Population dynamics of phlebotomine sand flies (Diptera: Psychodidae) in cutaneous leishmaniasis endemic areas of Kurunegala District, Sri Lanka.

Sand flies are the primary vectors of leishmaniasis. Disease management with effective vector control depends on the knowledge of vector population dynamics. In Sri Lanka, despite few isolated entomological collections, long-term studies are not reported to date. In the reported study, monthly entomological surveillance was conducted from May 2017 to December 2018 in Polpithigama, Maho, and Galgamuwa Medical Officer of Health (MOH) areas using standard entomological techniques. Climatic data were collected from the Department of Meteorology, Sri Lanka. Patient records were collected from each MOH office. A total of 38,339 sand flies were collected. The majority were Phlebotomus argentipes (99.50%, n = 38,147), while the rest was Sergentomyia punjabensis (0.50%, n = 192). Wind speed reduces sand fly abundance (r = -0.519, P < 0.05) evidently by limiting the movement. Rainfall increases sand fly abundance (r = 0.842, P <0.05, lag = 6 months) probably by improving favorable conditions in breeding sites. The actual effect of humidity is not conclusive without further research. The observed higher prevalence of leishmaniasis in the study sites could be due to the higher sand fly density in these areas. The number of patients shows a strong positive correlation (r = 0.516, P <0.05) to the sand fly abundance with a lag of 7 months. Systematic surveillance of sand flies as a part of general healthcare services is strongly recommended to identify and prevent possible outbreaks through timely planned vector control measures.

Detection of Leishmania donovani DNA within Field-Caught Phlebotomine Sand Flies (Diptera: Psychodidae) in Three Cutaneous Leishmaniasis Endemic Foci of Kurunegala District, Sri Lanka.

Leishmaniasis is a parasitic infection transmitted through the bite of female phlebotomine sand flies. Microscopy is the gold standard to detect parasites within the sand flies and for vector incrimination. However, molecular-based detection has become more popular nowadays in the identification of Leishmania parasites since it provides detection and species identification simultaneously with no need of laborious procedures. The entomological surveys were conducted monthly from May to October 2017 using standard entomological techniques. Field-caught sand flies were identified to the species level followed by DNA extraction. The polymerase chain reaction (PCR) was performed using species-specific primers to detect Leishmania donovani parasites. A total of 1,662 sand flies were encountered from the entomological surveys, and the majority of them were Phlebotomus argentipes (n = 1517; 91.27%), while others were Sergentomyia punjabiensis (n = 140; 8.72%). Leishmania donovani parasite DNA was detected only from P. argentipes (2.3%; n = 2). The detection of Leishmania DNA in P. argentipes suggests the possible role of this species as a vector for leishmaniasis in Sri Lanka.

Socioeconomic, demographic and landscape factors associated with cutaneous leishmaniasis in Kurunegala District, Sri Lanka.

BACKGROUND: Leishmaniasis is a neglected tropical disease that affects countries in the developing world. In Sri Lanka, cutaneous leishmaniasis is the most common form of the disease. It is prevalent in dry and intermediate zones, mostly associated with rural settings. Understanding basic risk factors is critical in the management of the disease with effective interventions. This study is focused on assessing the demographic, socioeconomic and landscape factors associated with leishmaniasis in Kurunegala District, Sri Lanka. METHODS: A descriptive cross-sectional study was conducted. Households of the past patients and randomly selected households, which had no history of leishmaniasis cases were interviewed. The clinical, socioeconomic, demographic, landscape and awareness-related data were obtained using a pre-tested, interviewer-administered questionnaire. RESULTS: A total of 101 patients and a similar number of controls were included in the study. All the patients had the cutaneous form of the disease. Housewives and personnel with monthly incomes less than Rs. 10,000 (56.76 USD) were 3.9- and 9.5-times more prone to the disease, respectively, according to multivariate analysis. Presence of decaying garbage, termite hills, unclear areas, wet soil and gardening areas were always associated with the increased odds of acquiring the disease. CONCLUSIONS: Demographic factors do not play a pivotal role in the prevalence of leishmaniasis in the area. Housewives, inhabitants with low incomes and individuals who live in areas with conditions suitable for sand fly breeding and resting are major groups with a higher risk of infection. Special attention must be given in raising awareness and environmental management in control activities.

Establishment of a Colony of Phlebotomus argentipes under Laboratory Conditions and Morphometric Variation between Wild-Caught and Laboratory-Reared Populations.

