Typing of Chlamydia trachomatis strains from urine samples by amplification and sequencing the major outer membrane protein gene (omp1).

OBJECTIVES: To develop a novel protocol for the extraction, amplification, and sequencing of Chlamydia trachomatis MOMP gene (omp1) from urine, a non-invasive source, and apply it to an epidemiological study on the distribution of C trachomatis strains in a population of pregnant women in Thailand. METHODS: The C trachomatis DNA was extracted from culture stocks and urine using a slightly modified commercially available kit, the High Pure PCR Template Preparation Kit (Roche Molecular Biochemicals, IN, USA). The PCR and sequencing primers used for the amplification and sequencing of the omp1 were designed based on the nucleotide sequence of multiple C trachomatis strains found in GenBank. The protocol for the extraction, amplification, and sequencing was tested on laboratory culture stocks of reference strains of all C trachomatis serovars and on urine samples collected in a cross sectional study designed to assess the prevalence of C trachomatis infections in the cities of Bangkok and Chiang Rai, Thailand. RESULTS: The omp1 gene was successfully amplified and sequenced from 18 laboratory C trachomatis reference strains and from 45 C trachomatis positive urine clinical samples collected from asymptomatic pregnant women. Among clinical samples, we found nine different C trachomatis genotypes: F (11, 25%), D (10, 22.6%), H (5, 11.7%), K (5, 11.7%), E (4, 9.3%), Ia (3, 7%), B (3, 7%), Ja (2, 4.5%), and G (1, 2.3%). One specimen generated an omp1 DNA sequence pattern indicating the presence of a mixed infection with at least two different serovars. CONCLUSIONS: Urine is a convenient and reliable source for genotyping C trachomatis strains. A clear advantage of urine over traditional samples, such as cervical swabs, is that urine is a non-invasive source which makes collection easier and thus facilitates the enrolment of patients in clinical and epidemiological studies. In addition to typing, urine is increasingly used for diagnosis of C trachomatis infection by several commercially available nucleic acid amplification assays which represents a distinct advantage for collecting, transport, storage, and laboratory handling of samples.

Diversity of HIV-1 subtype E in semen and cervicovaginal secretion.

OBJECTIVES: To characterize human immunodeficiency virus type 1 (HIV-1) subtype E variants in blood and genital fluid of infected Thai couples. STUDY DESIGN/METHODS: Blood and genital fluid were collected from 30 asymptomatic healthy HIV-1 subtype E infected couples from Bangkok, Thailand from 1995 to 1998. RESULTS: All 60 viruses in blood samples were identified as subtype E by heteroduplex mobility assay. The biotype of viruses founded in blood was syncytium-inducing (SI), whereas M-tropic and non-syncytium-inducing (NSI) isolates were predominantly detected in genital fluid. HIV-1 proviral DNA was detected in 43.33% and 56.67%, and viral RNA was detected in 93.33% and 56.67%, of semen (n = 30) and cervicovaginal secretion (n = 30) samples tested, respectively. A higher intersample genetic distance and more positive charge of the V3 loop were found in blood strains composed of genital fluid strains (22.30 +/- 5.92% and 17.96 +/- 6.3%), which was statistically significant (P = 0.003). The env V1-V4 intraperson variation of the HIV-1 subtype E in the blood and genital fluid of each individual was in the range 3.0%-5.7%. We also determined the intrasample variation of HIV-1 from blood and genital fluid by heteroduplex mobility assay. The mean heteroduplex mobility of the HIV-1, V1-V4 region of env gene, in blood (n = 8) and genital fluid (n = 8) was 0.59 +/- 0.06 and 0.74 +/- 0.11 (t test, p = 0.001), respectively. CONCLUSIONS: There was genetic and phenotypic compartmentalization of HIV-1 subtype E in blood and genital fluid with the presence of SI and NSI phenotypic variants as a common property of subtype E isolates from blood and genital fluid, respectively.

Lack of association between human immunodeficiency virus type 1 antibody in cervicovaginal lavage fluid and plasma and perinatal transmission, in Thailand.

To determine the association between human immunodeficiency virus type 1 (HIV)-specific antibody and RNA levels in cervicovaginal lavage (CVL) samples and plasma, zidovudine treatment, and perinatal transmission, HIV subtype E gp160-specific IgG and IgA were serially measured in a subset of 74 HIV-infected women in a placebo-controlled trial of zidovudine, beginning at 36 weeks of gestation. HIV IgG was detected in 100% of plasma and 97% of CVL samples; HIV IgA was consistently detected in 62% of plasma and 31% of CVL samples. Antibody titers in CVL samples correlated better with the RNA level in CVL samples than with plasma antibody titers. Zidovudine did not affect antibody titers. Perinatal HIV transmission was not associated with antibody in CVL samples or plasma. HIV-specific antibody is present in the cervicovaginal canal of HIV-infected pregnant women; its correlation with the RNA level in CVL fluid suggests local antibody production. However, there was no evidence that these antibodies protected against perinatal HIV transmission.

