Development of a controlled release formulation of an indigenous insect growth regulator, DPE-28, a substituted diphenylether, for controlling the breeding of Culex quinquefasciatus.

Background & objectives: DPE-28, a substituted diphenyl ether (2,6-ditertiarybutyl phenyl-2’,4’-dinitro phenyl ether) was reported to exhibit promising insect growth regulating activity against Culex quinquefasciatus, the vector of lymphatic filariasis. A controlled release formulation (CRF) of DPE-28 has been developed to control Cx. quinquefasciatus in its breeding habitats. Toxicity of DPE-28, safety to non-target mosquito predators and the release profile of the CRF of DPE-28 are studied and discussed. Methods: The acute oral and dermal toxicity was tested in male and female Wistar rats as per the Organization for Economic Cooperation and Development (OECD) guidelines 425 and 402 respectively. The toxicity of DPE-28 to non-target predators was tested as per the reported procedure from this laboratory. The CRF of DPE-28 was prepared by following the reported procedure developed at this laboratory earlier. The concentration of DPE-28 released from the CRF was monitored by HPLC by constructing a calibration graph by plotting the peak area in the Y-axis and the concentration of DPE-28 in the X-axis. Results: DPE-28 has been tested for acute oral toxicity and found to be moderately toxic with LD50 value of 1098 mg/kg body weight (b.w). The results of the acute dermal toxicity and skin irritation studies reveal that DPE-28 is safe and non-irritant. DPE-28 when tested at 0.4 mg/litre against non-target mosquito predators did not produce any mortality. The release profile of the active ingredient DPE-28 from the CRF by HPLC technique showed that the average daily release (ADR) of DPE-28 ranged from 0.07 to 5.0 mg/litre during first four weeks. Thereafter the matrix started eroding and the ADR ranged from 5 to 11 mg/litre during the remaining 5 wk. The cumulative release of active ingredient showed that > 90 per cent of the active ingredient was released from the matrix. Interpretation & conclusions: The controlled release matrix of DPE-28 was thus found to inhibit the adult emergence (>80%) of Cx. quinquefasciatus for a period of nine weeks. The CRF of DPE-28 may play a useful role in field and may be recommended for mosquito control programme after evaluating the same under field conditions.

Kinetics of microfilaraemia & antigenaemia status by Og(4)C(3) ELISA in bancroftian filariasis.

BACKGROUND & OBJECTIVE: Bancroftian filariasis caused by Wuchereria bancrofti is endemic in many parts of India. In recent years diagnosis of W. bancrofti infection has been revolutionized with the availability of filarial antigen tests, which is important in monitoring success of chemotherapy. We carried out this study to measure microfilariaemia and antigenemia levels in bancroftian microfilariae (mf) carriers at 1 yr follow up after chemotherapy, in lymphoedema patients and in endemic controls from a filariasis endemic area in Tamil Nadu State using Og(4)C(3) ELISA to identify the best marker to assess success of chemotherapy. METHODS: Serum samples were collected from 30 bancroftian microfilaremic (Mf) carriers pre-treatment and at sequential intervals (7,30,60,90,180 and 365 days) following treatment with diethylcarbamazine (DEC:6mg/kg body weight, single dose), 30 lymphoedema patients (without treatment) at periodic intervals, and 68 control subjects (24 endemic normal subjects in filariasis endemic area in Tamil Nadu State, 24 non-endemic normal subjects residing in Chandigarh, India; 5 brugian filariasis, 5 endemic control subject in brugian filariasis endemic area and 10 other disease controls). The circulating antigen of W. bancrofti was measured quantitatively using Og(4)C(3) ELISA kit. RESULTS: In Mf carriers, there was no significant difference in microfilariae count in pre- and post-treatment (PT) samples till day 30 while significant differences were observed in pre- and sequentially collected post-treatment (PT) samples day 60 to 180 (P<0.001), day 365 (P<0.005). However, there was no significant difference in antigenaemia levels between pre-treatment (day 0) and PT samples collected on day 7 onwards till day 365. Though of the 19 patients who could be followed up till 365 days PT, 4 (21%) were amicrofilaraemic, none became antigen negative. No significant difference was found in antigenaemia levels in sequentially collected samples from lymphoedema patients. Significant differences were observed in antigenaemia levels in samples collected at the start of study in mf carriers as compared to lymphoedema patients and endemic normal subjects (P<0.001). Subjects (non-endemic control) residing in filariasis free area (24), brugian endemic area (5), B.malayi infected patients (5) and patients with other parasitic diseases (10) were found antigen negative. INTERPRETATION & CONCLUSION: Annual single dose of DEC therapy alone may not result in complete clearance of infection and detection of antigenaemia rather than microfilaraemia may be taken into consideration as an indicator of successful chemotherapy. The study supports the earlier view that filarial antigenaemia is relatively common in amicrofilaraemic and asymptomatic subjects in endemic areas and further studies are needed to determine the clinical significance, prognosis and effective management of such infections in endemic areas.

