A human colorectal explant culture to evaluate topical microbicides for the prevention of HIV infection.

A human colorectal explant culture was developed to assess the safety and efficacy of topical microbicides proposed for use in humans. Because any product marketed for vaginal application will likely be used for anal intercourse, it is important to evaluate these products in colorectal explant tissue. Microbicides tested included cellulose acetate 1,2-benzenedicarboxylate (CAP), PRO 2000, SPL7013, Vena Gel, and UC781, along with their accompanying placebos. Colorectal tissues were exposed to microbicides overnight and either fixed in formalin to evaluate toxicity by histological analysis or placed in 1-(4,5-dimethylthiazol-2-yl)-3,5-diphenylformazan (MTT) to quantitatively determine tissue viability. Histological analysis showed minimal toxicity for CAP, UC781, and Vena Gel. Shedding of epithelium with intact lamina propria occurred for the PRO 2000 and SPL7013 products, and shedding of epithelium and necrosis of the lamina propria occurred in explants cultured with nonoxynol-9. The MTT assay confirmed these results for PRO 2000 (4% and 0.5%), SPL7013 (and placebo), and nonoxynol-9 but also demonstrated reduced viability for CAP. However, viability of tissues treated with all products was not significantly different from that of the medium control. Efficacy of the microbicides was evaluated by measuring human immunodeficiency virus type 1 (HIV-1) infection of explants in the absence or presence of products. All microbicide formulations tested were highly effective in preventing HIV infection. However, explants treated with some of the placebo formulations also exhibited a lower level of infection. Most of the products developed for vaginal application showed minimal toxicity and were effective in reducing HIV-1 infection in colorectal tissues. These results suggest that this model is useful for evaluating the safety and efficacy of topical microbicides when used rectally.

Detection of infectious human immunodeficiency virus type 1 in female genital secretions by a short-term culture method.

Infectious human immunodeficiency virus type 1 (HIV-1) is difficult to detect in female genital secretions by standard virus culture techniques. To improve detection of cell-free HIV-1 in female genital secretions, we adapted a short-term assay that uses the multinuclear-activation galactosidase indicator (MAGI) assay. When vaginal lavages from HIV-1-infected women were tested with the adapted MAGI assay, 25 (64%) of 39 lavages with detectable, cell-free HIV-1 RNA were shown to have infectious virus. No infectious virus was found in 10 vaginal lavages from HIV-1-infected women with undetectable vaginal viral loads. Significantly (P < 0.01) more lavages from HIV-1-infected women tested positive for infectious virus by the MAGI assay than by standard peripheral blood mononuclear cell (PBMC) coculture, which detected infectious virus in only 6 (17%) of 35 vaginal lavages. Lavages with viral loads of >10,000 copies per lavage yielded significantly (P < 0.01) more positive cultures than those with <10,000 copies by using the MAGI assay. Detection of infectious HIV-1 in vaginal lavages was not associated with the presence of genital tract infections or CD4(+)-T-cell counts. However, although the results were not significant (P = 0.08), the MAGI assay detected infectious virus from more vaginal lavages at a vaginal pH of >/=4.5 than at a pH of <4.5. These results indicate that the MAGI assay is more sensitive than PBMC culture methods for detecting infectious virus in female genital secretions. Accurate measurements of infectious virus in genital secretions will improve studies that evaluate sexual transmission of HIV-1.

Enhancement of disease and pathology by synergy of Trichuris suis and Campylobacter jejuni in the colon of immunologically naive swine.

Campylobacterjejuni, a leading cause of bacterial gastroenteritis, has different age distribution and disease expression in developing and developed countries, which may be due to the endemnicity of infection and the age of acquisition of immunity. Differences in disease expression are not solely dependent on the C. jejuni strain or virulence attributes. Another modulating factor in developing countries may be endemic nematode infections such as Trichuris, which drive type 2 cytokine responses and down-regulate type 1 immune responses. In this study, three-day-old germ-free pigs given dual infections with Trichuris suis and C. jejuni had more frequent, more severe diarrhea and severe pathology than pigs given no pathogens, only T. suis, or only C. jejuni. These pigs had significant hemorrhage and inflammatory cell infiltrates in the proximal colon where adult worms were found, and abscessed lymphoglandular complexes in the distal colon with intracellular C. jejuni. Pigs given only C. jejuni had mild clinical signs and pathology, and bacteria in feces or extracellular sites. Pigs given T. suis or no pathogens had no disease and minimal pathology. Thus, these agents synergized to produce significant disease and pathology, which was site specific.

Enhancement of disease and pathology by synergy of Trichuris suis and Campylobacter jejuni in the colon of immunologically naive swine.

Campylobacter jejuni, a leading cause of bacterial gastroenteritis, has different age distribution and disease expression in developing and developed countries, which may be due to the endemnicity of infection and the age of acquisition of immunity. Differences in disease expression are not solely dependent on the C jejuni strain or virulence attributes. Another modulating factor in developing countries may be endemic nematode infections such as Trichuris, which drive type 2 cytokine responses and down-regulate type 1 immune responses. In this study, three-day-old germ-free pigs given dual infections with Trichuris suis and C jejuni had more frequent, more severe diarrhea and severe pathology than pigs given no pathogens, only T. suis, or only C jejuni. These pigs had significant hemorrhage and inflammatory cell infiltrates in the proximal colon where adult worms were found, and abscessed lymphoglandular complexes in the distal colon with intracellular C jejuni. Pigs given only C jejuni had mild clinical signs and pathology, and bacteria in feces or extracellular sites. Pigs given T. suis or no pathogens had no disease and minimal pathology. Thus, these agents synergized to produce significant disease and pathology, which was site specific.