Brugia Malayi Infection Research Articles

Eosinophil-Mediated Immune Control of Adult Filarial Nematode Infection Can Proceed in the Absence of IL-4 Receptor Signaling.

Helminth infections are accompanied by eosinophilia in parasitized tissues. Eosinophils are effectors of immunity to tissue helminths. We previously reported that in the context of experimental filarial nematode infection, optimum tissue eosinophil recruitment was coordinated by local macrophage populations following IL-4R-dependent in situ proliferation and alternative activation. However, in the current study, we identify that control of chronic adult filarial worm infection is evident in IL-4Rα-deficient (IL-4Rα-/-) mice, whereby the majority of infections do not achieve patency. An associated residual eosinophilia was apparent in infected IL-4Rα-/- mice. By treating IL-4Rα-/- mice serially with anti-CCR3 Ab or introducing a compound deficiency in CCR3 within IL-4Rα-/- mice, residual eosinophilia was ablated, and susceptibility to chronic adult Brugia malayi infection was established, promoting a functional role for CCR3-dependent eosinophil influx in immune control in the absence of IL-4/IL-13-dependent immune mechanisms. We investigated additional cytokine signals involved in residual eosinophilia in the absence IL-4Rα signaling and defined that IL-4Rα-/-/IL-5-/- double-knockout mice displayed significant eosinophil deficiency compared with IL-4Rα-/- mice and were susceptible to chronic fecund adult filarial infections. Contrastingly, there was no evidence that either IL-4R-dependent or IL-4R-independent/CCR3/IL-5-dependent immunity influenced B. malayi microfilarial loads in the blood. Our data demonstrate multiplicity of Th2-cytokine control of eosinophil tissue recruitment during chronic filarial infection and that IL-4R-independent/IL-5- and CCR3-dependent pathways are sufficient to control filarial adult infection via an eosinophil-dependent effector response prior to patency.

Deteksi filariasis dan vektornya di wilayah kerja Dinas Kesehatan Kabupaten Bantul, Daerah Istimewa Yogyakarta

In Bantul, Yogyakarta Special Region there have been reported 6 elephantiasis cases and ware suspected as filariasis. Elephantiasis is classified into two, namely filarial elephantiasis and nonfilarial elephantiasis. Filarial elephantiasis caused by lymphatic filarial worms infection and nonfilarial elephantiasis can be caused by podoconiosis, leprosy, tuberculosis, or chlamydia infection. The aim of the study was to ascertain whether elephantiasis cases are caused by filariasis. Activities carried out in the work area of the Bantul District Health Office in July 2016 i.e.: (1) patient location survey, (2) interviews with patients, their families, and surrounding communities, (3) examination of patients, and (4) observe the environment around the patient's residence to ensure existence of vector mosquito breeding places. This study found two people with elephantiasis, patients from Depok, Gilangharjo, Pandak, Bantul was suspected elephantiasis due to Brugia malayi infection. Patient from Cawan, Argodadi, Sedayu, Bantul was suspected elephantiasis due to podoconiosis. In Cawan found many breeding sites for the Anopheles vagus mosquito.

Brugia malayi infection in ferrets - A small mammal model of lymphatic filariasis.

