peaked in caves with high organic content in soil. Roof material and ownership of fans were associated with significant risk of CL. Findings of this study present a list of potential breeding sites for sand flies and CL risk factors that can be targeted and implemented in control programs for CL. Further studies are required to elucidate the breeding behavior and preferences of these medically important vectors.

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BackgroundAfghan refugees in northern Pakistan have been resident for over 30 years and current information on malaria in this population is sparse. Understanding malaria risk and distribution in refugee camps is important for effective management both in camps and on return to Afghanistan.MethodsCross-sectional malariometric surveys were conducted in five Afghan refugee camps to determine infection and exposure to both Plasmodium falciparum and Plasmodium vivax. Factors associated with malaria infection and exposure were analysed using logistic regression, and spatial heterogeneity within camps was investigated with SatScan.ResultsIn this low-transmission setting, prevalence of infection in the five camps ranged from 0–0.2 to 0.4–9 % by rapid diagnostic test and 0–1.39 and 5–15 % by polymerase chain reaction for P. falciparum and P. vivax, respectively. Prevalence of anti-malarial antibodies to P. falciparum antigens was 3–11 and 17–45 % for P. vivax antigens. Significant foci of P. vivax infection and exposure were detected in three of the five camps. Hotspots of P. falciparum were also detected in three camps, only one of which also showed evidence of P. vivax hotspots.ConclusionsThere is low and spatially heterogeneous malaria transmission in the refugee camps in northern Pakistan. Understanding malaria risk in refugee camps is important so the malaria risk faced by these populations in the camps and upon their return to Afghanistan can be effectively managed.

We provide a comprehensive and updated review on the effects of elevation and climatic factors on distribution of sand fly (Diptera: Psychodidae) vectors of cutaneous leishmaniasis (CL) and visceral leishmaniasis (VL) across Pakistan. Our study was undertaken in view of the increasing risk of leishmaniasis, particularly resulting from climatic deviations and political instability in the region. Literature (published and unpublished) on suspected vectors of CL (Phlebotomus sergenti Parrot and Phlebotomus papatasi Scopoli) and VL (Phlebotomus major Yakimoff and Schokhor and Phlebotomus hindustanicus Theodor) was collected, sorted, and utilized in geospatial analysis. P. papatasi and P. sergenti co-occurred in majority of the referenced localities and were generally distributed toward the western and southern parts across a wide range of elevation, whereas P. major and P. hindustanicus were limited to high elevation in cool and humid northern areas of Pakistan. Conversely P. papatasi and P. sergenti exploited diverse ecological settings across the country, being particularly abundant in warm/hot and arid regions.

This study was conducted to investigate the diversity of sandflies (Psychodidae: Phlebotominae) and the incidence of leishmaniasis in three villages of North Waziristan Agency, Pakistan. Sandflies were sampled monthly during 2012, at dusk and dawn, in selected indoor habitats including both bedrooms and animal sheds using a knock-down spray catch method. A total of 3687 sandflies were collected, including 1444 individuals in Drezanda, 1193 in Damdil and 1050 in Dattakhel. This study revealed 14 species of two genera, Phlebotomus (Phlebotomus sergenti, Phlebotomus papatasi, Phlebotomus caucasicus, Phlebotomus kazeruni, Phlebotomus alexandri and Phlebotomus salehi) and Sergentomyia (Sergentomyia dentate, Sergentomyia baghdadis, Sergentomyia babu, Sergentomyia theodori, Sergentomyia sumbarica, Sergentomyia dreyfussitur kestanica, Sergentomyia hogsoni pawlowskyi and Sergentomyia fallax afghanica) (both: Diptera: Psychodidae). Phlebotomus sergenti was the most abundant species (42.1%), followed by S. dentata (17.7%) and S. baghdadis (17.4%). The number of males collected represented about twice that of female flies, and the maximum number was collected in July, followed by August. The determination of the species composition of sandfly populations, seasonal variations, relative abundances and estimations of infection in the vector population may provide information about the dynamics of leishmaniasis transmission that is useful in planning vector control activities.

The present study was aimed at elucidation of malaria epidemiology and comparing performance of several diagnostic procedures in Bannu, a highly endemic district of Khyber Pakhtunkhwa, Pakistan. Dried blood spots were collected from patients suspected of malaria visiting a hospital and two private laboratories in district Bannu and processed for species-specific PCR (rRNA). Patients were also screened for malaria through microscopy and RDT. A well-structured questionnaire was used to collect patient information to assess risk factors for malaria. Of 2033 individuals recruited, 21.1% (N = 429) were positive for malaria by at least one method. Overall, positivity detected by PCR was 30.5% (95/311) followed by 17.7% by microscopy (359/ 2033) and 16.4% by RDT (266/1618). Plasmodium vivax (16.9%, N = 343) was detected as the dominant species followed by Plasmodium falciparum (2.3%, N = 47) and mixed infections (1.2%, N = 39). Microscopy and RDT (Cohen's kappa k = 0.968, p = <0.0001, McNemar test p = 0.069) displayed significant agreement with each other. Satisfactory health, sleeping indoors, presence of health-care facility in vicinity (at an accessible range from home), living in upper middle class and in concrete houses significantly reduced malaria risk; whereas, low literacy level, presence of domestic animals indoors and malaria diagnosis recommended by clinician increased the disease risk. Overall, findings from the study provide reasonable basis for use of RDT as a cost-effective screening tool in field and for clinicians who can proceed with timely treatment of malaria patients. Appropriate management of identified risk factors could contribute to reduction of malaria prevalence in Bannu and its peripheries.

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