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Paired serial limiting dilution assays.

ABSTRACT: Serial limiting dilution (SLD) assays are a widely used tool in many areas of public health research to measure the concentration of target entities. This concentration can be estimated via maximum likelihood. Asymptotic as well as exact inference methods have been proposed for hypothesis testing and confidence interval construction in this one-sample problem. However, in many scientific applications, it may be of interest to compare the concentration of target entities between a pair of samples and construct valid confidence intervals for the difference in concentrations. In this paper, an exact, computationally efficient inferential procedure is proposed for hypothesis testing and confidence interval construction in the two-sample SLD assay problem. The proposed exact method is compared to an approach based on asymptotic approximations in simulation studies. The methods are illustrated using data from the University of North Carolina HIV Cure Center.

SUBMITTER: Li X

PROVIDER: S-EPMC9588648 | biostudies-literature | 2022 Oct

REPOSITORIES: biostudies-literature

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Paired serial limiting dilution assays.

Li Xiudi X May Susanne S Trumble Ilana M IM Archin Nancie M NM Hudgens Michael G MG

Statistics in medicine 20220817 24

Serial limiting dilution (SLD) assays are a widely used tool in many areas of public health research to measure the concentration of target entities. This concentration can be estimated via maximum likelihood. Asymptotic as well as exact inference methods have been proposed for hypothesis testing and confidence interval construction in this one-sample problem. However, in many scientific applications, it may be of interest to compare the concentration of target entities between a pair of samples ...[more]

PMID: 35975729

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Project description:Viral lysis of phytoplankton is one of the most common forms of death on Earth. Building on an assay used extensively to assess rates of phytoplankton loss to predation by grazers, lysis rates are increasingly quantified through dilution-based techniques. In this approach, dilution of viruses and hosts are expected to reduce infection rates and thus increase host net growth rates (i.e., accumulation rates). The difference between diluted and undiluted host growth rates is interpreted as a measurable proxy for the rate of viral lytic death. These assays are usually conducted in volumes ≥ 1 L. To increase throughput, we implemented a miniaturized, high-throughput, high-replication, flow cytometric microplate dilution assay to measure viral lysis in environmental samples sourced from a suburban pond and the North Atlantic Ocean. The most notable outcome we observed was a decline in phytoplankton densities that was exacerbated by dilution, instead of the increased growth rates expected from lowered virus-phytoplankton encounters. We sought to explain this counterintuitive outcome using theoretical, environmental, and experimental analyses. Our study shows that, while die-offs could be partly explained by a 'plate effect' due to small incubation volumes and cells adhering to walls, the declines in phytoplankton densities are not volume-dependent. Rather, they are driven by many density- and physiology-dependent effects of dilution on predation pressure, nutrient limitation, and growth, all of which violate the original assumptions of dilution assays. As these effects are volume-independent, these processes likely occur in all dilution assays that our analyses show to be remarkably sensitive to dilution-altered phytoplankton growth and insensitive to actual predation pressure. Incorporating altered growth as well as predation, we present a logical framework that categorizes locations by the relative dominance of these mechanisms, with general applicability to dilution-based assays.

Dataset Information

Paired serial limiting dilution assays.

Publications

Paired serial limiting dilution assays.

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