BackgroundThe lack of effective short-course therapies for treatment of the adult stage of filarial worms is a major limitation in the global effort to eliminate lymphatic filariasis. Studies using current small mammal models of lymphatic filariasis are limited by difficulties in quantifying adult worm numbers and in assessing lymphatic anatomy and function.Methodology/Principal findingsHere, we re-established Brugia malayi infection of ferrets as a model for lymphatic filariasis and demonstrated parasitological, immunological, and histological parallels with human infection. Subcutaneous injection of L3 larvae into a hind-footpad resulted in a mean of 18 adult worms recovered 16 weeks post-infection, primarily from the draining inguinal and femoral lymphatics of the injected limb. Infected ferrets developed microfilaremia, with patency lasting from 12–26 weeks post-infection. Quantitative PCR assessing cytokine transcription by antigen-stimulated lymph node cells demonstrated a mixed Th1/Th2 response occurring during early infection. Immunoregulation with production of down-regulatory cytokine IL-10 occurred just prior to peak microfilaremia. Histological analysis revealed progressive inflammation of the lymphatic vessel walls, with intimal thickening and disorganization of collagen fibers. Inflammation was observed as early as 8 weeks post-infection and extended into the perivascular and subcutaneous tissues by 16 weeks post-infection. Finally, we developed a novel ferret PET/CT lymphoscintigraphy method demonstrating substantial changes in lymphatic anatomy and function as early as 3 weeks post-infection, with progression over the course of infection.Conclusions/SignificanceB. malayi infection of ferrets is a robust model of human lymphatic filariasis that can be utilized to study efficacy of novel antifilarial agents against adult worms residing within lymphatic vessels. In conjunction with PET/CT lymphoscintigraphy, this model can also be used to investigate pathogenesis of lymphatic dysfunction in lymphatic filariasis and efficacy of medications aimed at reversing lymphatic dysfunction after clearance of adult worms.

Canine parasitic zoonoses in India: status and issues.

Dogs play valuable roles in human society. In addition to serving as pets and companions, dogs have also been important in hunting and, in recent times, as therapy animals. In India, the number of pet dogs is estimated to be around 5 million. The stray dog population in India is estimated to be 19 million and still increasing, due to ineffective control measures. Stray dogs pose substantial risks to public health due to injury and transmission of zoonoses such as rabies. Both pet and stray dogs may act as reservoirs of zoonotic parasites in India, which has a climate conducive to the environmental survival and transmission of many zoonotic parasites. At present, visceral larva migrans, cutaneous larva migrans and echinococcosis are the most important parasitic zoonoses in India. Leishmaniosis, dirofilariosis, Brugia malayi infection and giardiosis are potentially significant emerging parasitic zoonoses, and theleziosis, gnathostomiosis and dipylidiosis occur sporadically. Because of their biomedical and public health significance, and the lack of literature and compiled data on parasitic zoonoses of dogs in India, the authors provide a concise review on this topic along with potential control strategies.

Vaccination of Gerbils with Bm-103 and Bm-RAL-2 Concurrently or as a Fusion Protein Confers Consistent and Improved Protection against Brugia malayi Infection.

BackgroundThe Brugia malayi Bm-103 and Bm-RAL-2 proteins are orthologous to Onchocerca volvulus Ov-103 and Ov-RAL-2, and which were selected as the best candidates for the development of an O. volvulus vaccine. The B. malayi gerbil model was used to confirm the efficacy of these Ov vaccine candidates on adult worms and to determine whether their combination is more efficacious.Methodology and Principle FindingsVaccine efficacy of recombinant Bm-103 and Bm-RAL-2 administered individually, concurrently or as a fusion protein were tested in gerbils using alum as adjuvant. Vaccination with Bm-103 resulted in worm reductions of 39%, 34% and 22% on 42, 120 and 150 days post infection (dpi), respectively, and vaccination with Bm-RAL-2 resulted in worm reductions of 42%, 22% and 46% on 42, 120 and 150 dpi, respectively. Vaccination with a fusion protein comprised of Bm-103 and Bm-RAL-2 resulted in improved efficacy with significant reduction of worm burden of 51% and 49% at 90 dpi, as did the concurrent vaccination with Bm-103 and Bm-RAL-2, with worm reduction of 61% and 56% at 90 dpi. Vaccination with Bm-103 and Bm-RAL-2 as a fusion protein or concurrently not only induced a significant worm reduction of 61% and 42%, respectively, at 150 dpi, but also significantly reduced the fecundity of female worms as determined by embryograms. Elevated levels of antigen-specific IgG were observed in all vaccinated gerbils. Serum from gerbils vaccinated with Bm-103 and Bm-RAL-2 individually, concurrently or as a fusion protein killed third stage larvae in vitro when combined with peritoneal exudate cells.ConclusionAlthough vaccination with Bm-103 and Bm-RAL-2 individually conferred protection against B. malayi infection in gerbils, a more consistent and enhanced protection was induced by vaccination with Bm-103 and Bm-RAL-2 fusion protein and when they were used concurrently. Further characterization and optimization of these filarial vaccines are warranted.

