Lack of evidence for sexual transmission of genital Candida species among women who have sex with women: a mixed methods study.

OBJECTIVE: The contribution of sexual transmission to genital Candida infection remains unclear. This study sought to investigate whether sexual behaviours were associated with the presence of genital Candida species among a cohort of women who have sex with women (WSW) in addition to determining the genetic concordance of genital Candida spp. among WSW in sexual partnerships. METHODS: WSW ≥18 years of age presenting to the Mississippi State Department of Health STD Clinic during 2009-2010 completed a sexual behaviour survey. Culture of vaginal fluid was performed for Candida spp. identification; associations with participant characteristics were determined using logistic regression analysis. Random amplified polymorphic DNA (RAPD) PCR was performed on DNA extracted from yeast cultures of WSW in sexual partnerships in which both partners had isolates of Candida spp. identified and among a set of age/sexual behaviour matched controls. RAPD genetic fingerprints were evaluated by hierarchical cluster analysis for concordance. RESULTS: Genital Candida spp. were isolated in 105/196 (53.6%) of women: 13/105 (12.4%) had symptomatic vulvovaginal candidiasis while 92/105 (87.6%) had asymptomatic vaginal colonisation. Bisexual identity, sex with women and men during the past 12 months and numbers of male sexual partners during the past 12 months were the only significant predictors of genital Candida spp. in bivariate analysis. 13 pairs of WSW in sexual partnerships in which both partners had genital Candida spp. and 11 WSW with genital Candida spp. not in sexual partnerships were identified. Candida spp. RAPD banding patterns were discordant for all isolates among WSW within partnerships and in controls. CONCLUSIONS: This study found no evidence supporting sexual transmission of genital Candida spp. between women.

Detection of Trichomonas vaginalis DNA by use of self-obtained vaginal swabs with the BD ProbeTec Qx assay on the BD Viper system.

Trichomonas vaginalis is the most prevalent nonviral sexually transmitted infection worldwide, and improved diagnostic methods are critical for controlling this pathogen. Diagnostic assays that can be used in conjunction with routine chlamydia/gonorrhea nucleic acid-based screening are likely to have the most impact on disease control. Here we describe the performance of the new BD T. vaginalis Qx (TVQ) amplified DNA assay, which can be performed on the automated BD Viper system. We focus on data from vaginal swab samples, since this is the specimen type routinely used for traditional trichomonas testing and the recommended specimen type for chlamydia/gonorrhea screening. Vaginal swabs were obtained from women attending sexually transmitted disease or family planning clinics at 7 sites. Patient-collected vaginal swabs were tested by the TVQ assay, and the Aptima T. vaginalis (ATV) assay was performed using clinician-collected vaginal swabs. Additional clinician-collected vaginal swabs were used for the wet mount and culture methods. Analyses included comparisons versus the patient infection status (PIS) defined by positive results with the wet mount method or culture, direct comparisons assessed with κ scores, and latent class analysis (LCA) as an unbiased estimator of test accuracy. Data from 838 women, 116 of whom were infected with T. vaginalis, were analyzed. The TVQ assay sensitivity and specificity estimates based on the PIS were 98.3% and 99.0%, respectively. The TVQ assay was similar to the ATV assay (κ=0.938) in direct analysis. LCA estimated the TVQ sensitivity and specificity as 98.3 and 99.6%, respectively. The TVQ assay performed well using self-collected vaginal swabs, the optimal sample type, as recommended by the CDC for chlamydia/gonorrhea screening among women.

Prevalence of Gardnerella vaginalis among women with lactobacillus-predominant vaginal flora.

