Surveillance of Wuchereria bancrofti infection by anti-filarial IgG4 in urine among schoolchildren and molecular xenomonitoring in Sri Lanka: a post mass drug administration study.

BACKGROUND: Surveillance of hidden foci or resurgence of the bancroftian filariasis has high priority to maintain the elimination status in Sri Lanka. For the surveillance, two methods were applied in Matotagama, Matara, Sri Lanka; (i) molecular xenomonitoring (MX) by PCR to detect parasite DNA in the vector, Culex (Cx) quinquefasciatus and (ii) survey of anti-filarial IgG4 in urine samples from schoolchildren. RESULTS: Mosquitoes were collected monthly from index houses for 17 months (2013 to 2014) to confirm the existence of bancroftian parasite. Index houses in Matotagama had recorded microfilaria-positive cases in the recent past. Five schools were selected considering Matotagama as the catchment area and all students who presented on the day were tested for urine anti-filarial IgG4 in 2015. Wuchereria bancrofti DNA in Cx. quinquefasciatus pools were found in 14 of 17 months studied and ranged between 0 and 1.4%. The MX rate was greatly increased at least two times in the year following the driest months (March, August). A total of 735 schoolchildren were tested for urine anti-filarial IgG4. Three schools located closer to the MX area had higher positive rates, 3.4%, 3.6%, and 6.6%. Both highest positive rates of MX and urine were located in a nearer vicinity. CONCLUSION: Monthly collections to study lymphatic filariasis (LF) transmission by MX was conducted for the first time in Sri Lanka. We observed that the filarial DNA-positive rate had an association with seasonal cycle of precipitation. More than 1% filarial DNA and > 5% anti-filarial antibody rates confirmed ongoing transmission in Matotagama. The combination of two non-invasive surveys, the urine anti-filarial IgG4 levels of schoolchildren and MX of vector mosquitoes, would be a convenient package to monitor the ongoing transmission (hotspots) of LF in the surveillance.

A surveillance system for lymphatic filariasis after its elimination in Sri Lanka.

Lymphatic filariasis (LF) has been declared eliminated in Sri Lanka in September 2016. To maintain elimination status, a surveillance system to detect hidden endemic foci or LF resurgence is of highest priority. In this paper, we have reported an investigation of LF transmission in Trincomalee district where a surveillance program was not carried out due to 30 years of civil unrest. Proposed surveillance system included, measurement of anti-filarial IgG4 in urine of schoolchildren in areas where LF transmission could exist and assessment of circulating filarial antigen (CFA) and microfilaria (mf) in all urine antibody positive schoolchildren, their family members and 10-15 neighbours of each urine antibody positive household. Spatial distribution of the anti-filarial antibody titers in urine in a high antibody suspected area was analyzed using GPS logger data. Among 2301 school children from 11 schools studied, 41 (1.8%) urine antibody positives were found. The antibody positive rates of the schools ranged between 0 and 4.0%. Nine of the 630 (1.4%) examined became positive for CFA but were negative for mf. Although there were no mf positives, positive CFA and antibody results indicated the existence of Wuchereria bancrofti in Trincomalee. Highest antibody titres in an area correlated with the prevalences of urine antibodies and CFA. Spatial analysis showed LF transmission foci. Therefore, a combination of the non-invasive methods, urine ELISA and GPS mapping, will be a new effective surveillance system to identify hidden LF transmission foci.

Impact of two follow-up schemes on morbidity management and disability prevention (MMDP) programme for filarial lymphedema in Matara, Sri Lanka.

Alleviating morbidity due to lymphatic filariasis (LF)-especially in elderly patients who are rather ignorant-is presently the biggest challenge for the national filariasis campaign. We introduced two follow-up schemes and compared each other to address three key programmatic issues (1) locating patients, (2) educating patients, family members on practice of lymphoedema self-care (3) well sustained daily self-care. Hundred and seven lymphoedema patients were introduced to the new Community Home Based Care (CHBC) programme as a part of MMDP programme at their homes. Twenty seven of 107 patients were selected by purposive sampling and followed-up under two schemes, 14 in Daily follow-up (DFU) scheme and 13 in Monthly follow-up (MFU) scheme. Impact was assessed using a KAP score, number of entry lesions (EL) and number of ADL episodes, limb volume, its appearance, changes in the quality of life and gained benefits. Visiting patients in their homes to introduce lymphoedema care programme was a success. KAP scores of the more important activities on lymphoedema care were significantly higher in DFU scheme. Number of patients (51.9%; 14/27) who had EL/s at baseline reduced significantly to 18.5% (5/27) at one year follow-up. The mean numbers of ADL episodes/year reduced significantly in both schemes. Six photographs of 27 showed obvious improvement in lymphoedema and its grade. Mean volume of lymphoedema reduced significantly in both schemes at one year no significant difference between schemes. Benefit score at one year revealed that the patients in DFU scheme received significantly higher amount of benefits compared to MFU scheme. In conclusion daily instruction has significantly motivated the patient and his/her family bringing a new hope.

