Statistical analysis of outcomes
SECONDARY
The analysis of the collected data in the study will be performed with SAS v9.4. The primary analysis will be performed on the intention-to-treat population. Details regarding the defined populations and the statistical analysis will be provided in the statistical analysis plan (SAP). The SAP will be prepared before unblinding the trial.The analyses described below will be performed for the comparison of the probiotic and the placebo arms and the probiotic and the metformin arms, separately.Demographic and baseline characteristics will be summarized and compared between the groups descriptively. The data will be presented as summary tables and, where appropriate, as plots. Continuous variables will be presented as means, standard deviation, median, minimum and maximum, for categorical data frequencies and relative frequencies will be used.The primary outcome, changes in serum free testosterone concentration from baseline (screening visit) to 6 months (visit 2) will be compared between the probiotic and the placebo arms by analysis of covariance (ANCOVA) with serum free testosterone concentration at month 6 as dependent variable, and the baseline serum free testosterone concentration and treatment group as covariates. If the assumptions of the ANCOVA are not fulfilled, the changes in serum free testosterone concentration (i.e. difference visit 2 - baseline) will be compared between the groups by t-test or Mann-Whitney-U-test. A two-sided p-value of < 0.05 is considered to indicate statistical significance.For the secondary and exploratory endpoints, group comparisons will be performed using parametric or nonparametric methods for unpaired data (as appropriate). I.e. the continuous secondary endpoints will be analysed similarly to the primary endpoint. In addition, linear mixed models will be used to analyse the group difference over the three time points (baseline, visit 2 and follow-up visit).The collected stool samples will be sequenced with Illumina and the raw data will be uploaded on the Galaxy server of the Medical University of Graz. Microbiome analysis will be conducted using the QIIME 2 pipeline of the Galaxy server, the samples will be compared in respect to alpha- and beta-diversity using established methods. The results can be downloaded directly from the server.