OBJECTIVES: To determine the prevalence of Gardnerella vaginalis in women with normal vaginal flora. METHODS: Women without symptoms or signs of vaginal infection and five or fewer lifetime sexual partners were recruited for a longitudinal study of vaginal flora. Negative Amsel criteria and a Nugent score of 0-3 were required for enrolment. Vaginal specimens were self-collected daily for Gram stain and every 3 days for PCR for G vaginalis for 30 days. Women completed daily diaries recording sexual activity, symptoms and menses. RESULTS: Twenty women were recruited for the study with 19 completing all specimens and 1 lost to follow-up. During the 30-day study period, 13/19 (68.4%) of women had normal Nugent scores (0-3) whereas 6/19 (31.6%) of women had at least 2 days of Nugent scores in the intermediate range (p=0.09). Among the 19 women, 9 (47%) were negative for G vaginalis by PCR throughout the study period whereas 10 (53%) had at least one specimen that demonstrated the presence of G vaginalis by PCR. Of those women with intermediate flora on Gram stain during the course of the study 5/6 (83.3%) were positive for G vaginalis while 5/13 (38.5%) of those women with only normal Nugent scores were positive for G vaginalis. Thus, 61.5% of women with normal Nugent scores had no evidence of G vaginalis by serial PCR. CONCLUSIONS: Gardnerella may not be part of the normal flora in women with optimal vaginal health.

Diagnostic rates differ on the basis of the number of read days with the use of the InPouch culture system for Trichomonas vaginalis screening.

The InPouch Trichomonas vaginalis test is the gold standard for clinical culture for Trichomonas vaginalis screening. The current package insert recommends an examination period of 3 days. After review of 2,499 InPouch tests spanning 13 years, we observed that examination up to 3 days will detect only 82.8% (95% confidence interval [CI], 79.0% to 86.2%) of positive specimens.

Comparison of nucleic acid amplification assays with BD affirm VPIII for diagnosis of vaginitis in symptomatic women.

A commercially available, nonamplified, nucleic acid probe-based test system (BD Affirm VPIII) was compared with nucleic acid amplification (NAA)-based assays for determining the etiology of vaginitis in a cohort of 323 symptomatic women. First, a semiquantitative, multiplexed PCR assay (BV-PCR) and the Affirm VPIII Gardnerellavaginalis test were compared with a unified bacterial-vaginosis (BV) reference standard incorporating both Nugent Gram stain scores and Amsel clinical criteria. In the evaluable population of 305 patients, BV-PCR was 96.9% (191/197) sensitive and 92.6% specific (100/108) for BV, while Affirm VPIII was 90.1% sensitive (179/197) and 67.6% specific (73/108). Second, a multiplexed PCR assay detecting Candida albicans and Candida glabrata (CAN-PCR) was compared with the Affirm VPIII Candida test using a reference standard for vulvovaginal candidiasis (VVC) of yeast culture plus exclusion of alternate vaginitis etiologies. In the population evaluated (n = 102), CAN-PCR was 97.7% sensitive (42/43) and 93.2% specific (55/59) and Affirm VP III was 58.1% sensitive (25/43) and 100% specific (59/59) for VVC. Finally, the results of a commercial NAA test (GenProbe Aptima Trichomonas vaginalis assay; ATV) for T. vaginalis were compared with the Affirm VPIII Trichomonas vaginalis test. In the absence of an independent reference standard for trichomonal vaginitis (TV), a positive result in either assay was deemed to represent true infection. In the evaluable cohort of 388 patients, the sensitivity of ATV was 98.1% (53/54) versus 46.3% (25/54) for Affirm VPIII. The diagnostic accuracy of the combined NAA-based test construct was approximately 20 to 25% higher than that of the Affirm VPIII when modeled in populations with various prevalences of infectious vaginitis.

Trichomonas vaginalis infection among women receiving gynaecological care at an Alabama HIV Clinic.

