ABSTRACT
The Cqm1 α-glucosidase, expressed within the midgut of Culex quinquefasciatus mosquito larvae, is the receptor for the Binary toxin (Bin) from the entomopathogen Lysinibacillus sphaericus. Mutations of the Cqm1 α-glucosidase gene cause high resistance levels to this bacterium in both field and laboratory populations, and a previously described allele, cqm1REC, was found to be associated with a laboratory-resistant colony (R2362). This study described the identification of a novel resistance allele, cqm1REC-2, that was co-selected with cqm1REC within the R2362 colony. The two alleles display distinct mutations but both generate premature stop codons that prevent the expression of midgut-bound Cqm1 proteins. Using a PCR-based assay to monitor the frequency of each allele during long-term maintenance of the resistant colony, cqm1REC was found to predominate early on but later was replaced by cqm1REC-2 as the most abundant resistance allele. Homozygous larvae for each allele were then generated that displayed similar high-resistance phenotypes with equivalent low levels of transcript and lack of protein expression for both cqm1REC and cqm1REC-2. In progeny from a cross of homozygous individuals for each allele at a 1 : 1 ratio, analyzed for ten subsequent generations, cqm1REC showed a higher frequency than cqm1REC-2. The replacement of cqm1REC by cqm1REC -2 observed in the R2362 colony, kept for 210 generations, indicates changes in fitness related to traits that are unknown but linked to these two alleles, and constitutes a unique example of evolution of resistance within a controlled laboratory environment.
Subject(s)
Bacillaceae/pathogenicity , Culex/genetics , Culex/microbiology , Animals , Bacterial Toxins/metabolism , Bacterial Toxins/toxicity , Crosses, Genetic , Culex/enzymology , Evolution, Molecular , Female , Gene Frequency , Genes, Insect , Gram-Positive Bacterial Infections/genetics , Gram-Positive Bacterial Infections/microbiology , Insect Proteins/genetics , Insect Proteins/metabolism , Larva/enzymology , Larva/genetics , Larva/microbiology , Male , Mutation , Selection, Genetic , alpha-Glucosidases/genetics , alpha-Glucosidases/metabolismABSTRACT
Biological fitness components of a field-collected colony of Culex quinquefasciatus Say that was highly resistant to Bacillus sphaericus strain 2362 (resistance ratio greater than 163,000) after 46 generations of selection were compared to those of a susceptible colony (CqSF) that had originated from the same parental cohort but that had not been exposed to B. sphaericus. The effect of B. sphaericus on the fitness of Cx. quinquefasciatus was determined in terms of fecundity, fertility, and development time. The resistant colony (CqRL) showed significantly lower fecundity and fertility, and slower development than the susceptible colony. Development time from egg to egg showed a 20% increase in CqRL compared to CqSF. The generation time increased from 21.6 days to 26 days for highly resistant generations of CqRL.