Use of a Novel Portable Three-Dimensional Imaging System to Measure Limb Volume and Circumference in Patients with Filarial Lymphedema.

The World Health Organization's Global Program to Eliminate Lymphatic Filariasis (LF) has reduced LF transmission worldwide, but millions remain affected by filarial lymphedema. Tools for clinically monitoring lymphedema in developing nations are limited. We tested a novel, portable, infrared three-dimensional imaging system (3DIS) against water displacement (WD) and tape measurement of limb circumference (TMLC) among patients with filarial leg lymphedema in Galle, Sri Lanka. Outcomes were accuracy and reproducibility of imaging system measurements. In parallel, we also tested the reproducibility of skin thickness ultrasound (STU) measurements. We examined 52 patients (104 limbs) with lymphedema of stages 0-6 (N = 28, 19, 20, 21, 2, 4, and 10, respectively). 3DIS measurements correlated nearly perfectly with WD (r2 = 0.9945) and TMLC values (r2 > 0.9801). The median time required to acquire imaging system measurements for both legs was 2.1 minutes, compared with 17, 7, and 29 minutes, respectively, for WD, TMLC, and STU. Median interexaminer coefficients of variation (CVs) for volume measurements were 1.1% (interquartile range [IQR] 0.5-2.1%) for WD and 1.7% (IQR 1.2-2.4%) for the 3DIS. CVs for circumference measurements were 1.4% (IQR 0.8-2.4%) by TMLC and 1.3% (0.8-1.9%) by 3DIS. Median interexaminer CV for STU was 13.7% (IQR 8.5-21.3%). The portable imaging system noninvasively provided accurate and reproducible limb volume and circumference measurements in approximately 2 minutes per patient. This portable technology has the potential to greatly improve assessment and monitoring of lymphedema in the clinic and in the field.

Case report of Plasmodium ovale curtisi malaria in Sri Lanka: relevance for the maintenance of elimination status.

BACKGROUND: Following its recent certification as malaria-free, imported infections now pose the greatest threat for maintaining this status in Sri Lanka. Imported infections may also introduce species that are uncommon or not previously endemic to these areas. We highlight in this case report the increasing importance of less common malaria species such as Plasmodium ovale in elimination settings and discuss its relevance for the risk of malaria resurgence in the country. CASE PRESENTATION: A 41-year-old patient from southern Sri Lanka was diagnosed with malaria after 8 days of fever. Microscopy of blood smears revealed parasites morphologically similar to P. vivax and the rapid diagnostic test was indicative of non-P. falciparum malaria. He was treated with chloroquine over 3 days and primaquine for 14 days. He was negative for malaria at a one-year follow-up. Molecular testing performed subsequently confirmed that infection was caused by P. ovale curtisi. The patient gave a history of P. vivax malaria treated with chloroquine and primaquine. He also provided a history of travel to malaria endemic regions, including residing in Liberia from May 2012 to November 2013, throughout which he was on weekly malaria prophylaxis with mefloquine. He had also visited India on an eight-day Buddhist pilgrimage tour in September 2014 without malaria prophylaxis. CONCLUSIONS: It is crucial that every case of malaria is investigated thoroughly and necessary measures taken to prevent re-introduction of malaria. Accurate molecular diagnostic techniques need to be established in Sri Lanka for the screening and diagnosis of all species of human malaria infections, especially those that may occur with low parasitemia and are likely to be undetected using the standard techniques currently in use. In addition, ascertaining whether an infection occurred through local transmission or by importation is critical in the implementation of an effective plan of action in the country. This new era emphasizes the global nature of regional malaria elimination. Increasing global surveillance and tool development are necessary in order to "fingerprint" parasites and identify their origin.

A comparison of two tests for filarial antigenemia in areas in Sri Lanka and Indonesia with low-level persistence of lymphatic filariasis following mass drug administration.

