Search

Background: It is important to understand whether the potential impact of pyrethroid resistance on malaria control can be mitigated by switching between different pyrethroids or whether cross-resistance within this insecticide class precludes this approach. Methods: Here we assess the relationships among pyrethroids in terms of their binding affinity to, and depletion by, key cytochrome P450 enzymes (hereafter P450s) that are known to confer metabolic pyrethroid resistance in Anopheles gambiae (s.l.) and An. funestus, in order to identify which pyrethroids may diverge from the others in their vulnerability to resistance. We then investigate whether these same pyrethroids also diverge from the others in terms of resistance in vector populations. Results: We found that the type I and II pyrethroids permethrin and deltamethrin, respectively, are closely related in terms of binding affinity to key P450s, depletion by P450s and resistance within vector populations. Bifenthrin, which lacks the common structural moiety of most pyrethroids, diverged from the other pyrethroids tested in terms of both binding affinity to key P450s and depletion by P450s, but resistance to bifenthrin has rarely been tested in vector populations and was not analysed here. Etofenprox, which also lacks the common structural moiety of most pyrethroids, diverged from the more commonly deployed pyrethroids in terms of binding affinity to key P450s and resistance in vector populations, but did not diverge from these pyrethroids in terms of depletion by the P450s. The analysis of depletion by the P450s indicated that etofenprox may be more vulnerable to metabolic resistance mechanisms in vector populations. In addition, greater resistance to etofenprox was found across Aedes aegypti populations, but greater resistance to this compound was not found in any of the malaria vector species analysed. The results for pyrethroid depletion by anopheline P450s in the laboratory were largely not repeated in the findings for resistance in malaria vector populations. Conclusion: Importantly, the prevalence of resistance to the pyrethroids α-cypermethrin, cyfluthrin, deltamethrin, λ-cyhalothrin and permethrin was correlated across malaria vector populations, and switching between these compounds as a tool to mitigate against pyrethroid resistance is not advised without strong evidence supporting a true difference in resistance. [ABSTRACT FROM AUTHOR]

Background: It is important to understand whether the potential impact of pyrethroid resistance on malaria control can be mitigated by switching between different pyrethroids or whether cross-resistance within this insecticide class precludes this approach. Methods: Here we assess the relationships among pyrethroids in terms of their binding affinity to, and depletion by, key cytochrome P450 enzymes (hereafter P450s) that are known to confer metabolic pyrethroid resistance in Anopheles gambiae (s.l.) and An. funestus, in order to identify which pyrethroids may diverge from the others in their vulnerability to resistance. We then investigate whether these same pyrethroids also diverge from the others in terms of resistance in vector populations. Results: We found that the type I and II pyrethroids permethrin and deltamethrin, respectively, are closely related in terms of binding affinity to key P450s, depletion by P450s and resistance within vector populations. Bifenthrin, which lacks the common structural moiety of most pyrethroids, diverged from the other pyrethroids tested in terms of both binding affinity to key P450s and depletion by P450s, but resistance to bifenthrin has rarely been tested in vector populations and was not analysed here. Etofenprox, which also lacks the common structural moiety of most pyrethroids, diverged from the more commonly deployed pyrethroids in terms of binding affinity to key P450s and resistance in vector populations, but did not diverge from these pyrethroids in terms of depletion by the P450s. The analysis of depletion by the P450s indicated that etofenprox may be more vulnerable to metabolic resistance mechanisms in vector populations. In addition, greater resistance to etofenprox was found across Aedes aegypti populations, but greater resistance to this compound was not found in any of the malaria vector species analysed. The results for pyrethroid depletion by anopheline P450s in the laboratory were largely not repeated in the findings for resistance in malaria vector populations. Conclusion: Importantly, the prevalence of resistance to the pyrethroids α-cypermethrin, cyfluthrin, deltamethrin, λ-cyhalothrin and permethrin was correlated across malaria vector populations, and switching between these compounds as a tool to mitigate against pyrethroid resistance is not advised without strong evidence supporting a true difference in resistance. [ABSTRACT FROM AUTHOR]