OBJECTIVE: Trichomoniasis vaginalis is a risk factor for the acquisition and transmission of HIV. The objective of this study was to determine the prevalence of T vaginalis (using culture) among HIV-infected women receiving gynaecological care at an university HIV clinic in Alabama in addition to predictors of infection. METHODS: Electronic medical record review of women presenting to the clinic for gynaecological care during 2006-2012 was performed. Demographic and sexual history data was abstracted in addition to absolute CD4 cell count, HIV-1 viral load and sexually transmitted infection (STI) (including T vaginalis) testing results. Analysis was conducted using Stata V.12. RESULTS: T vaginalis was prevalent in 17.4% (83/478) of HIV-infected women; other STIs were less prevalent. Among these women, 384 presented for routine STI screening, of which 12% (46/384) were T vaginalis-infected. Younger age, African-American race, lifetime history of tobacco and drug abuse, lack of HIV therapy, HIV-1 viral load >400 copies/ml, and report of seeking gynaecological care for reasons other than routine STI screening (ie, having symptoms) were significant predictors of T vaginalis in univariate analysis. Age, African American race, and report of seeking gynaecological care for reasons other than routine STI screening remained associated with T vaginalis in multivariable analysis. CONCLUSIONS: T vaginalis remains highly prevalent among HIV-infected women, a proportion of which may be asymptomatic. If left undiagnosed and untreated, these women may be more likely to transmit HIV. Increased emphasis on screening for high risk sexual behaviours, testing for T vaginalis, and risk reduction counselling is necessary for all HIV-infected women.

Genotypic characterization of Trichomonas vaginalis isolates among women who have sex with women in sexual partnerships.

The random amplified polymorphic DNA technique was used to delineate the genetic relatedness of Trichomonas vaginalis isolates among 3 pairs of mutually infected women who have sex with women in sexual partnerships. One of the 3 pairs of women shared a T. vaginalis isolate with the same random amplified polymorphic DNA banding patterns. Shared use of washcloths to cleanse the vaginal area after receptive oral sex was the most likely method of T. vaginalis transmission among this pair of women.

Can mailed swab samples be dry-shipped for the detection of Chlamydia trachomatis, Neisseria gonorrhoeae, and Trichomonas vaginalis by nucleic acid amplification tests?

Dry-shipped and mailed vaginal swabs collected at home have been used in research studies for the detection of Chlamydia trachomatis (CT), Neisseria gonorrhoeae (GC), and Trichomonas vaginalis (TV) by nucleic acid amplification tests (NAATs) in screening programs. A verification study was performed to compare the limit of detection of CT, GC, and TV on swabs that were dry-shipped to paired swabs that were wet-shipped in transport media through the US mail. The Centers for Disease Control and Prevention prepared inocula in sterile water to mock simulated urogenital swabs with high to low concentrations of CT and GC. Replicate swabs were inoculated with 100 µL of dilutions and were dry transported or placed into commercial transport media ("wet") for mailing for NAAT testing. The University of Alabama prepared replicate concentrations of TV, which were similarly shipped and tested by NAAT. All paired dry and wet swabs were detectable for CT. For GC, all paired dry and wet swabs were detectable for GC at concentrations ≥ 10(3). At 10(2) and 10 CFU/mL, the 10 replicate GC results were variably positive. For TV, wet and dry shipped concentrations >10(2) TV/mL tested positive, while results at 10 TV/mL were negative for dry swabs. Holding replicate dry swabs at 55 (○)C 5 days before testing did not affect results. NAATs were able to detect CT, GC, and TV on dry transported swabs. Using NAATs for testing home-collected, urogenital swabs mailed in a dry state to a laboratory may be useful for outreach screening programs.

Development and validation of a semiquantitative, multitarget PCR assay for diagnosis of bacterial vaginosis.

