Fatal association between dieldrin-resistant and susceptible Australian sheep blowflies, Lucilia cuprina.

A novel phenomenon of interactions between genotypes of the dieldrin-resistance (Rdl) locus of the Australian sheep blowfly (Lucilia cuprina) is described. Susceptible adult flies exposed to dieldrin-resistant (Rdl/Rdl or Rdl/S) adults, raised from larvae grown on media containing sublethal concentrations of dieldrin, display mortality related to the concentration on which the resistant flies developed. The resistant flies excrete quantities of dieldrin that are toxic to susceptible flies. These observations provide an additional mechanism to those previously identified for the rapid evolution of resistance to dieldrin by L. cuprina.     

Genetic architecture and adaptation: quantitative analysis of sheep and refuse tip populations of the Australian sheep blowfly, Lucilia cuprina

Phenotypic differentiation between geographic areas and between sheep and adjacent refuse tip populations was assessed by quantitative analysis of population samples of L. cuprina from New South Wales (Lismore) and Victoria (Mansfield). In addition the genetic structure of populations has been defined and compared by biometrical analysis techniques. For all morphological and fitness characters examined significant phenotypic differentiation was observed both between geographic localities and between sheep and non-sheep populations of each locality. Diallel analysis of the populations revealed architectural differences between sheep and non-sheep populations for both fecundity and egg hatchability. Sheep populations only, regardless of locality, displayed dominant gene effects on these fitness traits. The results suggest that refuse tip populations may be other than transients and that the differentiation may reflect differing patterns of adaptation and history of selection of the populations. The relevance of such differentiation to the successful establishment of a chemical and/or autocidal control zone is considered.     

Selection of dieldrin-resistant strains of Lucilia cuprina (Diptera: Calliphoridae) after ethyl methanesulfonate mutagenesis of a susceptible strain.

After ethyl methanesulfonate (EMS) mutagenesis of a susceptible strain (SWT), selective screening of Lucilia cuprina (Wiedemann) resulted in four strains that were resistant to the insecticide dieldrin. Concentrations used for selection were greater than LC99 of susceptible phenotypes. No resistant variants were screened from the standard laboratory strain (SWT) not treated with EMS. The resistance phenotypes of the four resistant strains were similar to each other and to that of a field-selected resistant strain. The genetic basis of resistance is monogenic in all strains and the data are consistent with the same locus, Rdl, determining resistance status in each strain. The Rdl locus maps to chromosome V, approximately 3.5 map units distal to the Sut locus. Dieldrin resistance may be caused by less effective blocking of insect neuronal GABA receptors by the chemical in resistant strains. The data indicate that the evolution of resistance to an insecticide in the field may be constrained by a limited number of genetical and biochemical options if a monogenic response is selected for and that the spontaneous mutation rate to the Rdl allele is less than 1 in 10(6) in the laboratory.     

Predicting insecticide resistance: mutagenesis, selection and response

Strategies to manage resistance to a particular insecticide have usually been devised after resistance has evolved. If it were possible to predict likely resistance mechanisms to novel insecticides before they evolved in the field, it might be feasible to have programmes that manage susceptibility. With this approach in mind, single-gene variants of the Australian sheep blowfly, Lucilia cuprina, resistant to dieldrin, diazinon and malathion, were selected in the laboratory after mutagenesis of susceptible strains. The genetic and molecular bases of resistance in these variants were identical to those that had previously evolved in natural populations. Given this predictive capacity for known resistances, the approach was extended to anticipate possible mechanisms of resistance to cyromazine, an insecticide to which L. cuprina populations remain susceptible after almost 20 years of exposure. Analysis of the laboratory-generated resistant variants provides an explanation for this observation. The variants show low levels of resistance and a selective advantage over susceptibles for only a limited concentration range. These results are discussed in the context of the choice of insecticides for control purposes and of delivery strategies to minimize the evolution of resistance.     

