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The symbiont utilizes vertical (mother-to-offspring) and horizontal (sexual) transmission routes to propagate across anopheline populations.\n                      <jats:italic toggle=\"yes\">Microsporidia MB<\/jats:italic>\n                      occurs throughout the\n                      <jats:italic toggle=\"yes\">Anopheles<\/jats:italic>\n                      life cycle, with no evidence to date of the existence of a secondary host. In this study, we conducted a comprehensive molecular screening of\n                      <jats:italic toggle=\"yes\">Microsporidia MB<\/jats:italic>\n                      in aquatic macrofauna associated with\n                      <jats:italic toggle=\"yes\">Anopheles<\/jats:italic>\n                      larval habitats to determine whether they serve as secondary hosts for\n                      <jats:italic toggle=\"yes\">Microsporidia MB<\/jats:italic>\n                      . We collected aquatic macrofauna from anopheline breeding sites along the Ahero irrigation scheme, including insects from the orders Hemiptera (families Naucoridae, Micronectidae, Nepidae, and Notonectidae), Coleoptera (families Staphylinidae, Noteridae, Hydrophilidae, and Dytiscidae), Diptera (anopheline and non-anopheline larvae and pupae), Orthoptera, and Ephemeroptera; snails (Hygrophila); crustaceans (Podocopida); and frogs (Anura). We detected\n                      <jats:italic toggle=\"yes\">Microsporidia MB<\/jats:italic>\n                      in 15.04% of anopheline larvae, but not in non-anopheline mosquito larvae, pupae, or any other aquatic macrofauna, despite their continuous interaction with anopheline mosquitoes in the larval habitats. The absence of\n                      <jats:italic toggle=\"yes\">Microsporidia MB<\/jats:italic>\n                      detection across the diverse assemblage of aquatic macrofauna surveyed in this study suggests that these taxa are unlikely to constitute major secondary hosts or reservoirs of the symbiont. While broader surveys across additional taxa are needed to fully exclude alternative hosts, the consistent detection of\n                      <jats:italic toggle=\"yes\">Microsporidia MB<\/jats:italic>\n                      at high infection intensities in\n                      <jats:italic toggle=\"yes\">Anopheles<\/jats:italic>\n                      mosquitoes supports a strong host association. This reinforces its potential as a targeted, species-specific, and environmentally safe symbiont-based intervention for malaria control.\n                    <\/jats:p>\n                    <jats:sec>\n                      <jats:title>IMPORTANCE<\/jats:title>\n                      <jats:p>\n                        Malaria continues to pose a significant global health burden, particularly in sub-Saharan Africa, and causes thousands of deaths annually.\n                        <jats:italic toggle=\"yes\">Microsporidia MB<\/jats:italic>\n                        is an anopheline-associated symbiont that is vertically and horizontally transmitted in\n                        <jats:italic toggle=\"yes\">Anopheles<\/jats:italic>\n                        mosquitoes and shows great promise as a complementary malaria control tool due to its\n                        <jats:italic toggle=\"yes\">Plasmodium<\/jats:italic>\n                        blocking phenotype. However, its lifecycle is not fully understood. Understanding its lifecycle and host range is important for assessing the symbiont\u2019s ecological safety and feasibility for field release. This study found no evidence of\n                        <jats:italic toggle=\"yes\">Microsporidia MB<\/jats:italic>\n                        in a wide range of aquatic macrofauna using sensitive molecular screening, despite their frequent contact with infected mosquito larvae. While non-detection does not rule out rare or transient infections, the results suggest that these organisms are unlikely to serve as major hosts of the symbiont. This apparent host specificity reduces concerns about spreading to non-target species and supports the development of\n                        <jats:italic toggle=\"yes\">Microsporidia MB<\/jats:italic>\n                        as a targeted malaria-control intervention.\n                      <\/jats:p>\n                    <\/jats:sec>\n                  <\/jats:sec>","DOI":"10.1128\/spectrum.00679-26","type":"journal-article","created":{"date-parts":[[2026,9,2]],"date-time":"2026-09-02T16:44:20Z","timestamp":1788367460000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":0,"title":["Host range of\n                    <i>Microsporidia MB<\/i>\n                    in aquatic macrofauna inhabiting\n                    <i>Anopheles<\/i>\n                    breeding sites"],"prefix":"10.1128","volume":"14","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-6715-634X","authenticated-orcid":false,"given":"Cynthia Nyambura","family":"King\u2019ori","sequence":"first","affiliation":[{"name":"International Centre of Insect Physiology and Ecology (ICIPE)","place":["Kasarani, Kenya"]},{"id":[{"id":"https:\/\/ror.org\/015h5sy57","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Biochemistry, Jomo Kenyatta University of Agriculture and Technology","place":["Nairobi, Kenya"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Collins Odongo","family":"Okoth","sequence":"additional","affiliation":[{"name":"International Centre of Insect Physiology and Ecology (ICIPE)","place":["Kasarani, Kenya"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Joel Ltilitan","family":"Bargul","sequence":"additional","affiliation":[{"id":[{"id":"https:\/\/ror.org\/015h5sy57","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Biochemistry, Jomo Kenyatta University of Agriculture and Technology","place":["Nairobi, Kenya"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sevgan","family":"Subramanian","sequence":"additional","affiliation":[{"name":"International Centre of Insect Physiology and Ecology (ICIPE)","place":["Kasarani, Kenya"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2239-7275","authenticated-orcid":true,"given":"Jeremy Keith","family":"Herren","sequence":"additional","affiliation":[{"name":"International Centre of Insect Physiology and Ecology (ICIPE)","place":["Kasarani, Kenya"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4490-9449","authenticated-orcid":true,"given":"Thomas Ogao","family":"Onchuru","sequence":"additional","affiliation":[{"name":"International Centre of Insect Physiology and Ecology (ICIPE)","place":["Kasarani, Kenya"]},{"id":[{"id":"https:\/\/ror.org\/03p41j374","id-type":"ROR","asserted-by":"publisher"}],"name":"Department of Physical and Biological Sciences, Bomet University","place":["Bomet, Kenya"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"235","reference":[{"key":"e_1_3_3_2_2","unstructured":"World Health Organization. 2025. 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