Our purpose was to identify an experimental procedure using PCR that provides a reliable genotype at a microsatellite locus using only a few picograms of template DNA. Under these circumstances, it is possible (i) that one allele of a heterozygous individual will not be detected and (ii) that PCR-generated alleles or 'false alleles' will arise. A mathematical model has been developed to account for stochastic events when pipetting template DNA in a very dilute DNA extract and computer simulations have been performed. Laboratory experiments were also carried out using DNA extracted from a bear feces sample to determine if experimental results correlate with the mathematical model. The results of 150 typing experiments are consistent with the proposed model. Based on this model and the level of observed false alleles, an experimental procedure using the multiple tubes approach is proposed to obtain reliable genotypes with a confidence level of 99%. This multiple tubes procedure should be systematically used when genotyping nuclear loci of ancient or forensic samples, museum specimens and hair or feces of free ranging animals.

Pyrenean brown bears Ursus arctos are threatened with extinction. Management efforts to preserve this population require a comprehensive knowledge of the number and sex of the remaining individuals and their respective home ranges. This goal has been achieved using a combination of noninvasive genetic sampling of hair and faeces collected in the field and corresponding track size data. Genotypic data were collected at 24 microsatellite loci using a rigorous multiple-tubes approach to avoid genotyping errors associated with low quantities of DNA. Based on field and genetic data, the Pyrenean population was shown to be composed at least of one yearling, three adult males, and one adult female. These data indicate that extinction of the Pyrenean brown bear population is imminent without population augmentation. To preserve the remaining Pyrenean gene pool and increase genetic diversity, we suggest that managers consider population augmentation using only females. This study demonstrates that comprehensive knowledge of endangered small populations of mammals can be obtained using noninvasive genetic sampling.

BackgroundCOVID-19 may have a profound impact on sexual health, reproductive health and social life across the world. Shelter in place regulations that have extended across the globe may influence condomless sex, exacerbate intimate partner violence and reduce access to essential reproductive health services. Population representative research is challenging during shelter in place, leaving major gaps in our understanding of sexual and reproductive health during COVID-19. This International Sexual Health And ReproductivE health (I-SHARE) study protocol manuscript describes a common plan for online national surveys and global comparative analyses.MethodsThe purpose of this cross-sectional study is to better understand sexual and reproductive health in selected countries during the COVID-19 pandemic and facilitate multinational comparisons. Participants will be recruited through an online survey link disseminated through local, regional and national networks. In each country, a lead organisation will be responsible for organising ethical review, translation and survey administration. The consortium network provides support for national studies, coordination and multinational comparison. We will use multilevel modelling to determine the relationship between COVID-19 and condomless sex, intimate partner violence, access to reproductive health services, HIV testing and other key items. This study protocol defines primary outcomes, prespecified subanalyses and analysis plans.ConclusionThe I-SHARE study examines sexual and reproductive health at the national and global level during the COVID-19 pandemic. We will use multilevel modelling to investigate country-level variables associated with outcomes of interest. This will provide a foundation for subsequent online multicountry comparison using more robust sampling methodologies.