Insecticide resistance is a major impediment to the control of vectors and pests of public health importance and is a strongly selected trait capable of rapid spread, sometimes even between closely related species. Elucidating the mechanisms generating insecticide resistance in mosquito vectors of disease, and understanding the spread of resistance within and between populations and species are vital for the development of robust resistance management strategies. Here, we studied the mechanisms of resistance in two sympatric members of the Anopheles gambiae species complex-the major vector of malaria in sub-Saharan Africa-to understand how resistance has developed and spread in eastern Uganda, a region with some of the highest levels of malaria. In eastern Uganda, where the mosquitoes Anopheles arabiensis and An. gambiae can be found sympatrically, low levels of hybrids (0.4 %) occur, offering a route for introgression of adaptively important variants between species. In independent microarray studies of insecticide resistance, Gste4, an insect-specific glutathione S-transferase, was among the most significantly up-regulated genes in both species. To test the hypothesis of interspecific introgression, we sequenced 2.3 kbp encompassing Gste4. Whilst this detailed sequencing ruled out introgression, we detected strong positive selection acting on Gste4. However, these sequences, followed by haplotype-specific qPCR, showed that the apparent up-regulation in An. arabiensis is a result of allelic variation across the microarray probe binding sites which artefactually elevates the gene expression signal. Thus, face-value acceptance of microarray data can be misleading and it is advisable to conduct a more detailed investigation of the causes and nature of such signal. The identification of positive selection acting on this locus led us to functionally express and characterise allelic variants of GSTE4. Although the in vitro data do not support a direct role for GSTE4 in metabolism, they do support a role for this enzyme in insecticide sequestration. Thus, the demonstration of a role for an up-regulated gene in metabolic resistance to insecticides should not be limited to simply whether it can metabolise insecticide; such a strict criterion would argue against the involvement of GSTE4 despite the weight of evidence to the contrary. [ABSTRACT FROM AUTHOR]

Insecticide resistance in the malaria vector Anopheles gambiae s.l. (Diptera: Culicidae) threatens insecticide-based control efforts, necessitating regular monitoring. We assessed resistance in field-collected An. gambiae s.l. from Jinja, Uganda using World Health Organization (WHO) biosassays. Only An. gambiae s.s. and An. arabiensis (≅70%) were present. Female An. gambiae exhibited extremely high pyrethroid resistance (permethrin LT50 > 2 h; deltamethrin LT50 > 5 h). Female An. arabiensis were resistant to permethrin and exhibited reduced susceptibility to deltamethrin. However, while An. gambiae were DDT resistant, An. arabiensis were fully susceptible. Both species were fully susceptible to bendiocarb and fenitrothion. Kdr 1014S has increased rapidly in the Jinja population of An. gambiae s.s. and now approaches fixation (≅95%), consistent with insecticide-mediated selection, but is currently at a low frequency in An. arabiensis (0.07%). Kdr 1014F was also at a low frequency in An. gambiae. These frequencies preclude adequately-powered tests for an association with phenotypic resistance. PBO synergist bioassays resulted in near complete recovery of pyrethroid susceptibility suggesting involvement of CYP450s in resistance. A small number (0.22%) of An. gambiae s.s. × An. arabiensis hybrids were found, suggesting the possibility of introgression of resistance alleles between species. The high levels of pyrethroid resistance encountered in Jinja threaten to reduce the efficacy of vector control programmes which rely on pyrethroid-impregnated bednets or indoor spraying of pyrethroids. [ABSTRACT FROM AUTHOR]

The primary Afrotropical malaria mosquito vector Anopheles gambiae sensu stricto has a complex population structure. In west Africa, this species is split into two molecular forms and displays local and regional variation in chromosomal arrangements and behaviors. To investigate patterns of macrogeographic population substructure, 25 An. gambiae samples from 12 African countries were genotyped at 13 microsatellite loci. This analysis detected the presence of additional population structuring, with the M-form being subdivided into distinct west, central, and southern African genetic clusters. These clusters are coincident with the central African rainforest belt and northern and southern savannah biomes, which suggests restrictions to gene flow associated with the transition between these biomes. By contrast, geographically patterned population substructure appears much weaker within the S-form. [ABSTRACT FROM AUTHOR]

Malaria control depends on mosquito susceptibility to insecticides. We tested Anopheles gambiae mosquitoes from Côte d'Ivoire for resistance and screened a subset for target site mutations. Mosquitoes were resistant to insecticides of all approved classes. Such complete resistance, which includes exceptionally strong phenotypes, presents a major threat to malaria control. [ABSTRACT FROM AUTHOR]

Tsetse flies of the palpalis group are major vectors of Human African Trypanosomiasis in Africa. Accurate knowledge of species identity is essential for vector control. Here, we combine ribosomal internal transcribed spacer 1 ( ITS1), mitochondrial Cytochrome Oxidase 1 ( COI) and microsatellites to determine the population structure and phylogenetic relations of Glossina p. palpalis in Equatorial Guinea. CO1 sequence data suggest that G. p. palpalis in Equatorial Guinea is a distinct subspecies from previously described G. p. palpalis in West Africa and Democratic Republic of Congo. Glossina p. palpalis in Equatorial Guinea and DRC share a common ancestor which diverged from West African G. p. palpalis around 1.9 Ma. Previous ITS1 length polymorphism data suggested the possible presence of hybrids in Equatorial Guinea. However, ITS1 showed incomplete lineage sorting compared with clearly defined COI groups, and data from 12 unlinked microsatellites provided no evidence of hybridization. Microsatellite data indicated moderate but significant differentiation between the populations analysed (Rio Campo, Mbini and Kogo). Moreover, unlike previous studies of G. p. palpalis, there was no evidence for heterozygote deficiency, presence of migrants or cryptic population structure. Variance effective population size at Rio Campo was estimated at 501–731 assuming eight generations per year. This study of the population genetics of G. p. palpalis in central Africa provides the first estimate of genetic differentiation between geographically separated G. p. palpalis populations. [ABSTRACT FROM AUTHOR]