The field-based studies on sand flies are not adequate to uncover information required for the control of the leishmaniasis through reduction of vector populations. Therefore, establishment and maintenance of laboratory colonies of sand flies is an essential step in leishmaniasis research. In the current study, a colony of P. argentipes was established from wild-caught sand flies following standard procedures from the published literature. Morphological measurements of laboratory-reared and wild-caught individual sand flies were compared to assess the difference between two groups. The colony was successfully established under confined laboratory conditions. The comparison of morphometric parameters revealed that the laboratory-reared sand flies are significantly larger than those caught from wild, suggesting a possibility of increased fitness of sand flies under favorable environmental conditions which may cause higher prevalence in the disease. The current study reports the first successful attempt in colonizing sand flies under laboratory conditions. However, the colony data suggest that the conditions extracted from the published literature need to be optimized to suit local settings in order to achieve maximum population sizes within the available amount of resources.

Evaluation of rapid extraction and isothermal amplification techniques for the detection of Leishmania donovani DNA from skin lesions of suspected cases at the point of need in Sri Lanka.

BACKGROUND: Leishmaniasis is a disease caused by vector-borne protozoans. In Sri Lanka, the cutaneous form of the disease is predominant, which is usually diagnosed using Giemsa-stained slit skin smear examination and by histology. However, the sensitivity of slit skin smears and histology are reportedly low. Moreover, facilities for the highly sensitive polymerase chain reaction (PCR) are available only in a few highly-equipped parasitology laboratories. Therefore, there is a need for low cost, sensitive and specific screening tests for diagnosis of leishmaniasis at the point of need. RESULTS: In this study, a mobile suitcase laboratory applying novel extraction (SpeedXtract) and isothermal amplification and detection (recombinase polymerase amplification assay, RPA) methods were evaluated for the diagnosis of cutaneous leishmaniasis in Sri Lanka. First, the developed assay was applied to three different sample types (punch biopsy, slit skin smears and fine needle aspirates) at a local hospital. The results showed that the 2 mm punch biopsy sample produced the best exponential amplification curve and early fluorescence signal in the RPA assay. Secondly, punch biopsies were collected from 150 suspected cutaneous leishmaniasis cases and screened with SpeedXtract/RPA, RNAlater/PCR and ATL buffer/PCR, in addition to Giemsa-stained slit skin smears. Fifty-seven samples were negative in all detection methods. In total 93 samples were positive with assay sensitivities of 65.5% (SpeedXtract/RPA), 63.4% (RNAlater/PCR) and 92.4% (ATL buffer/PCR). The Giemsa-stained slit skin smear delivered the worst clinical sensitivity (32.2%). CONCLUSIONS: The SpeedXtract/RPA method under field conditions took 35 min, while almost 8 h were needed to finalize the extraction and detection by PCR in the laboratory. The SpeedXtract/RPA method produced similar sensitivity to samples preserved in RNAlater and subjected to PCR amplification, but both were less sensitive than ATL-preserved samples subjected to PCR amplification. There is a need for a standardization of sample collection and nucleic acid extraction methods.

The Economic Impact of Cutaneous Leishmaniasis in Sri Lanka.

Cutaneous leishmaniasis (CL) is a neglected tropical disease which affects mainly the poorest communities in developing countries. Considering the limited published information on economic impact of CL in Sri Lanka, the current study was conducted with the objective of revealing the nature and magnitude of the economic impact of CL in three selected disease endemic regions in Kurunegala District, Sri Lanka. The patient records of CL notified of relevant Medical Officer of Health (MOH) office during 2013- 2016 were obtained. Patient households were visited and data collection was done using an interviewed administered questionnaire. The majority of patients (57%) were economically active at the time of infection. Of them, 65% were the only contributors to household economy. Total median costs including both direct and indirect costs were 66.85 USD (Rs. 10,831) (IQR = 57.26 - 86.78 USD), while total median economic loss to households was 61.27 USD (Rs. 9,927) (IQR= 49.61- 75.04 USD). From provider perspective, total median cost per patient was 22.83 USD (Rs. 3,696). The mean total economic loss was denoted as 65.26 USD (Rs. 10,572) which is about 5.4% of the annual household income and 20.9% of the mean annual per capita income of the study population. Although economic impact of CL infection is not catastrophic according to current interpretation, the infection may have significant economic impacts on households when considering the mean economic loss to household as a percentage of the mean annual per capita income of the population.

The epidemiology of soil-transmitted helminths in Bihar State, India.

