Carbamate resistance in Anopheles albimanus: Penetration and metabolism of carbaryl in propoxur-selected larvae*

The roles of increased metabolism and reduced penetration as possible mechanisms of resistance to carbaryl were investigated in a strain of A. albimanus from El Salvador in which carbamate and organophosphorus resistance had been induced by laboratory selection with propoxur. Carbaryl was metabolized to N-hydroxymethylcarbaryl, 5,6-dihydro-5,6-dihydroxycarbaryl and an unidentified metabolite A. The rate of metabolism by the resistant strain was only slightly higher than that by the parental strain. The rate of penetration of carbaryl in the parental strain was nearly twice that in the resistant strain during the first hour of exposure. However, this difference did not persist for as long as 90 min. Because of the low level of carbamate metabolism detected, it was concluded that the high resistance of the strain must be due to the action of some as yet unidentified mechanism, possibly enhanced by factors controlling the penetration and metabolism of the toxicant.     

Laboratory studies on the behaviouristic resistance of Anopheles albimanus in Panama

The behaviouristic resistance of A. albimanus to DDT observed in populations on the Rio Chagres by Trapido in 1952 was re-investigated in 1958. Measurement of the excitatory effect of DDT in inducing flight showed that the mean excitation times for engorged adults from the Chagres populations of Gatuncillo and Santa Rosa were respectively significantly and almost significantly less than those for a population from an untreated area or for a laboratory population which had never encountered DDT. There was no difference in the susceptibility levels between the Gatuncillo, the untreated and the laboratory strains when they were confined to DDT deposits for one hour.     

DDT-resistance and dieldrin-resistance in Anopheles albimanus

The nature and mode of inheritance of the component resistances of a multi-resistant strain of Anopheles albimanus from El Salvador are described. Dieldrin-resistance is shown to be dependent on a single, semi-dominant, genetic factor and imparts cross-resistance to other cyclodiene chlorinated hydrocarbons and to γ-BHC. DDT-resistance imparts cross-resistance to DDT-analogues and is dependent on a single, recessive, genetic factor. The two resistances are separate entities and can, in fact, be separated. The modes of inheritance of the two types of resistance are the same in both larvae and adults.     

Recent experiments on possible resistance to DDT by Anopheles albimanus in Panama

Following the observation of a change in the effectiveness of DDT residual house-spraying for Anopheles albimanus control at two experimental villages on the Chagres River in Panama, laboratory experiments were made to measure possible physiological resistance in this mosquito.Blood-engorged albimanus from (a) a stock colony which had never had contact with DDT or any other chlorinated-hydrocarbon insecticide, (b) a stock-colony strain which had been exposed to DDT for more than 70 generations, and (c) two villages where 5% DDT in kerosene had been frequently applied, were exposed for 12 minutes to 0.5% p,p'-DDT in mineral oil.The results showed that both the wild albimanus and the exposed stock-colony strain had essentially the same susceptibility to DDT as the unexposed stock-colony mosquitos. Any difference in the effectiveness of the insecticide for albimanus is therefore considered to be due to a behaviour change rather than to a change in the intrinsic toxicity of DDT for this mosquito.     

Recessive lethal mutations in Anopheles albimanus

Six recessive lethal mutants of Anopheles albimanus are described. Homozygotes for three of the autosomal mutants, viz., bar eye, dot eye and hairy, die during the last larval or early pupal stages; complete linkage data are lacking for bar eye and hairy but dot eye is tightly linked to red eye on the right arm of chromosome 2. Larval and pupal mortality is high for homozygotes of the other two autosomal mutants, diseased larva and lunate. Survival to the adult stage of the lunate type is about 60% for both sexes, and for diseased larva the rates differed according to sex, 15% for males and 3% for females. A general lack of vitality of the surviving adults has prevented the establishment of homozygous stocks. From analyses of linkage data, diseased larva and lunate were assigned to chromosome 2, most probably on the left arm. The mutant, bubble head, is on the X chromosome, and therefore is expressed only in the hemizygous male, which usually dies during the early pupal stage.     