Real-time PCR detection of the HhaI tandem DNA repeat in pre- and post-patent Brugia malayi Infections: a study in Indonesian transmigrants.

BackgroundLymphatic filariasis caused by Wuchereria bancrofti or Brugia spp. is a public health problem in developing countries. To monitor bancroftian filariasis infections, Circulating Filarial Antigen (CFA) test is commonly used, but for brugian infections only microfilariae (Mf) microscopy and indirect IgG4 antibody analyses are available. Improved diagnostics for detecting latent infections are required.MethodsAn optimized real-time PCR targeting the brugian HhaI repeat was validated with plasma from microfilariae negative Mongolian gerbils (jirds) infected with B. malayi. Plasma samples from microfilaremic patients infected with B. malayi or W. bancrofti were used as positive and negative controls, respectively. PCR results of plasma samples from a transmigrant population in a B. malayi endemic area were compared to those of life-long residents in the same endemic area; and to IgG4 serology results from the same population. To discriminate between active infections and larval exposure a threshold was determined by correlation and Receiver-Operating Characteristics (ROC) curve analyses.ResultsThe PCR detected HhaI in pre-patent (56 dpi) B. malayi infected jirds and B. malayi Mf-positive patients from Central Sulawesi, Indonesia. HhaI was also detected in 9/9 elephantiasis patients. In South Sulawesi 87.4% of the transmigrants and life-long residents (94% Mf-negative) were HhaI PCR positive. Based on ROC-curve analysis a threshold for active infections was set to >53 HhaI copies/μl (AUC: 0.854).ConclusionsThe results demonstrate that the HhaI PCR detects brugian infections with greater sensitivity than the IgG4 test, most notably in Mf-negative patients (i.e. pre-patent or latent infections).

Antifilarial activity of marine sponge Haliclona oculata against experimental Brugia malayi infection

The present study incorporates the findings on in vitro and in vivo antifilarial activity in the marine sponge, Haliclona oculata using an experimental rodent infection of human lymphatic filarial parasite, Brugia malayi. The in vitro antifilarial action was determined on both adult female worms as well as microfilariae using two parameters viz. adverse effect on motility and inhibition in MTT reduction by the treated adult parasite over control worm. The antifilarial activity could be located in the methanol extract and one of its four fractions (chloroform). Bioactivity guided fractionation of chloroform fraction led to localization of in vitro activity in one of its eight chromatographic fractions. Methanol extract, chloroform fraction and one of the chromatographic fractions revealed IC50 values of 5.00, 1.80, and 1.62μg/ml, respectively when adult B. malayi were exposed to these test samples for 72h at 37°C. Under similar exposure conditions, the IC50 values for microfilariae were 1.88, 1.72 and 1.19μg/ml, respectively. The active test samples were found to be safe revealing >10 selectivity indices (SI) on the basis of cytotoxicity to Vero cells (monkey kidney cells) and therefore selected for in vivo evaluation against primary (adult B. malayi intraperitoneal transplanted jird) and secondary (subcutaneous infective larvae induced mastomys) screens. In primary jird model, the three test samples at 100mg/kg for five consecutive days by subcutaneous route demonstrated significant macrofilaricidal efficacy to the tune of 51.3%, 64% and 70.7% by methanol extract, chloroform and chromatographic fraction, respectively. The three samples demonstrated 45–50% macrofilaricidal activity with moderate embryostatic effect in secondary model at 5×500, 5×250 and 5×125mg/kg by oral route. Chromatographic fraction possessing highest antifilarial action was primarily found to be a mixture of four alkaloids Mimosamycin, Xestospongin-C, Xestospongin-D and Araguspongin-C in addition to few minor compounds.

