Imputation for exposure histories with gaps, under an excess relative risk model

An Onchocerca volvulus expression library was differentially screened to identify a molecular marker distinguishing sowda (lichenified onchodermatitis) from other onchocerciasis forms. One clone, PG3, was recognized by pooled sera from patients with sowda, but not by pooled sera from patients with generalized onchocerciasis; it was not recognized by sera from patients with lymphatic filariasis or other helminth infections. The DNA of PG3 hybridized strongly with O. volvulus Eco RI-digested DNA, but not with DNA from Brugia spp., Trichinella spp., and humans. A weak reaction was observed with DNA from O. gibsoni and Acanthocheilonema viteae. The PG3 DNA sequence showed a high homology with both human and nematode collagens. Confirmation of the collagen-like nature of the sowda-specific PG3 product was obtained by amino terminal sequencing of the PG3 expression product, as well as by demonstrating its susceptibility to collagenase digestion. The characteristic recognition of the O. volvulus collagen specified by clone PG3 was confirmed by measuring antibody levels to the expressed product in individual sowda and generalized onchocerciasis sera, respectively. Identification of a nematode collagen antigen mainly recognized in sowda patients raises the possibility that this extreme form of dermatitis might arise through cross-reactivity between anti-O. volvulus collagen antibodies and human collagen. However, a relationship between the PG3 recognition by antibodies and the sowda pathogenesis could not be demonstrated.     

Immunodiagnostic studies on Onchocerca volvulus and Mansonella perstans infections using a recombinant 33 kDa O. volvulus protein (Ov33)

A total detergent-soluble extract of adult female Onchocerca volvulus (OvAg) and a recombinant O. volvulus protein (Ov33) linked to glutathione-S-transferase (GST) were compared with regard to their serodiagnostic suitability for differentiating between O. volvulus and Mansonella perstans infections in a region endemic for both filarial worms in western Uganda. Using OvAg in an enzyme-linked immunosorbent assay (ELISA), 98.8% sensitivity was obtained examining 84 O. volvulus microfilariae (mf) carriers living in the hyperendemic area. However, 5 of 18 (28%) sera from M. perstans mf carriers without O. volvulus mf, from another area hypoendemic for O. volvulus, cross-reacted with OvAg. Using the recombinant antigen Ov33-GST in an ELISA and Western blot assay, sensitivity for O. volvulus remained high (97.2% and 98.8% respectively) while none of 90 sera from M. perstans mf carriers reacted positively. Both antigens were used to examine a batch of sera from 260 persons living in the onchocerciasis hyperendemic area who did not have mf in their skin snips (9.5% of 2728 persons examined); 116 of these sera (44.6%) were positive in the OvAg ELISA, compared to 85 (32.7%) and 69 (26.5%) which were positive in Ov33-GST ELISA and Ov33-GST Western blot, respectively. Reaction with GST alone was minimal. The recombinant antigen Ov33 efficiently differentiates between O. volvulus and M. perstans infections, and is sensitive when used to detect patent and prepatent or low-level O. volvulus infections.     

Studies on Bancroftian filariasis in Liberia, West Africa. IV. Notes on side effects observed during a diethylcarbamazine treatment campaign in a rural area endemic for Wuchereria bancrofti and Onchocerca volvulus

464 persons of whom 189 proved to be infected with microfilariae of Wuchereria bancrofti and/or Onchocerca volvulus were examined for adverse reactions due to diethylcarbamazine, which was administered during a filariasis control campaign. Persons older than 20 years of age were significantly more affected than younger persons and men showed distinctly more side effects than women. The most frequent complaints were skin reactions followed by gastrointestinal complications. The significantly highest rate of adverse reactions was observed in persons being infected with microfilariae of O. volvulus, whereas no significant difference was registered between side effect rates of bancroftian microfilaria carriers and non-infected persons. It is concluded that in areas where both filarial species are endemic, infections with O. volvulus are a limiting factor for the control of bancroftian filariasis by means of mass treatment with diethylcarbamazine.     