Quantitative PCR assays were developed for 4 organisms reported previously to be useful positive indicators for the diagnosis of bacterial vaginosis (BV)--Atopobium vaginae, Bacterial Vaginosis-Associated Bacterium 2 (BVAB-2), Gardnerella vaginalis, and Megasphaera-1--and a single organism (Lactobacillus crispatus) that has been implicated as a negative indicator for BV. Vaginal samples (n = 169), classified as positive (n = 108) or negative (n = 61) for BV based on a combination of the Nugent Gram stain score and Amsel clinical criteria, were analyzed for the presence and quantity of each of the marker organisms, and the results were used to construct a semiquantitative, multiplex PCR assay for BV based on detection of 3 positive indicator organisms (A. vaginae, BVAB-2, and Megasphaera-1) and classification of samples using a combinatorial scoring system. The prototype BV PCR assay was then used to analyze the 169-member developmental sample set and, in a prospective, blinded manner, an additional 227 BV-classified vaginal samples (110 BV-positive samples and 117 BV-negative samples). The BV PCR assay demonstrated a sensitivity of 96.7% (202/209), a specificity of 92.2% (153/166), a positive predictive value of 94.0%, and a negative predictive value of 95.6%, with 21 samples (5.3%) classified as indeterminate for BV. This assay provides a reproducible and objective means of evaluating critical components of the vaginal microflora in women with signs and symptoms of vaginitis and is comparable in diagnostic accuracy to the conventional gold standard for diagnosis of BV.

Prevalence of bacterial vaginosis and vulvovaginal candidiasis mixed infection in a southeastern american STD clinic.

Bacterial vaginosis (BV) was diagnosed in 72.5% of female participants. Among women with BV, 33.1% were colonized with yeast. Vulvovaginal candidiasis was observed in 15.7% of participants irrespective of BV status. Overall, the prevalence of BV/vulvovaginal candidiasis mixed infections among young women was observed to be 4.4%.

Potential nonresponse bias in a clinical examination after initial screening using iron phenotyping and HFE genotyping in the hemochromatosis and iron overload screening study.

BACKGROUND: Little is known about the factors affecting participation in clinical assessments after HEmochromatosis and IRon Overload Screening. METHODS: Initial screening of 101,168 primary care patients in the HEmochromatosis and IRon Overload Screening study was performed using serum iron measures and hemochromatosis gene (HFE) genotyping. Using iron phenotypes and HFE genotypes, we identified 2256 cases and 1232 controls eligible to participate in a clinical examination. To assess the potential for nonresponse bias, we compared the sociodemographic, health status, and attitudinal characteristics of participants and nonparticipants using adjusted odds ratios (ORs) and 95% confidence interval (CI). RESULTS: Overall participation was 74% in cases and 52% in controls; in both groups, participation was highest at a health maintenance organization and lowest among those under 45 years of age (cases: OR = 0.68; 95% CI 0.53, 0.87; controls: OR = 0.59; 95% CI 0.44, 0.78). In controls only, participation was also lower among those over 65 years of age than the reference group aged 46-64 (OR = 0.64; 95% CI 0.47, 0.88). Among cases, participation was higher in HFE C282Y homozygotes (OR = 3.98; 95% CI 2.60, 6.09), H63D homozygotes (OR = 2.79; 95% CI 1.23, 6.32), and C282Y/H63D compound heterozygotes (OR = 1.82; 95% CI 1.03, 3.22) than in other genotypes, and lower among non-Caucasians and those who preferred a non-English language than in Caucasians and those who preferred English (p < 0.0001). CONCLUSIONS: Subjects with greatest risk to have iron overload (C282Y homozygotes; cases > or =45 years; Caucasians) were more likely to participate in a postscreening clinical examination than other subjects. We detected no evidence of strong selection bias.

Health-related quality of life in a racially diverse population screened for hemochromatosis: results from the Hemochromatosis and Iron Overload Screening (HEIRS) study.

PURPOSE: The HEIRS Study screened 101,168 primary care participants for iron overload with serum transferrin saturation (TS), serum ferritin (SF), and C282Y and H63D mutations of the HFE gene. The objective of this study was to evaluate the impact of screening on participants' well-being. METHODS: All C282Y homozygotes, participants with an elevated TS and SF concentration, and a control group of phenotype-genotype negative persons, with neither C282Y nor H63D mutations in the HFE gene were recalled for a clinical evaluation. Health-related quality of life was assessed before screening and approximately 1 week after receipt of the results. Health worries were assessed only at follow-up. RESULTS: Participants (N = 1478) completed both initial and follow-up surveys. After adjusting for model covariates, phenotype and genotype combinations were statistically significant predictors of changes in psychological well-being (P = 0.0001) and general health (P = 0.0014). C282Y homozygotes with transient elevations in TS or SF were significantly more likely to worry about their health compared to study controls. Race, ethnicity, and preferred language subgroups differed on psychological well-being, general health, and health worry. CONCLUSION: Iron phenotype and HFE genotype are associated with health-related quality of life. Health worry was greatest among those considered genetically "at risk. " This may have important implications for multi-ethnic population-based screening studies in which genotype and phenotype are communicated.