Antifilarial drugs, in the doses employed in mass drug administrations by the Global Programme to Eliminate Lymphatic Filariasis, reverse lymphatic pathology in children withBrugia malayiinfection

Lymphatic filariasis is increasingly viewed as the result of an infection that is often acquired in childhood. The lymphatic pathology that occurs in the disease is generally believed to be irreversible. In a recent study in India, Doppler ultrasonography and lymphoscintigraphy were used to explore subclinical pathology in 100 children from an area endemic for Brugia malayi infection. All the children investigated showed some evidence of current or previous filarial infection. Some were microfilaraemic but asymptomatic, some were amicrofilaraemic but had filarial disease or a past history of microfilaraemia and/or filarial disease, and the rest, though amicrofilaraemic, asymptomatic and without any history of microfilaraemia or filarial disease, were seropositive for antifilarial IgG(4) antibodies. All the children were treated every 6 months, with a single combined dose of diethylcarbamazine (6 mg/kg) and albendazole (400 mg), and followed up for 24 months. By the end of this period all but one of the children were amicrofilaraemic and the 'filarial dance sign' could not be detected in any of the 14 children who had initially been found positive for this sign. Although lymphoscintigraphy revealed lymph-node and lymph-vessel damage in 82% of the children at enrolment, in about 67% of the children this pathology was markedly reduced by the 24-month follow-up. These results indicate that the drug regimens used in the mass drug administrations run by the Global Programme to Eliminate Lymphatic Filariasis are capable of reversing subclinical lymphatic damage and can provide benefits other than interruption of transmission in endemic areas. The implications of these findings are presented and discussed.

The n-hexane and chloroform fractions of Piper betle L. trigger different arms of immune responses in BALB/c mice and exhibit antifilarial activity against human lymphatic filarid Brugia malayi

Modulation of immune functions by using herbal plants and their products has become fundamental regime of therapeutic approach. Piper betle Linn. (Piperaceae) is a widely distributed plant in the tropical and subtropical regions of the world and has been attributed as traditional herbal remedy for many diseases. We have recently reported the antifilarial and antileishmanial efficacy in the leaf extract of Bangla Mahoba landrace of P. betle which is a female plant. The present report describes the in vivo immunomodulatory efficacy of the crude methanolic extract and its n-hexane, chloroform, n-butanol fractions of the female plant at various dose levels ranging between 0.3 and 500 mg/kg in BALB/c. Attempts were also made to observe antifilarial activity of the active extracts and correlate it with the antigen specific immune responses in another rodent Mastomys coucha infected with human lymphatic filarial parasite Brugia malayi. The crude methanol extract and n-hexane fraction were found to potentiate significant ( p < 0.001) enhancement of both humoral (plaque forming cells, hemagglutination titre) as well as cell-mediated (lymphoproliferation, macrophage activation, delayed type hypersensitivity) immune responses in mice. The flow cytometric analysis of splenocytes of treated mice indicated enhanced population of T-cells (CD4 +, CD8 +) and B-cells (CD19 +). The n-hexane fraction (3 mg/kg) was found to induce biased type 2 cytokine response as revealed by increased IL-4 + and decreased IFN-γ + T-cell population while the chloroform fraction (10 mg/kg) produced a predominant type 1 cytokines. Crude methanolic extract (100 mg/kg) demonstrated a mixed type 1 and type 2 cytokine responses thus suggesting a remarkable immunomodulatory property in this plant. The induction of differential T-helper cell immune response appears ideal to overcome immunosuppression as observed in case of lymphatic, filarial Brugia malayi infection which may also be extended to other infections as well.