T cell responses in coinfection with Onchocerca volvulus and the human immunodeficiency virus type 1

Onchocerca volvulus and the human immunodeficiency virus (HIV) are two immunocompromising infectious agents of major public health concern in Uganda. To examine the effect of coinfection with O. volvulus and HIV on cellular immune responses, lymphocyte proliferative responses and cytokine production of peripheral blood mononuclear cells (PBMC) from persons infected with O. volvulus with and without HIV type 1 infection were compared. Proliferation of PBMC to PHA and tuberculin (PPD) in coinfection was less (P = 0.08, P < 0.01) than in O. volvulus infection. O. volvulus extract stimulated lymphocyte proliferation in microfilaria-negative and HIV-negative O. volvulus infection while only an inconspicuous response was observed in microfilaria-negative coinfection. After stimulation of PBMC with PPD, the production of interferon-gamma (IFN-gamma), interleukin (IL)-4 and IL-5-demonstrated in O. volvulus infection-were reduced in coinfection with HIV (P < 0.01). While both groups failed to produce IFN-gamma in response to O. volvulus extract, only O. volvulus infected persons generated pronounced IL-5 and low IL-4 levels (0.01 > P = 0.02). The cellular immune responses in coinfection suggested an HIV-related lack of specific reactivity to O. volvulus antigen and impairment of IL-4 and IL-5 production in addition to the lack of IFN-gamma response on antigenic stimulation.     

Onchocerca volvulus provides ligands for the stimulation of human gamma/delta T lymphocytes expressing V delta 1 chains

Peripheral blood mononuclear cells (PBMC) from 8 onchocerciasis patients, treated or not with ivermectin, were analyzed for phenotypic cell surface markers. A significant increase (P < .05) in gamma/delta T cells expressing the V delta 1 chain compared with normal and endemic controls was detected in all patients. PBMC populations from onchocerciasis patients were not expanded after restimulation with Onchocerca volvulus antigens in vitro, but both V delta 1 and V delta 2 T cells from normal donors were increased significantly in response to O. volvulus and Mycobacterium tuberculosis (P < .05), respectively. Frozen sections of all 5 onchocerca nodules tested demonstrated an increased number of CD3+ cells in the vicinity of the adult worm, in all cases expressing the alpha/beta T cell receptor and in 2 patients also expressing the gamma/delta T cell receptor; 60% of T cells expressed the activation marker Ki67. These data suggest that O. volvulus provides ligands to V delta 1 T cells.     

Effect of hemodialysis on leukocyte adhesion receptor expression

Onchocerciasis is associated with blindness and gross skin changes, believed to be a consequence of the immune response to antigens released from the offspring of the female worm of Onchocerca volvulus, the microfilariae (mf). An effective microfilaricidal drug is now available which quickly reduces the mf burden without affecting the adult worm. There exist foci in onchocerciasis endemic areas where some of the patients have many mf in their skin but relatively few clinical symptoms. This state of hyposensitivity is believed to be due to immunosuppression. The aim of this study was to address the question of the basis of, and the effect of ivermectin treatment on this immunosuppression. Female adult worms of O. volvulus were used as whole or fractionated antigens to stimulate peripheral blood mononuclear cells. Microfilariae are found in the reproduction tract of the female worms, and thus an antigen preparation of the female adult O. volvulus contains both exclusive adult antigens as well as antigens from microfilariae. Cells were obtained from onchocerciasis patients, individuals of similar socio-economic status living in the same Ghanaian village, but who showed no parasitological or clinical evidence of onchocerciasis (exposed endemic controls), healthy Ghanaians living in areas where transmission of onchocerciasis does not seem to occur (non-exposed endemic controls) and unexposed healthy Swedish donors. As a group, cells from onchocerciasis patients proliferated to a lesser degree than cells from the exposed endemic control and the non-exposed endemic control groups to the whole worm antigen, whereas the phytohaemagglutinin (PHA) response was strongest in the patients. Proliferative responses of above 1000 ct/min to fractions of the worm extract were only evident in the cells from a few individuals in each of the various groups. However, 28 days following ivermectin treatment, cells from all onchocerciasis patients were able to mount significantly enhanced proliferation to a fraction of approximately 96 kD (fraction 3), while only four of nine of this group showed an increased response to the whole worm antigen. The proportional increase in the response to the whole organism in these individuals was of a much lower magnitude than the increased response to fraction 3. The O. volvulus antigen-specific immunosuppression observed in these onchocerciasis patients appears to be due to suppressive antigens which have the capacity to mask the potential response to selected antigens of O. volvulus, and ivermectin treatment possibly modulates the immune response, allowing for stepwise recognition of such antigens. Since ivermectin treatment kills only the microfilariae and not the adult worm, the putative suppressive antigens would be expected to be from the microfilariae.     

