Wol­bachia: The Dif­fer­ence Be­tween a Nui­sance and a Threat?

by Jamie Schafer

No sum­mer evening out­doors seems com­plete with­out a mos­quito bite (or ten). While these bites are of­ten just a nui­sance, they can also serve as the point of trans­mis­sion for many mos­quito-borne dis­eases: West Nile virus, dengue virus, yel­low fever virus, malaria, and the fi­lar­ial ne­ma­todes that cause ele­phan­ti­a­sis, to name a few. Ef­forts to con­trol the spread of these dis­eases, many of which can­not be curbed by vac­ci­na­tion, have lately looked to the in­tra­cel­lu­lar bac­terium Wol­bachia pip­i­en­tis for help. Wol­bachia nat­u­rally in­fect many in­sects in­clud­ing some mos­qui­toes, and strains have re­cently been de­vel­oped to in­fect Aedes ae­gypti, the vec­tor for dengue, and Anophe­les gam­biae, malaria's ve­hi­cle of choice. In­fec­tion by Wol­bachia may limit the spread of these dis­eases by ef­fi­ciently be­com­ing es­tab­lished in in­sect pop­u­la­tions, where they shorten the lifes­pan of their hosts or di­rectly in­ter­fere with the pathogens the in­sects carry.

The par­a­sitoid wasp N. vit­ripen­nis is a Wol­bachia host in which the de­gree of cy­to­plas­mic in­com­pat­i­bil­ity cor­relates with bac­te­r­ial den­sity. Credit: Oliver Niehuis. Source

Wol­bachia spread very ef­fec­tively through in­sect pop­u­la­tions. For ex­am­ple, when re­searchers re­leased in­fected A. ae­gypti mos­qui­toes into a wild pop­u­la­tion, nearly all mos­qui­toes in the pop­u­la­tion be­came in­fected in just a few months. This ef­fi­ciency is due largely to a phe­nom­e­non called cy­to­plas­mic in­com­pat­i­bil­ity. Upon Wol­bachia in­fec­tion, dra­matic changes oc­cur in the host insect's ga­metes such that in­fected fe­males can re­pro­duce by mat­ing with ei­ther in­fected or un­in­fected males, but un­in­fected fe­males who mate with in­fected males have no off­spring. Since the bac­terium is passed on ma­ter­nally from in­fected fe­males to their eggs, this in­com­pat­i­bil­ity gives a re­pro­duc­tive ad­van­tage to in­fected fe­males, thereby fa­vor­ing the spread of Wol­bachia. Al­though its cause has re­mained some­what elu­sive, the de­gree of cy­to­plas­mic in­com­pat­i­bil­ity cor­re­lates with bac­te­r­ial den­sity. Re­cently it was also shown to in­versely cor­re­late with in­fec­tion of the Wol­bachia with bac­te­rio­phage WO‑B in the par­a­sitoid wasp Na­so­nia vit­ripen­nis. WO‑B lytic de­vel­op­ment would cause lower bac­te­r­ial den­sity, and may  ex­plain how WO‑B in­fec­tion causes dif­fer­ences in the strength of cy­to­plas­mic in­com­pat­i­bil­ity ob­served in some Wol­bachia strains.

Wol­bachia in­fect­ing the testes of the wasp N. vitri­pennis. Ar­row­heads = phage par­ti­cles in­side Wol­bachia. Field of view ~0.6 μm wide. Adapted from source.

Many strains of Wol­bachia also af­fect the lifes­pan of their hosts. One of these strains, wMelPop, was adapted from its orig­i­nal host, Drosophila melanogaster, to be able to in­fect the dengue vec­tor A. ae­gypti. The goal here was to re­duce the num­ber of mos­qui­toes that reach ma­tu­rity. Ma­ture mos­qui­toes are more likely to pass on dengue virus, since this virus must in­cu­bate in the mos­quito for sev­eral days be­fore be­ing trans­mis­si­ble. In­ter­est­ingly, the as­so­ci­a­tion be­tween wMelPop in­fec­tion and early mor­tal­ity is only ob­served in mos­qui­toes that have fed on blood. Upon closer in­ves­ti­ga­tion, it was found that for wMelPop-in­fected A. gam­biae, 80% die within 3 days of feed­ing on blood, com­pared to about 20% mor­tal­ity for un­in­fected mos­qui­toes. Since con­sum­ing blood in­creases the amount of iron in the in­sects, and Wol­bachia have been shown else­where to in­flu­ence iron me­tab­o­lism, the study au­thors sug­gest that the iron may be the cause of post-blood­meal mor­tal­ity. Re­ac­tive oxy­gen species could in­crease to fa­tal lev­els af­ter iron in­take be­cause Wol­bachia in­fec­tion im­pairs iron me­tab­o­lism. Al­though the ini­tial hope was to hin­der dis­ease trans­mis­sion by short­en­ing the lifes­pan of the in­sect hosts, this strat­egy may also ham­per the spread of in­fec­tion within wild pop­u­la­tions. The se­lec­tive ad­van­tage of Wol­bachia in­fec­tion due to cy­to­plas­mic in­com­pat­i­bil­ity may be out­weighed by the dis­ad­van­tage of a shorter lifes­pan.