Detection of day blood filarial antigens by Og4C3 ELISA test using filter paper samples.

BACKGROUND: The launching of the global filariasis elimination programme has necessitated the use of highly sensitive and specific diagnostic tests. The Og4C3 monoclonal antibody-based ELISA test has been found to be highly specific and sensitive for the diagnosis of filariasis using night blood samples. However, it requires a serum sample which poses problems of transport and storage. Collection of blood samples on filter paper the will greatly circumvent these problems. Therefore, we evaluated the utility of the Og4C3 assay on filter paper samples collected during daytime. METHODS: Blood samples were collected from 63 microfilariae (mf) carriers during different time periods in a day on filter paper discs as well as venous blood for sera. The mf carriers and chronic (hydrocele n = 20; lymphoedema n = 120) and acute filariasis (adenolymphangitis n = 39) patients were from the endemic areas and the non-endemic normals were from Uthagamandalam district of Tamil Nadu, India. The filarial antigens in the samples were determined using the Og4C3 filarial antigen assay as per the manufacturer's instructions (JCU TrapBio, Australia). The sensitivity of the assay on sera and filter paper samples collected during night and also on filter paper samples collected during different time intervals of the day were compared with those of the membrane filtration technique, which was used as a gold standard. RESULTS: The geometric mean titre of the sera samples collected during night was 11 units/ml for non-endemic normals and 601.2 units/ml for mf carriers. The specificity of the assay on sera samples collected during night was 100% and the sensitivity 96.8% and the positive and negative values were 100% and 95.2%, respectively. The antigen positivity of the filter paper samples collected during morning hours was 93.3% while it was 76.6% and 86.7% for afternoon and evening hours. A significant association was observed between antigenaemia levels and mf density in the blood samples collected during the night. CONCLUSION: The samples collected on filter paper during the day can be used as an alternative to sera samples for detection of filarial antigens employing Og4C3 ELISA. Also, samples collected during morning hours yield a higher positivity. The assay when applied to serum samples will be useful especially when quantitative results are required.

Tropical vaginal hydroceles: are they all filarial in origin?

Hydrocele of the tunica vaginalis testis has been conventionally used as an absolute indicator of filarial disease in most clinical surveys. The prevalence of filarial etiology in 100 consecutive hydroceles was studied using clinical, parasitological, histopathological and immunological parameters. Filarial etiology could be proved in 57% of hydrocele cases using major criteria: presence of microfilaria in hydrocele fluid, presence of chyle in hydrocele fluid, demonstration of adult worm in tunica, ratio of fluid antibody titer to serum antibody titer more than 2 and presence of filarial antigen in hydrocele fluid. The results of other tests in these 57 cases were used to define the minor criteria. In the other 43 cases, based on the minor criteria, 12 hydroceles could be classified as likely to be due to filariasis and the rest were probably non-filarial. Thus only 69% of hydroceles were definitely or probably filarial.