The advantages of mosquito biocontrol by stocking edible fish in rice paddies.

Edible fish stocked in rice fields at a density of 600-800 fry per mu (1 mu = 1/15 hectare) for 150-170 days may act as an effective mosquito biocontrol agent. Common carp (Cyprinus carpio), grass carp Ctenopharyngodon idella) and Tilopia spp. killed late stage larvae and pupae of Anopheles sinensis and Culex tritaeniorrhyncus in laboratory and field trials. Stocking of fish in experimental rice fields decreased larval numbers significantly in comparison with control areas. Expansion of fish stocking in rice fields on a large scale over several years correlated with a marked decrease in malaria transmission. The addition of fish to the rice fields also resulted in increased yields. A ditch-ridge system of field arrangements is described for optimization of fish handling. Preliminary cost-benefit analysis indicates that this approach to mosquito control conveys considerable economic advantage and thus provides incentive to the community to participate in vector control programs. Farmers' experience in Guangxi over a number of years indicates that the use of edible fish for this purpose can be carried on a large, commercially viable scale.

Antigenic determinants of the chlamydial major outer membrane protein resolved at a single amino acid level.

Antigenic determinants were identified from seven chlamydial major outer membrane proteins by using overlapping hexapeptides and polyclonal antisera. Sixty-one determinants were detected, and 30 were surface exposed on the native organisms. The two negatively charged residues, aspartic acid and glutamic acid, were found most often in determinants. Thirteen antigenic sites were further characterized by alanine substitution. Differences in fine specificities of these linear determinants were observed in alanine substitution profiles. Five determinants had adjacent critical residues, while eight had critical residues alternated with noncritical residues. Complete replacement analysis of two antigenic determinants provided more detailed information for elucidating the structural basis of the specificity of antigen-antibody interaction and suggested a correlation between sequence conservation and tolerance to amino acid substitution for antigenic sites subject to intense immune selection pressure.

Immunologic characterization of a cloned fragment containing the species-specific epitope from the major outer membrane protein of Chlamydia trachomatis.

A 183-bp fragment encoding variable domain IV (VD IV) of Chlamydia trachomatis serovar B major outer membrane protein (MOMP) (amino acids 273 to 333) and containing the species-specific epitope was cloned and expressed in Escherichia coli as a fusion protein with Schistosoma japonicum glutathione S-transferase (GST-VD IV). The fusion protein was affinity purified under nondenaturing conditions and used to immunize rabbits. Antisera were characterized by microimmunofluorescence, immunoblot, dot blot, peptide enzyme-linked immunosorbent, and in vitro neutralization assays. Antisera recognized MOMP from all 12 tested serovars of C. trachomatis but not from Chlamydia psittaci. In a dot blot assay, antisera bound to elementary bodies of serovars B, D, E, L2, and K in a strong fashion and to elementary bodies of serovars F, G, A, and H in a weak fashion but not to elementary bodies of serovars C, J, and I. High-resolution peptide mapping with synthetic overlapping serovar B MOMP peptides in a solid-phase enzyme-linked immunosorbent assay showed that immunization with GST-VD IV produced a serologic response that closely mimicked the response produced with purified serovar B elementary bodies. Antipeptide antibodies with strong binding to species- and subspecies-specific epitopes were elicited. Antisera were able to neutralize only those C. trachomatis serovars that bound antibodies in the dot blot assay. These results suggest that antigenic fragments from VD IV containing the species-specific epitope may be useful in the construction of a chlamydial vaccine for some but not all C. trachomatis serovars.

Immunoaccessible peptide sequences of the major outer membrane protein from Chlamydia trachomatis serovar C.

The antigenicity of the major outer membrane protein of Chlamydia trachomatis serovar C was assessed by using overlapping hexapeptide homologs of serovar C major outer membrane protein and rabbit antisera in a peptide enzyme-linked immunosorbent assay. Five immunogenic sites were found distributed within variable sequences of the protein: four were immunodominant and three were surface exposed on native elementary bodies of serovar C. None was surface exposed on serovars H, I, and J.

Mapping antigenic sites on the major outer membrane protein of Chlamydia trachomatis with synthetic peptides.

The antigenicity of the major outer membrane protein (MOMP) of Chlamydia trachomatis was comprehensively evaluated by using overlapping hexapeptide homologs of serovar B MOMP and polyclonal rabbit antisera in a peptide enzyme-linked immunosorbent assay. Of 367 hexapeptides, 152 showed reactivities with at least one antiserum. Seven hexapeptides located within variable domain (VD) IV (residues 288 to 316) were found to be most reactive in terms of their binding titer and frequency, suggesting that VD IV is the immunodominant region within the MOMP as detected by this assay. Peptide-reactive antibodies could also recognize corresponding epitopes on either viable or acetone-permeabilized organisms. The antigenic specificity and immunoaccessibility of epitopes located in VD IV were resolved by absorbing antisera with chlamydial elementary bodies. Six antigenic sites were found in this region and included a B-type-specific site (S1), four subserogroup-specific sites (S2 and S4 to 6), and one species-specific site (S3), each displaying varying degrees of surface exposures on elementary bodies from different C. trachomatis serovars.