Short-course antenatal zidovudine reduces both cervicovaginal human immunodeficiency virus type 1 RNA levels and risk of perinatal transmission. Bangkok Collaborative Perinatal HIV Transmission Study Group.

Human immunodeficiency virus (HIV) levels in cervicovaginal lavage (CVL) and plasma samples were evaluated in relation to perinatal transmission in a randomized placebo-controlled trial of brief antenatal zidovudine treatment. Samples were collected at 38 weeks' gestation from 310 women and more frequently from a subset of 74 women. At 38 weeks, after a 2-week treatment period, CVL HIV-1 was quantifiable in 23% and 52% of samples in the zidovudine and placebo groups, respectively (P<.001). The perinatal transmission rate was 28.7% among women with quantifiable CVL HIV-1 and high plasma virus levels (>10,000 copies/mL) and 1% among women without quantifiable CVL HIV-1 and with low plasma virus levels (P<.001). A 1-log increase in plasma HIV-1 increased the transmission odds 1.8 and 6.1 times (95% confidence interval, 0.9-3.5 vs. 2.4-15.4) for women with and without quantifiable CVL HIV-1, respectively (P=.03). CVL HIV-1 is an independent risk factor for perinatal HIV-1 transmission.

Rapid assessment of sexually transmitted diseases in a sentinel population in Thailand: prevalence of chlamydial infection, gonorrhoea, and syphilis among pregnant women--1996.

OBJECTIVE: To determine the prevalence of sexually transmitted diseases (STDs) among pregnant women in Thailand, where case reporting suggests a marked decrease in STDs following a campaign promoting condom use during commercial sex. DESIGN: Cross sectional study of women at their first visit to the study hospitals' antenatal clinics in Chiang Rai (n = 500) and Bangkok (n = 521). METHODS: First catch urine specimens were tested for Chlamydia trachomatis and Neisseria gonorrhoeae using the Amplicor CT/NG polymerase chain reaction assay. Syphilis and HIV serological testing were performed in the study hospitals' laboratories. RESULTS: The prevalence of chlamydial infection was 5.7%, gonorrhoea 0.2%, and syphilis 0.5% (all VDRL or RPR titres were < or = 1:4). The prevalence of HIV infection was 7.1% in Chiang Rai and 2.9% in Bangkok. In a multivariate logistic regression analysis, chlamydial infection was associated with younger age and with higher gestational age at first antenatal clinic visit, but was not associated with marital status, gravidity, city of enrollment, or HIV infection status. CONCLUSIONS: There was a low prevalence of gonorrhoea and syphilis among these pregnant women in Thailand. Chlamydial infection was detected at a higher prevalence, especially among younger women and women registering later for antenatal care. Testing of pregnant women using easily collected urine specimens and a sensitive nucleic acid amplification assay is a feasible method of rapidly assessing chlamydial and gonococcal prevalence.

Chlamydial and gonococcal cervicitis in HIV-seropositive and HIV-seronegative pregnant women in Bangkok: prevalence, risk factors, and relation to perinatal HIV transmission.

