Statistical Methods for Standard Membrane-Feeding Assays to Measure Transmission Blocking or Reducing Activity in Malaria

ABSTRACT

Transmission blocking vaccines for malaria are not designed to directly protect vaccinated people from malaria disease, but to reduce the probability of infecting other people by interfering with the growth of the malaria parasite in mosquitoes. Standard membrane-feeding assays compare the growth of parasites in mosquitoes from a test sample (using antibodies from a vaccinated person) compared to a control sample. There is debate about whether to estimate the transmission reducing activity (TRA) which compares the mean number of parasites between test and control samples, or transmission blocking activity (TBA) which compares the proportion of infected mosquitoes. TBA appears biologically more important since each mosquito with any parasites is potentially infective; however, TBA is less reproducible and may be an overly strict criterion for screening vaccine candidates. Through a statistical model, we show that the TBA estimand depends on μ_c, the mean number of parasites in the control mosquitoes, a parameter not easily experimentally controlled. We develop a standardized TBA estimator based on the model and a given target value for μ_c which has better mean squared error than alternative methods. We discuss types of statistical inference needed for using these assays for vaccine development. Supplementary materials for this article are available online.

KEYWORDS:

• Assay normalization
• Negative binomial
• Plasmodium falciparum
• Standardization
• Zero inflation

Supplementary Materials

Supplementary materials include a PDF with supplemental sections S1-S6, a vignette for R code detailing the RCM estimation and ad-hoc TRA confidence intervals, as well as datasets, R code, and SAS code files.

Acknowledgments

The authors thank Carole Long, Erin Gabriel, and Dean Follmann for discussions on the statistical analysis of standard membrane-feeding assays.

Additional information

Funding

This study was supported in part by the Intramural Research Program of the National Institute of Allergy and Infectious Diseases, NIH and also by the PATH Malaria Vaccine Initiative. This work used the computational resources of the NIH HPC Biowulf cluster (http://hpc.nih.gov).