BACKGROUND: Soil-transmitted helminths (STHs) infect over a billion individuals worldwide. In India, 241 million children are estimated to need deworming to avert the negative consequences STH infections can have on child health and development. In February-April 2011, 17 million children in Bihar State were dewormed during a government-led school-based deworming campaign. Prior to programme implementation, a study was conducted to assess STH prevalence in the school-age population to direct the programme. The study also investigated risk factors for STH infections, including caste, literacy, and defecation and hygiene practices, in order to inform the development of complementary interventions. METHODS: A cross-sectional survey was conducted among children in 20 schools in Bihar. In addition to providing stool samples for identification of STH infections, children completed a short questionnaire detailing their usual defecation and hand-hygiene practices. Risk factors for STH infections were explored. RESULTS: In January-February 2011, 1279 school children aged four to seventeen provided stool samples and 1157 children also completed the questionnaire. Overall, 68% of children (10-86% across schools) were infected with one or more soil-transmitted helminth species. The prevalence of ascariasis, hookworm and trichuriasis was 52%, 42% and 5% respectively. The majority of children (95%) practiced open defecation and reported most frequently cleansing hands with soil (61%). Increasing age, lack of maternal literacy and certain castes were independently associated with hookworm infection. Absence of a hand-washing station at the schools was also independently associated with A. lumbricoides infection. CONCLUSIONS: STH prevalence in Bihar is high, and justifies mass deworming in school-aged children. Open defecation is common-place and hands are often cleansed using soil. The findings reported here can be used to help direct messaging appropriate to mothers with low levels of literacy and emphasise the importance of water and sanitation in the control of helminths and other diseases.

Cluster-randomised trial of the impact of school-based deworming and iron supplementation on the cognitive abilities of schoolchildren in Sri Lanka's plantation sector.

OBJECTIVE: To assess the impact of deworming and iron supplementation on the cognitive abilities and educational achievement of school-age children in Sri Lanka. METHODS: Prospective, placebo-controlled randomised study. The treatment group received deworming and weekly iron supplementation for 6 months; the control group received placebo for both the anthelmintic and iron. A mixed effects regression model was used to answer the main research question. To increase the precision of this study's estimates, various background variables were controlled for that were not related to treatment but could have some impact on the outcome. RESULTS: The prevalence of soil-transmitted helminth (STH) infection was reduced in the treatment group (n = 615), with significant differences between treatment and control groups (n = 575) in the levels of Ascaris and Trichuris. No impact was found on haemoglobin (Hb) levels, nor any significant impact on concentration levels or on educational test scores. CONCLUSION: Decline in STH prevalence alone, in the absence of improved Hb status, produced no evidence of impact on concentration levels or educational test scores.

Evaluation of PCR-ELISA as a tool for monitoring transmission of Wuchereria bancrofti in District of Gampaha, Sri Lanka.

OBJECTIVE: To compare Wuchereria bancrofti (W. bancrofti) infection rates of Culex quinquefasciatus, using dissection and PCR-ELISA in two consecutive time periods (from 2007 to 2008 and from 2008 to 2009). METHODS: Mosquitoes were collected in 30 sentinel and 15 non-sentinel sites in 15 Medical Officer of Health areas of Gampaha District known for the presence of W. bancrofti transmission in two consecutive time period of 2007 to 2008 and 2008 to 2009. Captured mosquitoes were dissected to determine the W. bancrofti larvae (L1, L2, L3). PCR was carried out using DNA extracted from mosquito pools (15 body parts/pool) utilizing the primers specific for Wb-SspI repeat. PCR products were analyzed by hybridization ELISA using fluorescein-labeled wild type specific probes. The prevalence of infected/infective mosquitoes in PCR pools (3 pools/site) was estimated using the PoolScreen™ algorithm and a novel probability-based method. RESULTS: Of 45 batches of mosquitoes dissected, W. bancrofti infected mosquitoes were found in 19 and 13 batches, with an infection rate of 13.29% and 3.10% with mean larval density of 8.7 and 1.0 larvae per mosquito for two study periods in the Gampaha District. Total of 405 pools of head, thorax and abdomen were processed by PCR-ELISA for each year. Of these, 51 and 31 pools were positive for W. bancrofti in the two study periods respectively. The association of dissection based prevalence rates with PCR based rates as determined by the Pearson correlation coefficient were 0.176 and 0.890 respectively for the two periods. CONCLUSIONS: Data indicate that PCR-ELISA is more sensitive than the traditional dissection techniques for monitoring transmission intensity.