Diazinon resistance, fluctuating asymmetry and fitness in the Australian sheep blowfly, lucilia cuprina

Genetic evidence suggests that the evolution of resistance to the insecticide diazinon in Lucilia cuprina initially produced an increase in asymmetry. At that time resistant flies were presumed to be at a selective disadvantage in the absence of diazinon. Subsequent evolution in natural populations selected modifiers to ameliorate these effects. The fitness and fluctuating asymmetry levels of resistant flies are currently similar to those of susceptibles. Previous genetic analyses have shown the fitness modifier to co-segregate with the region of chromosome III marked by the white eyes, w, locus, unlinked to the diazinon resistance locus, Rop-1, on chromosome IV. This study maps the asymmetry modifier to the same region, shows, as in the case of the fitness modifier, its effect to be dominant and presents data consistent with the fitness/asymmetry modifier being the same gene (gene complex). These results suggest changes in fluctuating asymmetry reflect changes in fitness.     

Suppression of populations of Australian sheep blowfly, Lucilia cuprina (Wiedemann) (Diptera: Calliphoridae), with a novel blowfly trap

The Australian sheep blowfly, Lucilia cuprina , initiates more than 85% of fly strikes on sheep in Australia with an estimated average annual cost of A$280 million to the Australian sheep industry. LuciTrap^® is a commercially available, selective trap for L. cuprina consisting of a plastic bucket with multiple fly entry cones and a synthetic attractant. The impact of LuciTrap on populations of L. cuprina on sheep properties in five Australian states was evaluated by comparing L. cuprina populations on paired properties with and without LuciTraps over seasons when significant fly populations could be expected. Twenty-four comparisons (trials) were conducted over 4 years. During times of 'higher fly density' (when the 48 h geometric mean of trap catches on the control property was greater than five L. cuprina ), the overall geometric mean trap catches for control and trapped properties differed significantly ( P  < 0.001) with mean trap catches of 19.4 and 7.74 L. cuprina , respectively. The selectivity of the LuciTrap was confirmed with 59% of all trapped flies being L. cuprina . Chrysomya spp. and Calliphora spp. constituted 9.3% and 1.1% of the catches with a variety of other flies (mainly Sarcophagidae and Muscidae ) providing the remainder (31%). Lucilia sericata was only trapped in Tasmania and made up 7.7% of the Lucilia spp. catch in that state. Seventy-two per cent of the trapped L. cuprina were female. The deployment of LuciTrap on sheep properties at one trap per 100 sheep from the beginning of the anticipated fly season suppressed the populations of L. cuprina by 60% compared with matched control properties. The LuciTrap is a selective and easy to use fly trap and constitutes an effective, non-insecticidal tool for use in integrated management programs for L. cuprina .     

Monitoring and selection of resistance to pyrethroids in the Australian sheep blowfly, Lucilia cuprina

Field and laboratory populations of the Australian sheep blowfly, Lucilia cuprina (Wiedemann) (Calliphoridae), were surveyed by bioassay for possible resistance to the synthetic pyrethroids, a group of insecticides under development for blowfly control. A normal distribution of LC50 values was found using deltamethrin, the test pyrethroid, with no indication of specific resistance despite widespread use of deltamethrin on sheep to control the sheep body louse, Damalinia ovis (Schrank) (Trichodectidae). There was no cross-resistance to deltamethrin from existing organophosphate (OP) resistance nor from previous use of DDT. Selection with deltamethrin on a combined field strain, CSF85, increased the LC50 gradually over the first twenty generations until it stabilized at approximately 25x that of the unselected CSF85. This laboratory-induced resistance extended to other pyrethroids, cypermethrin (16x), cyhalothrin (25x) and cycloprothrin (10x), and increased the existing resistance of CFS85 to the OP diazinon (11x) and the carbamate, butacarb (83x).     

Stress and asymmetry during arrested development of the Australian sheep blowfly.

The dieldrin and diazinon resistance systems of the Australian sheep blowfly (Lucilia cuprina) have been used previously to relate stress, departures from bilateral symmetry, developmental stability and relative fitness. These systems are now used to consider stress and asymmetry in a developmental context. Larval to adult development is shown to be significantly impaired after arrested development at 8 degrees C, however the asymmetry score of adults of a given genotype is similar after arrested or continuous development. Selection against dieldrin-resistant and unmodified diazinon-resistant genotypes occurs during arrested development because greater proportions of these genotypes pupae at 8 degrees C than do susceptible or modified diazinon-resistant genotypes. Pre-pupae of all genotypes complete development equally successfully when transferred from 8 degrees C to 27 degrees C. Adults fail to emerge when pupae formed at 8 degrees C undergo this temperature transition. Temperature-shift experiments show the asymmetry score is determined between pre-pupal and pupal stages of the life cycle. This stage occurs at 27 degrees C in arrested and continuously developing cultures providing an explanation for the independence of stress, selective mortality during developmental arrest and asymmetry score. The results emphasize the need for genetic, environmental and developmental data before an asymmetry phenotype can be directly related to developmental stability and relative fitness.     