OBJECTIVES: To determine the prevalence and risk factors associated with cervicitis caused by Chlamydia trachomatis and Neisseria gonorrhoeae in human immunodeficiency virus (HIV) type 1-seropositive and HIV-seronegative pregnant women in Bangkok, and the relation to perinatal HIV transmission. METHODS: As part of a multicenter perinatal HIV transmission study in an antenatal population with 2% HIV seroprevalence, endocervical swabs obtained at mid-pregnancy from a consecutive sample of 222 HIV-seropositive and 219 HIV-seronegative pregnant women at two large hospitals in Bangkok were tested for the presence of C. trachomatis and N. gonorrhoeae by DNA hybridization probe (Gen-Probe). Clinical risk factors and DNA probe results were analyzed in relation to the women's and newborns' HIV infection status. RESULTS: The prevalence of C. trachomatis was 16.2% in HIV-seropositive pregnant women and 9.1% in HIV-seronegative pregnant women (P = 0.03). The prevalence of N. gonorrhoeae was 2.7% in HIV-seropositive pregnant women and 1.4% in HIV-seronegative pregnant women (P = 0.5). The overall population prevalence estimate was 9.2% for C. trachomatis and 1.4% for N. gonorrhoeae. Women with gonococcal infection were more likely to be positive for C. trachomatis (RR(MH) = 5.2, P < 0.01). Young age (<21 years) and primigravid status were associated with C. trachomatis infection among HIV-seropositive women; history of multiple sex partners (>1) were associated with C. trachomatis infection among HIV-seronegative women. For HIV-seropositive women, primigravida status also was associated with C. trachomatis infection. The perinatal HIV transmission rates were similar for those with and without C. trachomatis (24.1% and 23.2%, P = 0.9) and among those with and without N. gonorrhoeae (20% and 23.5%, P = 1.0). CONCLUSIONS: Among pregnant women in Bangkok, C. trachomatis infection was considerably more common than N. gonorrhoeae infection and was associated with HIV infection, young age and first pregnancy (HIV-seropositive women), and multiple partners (HIV-seronegative women). Our data do not suggest an association between perinatal HIV transmission and maternal C. trachomatis or N. gonorrhoeae infection identified and treated during pregnancy. The high prevalence of C. trachomatis found using a test not readily available in Thailand emphasizes the need for improved, inexpensive ways to screen for and diagnose these sexually transmitted infections in developing countries.

PIP: Numerous studies have suggested that Chlamydia trachomatis and Neisseria gonorrhoeae facilitate heterosexual HIV transmission; the impact of these sexually transmitted diseases (STDs) on perinatal HIV transmission is unknown, however, due to the expense of routine screening for STDs during pregnancy in developing countries. As part of a multicenter perinatal HIV transmission study, 222 HIV-positive and 219 HIV-negative women presenting for prenatal care at 2 hospitals in Bangkok, Thailand, during 1993-94 were enrolled. At mid-pregnancy, endocervical swabs were obtained and tested for the presence of C. trachomatis and N. gonorrhoeae by DNA hybridization probe. There were 36 cases (16.2%) of C. trachomatis infection among HIV-positive women and 20 cases (9.1%) among HIV-negative women. There were 6 cases (2.7%) of N. gonorrhoeae among HIV-positive women and 3 cases (1.4%) among HIV-negative women. Based on an estimated antenatal HIV seroprevalence of 2%, these findings imply a general antenatal prevalence of 9.2% for C. trachomatis and 1.4% for N. gonorrhoeae. Women with gonococcal infection were more likely (relative risk, 5.2) to be positive for C. trachomatis as well. C. trachomatis infection among HIV-infected pregnant women was associated with age under 21 years and primigravidity. The overall perinatal HIV transmission rate was 24.2%, with no significant difference according to STD infection status. However, since all women diagnosed with STDs received treatment by the mid-third trimester of pregnancy, it remains possible that untreated STDs facilitate perinatal HIV transmission. The high prevalence of C. trachomatis detected in this study through use of a test not readily available in Thailand emphasizes the need for inexpensive, reliable methods to screen for STDs among pregnant women in developing countries.

Shedding of HIV-1 subtype E in semen and cervico-vaginal fluid.

The uneven expansion of HIV-1 subtypes in each transmitted group raises the possibility that some viruses have less/more potential by qualitative/quantitative for heterosexual transmission compared to others. In Thailand, HIV-1 subtype E is mainly spread via heterosexual route and accounts for about 95 per cent of the infected cases. To determine whether high sexual infectivity of HIV-1 subtype E is due to the presence of a virus in genital fluid, we conducted a study to characterize shedding of HIV-1 in seminal and cervico-vaginal fluids of 30 HIV-1 subtype E infected Thai couples by PCR and virus isolation methods. All subjects had no HIV-associated diseases and other sexually transmitted diseases. HIV-1 subtype E DNA was detected in 22/30 (77.33%) of cervico-vaginal and also 22/30 (77.33%) of seminal fluid samples. The isolation rate of HIV-1 from semen and cervico-vaginal secretion was 36.67 per cent and 16.67 per cent, respectively. Number of HIV-1 subtype E DNA copies in the blood is reversely correlated with the number of blood CD4+ T cells, while that in genital fluid was not related to CD4+ T cell count. An increase in shedding of HIV- DNA subtype E in female genital tract compared to other HIV subtypes reported by other investigators might be one reason to explain the rapid spread of subtype E by heterosexual transmission in Thailand.