Vaccination with Setaria cervi 175 kDa collagenase induces high level of protection against Brugia malayi infection in jirds

A zinc containing metalloprotease, 175 kDa collagenase, purified from adult female Setaria cervi showed strong cross-reactivity with sera from putatively immune (PI) individuals (unpublished observation) and induced cytotoxicity to B. malayi L3 larvae and microfilariae by ADCC mechanism [Srivastava Y, Bhandari YP, Reddy MVR, Harinath BC, Rathaur S. An adult 175 kDa collagenase antigen of Setaria cervi in immunoprophylaxis against Brugia malayi. J Helminth 2004;78:347–52]. These preliminary observations suggested the immunoprotective nature of collagenase. To confirm the vaccine potential of this protease, a vaccine trial was conducted in jirds ( Meriones unguiculatus) against human filarial parasite B. malayi. The vaccination resulted into a mean protection level of 75.86% and produced high level of protease neutralizing antibodies. Cytokine analysis in immune jirds sera suggested a mixed Th1/Th2 type cellular immune response whereas ELISA, immunoblotting and enzyme antibody inhibition assay revealed the presence of specific anti-collagenase antibodies. Taken together, all these results suggest that S. cervi 175 kDa collagenase could form the basis of an effective molecular vaccine against human lymphatic filariasis.

Protection against L 3 induced Brugia malayi infection in Mastomys coucha pre-immunized with BmAFII fraction of the filarial adult worm

The present study was aimed at investigating protective efficacy of BmAFII (Sephadex G-200 eluted fraction of Brugia malayi adult worm extract) against establishment of infective larvae (L 3)-induced B. malayi infection in Mastomys coucha and to delineate immunological responses induced in the host. Healthy male M. coucha were immunized with BmAFII and subsequently inoculated with B. malayi L 3. Specific IgG and cell mediated immune responses (cellular proliferation) including release of tumor necrosis factor-α (TNF-α), interferon-γ (IFN-γ), transforming growth factor-β (TGF-β) and nitric oxide (NO) from host's cells stimulated with BmAFII or lipopolysaccharide (LPS)/concanavalin A (Con A) in vitro were determined. Immunization with BmAFII reduced the adult worm recovery by 85.7% ( P < 0.001) and microfilaraemia by 77–95% of unimmunized controls ( P < 0.05–0.01). Immunization alone resulted in downregulation of responses of cellular proliferation, IFN-γ, TNF-α and NO production ( P < 0.01) but increased TGF-β release ( P < 0.001) whereas the converse was seen after L 3 inoculation in these animals. In unimmunized + L 3 inoculated animals all the above parameters were found downregulated ( P < 0.01–0.001). The cell proliferative response of BmAFII immunized + L 3 challenged animals was larger for Con A ( P < 0.001) but not for BmAFII. Specific IgG levels were higher in immunized, immunized + L 3 inoculated and unimmunized + L 3 inoculated groups ( P < 0.001) compared to unimmunized animals, the highest level being shown by immunized + L 3 inoculated group. In conclusion, immunization with BmAFII suppresses establishment of L 3-induced infection in M. coucha by stimulating proinflammatory responses to L 3.

Brugia malayi infection in Meriones unguiculatus: antibody response to recombinant BmR1

BmR1 recombinant antigen has previously been shown to demonstrate high sensitivity and specificity in the serological diagnosis of brugian filariasis in humans. In this study, the pattern of recognition of antibody to BmR1 during Brugia malayi infection was investigated by employing Meriones unguiculatus as the experimental model. Thirty two gerbils were infected subcutaneously with 120 L 3; and two control groups each comprising 25 animals were employed. ELISA using BmR1 was used to detect filaria-specific IgG antibodies elicited by the gerbils; using sera collected from the day 1 until day 150 post-inoculation (p.i.). The results showed that BmR1 detected B. malayi infection in gerbils harboring adult worms irrespective of the presence of circulating microfilaria, and was exemplified by positive ELISA results in nine a microfilaraemic animals that harbored live adult worms. The initial time of the antibody recognition was at day 8 p.i. and the antibody titre showed some correlation with adult worm burden.

A survey of Brugia malayi infection on the Heugsan Islands, Korea.