Association of ferroportin Q248H polymorphism with elevated levels of serum ferritin in African Americans in the Hemochromatosis and Iron Overload Screening (HEIRS) Study.

The ferroportin (FPN1) Q248H polymorphism has been associated with increased serum ferritin (SF) levels in sub-Saharan Africans and in African Americans (AA). AA participants of the HEIRS Study who did not have HFE C282Y or H63D who had elevated initial screening SF (> or =300 microg/L in men and >= or =200 microg/L in women) (defined as cases) were frequency-matched to AA participants with normal SF (defined as controls) to investigate the association of the Q248H with elevated SF. 10.4% of cases and 6.7% of controls were Q248H heterozygotes (P=0.257). Q248H homozygosity was observed in 0.5% of the cases and none of the controls. The frequency of Q248H was higher among men with elevated SF than among control men (P=0.047); corresponding differences were not observed among women. This appeared to be unrelated to self-reports of a previous diagnosis of liver disease. Men with elevated SF were three times more likely than women with elevated SF to have Q248H (P=0.012). There were no significant differences in Q248H frequencies in men and women control participants. We conclude that the frequency of the FPN1 Q248H polymorphism is greater in AA men with elevated SF than in those with normal SF.

Allele frequencies of hemojuvelin gene (HJV) I222N and G320V missense mutations in white and African American subjects from the general Alabama population.

BACKGROUND: Homozygosity or compound heterozygosity for coding region mutations of the hemojuvelin gene (HJV) in whites is a cause of early age-of-onset iron overload (juvenile hemochromatosis), and of hemochromatosis phenotypes in some young or middle-aged adults. HJV coding region mutations have also been identified recently in African American primary iron overload and control subjects. Primary iron overload unexplained by typical hemochromatosis-associated HFE genotypes is common in white and black adults in Alabama, and HJV I222N and G320V were detected in a white Alabama juvenile hemochromatosis index patient. Thus, we estimated the frequency of the HJV missense mutations I222N and G320V in adult whites and African Americans from Alabama general population convenience samples. METHODS: We evaluated the genomic DNA of 241 Alabama white and 124 African American adults who reported no history of hemochromatosis or iron overload to detect HJV missense mutations I222N and G320V using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique. Analysis for HJV I222N was performed in 240 whites and 124 African Americans. Analysis for HJV G320V was performed in 241 whites and 118 African Americans. RESULTS: One of 240 white control subjects was heterozygous for HJV I222N; she was also heterozygous for HFE C282Y, but had normal serum iron measures and bone marrow iron stores. HJV I222N was not detected in 124 African American subjects. HJV G320V was not detected in 241 white or 118 African American subjects. CONCLUSIONS: HJV I222N and G320V are probably uncommon causes or modifiers of primary iron overload in adult whites and African Americans in Alabama. Double heterozygosity for HJV I222N and HFE C282Y may not promote increased iron absorption.

Genotypic and phenotypic heterogeneity of African Americans with primary iron overload.