Cross-resistance between dieldrin and fipronil in German cockroach (Dictyoptera: Blattellidae)

The toxicity of fipronil and dieldrin was determined in one susceptible laboratory strain and seven insecticide-resistant field-collected strains of Blattella germanica (L). The Zo960302 and Ga021001 strains were 1,270- and 2,030-fold resistant to dieldrin and 15- and 14-fold resistant to fipronil. The Su960304 and Od010803 strains were 15- and 13-fold resistant to dieldrin and two- and four-fold resistant to fipronil. Three strains showed no or a low level of resistance to dieldrin and fipronil. Crosses were performed between the susceptible strain Danish Pest Infestation Laboratory (DPIL)-SUS and the resistant strains Zo960302 and Su960304 and resistance to dieldrin and fipronil were intermediate compared with the susceptible and the resistant strains. Backcrosses to both of the parental strains showed cosegregation of dieldrin and fipronil resistance. The toxicity of dieldrin and fipronil was correlated when compared at LD50, and 93% of the observed variation in LD50 of fipronil can be ascribed to variation among predictions based on the value of LD50 of dieldrin. The frequency of the A302S substitution in the resistance to dieldrin (Rdl) gene in the highly dieldrin- and fipronil-resistant strains Zo960302 and Ga021001 and the moderately resistant Su960304 was 0.97, 1.0, and 0.38, respectively. We consider the connection between the frequency of the Rdl mutation and dieldrin and fipronil resistance a causal connection and not merely a coincidence.     

Prion gene sequence variation within diverse groups of U.S. sheep,beef cattle,and deer

Prions are proteins that play a central role in transmissible spongiform encephalopathies in a variety of mammals. Among the most notable prion disorders in ungulates are scrapie in sheep, bovine spongiform encephalopathy in cattle, and chronic wasting disease in deer. Single nucleotide polymorphisms in the sheep prion gene (PRNP) have been correlated with susceptibility to natural scrapie in some populations. Similar correlations have not been reported in cattle or deer; however, characterization of PRNP nucleotide diversity in those species is incomplete. This report describes nucleotide sequence variation and frequency estimates for the PRNP locus within diverse groups of U.S. sheep, U.S. beef cattle, and free-ranging deer (Odocoileus virginianus and O. hemionus from Wyoming). DNA segments corresponding to the complete prion coding sequence and a 596-bp portion of the PRNP promoter region were amplified and sequenced from DNA panels with 90 sheep, 96 cattle, and 94 deer. Each panel was designed to contain the most diverse germplasm available from their respective populations to facilitate polymorphism detection. Sequence comparisons identified a total of 86 polymorphisms. Previously unreported polymorphisms were identified in sheep (9), cattle (13), and deer (32). The number of individuals sampled within each population was sufficient to detect more than 95% of all alleles present at a frequency greater than 0.02. The estimation of PRNP allele and genotype frequencies within these diverse groups of sheep, cattle, and deer provides a framework for designing accurate genotype assays for use in genetic epidemiology, allele management, and disease control.     

Enzymes and Reproduction in Natural Populations of DROSOPHILA EURONOTUS

Populations of Drosophila euronotus, one from southern Louisiana )3 samples), and one from Missouri (2 samples), were classified for allele frequencies at alkaline phosphatase (APH) and acid phosphatase (ACPH) loci. The two populations differed consistently in allele frequencies at both loci. The APH locus is on the inversion-free X chromosome; the chromosomal locus of the autosomal ACPH is unknown, and could involve inversion polymorphism. Wild females from Missouri and Louisiana populations heterozygous at the APH locus carried more sperm at capture than did the corresponding homozygotes. This heterotic association was significant for the combined samples, and whether it was the result of heterosis at the enzyme locus studied, or due to geographically widespread close linkage with other heterotic loci, it should help to maintain heterozygosity at the APH locus. In a Louisiana collection which included large numbers of sperm-free females, simultaneous homozygosity at both enzyme loci was significantly associated with lack of sperm. It is suggested that the latter association is the result of young heterozygous females achieving sexual maturity earlier than do the double homozygotes. The average effective sperm load for 225 wild females was only 29.4, suggesting the necessity for frequent repeat-mating in nature to maintain female fertility. A comparison of the sex-linked APH genotypes of wild females with those of their daughters indicated that among 295 wild-inseminated females from five populations, 35% had mated more than once, and of this 35%, six females had mated at least three times. Because of ascertainment difficulties, it is clear that the true frequency of multiple-mating in nature must have been much higher than the observed 35%. Laboratory studies indicate that multiple-mating in this species does not involve sperm displacement, possibly due to the small number of sperms transmitted per mating, and the fact that the sperm receptacles are only partially filled by a given mating.     