Control of rubella and congenital rubella syndrome (CRS) in developing countries, Part 1: Burden of disease from CRS

A pool of sera from individuals classified as putatively immune (PI) to Onchocerca volvulus infection was employed in the screening of a fourth-stage larval cDNA expression library. A highly immunogenic clone, encoding the Ov 53/80 protein, was identified. The full length cDNA of clone 4.21 contained 2527 nucleotides encoding 769 amino acids of which 100 are glutamine residues (13%). Antibodies raised against recombinant protein encoded by a partial cDNA sequence (clone 73-k) recognized a 53 and 80 kDa protein in O. volvulus larval and adult parasite extracts, respectively. The antibodies localized the native protein in the cuticle, hypodermis, secretory vesicles and in granules of the glandular esophagus of larvae and in the hypodermis and the cuticle of adult worms. The recombinant 73-k polypeptide (r73) was recognized by 90-100% of sera from PI and infected individuals from Liberia, but only by 67% of similar groups from Ecuador. r73 specific IgG2 and IgG3 levels in the PI from Liberia and Ecuador, respectively, were significantly lower than in the infected, whereas the r73 specific IgG1/IgG3 or IgG1/IgG2 in the PI and the infected individuals from Liberia or Ecuador, respectively, were similar. The IgG4 specific antibody response in the PI from Liberia and Ecuador were lower than in the infected. The T-cell proliferative responses to r73 in infected individuals from Cameroon were found to be inversely correlated with their levels of microfilariae.     

Restoration of intraocular pressure after streptococcus endophthalmitis with vitrectomy

We have evaluated the endemicity of the onchocerciasis in two villages of the district of Amou in Togo, Okpahoué et Onglo. We have studied 294 subjects including 136 males and 158 females representing the quarter of the population of the two villages (clinical investigation and exsanguine cutaneous biopsy): 111 persons are parasited by Onchocerca volvulus that means a prevalence rate of 37.7%. We can therefore say that the villages are in a state of "meso-endemicity". We have been able to find nodules by 20 persons among the 111 that give a percentage of 6.8 of the examined persons. Among children of 3 to 9 years old, the prevalence and the average microfilaremia are respectively 8.86 and 3.54%. Among the persons of 50 or older than 50, we found a prevalence rate of 65.95% and an average microfilaremia of 28.37.     

PCR and DNA hybridization indicate the absence of animal filariae from vectors of Onchocerca volvulus in Uganda

In order to identify Onchocerca volvulus larvae from vectors, DNA of filaria larvae from dissected blackflies was isolated, and a 150-bp long tandemly repeated DNA sequence (0-150), which occurs in many Onchocerca species, was amplified using polymerase chain reaction (PCR). Subsequently, the PCR product was blotted onto a nylon membrane and hybridized with DNA probes specific for O. volvulus or Onchocerca ochengi. Filaria larvae from 395 infected Simulium neavei were examined and 259 samples produced detectable PCR products. Among these samples, 239 (92%) reacted with an O. volvulus-specific oligonucleotide. A sample of 69 PCR products was tested using an O. ochengi DNA probe, but all failed to hybridize. Filaria larvae from 64 infected Simulium damnosum, presumably of the cytotypes "Nyamagasani" and "Nkusi" were studied and 0-150 was amplified from 38 samples. From these samples, 35 (92%) hybridized specifically with an O. volvulus probe but none with the O. ochengi-specific DNA sequence. Nonamplified samples were obtained mainly from blackflies that contained only 1 or 2 filaria larvae, and therefore, an insufficient DNA extraction was assumed. It can be concluded that few, if any, filaria species of animal origin were transmitted by S. neavei and S. damnosum s.l. in Kabarole and Kasese districts in Uganda.     