Primary iron overload may be relatively common in African Americans, but its cause is incompletely understood. Thus, we evaluated genotype and phenotype characteristics of unselected African American index patients with primary iron overload who reside in central Alabama. All had hepatic iron concentration > or =30 micromol/g dry wt or > or =2.0 g of iron mobilized by phlebotomy to achieve iron depletion. Genotype analyses were performed in African American control subjects from the same region. There were 23 patients (19 men, 4 women); mean age at diagnosis was 52 +/- 12 years (1 SD) (range 32-69 years). Nine (39.1%) reported that they consumed > or =45 g of ethanol daily; five had chronic hepatitis C. Eight had some form of hemoglobinopathy or thalassemia. Mean serum transferrin saturation was 56 +/- 28% (range 15-100%). The geometric mean serum ferritin at diagnosis was 1076 ng/mL [95% confidence interval 297-3473 ng/mL]. Increased stainable liver iron was observed in hepatocytes only in 4 patients, in macrophages only in 8 patients, and in hepatocytes and macrophages in 8 patients. The mean quantity of iron mobilized by phlebotomy (corrected for iron absorbed during treatment) was 5.3 +/- 2.0 g (range 4.0-8.4 g). Iron removed by phlebotomy was greater in patients with hemoglobinopathy or thalassemia than in those without these forms of anemia (6.6 +/- 1.3 g vs 3.9 +/- 1.6 g, respectively; P = 0.0144). Daily consumption of > or =45 g of ethanol or chronic hepatitis C was not associated with an increased or decreased amount of phlebotomy-mobilized iron, on the average. The percentage of index patients positive for HFE C282Y was greater than that of controls (P = 0.0058). The respective percentages of phenotype positivity for HFE H63D, D6S105(8), and HLA-A*03 were similar in patients and controls. HFE S65C, I105T, and G93R were not detected in index or control subjects. Two of 13 patients were heterozygous for the ferroportin allele nt 744 G-->T (Q248H), although the phenotype frequency of this allele was similar in patients and 39 controls. Synonymous ferroportin alleles were also detected in some patients. The ceruloplasmin mutation nt 1099C-->T (exon 6; Arg367Cys) was detected in 1 of 2 patients tested. Abnormal alleles of beta-2 microglobulin, Nramp2, TFR2, hepcidin, or IRP2 alleles were not detected in either of the 2 patients so tested. We conclude that primary iron overload in African Americans is not the result of the mutation of a single gene. HFE C282Y, ferroportin 744 G-->T, and common forms of heritable anemia appear to account for increased iron absorption or retention in some patients.

Juvenile hemochromatosis in the southeastern United States: a report of seven cases in two kinships.

We report clinical and genetic characteristics of seven juvenile hemochromatosis (JH) patients (six females, one male) in two unrelated kinships from the southeastern U.S. All had severe iron overload. Mean age at diagnosis was 20 +/- 5 years (range 8-23 years). In six patients, the mean age at onset of signs and symptoms attributable to iron overload was 15 +/- 2 years (12-18 years); an 8-year-old girl had no symptoms. Six of the seven patients had hypogonadotrophic hypogonadism, two had severe cardiomyopathy, seven had hepatomegaly, two had hepatic cirrhosis, and five had hyperpigmentation. Two of four siblings with JH also had Hashimoto thyroiditis. One patient with severe cardiomyopathy improved with therapeutic phlebotomy, medical therapy for congestive heart failure, and a permanent pacemaker; the other died before phlebotomy was initiated. Estimates of average daily iron absorption before phlebotomy-induced iron depletion were 2.3, 3.1, and 1.7 mg in a male and two females, respectively. Both parents of four siblings with JH were heterozygous at two Ch1q loci (D1S1156, D1S2344); each of the four affected siblings was homozygous at both loci. An unaffected sib was heterozygous at D1S1156. One patient was heterozygous for HFE H63D, five others did not have HFE C282Y or H63D, and one was unavailable for testing. We conclude that JH occurs in the southeastern U.S. It is likely that JH allele(s) in at least one of the present kinships occur(s) on Ch1q, and presumably this represents a mutation(s) of the same gene localized to Ch1q in Italian and Greek JH kindreds. The present cases do not have HFE genotypes typical of hemochromatosis diagnosed in adults. Hashimoto thyroiditis, linked to Ch6p in many kinships, did not segregate with JH alleles on Ch1q in the present kinship.