Review 1: "The Plasmodium Transmission-Blocking Symbiont, Microsporidia MB, is Vertically 1 Transmitted through Anopheles Arabiensis Germline Stem Cells"

Steve Perlman

doi:doi:10.1162/2e3983f5.e4f849a7

The Plasmodium transmission-blocking symbiont, Microsporidia MB, is vertically transmitted through Anopheles arabiensis germline stem cells

by Thomas O. Onchuru, Edward E. Makhulu, Purity C. Ronnie, Stancy Mandere, Fidel G. Otieno, Joseph Gichuhi, and Jeremy K. Herren

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Published on Jun 17, 2024
www.biorxiv.org

Description

Abstract Microsporidia MB is a promising candidate for developing a symbiont-based strategy for malaria control because it disrupts the capacity of An. arabiensis to transmit the Plasmodium parasite. The symbiont is predominantly localized in the reproductive organs and is transmitted vertically from mother to offspring and horizontally (sexually) during mating. Due to the contribution of both transmission routes, Microsporidia MB has the potential to spread through target vector populations and become established at high prevalence. Stable and efficient vertical transmission of Microsporidia MB is important for its sustainable use for malaria control, however, the vertical transmission efficiency of Microsporidia MB can vary. In this study, we investigate the mechanistic basis of Microsporidia MB vertical transmission in An. arabiensis. We show that vertical transmission occurs through the acquisition of Microsporidia MB by Anopheles cystocyte progenitors following the division of germline stem cells. We also show that Microsporidia MB replicates to increase infection intensity in the oocyte of developing eggs when mosquitoes are given a blood meal suggesting that symbiont proliferation in the ovary is coordinated with egg development. The rate of Microsporidia MB transmission to developing eggs is on average higher than the recorded (mother to adult offspring) vertical transmission rate. This likely indicates that a significant proportion of An. arabiensis offspring lose their Microsporidia MB symbionts during development. The stability of germline stem cell infections, coordination of symbiont proliferation, and very high rate of transmission from germline stem cells to developing eggs indicate that Microsporidia MB has a highly specialized vertical transmission strategy in An. arabiensis, which may explain host specificity.Author Summary Mosquito vectors of diseases are associated with a broad range of microbes. Some of the microbes significantly affect vector biology including pathogen transmission efficiency. Anopheles mosquitoes, which transmit the malaria parasite, Plasmodium falciparum, harbor a native microbe known as Microsporidia MB. This microbe interferes with the formation of transmissible stages of the parasite that are transferred to humans by female mosquitoes when taking a blood meal. This phenotype can be exploited to develop a novel strategy for controlling malaria similar to the control of dengue fever using Aedes mosquitoes carrying Wolbachia bacteria. Mother-to-offspring transmission of protective microbes is important in sustainable application of microbe-based technologies to control vector-borne diseases because it ensures maintenance of the microbe in target vector populations across many generations. Here, we investigated stability of Microsporidia MB infections and efficiency of mother-to-offspring transmission during early stages of egg formation and development. We found that this microbe has a specialized transmission mechanism that involves infecting the germline cells that are important in egg production. We also demonstrated a very high transmission rate (97%) of the Microsporidia MB from infected germline cells into daughter cells during cell division. As the germline daughter cells developed into eggs, Microsporidia MB established itself in the egg yolk through active replication which only occured after the female mosquitoes had a blood meal. Our study gives insights into an efficient mother-to-offspring transmission route of Microsporidia MB that can be utilized sustainably in microbe-based intervention to control malaria.

RR:C19 Evidence Scale rating by reviewer:

Strong. The main study claims are very well-justified by the data and analytic methods used. There is little room for doubt that the study produced has very similar results and conclusions as compared with the hypothetical ideal study. The study’s main claims should be considered conclusive and actionable without reservation.

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Review: The authors recently discovered Microsporidia MB, a microsporidian microbe that infects Anopheles arabiensis mosquitoes, and is transmitted vertically (from infected mothers to their offspring), and sexually. This microbe also interferes with transmission of Plasmodium, and is therefore an exciting candidate for malaria control.

Here, the authors perform a detailed microscopy study of Microsporidia MB vertical (i.e. maternal) transmission, using Fluorescence in situ hybridization (FISH). They show that it targets and infects germ cells in the mosquito ovariole with extremely high efficiency; such efficient targeting of germline is reminiscent of Wolbachia, a highly abundant maternally transmitted bacterial symbiont of invertebrates, including mosquitoes. They also observe actively dividing Microsporidia cells following a mosquito bloodmeal, suggesting that microsporidian growth is coordinated with mosquito reproduction (although it’s missed whether the authors looked for replication in control mosquitoes that did not feed on blood). The authors also followed maternal transmission efficiency of field-caught mosquitoes. Interestingly, although almost all (97%) egg cells were infected, only about 60% of emerging adult offspring harbored Microsporidia MB. So an important outstanding question will be to understand when and why there is such a large decline in infection (authors could had screened mosquito larvae!).

This study is an important contribution to the biology of Microsporidia MB, and to vertically transmitted microsporidians, which are much less studied than their bacterial counterparts. It would be helpful to explain where Microsporidia MB belongs in the Microsporidia phylogeny, as well as the other lineages that the authors mention and compare with respect to transmission. Understanding transmission is critical for incorporating this microbe in potential malaria control strategies, and in understanding the ecology of mosquito-microsporidian-malaria interactions in general. The microscopy is detailed and beautiful. Finally, in addition to screening larvae, the authors could present more information about the screening experiments. How many isofemale lines were screened, and how much variation was there between lines? Was there a difference in infection in male and female offspring?

Review 1: "The Plasmodium Transmission-Blocking Symbiont, Microsporidia MB, is Vertically 1 Transmitted through Anopheles Arabiensis Germline Stem Cells"

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