BACKGROUND: Filarial antigen tests are key tools for mapping the distribution of bancroftian filariasis and for detecting areas with persistent infections following mass drug administration (MDA). A recent study showed that the new Alere Filariasis Test Strip (FTS) has better analytical sensitivity than the BinaxNOW Filariasis card test (Card Test) for detecting circulating filarial antigen, and the FTS detected more positive results than the Card Test in a field study performed in a highly endemic area in Liberia. METHODS: The present study compared the performance of the FTS and the Card Test in community surveys that were conducted in southern Sri Lanka and in Indonesia (Central Java) in areas with low-level persistence of LF following multiple rounds of MDA with diethylcarbamazine plus albendazole. The studies were performed in densely populated semi-urban areas where Wuchereria bancrofti is transmitted by Culex quinquefasciatus. RESULTS: Antigenemia rates by FTS were 138% higher in the Sri Lanka study (43/852 vs. 18/852) and 21% higher in the Indonesia study (50/778 vs. 41/778) than antigenemia rates by Card Test. Antigenemia rates were significantly higher in males than in females and higher in adults than in children in both study sites. Although overall antigenemia rates and test scores were significantly higher by FTS than by Card Test in both study areas, rates in young children were similar with both tests in both areas. CONCLUSIONS: These results extend the previously reported superior sensitivity of the FTS to areas with low residual infection rates following MDA, and this could affect mapping and post-MDA survey results in adults. However, our findings suggest that results of transmission assessment surveys (TAS) performed in school-aged children are likely to be similar with both tests.

Rapid assessment procedures to detect hidden endemic foci in areas not subjected to mass drug administration in Sri Lanka.

For the declaration of elimination of lymphatic filariasis, reliable epidemiological data in all parts of a country are required. In Sri Lanka, due to social disturbance, there are 3 provinces whose endemicity has been declared unknown. Further, a recent report revealed an endemic pocket, which is on the border with the district that was not covered by the national elimination program. These facts indicate the necessity of more extensive studies to discover hidden endemic foci. To facilitate such studies, we evaluated 2 methods of Rapid Assessment Procedure (RAP) in Hambantota district, where the filariasis endemicity was low: (1) indirect questioning by mailing a questionnaire to each local leader (IndQ), asking about the presence of clinical cases, and (2) focus group discussion (FGD) by villagers. The information given by people was validated with clinical examination by doctors (CE) and IgG4 ELISA using urine samples. In the results: there was a strong positive correlation between CE and ELISA rates. The hydrocele rates obtained by FGD or IndQ were associated significantly with CE rates. The rates by FGD or Cluster-IndQ ('modified' IndQ) were also associated significantly with ELISA rates. The IndQ was most cost-effective. Based on these findings, we have concluded that screening by IndQ and confirmation by the ELISA would be an effective and practical way in Sri Lanka to locate endemic foci in hitherto unsurveyed districts.

Visual detection of filaria-specific IgG4 in urine using red-colored high density latex beads.

The use of urine for the immunodiagnosis of lymphatic filariasis has a definite advantage: the sample collection is not invasive and thus well accepted by people. Urine-based ELISA to detect filaria-specific IgG4 has been used successfully. However, ELISA requires equipment such as a microplate reader, which is often not available in most endemic areas. We have developed a new visual immunodiagnosis that detects urinary IgG4 using red-colored latex beads (bead test). The sensitivity was 87.2% when ICT antigen test positive people were regarded as the standard (136/156), and the specificity was 97.2% with the non-endemic people in Japan and Bangladesh, and the urine ELISA negatives in Sri Lanka (1264/1300). In a prevalence study, the bead test could detect filarial infection more effectively than ICT test among young children in Sri Lanka, indicating the usefulness of the visual test in epidemiological studies.

Development of loop-mediated isothermal amplification method for detecting Wuchereria bancrofti DNA in human blood and vector mosquitoes.

We have developed loop-mediated isothermal amplification (LAMP) method to detect Wuchereria bancrofti DNA. The sensitivity and specificity of LAMP method were equivalent to those of PCR method which detects SspI repeat sequence in W. bancrofti genomic DNA: both methods detected one thousandth of W. bancrofti DNA from one microfilaria (Mf), and did not cross-react with DNAs of Brugia malayi, B. pahangi, Dirofilaria immitis, human and Culex quinquefasciatus. We also examined the sensitivity of LAMP using the mimic samples of patient's blood or blood-fed mosquitoes containing one W. bancrofti Mf per sample. The LAMP method was able to detect W. bancrofti DNA in 1000 µl of blood or in a pool of 60 mosquitoes, indicating its usefulness in detecting/monitoring W. bancrofti infection in humans and vector mosquitoes in endemic areas.

Effects of 5 rounds of mass drug administration with diethylcarbamazine and albendazole on filaria-specific IgG4 titers in urine: 6-year follow-up study in Sri Lanka.

