Effect of Seminal Plasma on Chlamydia trachomatis Strain LB-1 in Cell Culture

Abstract

Human seminal plasma inhibited formation of strain LB-1 chlamydial inclusions in McCoy cells proportional to the concentration of seminal plasma added after chlamydial adsorption.Chlamydia are among the most common sexually transmitted organisms and are probably responsible for at least one-half of the cases of nongonococcal urethritis in males. We questioned whether genital secretions might influence the infectivity of such agents (6) or the susceptibility to infection of the host cells. We describe here the effect of seminal plasma (SP) on standardized infections of Chlamydia trachomatis strain LB-1 in cultured cells in vitro and a few parameters which determined inclusion formation in such cells.Celi cultures. McCoy cells were grown on cover slips in Leighton tubes in Eagle minimum essential medium containing 10% fetal calf serum plus gentamicin (50 ,tg/ml) and fungizone (2.5 ,ug/ml). The maintenance medium (MS) contained gentamicin with 2% glucose plus cycloheximide (1 ,ug/ml). One-milliliter volumes of cell suspensions containing 3 x 105 to 3.5 x 105 cells were added to Leighton tubes and incubated at 350C. Cover slips were removed after 50 to 55 h of incubation at 350C, fixed in absolute methanol, and stained with Giemsa stain. Triplicate preparations were examined, and inclusions in 30 random fields per cover slip (40 to 50 cells per field) were counted by two observers with a 40x oil immersion objective (final magnification, 480x) (2). Counts of inclusions formed in the presence of SP were expressed as a percentage of the number of inclusions in control monolayers in the absence of SP in the same experiment.Chlamydial inoculum. C. trachomatis strain LB-1 (immunotype L2) was diluted in MS to produce 80 to 120 inclusion-forming units per 30 microscopic fields, with about 10% of the cells being infected.SP. Human SP was collected from normal donors, pooled, centrifuged at 300 x g for 10 min to remove sperm, filtered (0.45-,um pore size; Millipore Corp.), and stored frozen at -20°C in aliquots until tested (7). Dilutions were made in minimum essential medium. No antichlamydial antibodies were detected in the pooled SP by indirect microimmunofluorescence employing anti-human whole serum globulin (1).Procedure A. Medium was removed from cell monolayers, and 1 ml of LB-1 dilution was added and adsorbed for 3 h at 350C. Inoculum was removed, cells were washed once with Hanks balanced salt solution (BSS), and 1 ml of MS with or without diluted SP was added. Tubes were incubated at 350C for 3 h. Since high concentrations of SP were toxic if left on the cells, fluid was removed, the monolayer was again washed once with BSS, and fresh MS was added. Monolayers were then incubated at 350C for a total of 50 to 55 h. Procedure B. Medium was removed from cell monolayers, 0.1 ml (volume sufficient to cover the cell sheet) of diluted SP was added, and monolayers were incubated at 350C. After 30 min, 1 ml of diluted LB-1 was added and adsorbed at 35°C for 3 h. The SP and LB-1 were removed, cells were washed once with BSS, 1 ...

Diagnostic considerations and interpretation of microbiological findings for evaluation of chronic prostatitis

“…During the last 2 decades there have been many valuable studies of chronic bacterial prostatitis. These efforts have improved our understanding of both specific immunological factors and nonspecific host defenses of the male lower urogenital tract, such as pH and the prostatic antibacterial factor (3,4,6,8,14,17).…”

Chronic prostatitis, chlamydia trachomatis and infertility

“…The definitive demonstration of a chlamydial aetiology is based on the isolation of C. trachomatis from the mucous membranes. This task may be difficult since it has been demonstrated that seminal factors have some undetermined inhibitory effect on chlamydial inclusion formation in cell culture (a method used for isolation of C. trachomatis) (Mirdh et al 1980;Hanna et al 1981).…”

Changes in serum and prostatic zinc concentrations in rats after intraprostatic injection of zinc: Comparison of two forms of zinc delivery

“…concluded that PAF is free zinc. In addition to bactericidal activity against gram‐negative organisms, zinc may inhibit the growth of viruses, 8,9 yeats, 10 protozoa 11 and Chlamydia trachomatis 12,13 …”