Two field-collected Culex quinquefasciatus colonies were subjected to selection pressure by three strains of Bacillus sphaericus, C3-41, 2362, and IAB59, under laboratory conditions. After 13 and 18 generations of exposure to high concentrations of C3-41 and IAB59, a field-collected low-level-resistant colony developed >144,000-and 46.3-fold resistance to strains C3-41 and IAB59, respectively. A field-collected susceptible colony was selected with 2362 and IAB59 for 46 and 12 generations and attained >162,000-and 5.7-fold resistance to the two agents, respectively. The pattern of resistance evolution in mosquitoes depended on continuous selection pressure, and the stronger the selection pressure, the more quickly resistance developed. The resistant colonies obtained after selection with B. sphaericus C3-41 and 2362 showed very high levels of cross-resistance to B. sphaericus 2362 and C3-41, respectively, but they displayed only low-level cross-resistance to IAB59. On the other hand, the IAB59-selected colonies had high cross-resistance to both strains C3-41 and 2362. Additionally, the slower evolution of resistance against strain IAB59 may be explained by the presence of another larvicidal factor. This is in agreement with the nontoxicity of the cloned and purified binary toxin (Bin1) of IAB59 for 2362-resistant larvae. We also verified that all the B. sphaericus-selected colonies showed no cross-resistance to Bacillus thuringiensis subsp. israelensis, suggesting that it would be a promising alternative in managing resistance to B. sphaericus in C. quinquefasciatus larvae.

Investigations on the inheritance and mechanism of resistance to Bacillus sphaericus Neide in Culex quinquefasciatus Say colonies, selected with strains C3-41 (RLCq1/C3-41) and 2362 (CqRL1/2362), were performed in China and Brazil, respectively. The progeny of reciprocal F1 crosses (susceptible female x resistant male and vice versa) from both resistant colonies responded alike in bioassays, indicating recessive inheritance. Data on larvae susceptibility from the backcross offspring between F1 and their respective susceptible and resistant parental colonies are consistent with a monofactorial and autosomal mode of inheritance. In vitro binding assays between 125I binary (Bin2) toxin and the brush border membrane fractions (BBMF) from CqRL1/2362 and RLCq1/C3-41 larvae showed that resistance, in both colonies, is caused by a failure in the binding step of the B. sphaericus Bin2 toxin to its specific midgut receptor. The specific and saturable binding of Bin2 toxin to BBMF from F1 larvae (CqRL1/2362 X susceptible counterpart) confirms the recessive inheritance of the resistance gene. Further studies are needed to advance understanding of B. sphaericus resistance.

Two colonies of Culex quinquefasciatus Say (Diptera: Culicidae) were selected with Bacillus sphaericus strains C3-41 and IAB59 in the laboratory for 13 and 18 generations; they attained 145,000- and 48.3-fold resistance, respectively, in comparison with a susceptible laboratory colony (SLCq) and showed very high levels of cross-resistance (8500- to 145,000-fold) to B. sphaericus strains C3-41, 1593, 2297 and 2362. They were relatively susceptible to B. sphaericus strains LP1-G and 47-6B (only 0.8- to 2.8-fold tolerance), with 24.8- to 48.3-fold cross-resistance to strain IAB59. B. sphaericus-resistant mosquito colonies remained highly susceptible to B. thuringiensis israelensis, suggesting that B.t.i. would be of value in the management of B. sphaericus-resistant Cx. quinquefasciatus colonies. The demonstration of low or no cross-resistance of two selected resistant Cx. quinquefasciatus colonies to IAB59, LP1-G and 47-6B strains of B. sphaericus and the finding of a major 49 kDa protein in these strains suggest that there is likely to be another mosquitocidal factor in the three strains.