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The rapidly expanding use of wastewater for public health surveillance requires new strategies to protect privacy rights, while data are collected at increasingly discrete geospatial scales, i.e., city, neighborhood, campus, and building-level. Data collected at high geospatial resolution can inform on labile, short-lived biomarkers, thereby making wastewater-derived data both more actionable and more likely to cause privacy concerns and stigma- tization of subpopulations. Additionally, data sharing restrictions among neighboring cities and communities can complicate efforts to balance public health protections with citizens’ privacy. Here, we have created an encrypted framework that facilitates the sharing of sensitive population health data among entities that lack trust for one another (e.g., between adjacent municipalities with different governance of health monitoring and data sharing). We demonstrate the utility of this approach with two real-world cases. Our results show the feasibility of sharing encrypted data between two municipalities and a laboratory, while performing secure private com- putations for wastewater-based epidemiology (WBE) with high precision, fast speeds, and low data costs. This framework is amenable to other computations used by WBE researchers including population normalized mass loads, fecal indicator normalizations, and quality control measures. The Centers for Disease Control and Pre- vention’s National Wastewater Surveillance System shows ~8 % of the records attributed to collection before the wastewater treatment plant, illustrating an opportunity to further expand currently limited community-level sampling and public health surveillance through security and responsible data-sharing as outlined here. 

Abstract A major technical hurdle for T cell immune profiling is the time and cost to accurately genotype the Human Leukocyte Antigen (HLA) loci from peripheral blood. Here, we developed a rapid, highly multiplexed approach for HLA typing using RNA from <100,000 peripheral blood mononuclear cells with the Oxford Nanopore Technology (ONT) Minion sequencer. This method uses selective reverse transcription of mRNA of six HLA loci (A,B,C, DRB1, DQB1, DPB1), followed by PCR amplification. The individual amplified HLA cDNA was multiplexed in a single sequencing pool using primers with unique molecular identifiers, designed to permit sequencing errors for enhanced data capture. Pooled HLA amplicons were sequenced using the ONT Minion MK1B and R10.4 flowcells, with sequence Q scores> 20. Total RNA was extracted from PBMC samples from 12 individuals, reverse transcribed and amplified using the designed HLA loci specific primers. The pooled, amplified cDNA was then sequenced for 16 hours on the ONT Minion sequencer. The resulting sequencing data was analyzed and an average depth of coverage of 6000x was observed per sample. An average per loci depth of coverage of 1000x was observed. This method is designed to permit rapid (<24h), low-cost, portable HLA sequencing for T cell immune monitoring and epitope identification for immunologic studies. Arizona Piper Foundation 

Abstract Because errors at the DNA level power pathogen evolution, a systematic understanding of the rate and molecular spectra of mutations could guide the avoidance and treatment of infectious diseases. We thus accumulated tens of thousands of spontaneous mutations in 768 repeatedly bottlenecked lineages of 18 strains from various geographical sites, temporal spread, and genetic backgrounds. Entailing over ∼1.36 million generations, the resultant data yield an average mutation rate of ∼0.0005 per genome per generation, with a significant within-species variation. This is one of the lowest bacterial mutation rates reported, giving direct support for a high genome stability in this pathogen resulting from high DNA-mismatch-repair efficiency and replication-machinery fidelity. Pathogenicity genes do not exhibit an accelerated mutation rate, and thus, elevated mutation rates may not be the major determinant for the diversification of toxin and secretion systems. Intriguingly, a low error rate at the transcript level is not observed, suggesting distinct fidelity of the replication and transcription machinery. This study urges more attention on the most basic evolutionary processes of even the best-known human pathogens and deepens the understanding of their genome evolution. 

Analysis of municipal wastewater, or sewage for public health applications is a rapidly expanding field aimed at understanding emerging epidemiological trends, including human and disease migration. The newly gained ability to extract and analyze genetic material from wastewater poses important societal and ethical questions, including: How to safeguard data? Who owns genetic data recovered from wastewater? What are the ethical and legal issues surrounding its use? In the U.S., both corporate and legal policies regarding privacy have been historically reactive instead of proactive. In wastewater-based epidemiology (WBE), the pace of innovation has outpaced the ability of social and legal mechanisms to keep up. To address this discrepancy, early and robust discussions of the research, policies, and ethics surrounding WBE analysis and genetics is needed. This paper contributes to this discussion by examining ownership issues for human genetic data recovered from wastewater and the uses to which it may be put. We focus particularly on the risks associated with personally identifiable data, highlighting potential risks, relevant privacy-enhancing technologies, and appropriate ethics. The paper proposes an approach for people conducting WBE studies to help them systematically consider the ethical and privacy implications of their work.