Onchocerciasis

An estimated 17.7 million persons, most of them in Africa, are infected with the parasite Onchocerca volvulus. Onchocerciasis has caused blindness in 270,000 and left another 500,000 with severe visual impairment. Onchocerciasis also can cause disfiguring skin changes, musculoskeletal complaints, weight loss, changes in immune function, epilepsy, and growth arrest. The development of ivermectin and its donation by the manufacturer has reduced transmission in many parts of Africa and Latin America. Progress in developing a drug that can destroy adult worms has been slow, however. The immediate need is to extend ivermectin mass distribution to as many endemic areas as possible on a continuing basis, with steps toward community ownership of such programs. This article presents a comprehensive review of the literature on the epidemiology, parasitology, vector, symptoms, diagnosis, and treatment and control of onchocerciasis. Future research aims include fuller understanding of the parasite and its relationship with the host, the nature of the systemic effects of O. volvulus infection, the natural history of skin disease, and a better appreciation of the social and economic consequences of this parasitic disease.     

The indirect hemagglutination test for onchocerciasis performed with blood collected on filter paper

In Guatemala, an indirect hemagglutination (IHA) test for onchocerciasis, performed with a crude saline extract of Onchocerca volvulus and blood samples taken on filter paper, showed a high level of sensitivity and specificity. IHA titers of blood samples from the ear lobe taken on filter paper and of sera obtained by venipuncture showed a high correlation.     

Tropomyosin implicated in host protective responses to microfilariae in onchocerciasis

A cDNA from adult female Onchocerca volvulus encoding the C-terminal portion of a tropomyosin isoform (termed MOv-14) has been shown previously to confer protective immunity in rodent models of onchocerciasis. The full-length sequence (designated Ov-tmy-1) obtained by PCR amplification, codes for a protein of 33 kDa and shares 91% identity with tropomyosins from other nematodes, falling to 57% identity with human alpha-tropomyosin. Ov-TMY-1 migrates with an apparent molecular mass of 42 kDa on SDS/PAGE and is present in all life-cycle stages, as determined by immunoblotting. Immunogold electron microscopy identified antigenic sites within muscle blocks and the cuticle of microfilariae and infective larvae. Anti-MOv14 antibodies were abundant in mice exhibiting serum-transferable protection against microfilariae conferred by vaccination with a PBS-soluble parasite extract. In contrast, little or no MOv14-specific antibody was present in mice inoculated with live microfilariae, in which resistance is mediated by antibody-independent mechanisms. In human infections, there was an inverse correlation between anti-tropomyosin IgG levels and densities of microfilariae in the skin. Seropositivity varied with the relative endemicity of infection. An immunodominant B cell epitope within Ov-TMY-1 (AQLLAEEADRKYD) was mapped to the N terminus of the MOv14 protein by using sera from protectively vaccinated mice. Intriguingly, the sequence coincides with an IgE-binding epitope within shrimp tropomyosin, believed to be responsible for hypersensitivity in individuals exhibiting allergy to shellfish. IgG and IgE antibodies reacting with the O. volvulus epitope were detected in human infections. It is concluded that antibody responses to tropomyosin may be important in limiting microfilarial densities in a proportion of individuals with onchocerciasis and have the potential to mediate hypersensitivity reactions to dead microfilariae, raising the possibility of a link with the immunopathology of infection.     

Antibody to the Dirofilaria immitis aspartyl protease inhibitor homologue is a diagnostic marker for feline heartworm infections