Resistance to two organophosphorus insecticides in New Zealand populations of the Australian sheep blowfly, Lucilia cuprina

Abstract. A treated-surface test was used in a bioassay to detect insecticide resistance to the organophosphorus compounds diazinon and propetamphos, in larvae of field strains of Lucilia cuprina in New Zealand during 1990/91 and 1991/92 (December-April). Mean LC₅₀s for both diazinon and propetamphos showed a significant increase between seasons although this is not necessarily indicative of a trend. Mean LC₅₀s for diazinon increased from 1.16 to 1.60 mg/1 (P< 0.001) and for propetamphos from 0.49 to 0.59 mg/1 (P< 0.005) in pooled populations. Diazinon and propetamphos resistance was significantly correlated in both seasons, suggesting side-resistance between the two insecticides.     

Thermal kinetic differences between allelic isozymes of malate dehydrogenase (Mdh-B locus) of largemouth bass,Micropterus salmoides

Two alleles are encoded at the malate dehydrogenase locus in largemouth bass, Micropterus salmoides. Populations in the extreme northern areas of the range of this fish are fixed or nearly fixed for the B1 allele, whereas populations in Florida are fixed for the alternative allele, B2. The MDH-B1B1 and MDH-B2B2 allelic isozymes were isolated by preparative starch gel electrophoresis and subjected to in vitro kinetic analyses. The apparent Km (oxaloacetate) for each of these allelic isozymes was determined at 25, 30, and 35 degrees C. The Km values for both isozymes increased with increasing temperature and were not significantly different from each other at 25 and 35 degrees C. However, at 30 degrees C the Km value for the MDH-B1B1 allelic isozyme was higher than that for the MDH-B2B2 isozyme (i.e., 5.4 X 10(-5) vs 3.3 X 10(-5)). These results are consistent with the hypothesis that the different environmental temperatures at different latitudes may be at least partially responsible for the north-south cline in Mdh-B allele frequencies.     

Population structure and Mdh-1 locus variation in Apis mellifera ligustica

In a wide area of the Piedmont of Italy the apiaries of Apis mellifera ligustica Spin., (the Italian bee) show homogeneous allelic frequency distributions at the Mdh-1 locus, the only one known to be polymorphic in worker bees. This can be explained by considering that an apiary is not a closed genetic system and that among apiaries gene flow is sufficient to overcome the different forces of inbreeding and random genetic drift. Nevertheless there is some evidence for partial subdivision because the pooled samples show a weak Wahlund effect. Moreover, the M allele at the same locus can be used as a diagnostic marker to distinguish A. m. ligustica populations (M absent or at very low frequencies) from A. m. mellifera French populations (monomorphic for M). The two honey-bee varieties, almost entirely separated by the Alps, hybridize with each other in very limited alpine areas. Hybrid populations show intermediate M frequencies.     

A General Model of Negative Frequency Dependent Selection Explains Global Patterns of Human ABO Polymorphism

The ABO locus in humans is characterized by elevated heterozygosity and very similar allele frequencies among populations scattered across the globe. Using knowledge of ABO protein function, we generated a simple model of asymmetric negative frequency dependent selection and genetic drift to explain the maintenance of ABO polymorphism and its loss in human populations. In our models, regardless of the strength of selection, models with large effective population sizes result in ABO allele frequencies that closely match those observed in most continental populations. Populations must be moderately small to fall out of equilibrium and lose either the A or B allele (N_e ≤ 50) and much smaller (N_e ≤ 25) for the complete loss of diversity, which nearly always involved the fixation of the O allele. A pattern of low heterozygosity at the ABO locus where loss of polymorphism occurs in our model is consistent with small populations, such as Native American populations. This study provides a general evolutionary model to explain the observed global patterns of polymorphism at the ABO locus and the pattern of allele loss in small populations. Moreover, these results inform the range of population sizes associated with the recent human colonization of the Americas.     