Role of endogenous gamma interferon in host defense against Chlamydia trachomatis infections.

BALB/c mice (6 to 8 weeks old) infected with Chlamydia trachomatis serovar L1 were sacrificed, and the yield of Chlamydia inclusion-forming units from the liver and lungs was measured in HeLa 229 cells. The yield of inclusion-forming units reached a peak at 3 days postinfection and then progressively declined. The mice infected with C. trachomatis had no detectable levels of gamma interferon (IFN-gamma) in their sera. However, stimulation of their spleen cells with either concanavalin A or heat-killed C. trachomatis resulted in the release of high levels of IFN-gamma (600 to 900 IU/ml) at 5 to 8 days postinfection. The increased release of IFN-gamma from the spleen cells paralleled the clearance of chlamydia from the liver and lungs. Sera and spleen cells from animals immunized with live C. trachomatis were transferred to recipient mice that were subsequently challenged with C. trachomatis. Transfer of spleen cells resulted in a reduction of the infection in the recipient animal as measured by the yield of chlamydia from the spleen, but transfer of the sera did not confer protective immunity. In addition, mice infected with C. trachomatis serovar L1 were treated with a hamster neutralizing monoclonal antibody to recombinant murine IFN-gamma (MAb-MuIFN-gamma). In the animals receiving the MAb-MuIFN-gamma, the yield of chlamydia from the lungs, spleen, and liver was significantly higher than from the control groups of mice. Histopathological analysis of tissues from the chlamydia-infected mice showed that the animals treated with the MAb-MuIFN-gamma had a significantly more extensive inflammatory reaction in their lungs, liver, and spleen.

Activation of mouse peritoneal macrophages in vitro or in vivo by recombinant murine gamma interferon inhibits the growth of Chlamydia trachomatis serovar L1.

Peritoneal mouse macrophages activated in vitro with recombinant murine gamma interferon (10 ng/ml) or in vivo (10 micrograms per mouse) showed a significant decrease in the growth and yield of Chlamydia trachomatis. The restriction of the growth of C. trachomatis paralleled the expression of Iad on the macrophages. Mice that received macrophages activated in vitro with recombinant murine gamma interferon showed a significant decrease in the yield of chlamydial infection-forming units from their spleens and peritoneal fluids.

Protective role of magnesium in the neutralization by antibodies of Chlamydia trachomatis infectivity.

Neutralization of the infectivity of Chlamydia trachomatis was assessed by using polyclonal antisera and monoclonal antibodies (MAbs). Polyclonal antisera and a species-reactive MAb as well as a subspecies-specific MAb, both of which were directed toward the major outer membrane protein of C. trachomatis, reduced the number of chlamydial inclusion-forming units in an in vitro assay. Neutralization was dependent on the presence of complement. The species-specific MAb reacted with all 15 serovars by a microimmunofluorescence assay and a dot blot enzyme-linked immunosorbent assay with heat-treated elementary bodies. On the other hand, this same MAb reacted with all serovars, except those in the C complex, by the dot blot enzyme-linked immunosorbent assay with viable organisms and neutralized in vitro all 10 serovars tested, except those in the C complex. When neutralization assays were performed in a solution containing Mg2+, neutralization by both polyclonal antisera and MAbs was significantly reduced. A dose response to Mg2+ supplied as MgSO4 revealed that all concentrations tested from 50 to 800 microM had some effect. Concentrations of greater than or equal to 400 microM MgSO4 completely abolished neutralization at the lowest dilution of polyclonal antisera and species-reactive MAb tested. Although Mg2+ also blocked the neutralization effect of the subspecies-specific MAb, this neutralization was not as complete as that observed with the species-reactive MAb. Addition of Mg2+ to the assay over the initial 45 min of incubation of C. trachomatis with MAb and complement showed that the organisms could be rescued to some extent over the first 30 min of incubation, after which time neutralization of infectivity could not be reversed. C. trachomatis treated with Mg2+, the species-reactive MAb, and complement were lethal to mice in an in vivo toxicity and infectivity assay, whereas mice injected with organisms incubated with the same MAb and complement without Mg2+ survived.

Recombinant murine gamma interferon inhibits Chlamydia trachomatis serovar L1 in vivo.

Chlamydia trachomatis serovar L1 injected intravenously in mice resulted in systemic nonlethal infections of the animals. Treatment of mice with recombinant murine gamma interferon resulted in a decrease in the number of infectious C. trachomatis organisms recovered from the lungs, spleens, and livers as well as in a decrease of the inflammatory reaction in those organs when assessed 3 and 5 days after challenge.