Evaluation of PCR-ELISA as a tool for monitoring transmission of Wuchereria bancrofti in District of Gampaha, Sri Lanka

Objective: To compare Wuchereria bancrofti (W. bancrofti) infection rates of Culexquinquefasciatus, using dissection and PCR-ELISA in two consecutive time periods (from 2007 to 2008 and from 2008 to 2009). Methods: Mosquitoes were collected in 30 sentinel and 15 non-sentinel sites in 15 Medical Officer of Health areas of Gampaha District known for the presence ofW. bancrofti transmission in two consecutive time period of 2007 to 2008 and 2008 to 2009. Captured mosquitoes were dissected to determine the W. bancrofti larvae (L1, L2, L3). PCR was carried out using DNA extracted from mosquito pools (15 body parts/pool) utilizing the primers specific for Wb-SspI repeat. PCR products were analyzed by hybridization ELISA using fluorescein-labeled wild type specific probes. The prevalence of infected/infective mosquitoes in PCR pools (3 pools/site) was estimated using the PoolScreenTM algorithm and a novel probability-based method.Results:Of 45 batches of mosquitoes dissected, W. bancrofti infected mosquitoes were found in 19 and 13 batches, with an infection rate of 13.29% and 3.10% with mean larval density of 8.7 and 1.0 larvae per mosquito for two study periods in the Gampaha District. Total of 405 pools of head, thorax and abdomen were processed by PCR-ELISA for each year. Of these, 51 and 31 pools were positive for W. bancrofti in the two study periods respectively. The association of dissection based prevalence rates with PCR based rates as determined by the Pearson correlation coefficient were 0.176 and 0.890 respectively for the two periods. Conclusions: Data indicate that PCR-ELISA is more sensitive than the traditional dissection techniques for monitoring transmission intensity.

Soil-transmitted helminth infections among plantation sector schoolchildren in Sri Lanka: prevalence after ten years of preventive chemotherapy.

BACKGROUND: The plantation sector in Sri Lanka lags behind the rest of the country in terms of living conditions and health. In 1992, a sector-wide survey of children aged 3-12 years and women of reproductive age showed >90% prevalence of soil-transmitted helminth infections. Biannual mass de-worming targeting children aged 3-18 years started in 1994 and was continued until 2005. The present study was carried out to assess the status of infection four years after cessation of mass de-worming. METHODS/FINDINGS: A school-based cross-sectional survey was carried out. Faecal samples from approximately 20 children from each of 114 schools in five districts were examined using the modified Kato-Katz technique. Data regarding the school, the child's family and household sanitation were recorded after inspection of schools and households. Multivariate analysis was carried out using logistic regression, to identify risk factors for infection. Faecal samples were obtained from 1890 children. In 4/5 districts, >20% were infected with one or more helminth species. Overall combined prevalence was 29.0%; 11.6% had infections of moderate-heavy intensity. The commonest infection was Ascaris lumbricoides, present in all five districts, as was Trichuris trichiura. Hookworm was not detected in two districts. Multivariate analysis identified low altitude and maternal under-education as risk factors for all three infections. Poor household sanitation was identified as a risk factor for A. lumbricoides and hookworm, but not T. trichiura infections. CONCLUSIONS/SIGNIFICANCE: The results indicate that regular mass de-worming of plantation sector children should be resumed along with more emphasis on better sanitation and health education. They show that even after 10 years of mass chemotherapy, prevalence can bounce back after cessation of preventive chemotherapy, if the initial force of transmission is strong and other long-term control measures are not concomitantly implemented.

Social mobilisation, drug coverage and compliance and adverse reactions in a Mass Drug Administration (MDA) Programme for the Elimination of Lymphatic Filariasis in Sri Lanka.

BACKGROUND: In Sri Lanka filariasis is endemic in Southern, Western and North Western provinces covering eight districts designated as implementation units in the Programme for the Elimination of Lymphatic Filariasis (PELF). Despite control activities over sixty years including multidose diethylcarbamazine, 6 mg/kg treatment microfilaria rates had persisted at low levels. Following systematic social mobilisation the first MDA with DEC albendazole combination was conducted in 2002. METHODS: We investigated the extent social mobilisation had reached the people, their drug compliance and adverse reactions. Three localities were selected from each district to pick target population samples for pre-tested questionnaire. Three teams each with six people visited one district each day. One team worked from three starting points in one locality. A member applied eight part questionnaire to one family member totalling 150-160 people from one locality. Questions included social mobilisation, drug compliance and adverse reactions. RESULTS: Information was disseminated by television, radio, banners and leaflets, to a lesser extent by people. Information reached more people in the periphery than in Colombo. 35.2% from Colombo municipality were unaware of the MDA. Drug coverage was 79.6%, home delivery 71.7% and delivery centres 7.9%. 35.6% in Colombo district and 53.4% from Colombo municipality did not receive drugs. Drugs were consumed by 71.4%. 28.6% who did not comply included 20.4% who did not receive them. 91.4% showed no adverse reactions, 7.5% were mild, 1.1% recovered with home remedies. CONCLUSION: Drug compliance showed significant positive correlation with awareness of the MDA. Door to door delivery was more successful than delivery from centres. More delivery centres conveniently located would have rectified this disparity. Poor awareness and compliance in Colombo and urban areas could be rectified with separate strategy for urban areas. More time for MDA and trained adequate manpower would ensure coverage to achieve elimination.