Microplate assay analysis of the distribution of organophosphate and carbamate resistance in Guatemalan Anopheles albimanus

Simple microplate assay methods for determining the frequency of insecticide resistance in single mosquitos were used to study the distribution and localization of organophosphate and carbamate resistance in field populations of Anopheles albimanus Weidemann in Guatemala, where such resistance, caused by heavy use of agricultural pesticides, has long been assumed to be widespread. Areas of complete susceptibility to organophosphates and carbamates were observed, as well as areas where the resistant phenotypes represented up to 98% of the population. Overall, the resistance levels were lower and more localized than expected. Two mechanisms of resistance were identified by the microassay methods. These were the elevated esterase (nonspecific esterase) and insensitive acetylcholinesterase mechanisms which were selected independently, the former (documented for the first time in Central American anophelines) being predominant. These methods represent a promising new technology for the detection and assessment of resistance and will facilitate improved control strategy decisions.     

Genetic structure of natural populations of Anopheles albimanus in Colombia.

Electrophoretic and cytogenetic studies were undertaken on the population structure of Anopheles albimanus from 11 localities in Colombia, 3 from northern (Atlantic coast) and 8 from southern (Pacific coast) regions. Of the 25 allozyme loci examined, significant allele frequency differences were observed at 4 loci: hydroxy acid dehydrogenase (Had-1) and 3 esterases (Est-2, Est-4 and Est-6). The northern populations had higher variability, with 55% polymorphic loci, a mean heterozygosity of 20.4% and a mean of 3.0 alleles per locus. These values for southern populations were 24%, 9.1% and 1.5%, respectively. There were neither diagnostic loci nor clinal effect on frequencies of allozymes. Except for a small inversion on the X chromosome in low frequency in certain populations, all populations were homosequential in chromosomal banding patterns. Hybrids from matings between natural populations and the Gainesville laboratory strain were fully fertile. Estimates of genetic similarities (0.95-0.97 among southern and 0.99-1.00 among northern populations) suggest a lack of significant genetic differentiation among distant populations in this species. Based on the chromosomal, hybridization and electrophoretic data, we concluded that mosquitoes from the 11 collections were conspecific populations of An. albimanus.     

Effect of temperature on an enzyme assay to detect fenitrothion resistance in Anopheles albimanus mosquitos

A laboratory strain of Anopheles albimanus Wiedemann of known fenitrothion resistance was used in the field to compare the results of the WHO test for determining fenitrothion resistance in mosquitos with those of an enzyme microplate assay. The level of resistance obtained with the enzyme assay increased with the ambient temperature, and in order to compensate for this temperature effect, the incubation time was reduced. With the adjusted incubation times, the results for the microassay from 23 degrees C to 32 degrees C were the same as those found with the WHO test. The fenitrothion resistance of a field population of A. albimanus mosquitos determined between 27 degrees C and 31 degrees C using the adjusted enzyme microassay or the WHO test did not differ in a statistically significant way.     

Heat-shock mortality and induced thermotolerance in larvae of the mosquito Anopheles albimanus.

Temperature effects on Anopheles albimanus larval survival were investigated. Larvae were exposed to 30 min heat shocks at various temperatures. Almost no mortality was observed at 40 degrees C, but was complete at 43 degrees C. Increased larval thermotolerance could be induced by higher rearing temperature or by a 30 min exposure to 37 degrees C.     

The biting and resting behavior of Anopheles albimanus in northern Haiti

A one-year study of the biting and resting habits of the malaria vector Anopheles albimanus was carried out in four rural villages of northern Haiti. Man-biting rates and nightly biting cycles were determined by the use of all-night man-biting captures inside and outside houses. Seasonal changes in density and behavior were determined by repeating the captures on a bimonthly basis throughout one year. Exophily was demonstrated in these anopheline populations by a comparison of inside-biting with inside-resting densities. These behavior characteristics are discussed in relation to malaria transmission and to the choice of malaria control methods.     