Feline heartworm disease, caused by the filarial nematode Dirofilaria immitis, has been diagnosed with increased frequency in areas endemic for canine heartworm infection. The routine methods for determining the infection status of dogs, such as identification of circulating microfilariae in blood or identification of circulating antigen in serum, plasma or blood, have proven inadequate for screening cats. The inadequacies are due to the likelihood of single-sex infections and clinical disease during prepatent infections. Current antibody detection methodologies rely on crude or partially purified worm antigen preparations that may result in poor specificity. This report describes the cloning, expression, and diagnostic utility of the D. immitis homologue (PDi33) of the Onchocerca volvulus aspartyl protease inhibitor (Ov33). PDi33 is present in all stages that occur in the mammalian host (microfilariae, L3, L4, adult males, and females) and is released by adults cultured in vitro. An indirect enzyme-linked immunosorbent assay (ELISA) using antibody to recombinant PDi33 as a diagnostic marker for infection in cats was very sensitive and was useful for identifying prepatent infections. Testing of sera from cats infected with common gastrointestinal parasites also indicated excellent specificity. The same ELISA in dogs, although demonstrating reasonable sensitivity and specificity, appeared to be of less value as compared with the currently accepted antigen detection methodologies.     

HIV type 1 subtypes in Malaysia, determined with serologic assays: 1992-1996

We investigated the molecular epidemiology of HIV-1 subtypes in Malaysia among injecting drug users (IDUs) and sexual transmission risk groups, using serologic and genetic techniques. Frozen sera collected at a general hospital, a blood bank, several drug treatment centers, and an STD clinic in Kuala Lumpur, between 1992 and 1996, were investigated retrospectively. V3 peptide serotyping and monomeric gp120 capture serotyping were used to study 89 known HIV-1-infected subjects. The methods differentiate subtypes B, E, and C. V3 peptide and gp120 capture results were comparable. No subtype C-specific reactive sera were found; one specimen was dually reactive for subtypes C and B, using the V3 peptide ELISA; and four were durally reactive for subtypes E and C using this assay. Genotypic analysis of HIV-1 gag RNA in serum was done on a subset of subjects and confirmed serologic findings. HIV-1 subtypes differed significantly by risk category: of 53 IDUs, 29 (55%) were infected with subtype B and 19 (36%) were infected with subtype E, 3 (6%) were dually reactive, and 2 (4%) were not typable. Of 36 persons with heterosexual risks, 29 (81%) were infected with subtype E, 5 (14%) were infected with subtype B, and 2 (5%) were not typable. Persons with IDU risks were significantly more likely to be infected with subtype B than were those with sexual risks (OR 5.89; 95% CI, 1.94-18.54; p < 0.001). Subtypes B and E of HIV-1 appear to predominate in Malaysia; subtype B was more prevalent among IDUs; subtype E was more prevalent among all other groups. These results may have important HIV-1 vaccine implications.     

Onchocerciasis: invasion of deep organs by Onchocerca volvulus

Autopsies of two patients with onchocerciasis in the Republic of Za?re are reported. In one patient elephantiasis who died following diethylcarbamazine therapy there were large numbers of microfilariae of Onchocerca volvulus in the kidney, liver, pancreas, and lung. In the second patient, who also had leprosy and hyperinfection strongyloidiasis, we found an encapsulated adult O. volvulus in the wall of the thoracic aorta.     

Observations on Onchocerca volvulus in experimentally infected chimpanzees

The paper describes observations made on 32 chimpanzees experimentally infected with Onchocerca volvulus. The mean pre-patent intervals for the Cameroon forest and the Guatemalan strains of O. volvulus were 13-16 months and 12-15 months respectively. That for the Cameroon Sudan-savanna strain was much longer, i.e. 22-23 months. The numbers of microfilariae found in the skins of animals infected with the Cameroon Sudan-savanna strain were also much lower than in animals infected with the other two strains. Long-term observations on infected animals showed that microfilarial infections had virtually died out 6.5-9 years after the last inoculation with infective larvae. Those animals which were inoculated with infective larvae in the head or above the waist tended to show a higher proportion of microfilariae in the upper parts of the body, than did those inoculated with infective larvae below the waist. In animals which showed adult worm-bundles on only one side of the body, the concentration of microfilariae was usually greater on that side of the body. Worm-bundles in the chimpanzee varied in size from 8 x 5 x 2 mm to 4 x 3 x 2 cm. Out of 47 worm-bundles found, only two were subcutaneous. The remainder lay deep in the tissues, most commonly adjacent to the posterior surface of the capsule of the hip joint. No onchocerciasis eye lesions were seen in any of the infected animals.     