Lymphatic filariasis due to Brugia malayi infection was endemic in several areas of South Korea. The infection was controlled, or disappeared, in most areas, with the exception of the remote southwestern islands of Jeonranam-do, including the Heugsan Islands. To discover its current situation, a small-scale survey was performed on the Heugsan Islands in September 2000. A total of 378 people, 151 male and 227 female, living in 8 villages (6 on Daeheugsan-do, 1 on Daejang-do, and 1 on Yeongsan-do) were subjected to a night blood survey for microfilaremia, and physical examination for elephantiasis on the extremities. There were 6 (1.6%) microfilaria positive cases, all in females aged 57-72 years, and from only two villages of the Daeheugsan-do area. There were 4 patients with lower leg elephantiasis, but they showed no microfilaremia. The results show that a low-grade endemicity of filariasis remains on the Daeheugsan-do.

Severe reactions to filarial chemotherapy and release of Wolbachia endosymbionts into blood

Wolbachia bacteria seem to have evolved as essential endosymbionts of their filarial nematode hosts. Studies in mice have suggested that these bacteria are associated with systemic inflammatory reactions to filarial chemotherapy. We took blood samples from 15 Indonesian patients before and after treatment with diethylcarbamazine for Brugia malayi infection, and recorded the severity of any post-treatment inflammatory reactions. Blood from all three patients with severe adverse reactions and from one of six with moderate reactions was positive for Wolbachia DNA 4-48 h after diethylcarbamazine treatment. We suggest that these severe inflammatory reactions are associated with the release of endosymbionts into the blood after treatment for filariasis.

Course of Brugia malayi Infection in C57BL/6J NOS2 +/+ and −/− Mice

Ganley, L., Babu, S., and Rajan, T. V. 2001. Course of Brugia malayi infection in C57BL/6J NOS2 +/+ and −/− mice. Experimental Parasitology98, 35–43. Previous results from our laboratory using pharmacological approaches suggested a role for nitric oxide (NO) in the host defense against the human filarial parasite, Brugia malayi. We sought to determine whether a complementary genetic approach, using mice homozygous for a targeted mutation in the gene encoding inducible nitric oxide-synthase (NOS2), would confirm our observation. We hypothesized that such mice would exhibit some deficit in their ability to clear B. malayi. Our data show that the course of infection in NOS2−/− mice is the same as in wild-type mice. Thus, peritoneal cellular responses to infection are similar in NOS2−/− and wild-type mice, with the exception that T cells form a higher percentage of total peritoneal cells in the former. We find virtually no serum IgE in NOS2−/− mice, suggesting a less robust Th2 response. In contrast, NOS2−/− mice demonstrate an early rise in IgG2a titers compared to B6 +/+ mice. Our data suggest that NO is not an obligate requirement for the elimination of B. malayi from the peritoneal cavities of mice.

Clustering of Brugia malayi infection in a community in South-Sulawesi, Indonesia.

The present study was undertaken in village Karondang in South-Sulawesi, Indonesia, to investigate the influences of genetic, household and environmental factors on Brugia malayi infection. Infection status was determined by measuring both microfilariae in night blood and anti-filarial IgG4, as a marker for detection of active filarial infection. A total of 171 residents participated in the study; familial relationships between subjects were registered to construct pedigrees and distances between households were measured. The data were analysed using a test statistic for familial aggregation. For distribution of microfilariae over the study population a genetic influence on infection susceptibility was favoured over the household and environmental effects. For anti-filarial IgG4, all 3 clustering models gave significant results, suggesting that genetic, household and/or environmental factors influence specific IgG4 antibodies.

Immunity in experimental murine filariasis: roles of T and B cells revisited.

We have reevaluated the contributions of T and B cells in Brugia malayi infection by utilizing knockout mice on a uniform background (C57BL/6J). We find that B-cell-deficient mice are more permissive to infection than T-cell-deficient mice.