Mor­tal­ity of wMelPop or con­trol cell ho­mogenate-in­­­jected mos­qui­toes af­ter feed­ing on P. fal­ci­parum in­fected or un­in­fected hu­man blood. Mos­qui­toes were col­lected every 24 hours pre-blood­meal or every 12 hours post-blood­meal. Af­ter blood­feed­ing, there is a dra­matic in­crease in mor­tal­ity of wMelPop-in­fected mos­qui­toes, re­sult­ing in ap­prox­i­mately 80% mor­tal­ity af­ter 3 days post-feed­ing. The slope of the mor­tal­ity tra­jec­tory of sur­viv­ing mos­qui­toes is sim­i­lar to con­trols. wMelPop treat­ments dif­fer from cell ho­mogenate treat­ments (P<0.0001), but Plas­mod­ium in­fec­tion sta­tus was not sig­nif­i­cant. Adapted from source.

Ef­forts have in­stead turned to other strains such as wMel and wAlbB that have less ef­fect upon the vec­tor insect's lifes­pan but can still di­rectly in­hibit dengue virus and Plas­mod­ium fal­ci­parum, re­spec­tively. The most dra­matic data come from a study with dengue that was com­pleted as ground­work for the open re­lease of Wol­bachia-in­fected mos­qui­toes pre­vi­ously men­tioned. Dengue virus-in­fected blood was fed to A. ae­gypti that were in­fected with the wMel Wol­bachia strain. Strik­ingly, dengue virus in­fec­tion was more con­trolled in mos­qui­toes that car­ried wMel Wol­bachia than in their Wol­bachia-free coun­ter­parts. Only 12.5% of the wMel-in­fected group had dengue virus through­out their body, as mea­sured by the pres­ence of the virus in their legs, com­pared to 82.6% of the non-Wol­bachia-in­fected group. Still more im­pres­sively, even those wMel-in­fected mos­qui­toes that have a dis­sem­i­nated dengue in­fec­tion of­ten do not have virus in their saliva and there­fore can­not pass on the virus dur­ing feed­ing. In a vi­ral plaque as­say us­ing pooled saliva from sev­eral in­di­vid­u­als, 29 of 36 pools from non-wMel mos­qui­toes formed plaques in­di­cat­ing the pres­ence of dengue virus, whereas for the wMel mos­qui­toes only 2 of 48 saliva pools did, and those formed only one plaque each. Fur­ther in­ves­ti­ga­tion re­vealed that one in­di­vid­ual in each of the plaque-form­ing wMel saliva pools was not ac­tu­ally in­fected with Wol­bachia, sug­gest­ing that per­haps none of the mos­qui­toes car­ry­ing wMel have de­tectable dengue virus in their saliva! Given the quick spread of wMel in­fec­tion in a wild pop­u­la­tion of A. ae­gypti and its ap­par­ent ef­fec­tive­ness in block­ing, or at least se­verely lim­it­ing, the pres­ence of dengue virus in mos­quito saliva, we will likely see more re­leases of Wol­bachia-in­fected mos­qui­toes to con­trol dengue in the fu­ture.

Pathogen in­ter­fer­ence with dengue virus (DENV). Mos­quitoes were fed DENV-2-in­fected blood orally. (a) qPCR of to­tal dengue virus in whole fe­male mos­qui­toes (n = 19–30). White bar = con­trol mos­qui­toes (wild-type, Wol­bachia-un­in­fected); blue bar = wMel-in­fected mos­qui­toes. (b) qPCR of dis­sem­i­nated dengue virus in the legs of in­di­vid­ual Wol­bachia-un­in­fected (black cir­cles) and wMel-in­fected (blue tri­an­gles) mos­qui­toes (n = 23–30). Adapted from source.

Mean­while, re­searchers con­tinue to de­velop a sim­i­lar promis­ing strat­egy to com­bat malaria. P. fal­ci­parum oocyst de­vel­op­ment is re­duced by 40 to 60% when its A. gam­biae vec­tors are in­fected with wAlbB or wMelPop Wol­bachia. How­ever, a ma­ter­nally-trans­mit­ted sta­ble in­fec­tion of Wol­bachia  was not achieved in A. gam­biae—a ma­jor hur­dle to be sur­mounted. Nev­er­the­less, it seems we may be draw­ing closer to the day when Wol­bachia may pro­tect us from dengue virus, and per­haps from other mos­quito-borne ill­nesses, so that a bite can be thought of as merely an itchy nui­sance rather than a threat of se­ri­ous dis­ease.

Ref­er­ences

Hughes GL, Koga R, Xue P, Fukatsu T, Ras­gon JL. (2011). Wol­bachia in­fec­tions are vir­u­lent and in­hibit the hu­man malaria par­a­site Plas­mod­ium fal­ci­parum in Anophe­les gam­biae. PLoS pathogens, 7 (5). PMID 21625582

Walker T, John­son PH, Mor­eira LA, Iturbe-Or­maetxe I, Fren­tiu FD, Mc­Meni­man CJ, Leong YS, Dong Y, Ax­ford J, Kries­ner P, Lloyd AL, Ritchie SA, O'Neill SL, Hoff­mann AA. (2011). The wMel Wol­bachia strain blocks dengue and in­vades caged Aedes ae­gypti pop­u­la­tions. Na­ture, 476 (7361), 450−453. PMID 21866159

Jamie is a stu­dent in the Ph.D. pro­gram in Vi­rol­ogy at Har­vard Uni­ver­sity. She stud­ies NK cells in SIV in­fec­tion.