Wolbachia strain w AlbB maintains high density and dengue inhibition following introduction into a field population of Aedes aegypti

Abstract

Aedes aegypti mosquitoes carrying the wAlbB Wolbachia strain show a reduced capacity to transmit dengue virus. wAlbB has been introduced into wild Ae. aegypti populations in several field sites in Kuala Lumpur, Malaysia, where it has persisted at high frequency for more than 2 years and significantly reduced dengue incidence. Although these encouraging results indicate that wAlbB releases can be an effective dengue control strategy, the long-term success depends on wAlbB maintaining high population frequencies and virus transmission inhibition, and both could be compromised by Wolbachia-host coevolution in the field. Here, wAlbB-carrying Ae. aegypti collected from the field 20 months after the cessation of releases showed no reduction in Wolbachia density or tissue distribution changes compared to a wAlbB laboratory colony. The wAlbB strain continued to induce complete unidirectional cytoplasmic incompatibility, showed perfect maternal transmission under laboratory conditions, and retained its capacity to inhibit dengue. Additionally, a field-collected wAlbB line was challenged with Malaysian dengue patient blood, and showed significant blocking of virus dissemination to the salivary glands. These results indicate that wAlbB continues to inhibit currently circulating strains of dengue in field populations of Ae. aegypti, and provides additional support for the continued scale-up of Wolbachia wAlbB releases for dengue control. This article is part of the theme issue 'Novel control strategies for mosquito-borne diseases'.

Keywords: Aedes aegypti; Wolbachia; dengue virus; vector control; virus blocking; wAlbB.

Figure 1.

Wolbachia density in whole adult females. Wolbachia density was determined at 5 and 10 days post adult eclosion by qPCR. Twenty-four females were analysed per group. Individual dots represent Wolbachia densities in individual females. A significant difference was found between wAlbB.MC and wAlbB.L densities at day 5 (pairwise t-test, p < 0.0001), although no difference was found at day 10 (p = 0.17). Boxplots show median and interquartile range.

Figure 2.

Wolbachia density in dissected tissues across mosquito lines and ages. Ovaries, midguts and salivary glands were dissected at days 5 and 10 post adult eclosion, and wAlbB density was determined by qPCR. Each of the five dots represents the Wolbachia density from pools of tissue from three individual female mosquitoes. No significant differences were observed in ovary density (line effect: F_1,16 = 0.97, p = 0.33; age effect: F_1,16 = 0.02, p = 0.89; line-by-age interaction: F_1,16 = 0.55, p = 0.47) or salivary gland density (line effect: F_1,16 = 0.004, p = 0.95; age effect: F_1,16 = 3.39, p = 0.08; line-by-age interaction: F_1,16 = 0.002, p = 0.96). A significantly higher Wolbachia density was observed in the midguts of the wAlbB.MC line at day 10 (line effect: F_1,16 = 26.77, p < 0.0001; age effect: F_1,16 = 14.45, p = 0.0016; line-by-age interaction: F_1,16 = 14.96, p = 0.0014), but not at day 5 (pairwise t-test, p = 0.6). Boxplots show median and interquartile range. Five biological replicates were performed for each treatment.

Figure 3.

DENV-2 inhibition in wAlbB-carrying Ae. aegypti lines. Females from the wAlbB.MC, wAlbB.L and wild-type lines were fed a bloodmeal spiked with DENV-2. After an incubation period of 12 days, salivary glands were dissected and the viral load was quantified by RT-qPCR. Ten, 20 and 27 females were analysed for the wild-type, wAlbB.L and wAlbB.MC groups, respectively. Black dots indicate salivary glands from individual mosquitoes. Red lines indicate median values. wAlbB.MC and wAlbB.L showed a significant reduction in viral titres (p < 0.004 for both comparisons, one-way ANOVA with Dunnett's). There was no difference in viral load between the wAlbB.MC and wAlbB.L lines (p = 0.88, one-way ANOVA with Dunnett's).

Figure 4.

DENV-1 inhibition in wAlbB lines challenged with patient-derived blood. Females from the wAlbB.SA, wAlbB.L and wild-type lines were fed on clinical isolates of DENV-1 infected patient blood. After incubation periods of 7 and 9 days, midgut and salivary gland tissues were dissected, and the viral load was quantified by RT-qPCR. For midguts, tissues from 7, 18 and 13 females were analysed at day 7, and tissues from 7, 15 and 9 females were analysed at day 9 for the wAlbB.SA, wAlbB.L and wild-type groups, respectively. For salivary glands, tissues from 7, 16 and 13 females were analysed at day 7, and tissues from 6, 14 and 9 females were analysed at day 9 for the wAlbB.SA, wAlbB.L and wild-type groups, respectively. Black dots indicate tissues from individual mosquitoes. Red lines indicate median values. For midgut tissue, viral RNA titres in wAlbB.SA and wAlbB.L were significantly different from wild-type at day 7 (p < 0.004 for both wAlbB.SA and wAlbB.L, one-way ANOVA with Dunnett's) and at day 9 (p < 0.002 for wAlbB.SA wAlbB.L, one-way ANOVA with Dunnett's). Similarly, for salivary gland tissue, viral RNA titres in wAlbB.SA and wAlbB.L were significantly different from wild-type at day 7 (p < 0.0001 for both wAlbB.SA and wAlbB.L, one-way ANOVA with Dunnett's) and at day 9 (p < 0.0002 for wAlbB.SA wAlbB.L, one-way ANOVA with Dunnett's).