Molecular characterization of a cDNA encoding an excretory–secretory antigen from Toxocara canis second stage larvae and its application to the immunodiagnosis of human toxocariasis

The cDNA encoding an excretory–secretory antigen from the second stage larvae of Toxocara canis has been characterized. Sequence analysis revealed an open reading frame encoding a protein of 226 amino acid residues ( M r=24 398). Sequence database searches showed similarities to regions corresponding to epidermal growth factor-like and lectin-like domains of the core proteins of vertebrate chondroitin sulfate proteoglycans, which are major components of the extracellular matrix. The T. canis core protein was expressed as a fusion protein with thioredoxin A using an Escherichia coli expression system, and then affinity purified on a metal affinity resin in the presence of 8 M urea. When the purified recombinant T. canis protein was used as an antigen, immunoblot analysis revealed the protein specifically reacted with sera from toxocariasis patients. The antigenic protein did not react with sera from patients with Brugia malayi infection, dirofilariasis, or ascariasis. In some cases of anisakiasis, cross-reactions were observed; however, the cross-reacting bands disappeared when anisakiasis sera preabsorbed with Anisakis antigen were used, indicating that the recombinant T. canis protein is very promising for use as an immunodiagnostic antigen for human toxocariasis.

PCR- elisa for the detection of Brugia malayi infection using finger-prick blood

A polymerase chain reaction assay based on the enzyme-linked immunosorbent assay (PCR- elisa) has been developed to detect Brugia malayi infection in an area of low endemicity in Malaysia. Blood samples from 239 subjects were tested: 192 amicrofilaraemic individuals, 14 microfilaraemic persons and 3 chronic elephantiasis cases from endemic areas and 30 city-dwellers (non-endemic controls). PCR products were examined by elisa and Southern hybridization. In the PCR- elisa, digoxigenin-labelled PCR products were hybridized to a biotin-labelled probe. This was followed by incubation in streptavidin-coated microtitre wells and detection using anti-digoxigenin-peroxidase and ABTS TM [2,2′-azinobis(3-ethylbenzthiazoline-6-sulphonic acid)]. All microfilaraemic samples were positive by PCR- elisa and Southern hybridization and all samples from non-endemic subjects and chronic elephantiasis patients were negative. The PCR- elisa detected 12 times as many B. malayi infections as did thick blood film examination. Nineteen of the 194 samples from the endemic area gave positive results by both PCR- elisa and Southern hybridization, and an additional 5 samples were positive by PCR- elisa only. The PCR- elisa was specific and sensitive, detected more infections, and was more reproducible than Southern hybridization.

Use of antifilarial IgG4-ELISA to detect Brugia malayi infection in an endemic area of Malaysia.

To evaluate the usefulness of antifilarial IgG4 antibody assay in detecting B. malayi infection in a filaria endemic area in Malaysia. A sandwich ELISA using B. malayi soluble antigen was employed to detect antifilarial IgG4 antibodies in serum samples of 330 individuals who comprised 88 healthy individuals from nonendemic areas, 15 B. malayi microfilaraemic cases, 22 individuals with soil-transmitted helminthiases, 9 elephantiasis cases and 196 residents from a B. malayi-endemic area. An O.D. value of > 0.420 at serum dilution of 1:400 was used as the cut-off point. This cut-off point was obtained by taking the mean optical density (0.252 + 4 S.E.) of 36 negative sera which had O.D. values greater than 0.1 at serum dilution of 1:400. All 15 microfilaraemic persons were positive for antifilarial IgG4 antibody. Non-endemic normals, soil-transmitted helminth infected persons and chronic elephantiasis cases were negative for antifilarial IgG4 antibody. Of the 196 individuals from the filaria endemic area, 37 (18.8%) demonstrated presence of antifilarial IgG4 antibodies; and only eight individuals (4.1%) were positive for microfilariae. All eight microfilaraemic individuals were also positive for antifilarial IgG4 antibodies. Antifilarial IgG4-ELISA could detect 4.6 times more positive cases than the microfilaria detection method. With appropriate cut-off values that eliminate cross-reactivities, this serological tool is very useful for Brugia malayi prevalence surveys and diagnosis.