ELISA for filaria-specific IgG4 in urine (urine ELISA) was applied to children in 7 schools in Sri Lanka, before and after 5 rounds of annual mass drug administration (MDA). The pre-treatment IgG4 prevalence in 2002 was 3.20%, which decreased to 0.91% in 2003 after the first MDA (P<0.001), and finally to 0.36% in 2007 after the 5th MDA. Among 5-10 year-old children, the prevalence decreased from 3.37% in 2002 to 0.51% in 2003 (P=0.009). A pattern of IgG4 titer distribution according to age and its yearly change could also provide useful information in drug efficacy analysis. In 2008, new samples from eleven 2006/07 urine ELISA-positive students and their family members (total n=56) were examined by ICT antigen test, microfilaria test, and urine ELISA. No infection was confirmed among them. Urine ELISA will be useful in monitoring elimination/resurgence in a post-MDA low endemic situation.

Presence and gradual disappearance of filaria-specific urinary IgG4 in babies born to antibody-positive mothers: a 2-year follow-up study.

A total of 14 Sri Lankan pregnant women, who were anti-Brugia pahangi urinary IgG4 positive, and their 14 newborn babies were followed up for the urinary antibody for 2 years by enzyme-linked immunosorbent assay. Eight babies showed positive IgG4 reaction, at least once within 4 months after birth. Urinary antibody titers of mothers and their babies measured around the perinatal period showed a significant positive correlation, suggesting that baby's IgG4 was transferred from the mother through the placenta. The IgG4 decreased gradually and became negative in all positive babies by day 339.3 after birth. The present result provides a basis to judge if a positive urine ELISA test among babies is due to a new filarial infection.

Distribution of filarial elephantiasis and hydrocele in Matara district, Sri Lanka, as reported by local leaders, and an immunological survey in areas with relatively high clinical rates.

To eliminate lymphatic filariasis by means of mass drug administration, it is essential to have reliable data on the disease distribution and prevalence in targeted areas. In Matara district, Sri Lanka, self-administered questionnaires were mailed to 2105 local leaders questioning the presence and the numbers of elephantiasis and hydrocele cases. The information provided by them revealed that elephantiasis was clearly aggregated in the southern part of the district along the coast, while hydrocele was distributed rather evenly in the whole district, including Deniyaya region where no endemic filariasis had been known. To confirm active transmission of filariasis in Deniyaya, Wuchereria bancrofti antigen and filaria-specific urinary IgG4 antibody were measured with 2436 subjects. The positive rates for antigen and antibody were 0.6% and 4.3%, respectively. The titer analysis of IgG4 according to age revealed that the youngest IgG4 positive was 3 years old, and that in 10 years old or less, there were 16 positives out of 607 children examined (2.6%). It was concluded that filarial transmission at a low level was going on in the region.

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.

Knowledge on lymphatic filariasis and the response to July 2002 mass treatment campaign in two communities in the Galle district.

OBJECTIVE: To ascertain the knowledge on filariasis and response to the July 2002 mass treatment campaign in two sample populations. DESIGN: Application of pre-tested questionnaire by direct interviews of individuals from randomly selected streets. METHODS: Study areas were a coastal community in Unawatuna (population sample 381), and an inland community in Baddegama (population sample 236) in the Galle district. They were interviewed twice, 4 weeks before the mass drug administration (MDA) and 4 to 7 days after. RESULTS: The sample population of Unawatuna had a greater awareness of the clinical and parasitological features of the disease (p = 0.0003) and the drug treatment (p = 0.00380 than that of Baddegama. Only 5.5% of the combined sample attributed the cause of filariasis to a parasitic worm. However, over 70.0% of them knew that transmission was through mosquito bites. Volunteers formed 87.5% of the work force used for drug distribution in Unawatuna and 70.1% in Baddegama. The balance work force were Grama Niladaris and Public Health Midwives. Drugs were received by 76.9% of the Unawatuna sample compared with 89.0% at Baddegama (p < 0.001). Among those who received the drugs, consumption was 91.8% in Unawatuna and 96.2% in Baddegama. Taking the two communities together adverse effects were experienced by 22.9%. These effects in night time drug consumers (10.4%) were significantly less than in day time consumers (19.8%) (p < 0.005). The adverse effects were sleepiness (37.8%), malaise (28.2%), headache (16.8%), vomiting (5.1%), nausea (4.5%) and fever (3.9%). The message of mass treatment was carried to the community by the people themselves on the eve and on the day of drug distribution. Over 95% agreed that in future programs drugs should be delivered to their homes.