Results of fenitrothion selection of Culex pipiens fatigans Wied. and Anopheles albimanus Wied

Laboratory selection of Culex pipiens fatigans (=C. quinquefasciatus) and Anopheles albimanus by fenitrothion pressure for 30 and 25 generations, respectively, resulted in only 2.2-fold and 1.1-fold increases in tolerance to this compound. Cross-resistance to arprocarb in Culex increased 12.2-fold but smaller increases in tolerance to fenthion, malathion, DDT, and dieldrin, may be manifestations of vigour tolerance. Since the essential conditions for selection are believed to have been fulfilled, the authors consider the negative results obtained to be of practical significance, as well as of academic interest.     

Evidence for an inactive transposable mariner-like element in Anopheles albimanus

Transposable elements may eventually be used as transformation vectors of anti-Plasmodium genes in natural populations of Anopheles mosquitoes. The mariner transposable element is widespread in most animal phyla and we wished to test for its presence in the important Latin American malaria vector, Anopheles albimanus. Degenerate primers were designed from the consensus of insect mariner elements and used in a polymerase chain reaction to amplify a fragment of the predicted size from An. albimanus. The fragment was cloned, sequenced, and determined to be a mariner-like element through multiple alignment with known insect mariner elements. Dot blot and Southern blot analyses showed only one or a few elements per haploid genome. The cloned fragment was used as a probe to isolate similar sequences from an An. albimanus genomic DNA library. A 659-base-pair clone was 57% similar at the DNA level with mariner elements from Anopheles gambiae, Chrysoperla ptorapunda, and Haematobia irritans. This low sequence similarity is comparable to that found among mariner-like elements in all insects. However, conserved amino acid motifs were not discovered nor was a single open reading frame found. These aspects suggest that the An. albimanus mariner-like element may represent an ancient transposition event but that the element is no longer active, the typical open reading frame having been disrupted through nucleotide insertions and deletions.     

Microplate assay analysis of reduced fenitrothion susceptibility in Haitian Anopheles albimanus

Reduced fenitrothion susceptibility in Haitian Anopheles albimanus is documented using time/mortality measurements from otherwise standard World Health Organization (WHO) bioassays. Survival beyond a time threshold in bioassays is shown to be highly correlated with elevated non-specific esterase levels. A shift in resistance incidence from less than 20 to over 60% in a six-month period is documented using both the bioassay and microassay procedures, showing the potential of microplate assay methods in early detection of resistance. Conventional 24-hour reading of WHO bioassay data failed to detect resistance until the level reached 60%. Resistance appeared to be focal, with an increase in intensity coinciding with a fenitrothion spray cycle for malaria control.     

Overnight access to sugar and response to DDT in Anopheles albimanus Wied

A study was made of the response of A. albimanus females to DDT after overnight access to sugar solutions. Tarsal contact with the solutions had a statistically significant protective effect proportional to the strength of the solution. Prior access to sugar can produce considerable experimental error and thus invalidate DDT susceptibility tests. Reports of DDT resistance among anophelines should be treated with caution when the tests were conducted on insects that had access to sugar.     

Field evaluation of methods for estimating carbamate resistance in Anopheles albimanus mosquitos from a microplate assay for insensitive acetylcholinesterase

The reliability of a published method to predict survivorship in the WHO propoxur-resistance bioassay (WHO test) from the results of a biochemical assay for detecting the insensitivity of acetylcholinesterase (AChE) is described. For biochemical assay data from three field populations of Anopheles albimanus mosquitos, the results obtained using the method did not correlate consistently with the findings of the WHO test. A modified method is then described that eliminates the effect on the assay of factors unrelated to pesticide resistance, and it is shown that this modification can be used to predict survivorship in the WHO test for mosquitos from three study sites in Guatemala. The results show that when scored visually, the insensitive AChE microplate assay is an accurate method of estimating survival in the WHO test, regardless of whether the mosquitos tested are blood-fed or not. Recommendations are given for the application and analysis of data from the insensitive AChE microplate assay for detecting and monitoring resistance to carbamate insecticides.