Molecular analysis of Shiga toxigenic Escherichia coli O111:H- proteins which react with sera from patients with hemolytic-uremic syndrome

Western blot analysis was used to assess the reactivity of convalescent-phase sera from patients who were associated with an outbreak of hemolytic-uremic syndrome (HUS) caused by fermented sausage contaminated with Shiga toxin-producing Escherichia coli (STEC). The predominant STEC isolated from HUS patients belonged to serotype O111:H-, and reactivity to O111:H- whole-cell lysates, treated or untreated with proteinase K, was examined. As expected, all five serum samples demonstrated a marked anti-lipopolysaccharide response, but several protein bands were also immunoreactive, particularly one with an apparent size of 94 kDa. One convalescent-phase serum sample was subsequently used to screen an O111:H- cosmid bank and 2 of 900 cosmid clones were found to be positive, both of which contained a similar DNA insert. Western blot analysis of one of these clones identified three major immunoreactive protein bands of approximately 94, 70, and 50 kDa. An immune response to the three proteins was detectable with all five convalescent-phase serum samples but not with healthy human serum. Immunoreactive 94- and 50-kDa species were produced by a deletion derivative of the cosmid containing a 7-kb STEC DNA insert. Sequence analysis of this region indicated that it is part of the locus for enterocyte effacement, including the eaeA gene which encodes intimin. The deduced amino acid sequence of the O111:H- intimin was 88.6% identical to intimin from O157:H7 STEC, and the most divergent region was the 200 residues at the carboxyl terminus, which were only 75% identical. Such variation may be antigenically significant as serum from a HUS patient infected only with the O111:H- STEC reacted with intimin from an enteropathogenic E. coli O111 strain, as well as several other eaeA-positive STEC isolates, but not with an eaeA-positive STEC belonging to serotype O157:H-. Sera from two of the other HUS patients also failed to react with intimin from this latter strain. However, intimin from O157:H- STEC did react with serum from a patient infected with both O111:H- and O157:H- STEC.     

Signal transduction of the erythropoietin receptor

Several studies have been carried out on the transmission of Onchocerciasis by Simulium damnosum s.l. in the forest zone of C?te d'Ivoire. This study, carried out in 1979-1980 was devoted to determine the risk of onchocerciasis transmission inside and outside the rain forest of Ta? (5 degrees 50' N-7 degrees 25' W). We present the vectorial capacity of S. sanctipauli in the region of Ta? before massive flow of refugees from areas of Liberia without any control Programme. The results of micromorphological technics for determination of S. damnosum adults, showed that mainly females of S. sanctipauli were present. The studied populations had low parturity rates: 39.2% outside and 30.9% of parous flies inside the rain forest. The parasitic rates (0.4% of infectious females outside and 0.1% inside) and their parasitic loads (15 and 3 infective larvae per 1000 parous female respectively outside and inside the rain forest) were low, consequently their vectorial capacity with Onchocerca volvulus was almost non-existent in natural conditions. Before massive flow of refugees including persons carrying microfilariae, there were no problem of onchocerciasis within and outside the rain forest of Ta?. However, the massive flow of refugees and the deforestation for growing crops can create situations favourable to the installation of more efficient vectors, increase man/vector contact and contribute to more intense onchocerciasis transmission. The monitoring of onchocerciasis transmission is necessary.     

An enzyme-linked immunosorbent assay (ELISA) for monitoring guineapigs and rabbits for Bordetella bronchiseptica antibodies

An enzyme-linked immunosorbent assay (ELISA) for monitoring antibodies specific to Bordetella bronchiseptica in guineapigs and rabbits was developed. In conventional and SPF colonies of guineapigs and rabbits, the ELISA was equally successful in detecting infected animals when compared to selective cultivation from the respiratory tract. The ELISA showed a sensitivity of 100% and a specificity of 90% in guineapigs. In rabbits the sensitivity and specificity of the ELISA were 97% and 91% respectively. In rabbit sera from infected colonies, ELISA activity showed a statistically significant correlation with titres obtained in the micro-agglutination test. Since serologically unrelated strains of the bacterium exist, the monitoring of animals for B. bronchiseptica infection by ELISA should be performed with various antigens.