Human infection with Toxoplasma gondii is presumed due to the ingestion of either tissue cysts containing bradyzoites or oocyst/sporozoites that are excreted in the feces of infected cats. The incidence of human infection in the general population by either of these routes is unknown. We have previously described unique stage-specific oocyst/sporozoite antigens identified by murine hybridoma monoclonal antibodies. We obtained acute and convalescent antitoxoplasma antisera from patients in an epidemiologically well-documented outbreak of oocyst-transmitted infection associated with the ingestion of contaminated water. An enzyme-linked immunosorbent assay comparing equal numbers of tachyzoites (invasive stage) and oocyst/sporozoite (excreted stage) indicated that these antisera recognized antigens from both life forms. Absorption of pooled antisera with purified oocyst/sporozoites reduced both the antioocyst immunoglobulin G (IgG) and immunoglobulin M (IgM) titer but had only minimal effect on the antitachyzoite titer. Absorption of the antisera with tachyzoites reduced the IgG and IgM antioocyst and antitachyzoite titer. A sodium dodecyl sulfate-polyacrylamide gel analysis of radioiodinated oocyst/sporozoites shows that the principal stage-specific surface proteins of the oocyst/sporozoite have approximate Mr of 67,000 and 25,000. Periodic acid and silver stain of purified oocyst/sporozoite identified bands of similar molecular weight not present in the tachyzoite preparation. Western blot analysis of purified parasites assayed with human antioocyst antisera identified specific oocyst/sporozoite antigens not present on the tachyzoites. At least two major stage-specific oocyst/ sporozoite antigens of approximate M, of 67,000 and 190,000 were identified by the infected patients' antisera and not by the normal controls. Reaction to these oocyst/sporozoite antigens was seen primarily in the IgM fraction of the acute phase and the IgG fraction of convalescent phase antisera. Neither absorption of the antisera with tachyzoites nor periodate treatment of the oocyst/sporozoites reduced the antibody recognition of these stage-specific antigens. These data suggest that individuals infected by a presumed oocyst-transmitted route develop antibodies against unique stage-specific oocyst/ sporozoite antigens.

Women can become infected with human immunodeficiency virus type 1 (HIV-1) after the heterosexual transmission of virus from an infected male partner. To understand the events that result in transmission of HIV-1 across the female reproductive tract, we characterized the life-cycle events of HIV-1 in primary cultures of human uterine epithelial cells and stromal fibroblasts. Epithelial cells and stromal fibroblasts released virus particles after exposure to either X4-or R5-tropic strains of HIV-1. Virus released by these cells was able to infect CD4 + T cells. When exposed to an X4-tropic strain of HIV-1, these cells supported HIV-1 reverse transcription, integration, and viral DNA transcription. When exposed to an R5-tropic strain, however, these cells released unmodified virus. These data suggest that uterine cells are targets for productive infection with X4-tropic strains and release unmodified R5-tropic viruses that would then be able to infect submucosal target cells, including T cells and macrophages.Although heterosexual transmission is the predominant mechanism by which women acquire HIV-1 infection [1][2][3][4], our knowledge about the viral and host factors that lead to infection is limited. Identifying the cell population initially infected within the female reproductive tract (FRT) and the putative mechanisms by which HIV-1 is disseminated to distal sites is important to our understanding of the pathogenesis of HIV-1. Moreover, defining the mechanisms and conditions that either promote or inhibit HIV-1 infection within the FRT is necessary for the design and development of preventative measures.Studies to identify cell populations within the FRT that become infected have focused on cell lines or primary cells and tissues from the lower FRT [5,6].

Deletions of colicin El (colEl) plasmid deoxyribonucleic acid (DNA) carrying the TnA transposon have been isolated. All except two were generated by nuclease digestion of plasmid DNA from its EcoRI-sensitive site. A plasmid containing about 16% of the ColEl DNA (6.5 x 105 daltons) was generated that also contained the part of the TnA transposon conferring ampicillin resistance. The extents of different deletions were determined by analysis of restriction endonuclease fragments generated by the restriction endonucleases HaeII, BamHI, and HincII. NaCl plus 0.015 M sodium citrate).Enzymatic digestions of DNA. The endonucleases used were EcoRI, HaeII, BamHI, and HincII, which were obtained from New England Biolabs. Most HaeII used was a generous gift of H. Ohmori and J. Tomizawa. DNA digestions were conducted overnight at 37°C. EcoRI digestions of DNA were 321 on July 31, 2020 by guest

A detailed immunological assessment of strain-specific antigens of Toxoplasma gondii has not been reported. We developed rabbit antisera against three strains of toxoplasma obtained from divergent sources. These strains included the frequently studied laboratory strain RH, strain C, obtained from a naturally infected kitten, and strain P, which is maintained by passage in mice. The rabbit antisera were used to identify unique strain-specific and commonly shared tachyzoite antigens by radioiodination followed by immunoprecipitation and Western blot analysis. Both qualitative and quantitative differences of a number of the major tachyzoite antigens were found in these assays. A parasite plaque reduction assay using parasiticidal monoclonal antibody showed marked differences in the ability to kill these three different tachyzoite subtypes, further supporting antigenic variation among T. gondii strains.

The complete sequence of P30, the major surface Ag of the protozoan parasite, Toxoplasma gondii, has been deduced through the cloning and analysis of its gene. Using polyclonal serum specific for P30, we have isolated a P30 cDNA clone from a lambda gt11 cDNA expression library derived from tachyzoites of T. gondii (RH strain). This clone produces a beta-galactosidase fusion protein which reacts with several anti-P30 mAb. In addition, polyclonal anti-serum raised to the fusion protein reacts with purified P30 protein and exclusively with P30 in a whole cell lysate of T. gondii. This cDNA clone was used to isolate near full-length cDNA molecules and a cosmid clone containing the P30 gene. Sequence analysis of the cDNA reveals a single open reading frame with coding capacity for 34.7 kDa of primary translation product (consistent with the apparent Mr of P30 on SDS-acrylamide gels) including a presumptive hydrophobic signal sequence. The P30 primary translation product also has a carboxy-terminal hydrophobic tail which is predictive of a posttranslational cleavage and modification with a glycolipid anchor. We have identified the apparent 5' and 3' ends of the P30 mRNA transcript which is extremely abundant, 1500 nucleotides in length, and polyadenylated. The P30 gene is single copy and contains no introns.