PIP: Preliminary evidence suggests that HIV subgroups differ in both their transmissibility and virulence. In Thailand, HIV-1 subtype E (accounting for almost 95% of total HIV cases) is transmitted primarily through heterosexual sex, with a predominance of female-to-male infection. This study characterized virus shedding patterns in seminal and cervico-vaginal fluids from 30 asymptomatic husband-wife pairs from Bangkok, Thailand, known to be infected with HIV-1 subtype E. HIV-1 subtype E was detected in 22 (77.3%) cervico-vaginal and 22 (77.3%) seminal fluid samples. HIV-1 subtype B, in contrast, is found in only 30-50% of cervico-vaginal specimens; detection of subtype B in seminal specimens (70-80%) is comparable to that identified for subtype E in the present study. The isolation rate of HIV-1 was 36.67% from semen and 16.67% from cervico-vaginal secretions. The number of HIV-1 subtype E DNA copies in blood--but not in genital fluids--was inversely correlated with the number of blood CD4+ T cells. The increased shedding of HIV-1 DNA subtype E compared with other subtypes in the female genital tract presumably accounts for the rapid spread of subtype E among heterosexuals in Thailand.

Carbon dioxide laser vaporization for genital wart.

The purpose of this study was to evaluate the effectiveness of carbon dioxide laser vaporization in the treatment of coalescent condyloma acuminata of the female lower genital tract. Between July 1990 and September 1992, 24 cases were enrolled in the study. There were 13 pregnant patients and 11 nonpregnant patients. We performed the procedure under colposcopic guidance on an out-patient basis. Carbon dioxide laser vaporization was carried out in continuity to an appropriate tissue depth with carbon dioxide brushing 1 cm around the lesion. Of these 24 cases, biopsy proven 2 residual lesions and 1 recurrence were documented at the 2nd week and the 2nd month postoperative, respectively. We found low rate of intraoperative and postoperative morbidity or complication, with mild to moderate vulvar pain in the patients and no gross scarring. It is concluded that carbon dioxide laser is effective for eradicating extensive condylomata in the female lower genital tract with low intraoperative and postoperative morbidity on an out-patient basis.

Impact of anaerobic growth conditions on toxic shock syndrome toxin-i production by Staphylococcus aureus.

OBJECTIVE: The impact of anaerobic growth conditions on the Staphylococcus aureus toxic shock syndrome toxin-1 (TSST-1) production was studied. METHODS: Ten strains of S. aureus derived from patients with toxic shock syndrome (TSS), 10 isolates of S. aureus, and documented TSST-1-producing strains recovered from patients with either staphylococcal septicemia or staphylococcal nongenital abscesses were grown under aerobic and anaerobic conditions. The bacterial growth was measured using optical density (OD) determinations at 520 nm. The toxin production was assayed using the TS-RPLA latex agglutination test. RESULTS: Both TSS and non-TSS strains of S. aureus grown under aerobic and anaerobic conditions exhibited comparable OD patterns of growth, and the levels of toxin production remained constant during the logarithmic phase. Toxin titers developed during the logarithmic growth phase and peaked after 24 h of incubation. When stationary-phase isolates grown initially under aerobic conditions were subjected to strict anaerobic conditions, subsequent toxin titers, compared with isolates grown in the continued presence of oxygen, were depressed 2-fold, peaking at a later time. CONCLUSIONS: TSST-1 production is diminished under continued anaerobic conditions.

HIV-1 infection among low income women attending a Siriraj sexually transmitted disease clinic: sociodemographic differentials.

From August 1993 to October 1994, 322 women attended or were referred to a female sexually transmitted disease clinic, were studied for the prevalence of HIV infection. No subject had a history of commercial sex work, injection drugs use or blood transfusion within the past 8 years. The majority of women belonged to the low socioeconomic stratum. HIV-1 antibody was found in the sera of 38 women (11.8%). HIV-1 seropositivity was not associated with any type of current sexually transmitted disease such as genital ulcers, serologic markers of syphilis or other sexually transmitted disease as well as history of past sexually transmitted disease within the past 2 years. Significant differential factors were found between the HIV-1 seropositive and seronegative women for self risk assessment and ability to communicate concerns with the husband or partner regarding HIV infection/AIDS. Programs are urgently needed for HIV/AIDS prevention and control to low-income communities and to determine what factors enable the HIV-1 seronegative women to be more assertive in their relationship and whether these skills can be enhanced to eliminate future episodes of STD and transfer these skills to the more vulnerable low-income women. Early diagnosis and prevention of HIV infection among women is a priority for public health interventions both in industrialized and in developing countries.

