Hypospadias

In rodents, AGD is measured from the anus to the posterior base of the genital tubercle (Gallavan et al., 1999 ). In contrast to rodents, the external genitalia are well developed at birth in humans with the genital tubercle transformed into the penis in males and clitoris in females. Investigators have used different landmarks to measure AGD in humans to replicate the measurement in rodents. In males, AGD has been measured from the anus to the perineoscrotal junction (anoscrotal distance) (Salazar-Martinez et al., 2004 (link)), to the posterior base or to the anterior base of the penis (Hsieh et al., 2008 ) (Fig.1). Measurements in females use the distance from the anus to the anterior fourchette (anofourchettal distance) (Salazar-Martinez et al., 2004 (link)) or to the base of the clitoris (anoclitoral distance) (Liu et al., 2014 (link)). The method described by Salazar-Martinez et al. is commonly used; it is more reliable and has a lower inter-observer variability (Dean and Sharpe, 2013 (link); Papadopoulou et al., 2013 (link); Salazar-Martinez et al., 2004 (link)). In this review, the term AGD describes ‘anoscrotal distance’ in males and ‘anofourchettal distance’ in females unless otherwise stated (Salazar-Martinez et al., 2004 (link)). Although AGD has been widely used as a marker of potential endocrine disruption in utero, its limitations include a lack of standardisation of methodology and information on reproducibility (Table-1) and insufficient data on normative references, including ethnic differences (Dean and Sharpe, 2013 (link)). AGD is associated with birth weight to a varying degree depending on the population studied (regression coefficient adjusted for gestation ranges from 1.5 to 3.0 mm/kg) (Papadopoulou et al., 2013 (link); Romano-Riquer et al., 2007 ; Salazar-Martinez et al., 2004 (link)) and there is no consensus for adjusting AGD for the variations in body size. In addition, low birth weight is itself a risk factor for TDS as it is associated with hypospadias, cryptorchidism, male infertility and TGCC (Francois et al., 1997 (link); Juul et al., 2014 (link); Michos et al., 2007 (link); Toppari et al., 2010 (link)).

Six thousand three hundred ninety-six men aged 15–49 years were enrolled into 2 trials of male circumcision for HIV and STI prevention in Rakai, Uganda. The design and results of the trials have been reported elsewhere [8 (link), 9 ]. In brief, eligible persons were informed of study procedures and risks, provided written informed consent before screening, and provided additional written consent for enrollment. Men were excluded from the trial if they had anemia, active genital infections, anatomical abnormalities (e.g., hypospadias), or medical indications or contraindications for surgery (e.g., severe phimosis). Men were randomly assigned to receive immediate circumcision or circumcision delayed for 24 months. Serological testing for HIV, HSV-2, and syphilis and physical examinations and interviews to ascertain sociodemographic characteristics and sexual risk behaviors were conducted at baseline and repeated at 6-, 12-, and 24-month follow-up visits. Samples were collected by trained staff, and serum was stored at −70°C. At each visit, all subjects were provided free HIV counseling and testing, health education, and condoms. All participants found to be HIV positive were referred to an HIV treatment program funded by the President’s Emergency Fund for AIDS Relief.
The trials were approved by 4 institutional review boards: the Science and Ethics Committee of the Uganda Virus Research Institute (Entebbe, Uganda), the HIV Subcommittee of the National Council for Research and Technology (Kampala, Uganda), the Committee for Human Research at Johns Hopkins University Bloomberg School of Public Health (Baltimore, MD), and the Western Institutional Review Board (Olympia, WA). The trials were overseen by independent data safety monitoring boards [8 (link), 9 ] and were registered with ClinicalTrials.gov (identifiers NCT00425984 and NCT00124878).
HSV-2 infection was determined by HSV-2 ELISA (Kalon Biological). The assay was performed according to the manufacturer’s protocol with minor modifications. Test samples were run in duplicate, and the mean index value was used to classify HSV-2 status. On the basis of prior evaluation of test performance among Ugandan serum samples, HSV-2–positive individuals were defined as those with an index value of ≥1.5 [10 (link)]. HSV-2 seroconversion was defined as negative serological results (index value <0.9) at enrollment followed by positive serological results during follow-up.
HIV status was determined using 2 separate ELISAs and was confirmed by HIV-1 Western blot, as described elsewhere [8 (link)]. Active Treponema pallidum infection was determined by a positive rapid plasma reagin test result (Becton Dickinson) or a toluidine red unheated serum test result (New Horizons Diagnostics) followed by a positive T. pallidum particle agglutination assay result (Serodia-TP PA Kit; Fujirebio).
For exploratory analyses, covariates associated with prevalent HSV-2 infection were assessed by characteristics and behaviors at enrollment. Because the timing of the initial HSV-2 infection was unknown and could have preceded enrollment by several years, we assessed long-term risk behaviors (such a lifetime number of sex partners) rather than behaviors reported in the year preceding enrollment. For analyses of incident HSV-2 infection, we assessed associations with fixed covariates (such as age, marital status, and education at enrollment) and by time-varying covariates (such as sexual risk behaviors— e.g., number of partners, nonmarital relationships, condom use, and alcohol use with sexual intercourse) reported at follow-up visits. Symptoms of STIs (such as GUD) reported at follow-up visits were not included in regression models of predictors of HSV-2 infection because these are likely to be consequences of HSV-2 infection rather than causes of HSV-2 acquisition. Risk factors with P < .15 in univariate analysis were entered into the multivariable model to adjust for possible confounding. The multivariable model adjusted for all covariates shown.
Prevalence ratios (PRs) were used to measure the associations between prevalent HSV-2 infection and the potential risk factors at enrollment. A Poisson model was used to estimate adjusted PRs (adjPRs).
Incidence rate ratios (IRRs) were used to evaluate the association between HSV-2 acquisition among individuals with a baseline Kalon index value <0.9. For incidence-rate and person-time calculations, it was assumed that HSV-2 infection occurred at the middle time point between the last negative and first positive serological test result. Time from enrollment was accumulated for the 24-month follow-up visit or the visit at which the last sample was available, and HSV-2 incidence was estimated per 100 person-years. Adjusted IRRs (adjIRRs) were estimated for fixed and time-varying covariates (to account for changes in risk behaviors over time).