Base-line susceptibility of the oriental latrine fly, Chrysomya megacephala (Diptera: Calliphoridae), to five insecticides

Susceptibility to five insecticides was assessed on seven colonies of the oriental latrine fly, Chrysomya megacephala (Fabricius), originating from Japan, Hong Kong, Philippines, Indonesia and Papua New Guinea, and on C. saffranèa (Bigot) from PNG. All the colonies were susceptible to fenitrothion, diazinon, dichlorvos, permethrin and gamma-HCH, although the Jakarta, Manila and Tokyo colonies of C.megacephala were least susceptible (most tolerant) to diazinon, dichlorvos and permethrin. Feral forms of both Chrysomya species were 4-5-fold more susceptible to permethrin at the LD50 level than were all six colonies of the synanthropic form of C.megacephala.     

The toxicity of some soil insecticides to carabid predators of the cabbage root fly (Erioischia brassicae (Bouché))

In laboratory tests concentrations of dieldrin in soil, below that required to control cabbage root fly, were found to be toxic to several species of carabid beetles which prey on the immature stages of this pest. These results confirm previous findings from field experiments that dieldrin residues in soil can, by reducing the populations of predaceous beetles, increase the survival of the cabbage root fly and the subsequent crop damage caused. The toxicity of selected organophosphorus insecticides to these beneficial insects was studied in soils of different moisture content. Thionazin was found to be highly toxic at all moisture levels as was diazinon in moist soil. Chlorfenvinphos and azinphos-methyl and, in dry soil diazinon, were very much less toxic.     

Mitochondrial iso-citrate dehydrogenase variation in the Australian bush-rat, Rattus fuscipes greyii

Four reproductively isolated populations of the Australian bush-rat, Rattus fuscipes greyii , are polymorphic for electrophoretic variants of the mitochondrial enzyme, NADP-dependent iso-citrate dehydrogenase (M-Idh). The electrophoretic patterns of M-Idh and a small amount of breeding data are in agreement with the hypothesis that the variation is controlled by two alleles at an autosomal locus, Idh-2. In three relatively large populations, the proportion of heterozygotes at the Idh-2 locus ranges from 0.07 to 0.23, while on a small off-shore island it is 0.60. In this latter population there is an excess of heterozygotes which is on the borderline of statistical significance, suggesting that heterotic selection may be maintaining the polymorphism. Populations on eight other small islands are monomorphic for the allele which is the most frequent in the large populations. In most species M-Idh is less variable than most other enzymic proteins and the results presented here constitute one of the few published examples of an extensive M-Idh polymorphism.     

Single-strand conformational polymorphism and sequence polymorphism of Mhc-DRB in Latxa and Karrantzar sheep: implications for Caprinae phylogeny

Single-strand conformational polymorphism analysis and DNA sequencing were used to characterize Mhc-DRB second exon variability in the Latxa and Karrantzar breeds of sheep. The presence of more than two sequences in some animals indicates that alleles of two different loci have been amplified. Six new alleles were identified by sequencing. The allele frequency distribution of the DRB1 gene is striking, with two alleles accounting for half of the gene pool in both breeds under study. The most frequent allele in both breeds was the same (named DRB1*0702), with some specific amino acids: Tyr in position 31 and Thr in 51. A species variability analysis was also performed including the entire set of sheep DRB exon 2 sequences. Based on the patchwork patterns of different alleles, interallelic recombination appears to be playing a significant role in the generation of allelic diversity at this locus in sheep. The phylogenetic tree of all known Caprinae DRB sequences shows that certain alleles from one species are more closely related to those from other species than they are to each other. Allele DRB1*0702 merits special attention due to its high similarity to the Mufflon allele. As this is the most frequent in both breeds analyzed, one can hypothesize that in sheep, both Mufflon and Argali have had different influences depending on the sheep breed under study and that the relationship between domestic sheep and Mufflon is greater than previously thought. The data generated in this study can serve as a basis for developing a typing assay for the sheep DRB genes in the Latxa and Karrantzar populations.