In vitro ability of the group B streptococci to inhibit gram-positive and gram-variable constituents of the bacterial flora of the female genital tract.

OBJECTIVE: The purpose of this study was to analyze the ability of septicemic and nonsepticemic isolates of group B streptococci (GBS) to inhibit in vitro the principal bacterial groups found in the normal bacterial flora of the female genital tract. METHODS: The target groups were composed of 1) 10 strains each of the following: viridans streptococci, nonhemolytic streptococci (not group B or D), group A streptococci, GBS, peptostreptococci, coagulase-negative staphylococci, Staphylococcus aureus, and Gardnerella vaginalis; 2) 9 strains of enterococci; 3) 9 strains of group C or G streptococci; 4) 7 strains of lactobacilli; and 5) 7 strains of diphtheroids. All target groups were tested for inhibition by a test panel of either a group of 10 or 41 GBS isolates. If the GBS isolates failed to inhibit a target group, that group was tested for its ability to inhibit the GBS test panel. RESULTS: The GBS test panel did not inhibit the growth of coagulase-negative staphylococci or S. aureus but uniformly inhibited groups A, B, C, and G streptococci, lactobacilli, and G. vaginalis. One of the 7 strains of diphtheroids was inhibited by 37 of the 41 GBS isolates; the other 6 strains of diphtheroids were uniformly inhibited. Variable inhibition by GBS was observed with viridans streptococci, nonhemolytic (not group B or D) streptococci, peptostreptococci, and enterococci; however, inhibition or noninhibition was uniform for a given target strain against the entire GBS test panel. The 23 GBS isolates obtained from septicemic neonates or adults did not differ from the 18 nonsepticemic isolates in their ability to inhibit other species of streptococci or other gram-positive or gram-variable constituents of the bacterial flora of the female genital tract. When converse testing was done, all 10 GBS isolates were uniformly inhibited by coagulase-negative staphylococci and by the majority of enterococci, but were not inhibited by S. aureus. CONCLUSIONS: These studies suggest that GBS may be significant regulators of other beta-hemolytic streptococci, diphtheroids, lactobacilli, and G. vaginalis within the bacterial flora of the female genital tract. Moreover, the absence of GBS in the vaginal flora may be the result of mediation by coagulase-negative staphylococci and selected strains of enterococci.

Chlamydia trachomatis in patients who used oral contraceptives and had intermenstrual spotting.

OBJECTIVE: To identify the incidence of positive assays for direct antigen detection of Chlamydia trachomatis in oral contraceptive (OC) users with and without intermenstrual spotting and in women seeking contraception. METHODS: We studied 65 women who had used OCs for more than 3 months and who presented with intermenstrual spotting for which no readily demonstrable cause could be identified. They were compared with 65 matched controls who were taking OCs without intermenstrual spotting and who had chlamydia testing because of one or more risk factors, and 65 matched controls seeking contraception. The incidence of positive tests for each group was compared by chi 2 test for independence using the Yates formula. The strength of the relationship was analyzed by determining the Craemer phi coefficient. RESULTS: Nineteen of the 65 women (29.2%) taking OCs for more than 3 months and experiencing bleeding had positive tests, in contrast to seven of 65 matched controls (10.7%) who were also on OCs and who had had chlamydia testing because of vaginitis or new or multiple sexual consorts, and four of 65 women (6.1%) who were screened for C trachomatis before initiation of contraception. The significance level for the chi 2 statistic was P < .01. CONCLUSION: Intermenstrual bleeding in women previously well regulated on OCs appears to be an added marker for chlamydial infection.

Neonatal Streptococcus pneumoniae septicemia and meningitis. A case report.

Neonatal septicemia/meningitis from Streptococcus pneumoniae occurred in a 36-hour-old infant. The mother had no overt evidence of infection. This case illustrates the pathogenic potential of this common bacterium in the neonate.

Potential use of C-reactive protein determinations in obstetrics and gynecology.

C-reactive protein (CRP) is an acute-phase protein synthesized by the liver during the course of a large number of diseases. During gestation and parturition, progressively increasing numbers of gravida develop elevated levels up to 32, 48 to 80 per cent, respectively). These elevations in pregnancy, the nonspecificity of the test, and the inability to function as a reliable marker of fetal and/or maternal infectious morbidity have obscured the pragmatic utility of CRP determinations in both obstetrics and gynecology. Because of its biophysical kinetics, C-reactive protein determinations are best used as a monitoring parameter after the documentation of infection rather than as a diagnostic indicator of infection. They can provide valuable end titration points for termination of parenteral antibiotic therapy.