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Background Robust population size estimates of female sex workers and other key populations in South Africa face multiple methodological limitations, including inconsistencies in surveillance and programmatic indicators. This has, consequently, challenged the appropriate allocation of resources and benchmark-setting necessary to an effective HIV response. A 2013-2014 integrated biological and behavioral surveillance (IBBS) survey from South Africa showed alarmingly high HIV prevalence among female sex workers in South Africa's three largest cities of Johannesburg (71.8%), Cape Town (39.7%), and eThekwini (53.5%). The survey also included several multiplier-based population size estimation methods. Objective The objective of our study was to present the selected population size estimation methods used in an IBBS survey and the subsequent participatory process used to estimate the number of female sex workers in three South African cities. Methods In 2013-2014, we used respondent-driven sampling to recruit independent samples of female sex workers for IBBS surveys in Johannesburg, Cape Town, and eThekwini. We embedded multiple multiplier-based population size estimation methods into the survey, from which investigators calculated weighted estimates and ranges of population size estimates for each city's female sex worker population. Following data analysis, investigators consulted civil society stakeholders to present survey results and size estimates and facilitated stakeholder vetting of individual estimates to arrive at consensus point estimates with upper and lower plausibility bounds. Results In total, 764, 650, and 766 female sex workers participated in the survey in Johannesburg, Cape Town, and eThekwini, respectively. For size estimation, investigators calculated preliminary point estimates as the median of the multiple estimation methods embedded in the IBBS survey and presented these to a civil society-convened stakeholder group. Stakeholders vetted all estimates in light of other data points, including programmatic experience, ensuring inclusion only of plausible point estimates in median calculation. After vetting, stakeholders adopted three consensus point estimates with plausible ranges: Johannesburg 7697 (5000-10,895); Cape Town 6500 (4579-9000); eThekwini 9323 (4000-10,000). Conclusions Using several population size estimates methods embedded in an IBBS survey and a participatory stakeholder consensus process, the South Africa Health Monitoring Survey produced female sex worker size estimates representing approximately 0.48%, 0.49%, and 0.77% of the adult female population in Johannesburg, Cape Town, and eThekwini, respectively. In data-sparse environments, stakeholder engagement and consensus is critical to vetting of multiple empirically based size estimates procedures to ensure adoption and utilization of data-informed size estimates for coordinated national and subnational benchmarking. It also has the potential to increase coherence in national and key population-specific HIV responses and to decrease the likelihood of duplicative and wasteful resource allocation. We recommend building cooperative and productive academic-civil society partnerships around estimates and other strategic information dissemination and sharing to facilitate the incorporation of additional data as it becomes available, as these additional data points may minimize the impact of the known and unknown biases inherent in any single, investigator-calculated method.