The coastal shrimp Crangon crangon is an ecologically and commercially important species but there is limited knowledge of its genetic population structure. We utilised amplified fragment length polymorphisms (AFLPs) to investigate population differentiation among eight sampling locations comprising paired sites from north- and south-western Britain, the eastern English Channel and the Baltic Sea. Initial AMOVA and cluster analysis suggested strong differentiation, but outlier analysis identified three loci that might be subject to selection, one of which showed significant latitudinal variation in allele frequencies. Following exclusion of these outlier loci, and also of a divergent, genetically-impoverished sample from the UK Bristol Channel, cluster analysis revealed three major groupings, corresponding to geographical regions: western Britain, the eastern English Channel and the Baltic Sea. AMOVA identified significant differentiation both within and among these regions, with similar variation explained by each hierarchical level. C. crangon shows greater genetic structuring than has been found in many decapod crustaceans studied to date, and our results are consistent with unstable population sizes and gene flow restricted by distance and probably also hydrographic features. Further investigation of temporal stability in population structure is required, but broad-scale homogeneity of fishery stocks should not be assumed. [ABSTRACT FROM AUTHOR]

Negative correlations between environmental temperature and body size are widespread in planktonic organisms, and ectotherms generally, but remain poorly understood. Here we evaluate experimentally two alternative hypotheses suggested to explain life history shifts induced by raised temperature using parthenogenetic clones from two Daphnia species. Explanation 1 proposes that the life history shifts could be adaptive if increased temperature is used as an indirect cue to indicate increased risk from size-selective predators. Explanation 2 proposes that at larger body size energy becomes more limiting as temperature increases because of a less favourable assimilation: metabolism balance. In a factorial laboratory experiment we examine the effects of three rearing temperatures on the growth and reproductive traits of Daphnia raised in water with fish kairomone, Chaoborus kairomone, or in uncontaminated water. None of the three predictions of explanation 1 were met by the data. In both D. pulex and D. curvirostris, and some other published studies, data suggested that at larger body sizes the sum of growth and reproduction was lower at high temperature, supporting our prediction from explanation 2. However, we propose a novel third explanation based on new evidence of temperature-dependence in both reproductive effort and cost in D. pulex. [ABSTRACT FROM AUTHOR]

Examines the antipredator reaction norms of Daphnia pulex. Effects between temperature and food level on the vulnerability of Daphnia to predation; Response of Daphnia to the concentration of fish kairomone; Vulnerability of Daphnia to fish predation.

Nitric oxide a potent antimicrobial agent and vasodilator, may be synthesized in the human mouth due to the bacterial reduction of salivary nitrate. The purpose of this study was to determine whether intra-oral pH changes produced by the oral flora may augment the production of nitric oxide. To test this hypothesis a sucrose mouthrinse was administered to human volunteers who were free from caries and periodontal disease and production of oral nitric oxide was related to intra-oral pH changes. Baseline measurements consisted of pooled supragingival plaque and anterior and posterior tongue scrapings for microbiological analysis. Additional samples of saliva were collected for nitrate and nitrite analysis. Intra-oral pH measurements were recorded with a microelectrode at selected sites and NO intra-oral levels were measured by chemiluminescence. In addition selected members of the oral flora were tested for their ability to reduce nitrate to nitrite. A decrease in pH produced by the sucrose rinse was significantly associated with an increase in the intra-oral generation of NO after 5 min (p=0.006). The most frequently recovered nitrate reducing organisms were Veillonella spp. and Actinomyces spp. whilst the acidogenic Streptococcus spp. were negative nitrate reducers. The oral generation of NO appears to be influenced by pH levels within the oral cavity. [ABSTRACT FROM AUTHOR]

The aim of this in vitro study was to investigate the coronal leakage of obligate anaerobes into root canals obturated with lateral condensation of cold gutta-percha with two root canal sealers. Sixty extracted human teeth with straight, single root canals were prepared using the modified double-flared technique with balanced force under copious irrigation until the master apical file was size 40. The teeth were divided randomly into experimental groups (40 teeth) and control groups (20 teeth). In the experimental groups, 20 teeth were obturated with lateral condensation of cold gutta-percha and AH26 sealer and 20 teeth were obturated with the same technique using TubliSeal EWT sealer. In the control groups, 10 teeth were obturated with the same technique either with AH26 or TubliSeal EWT sealer. These teeth were completely sealed to serve as negative controls. The remaining 10 teeth were not obturated and served as positive controls. The root surface of each tooth was sealed with nail varnish except the apical 2 mm. The coronal part of each root canal was sealed with the cut end of polypropylene tube and placed in a glass bottle containing sterile Fastidious Anaerobe Broth (FAB). Aliquots of 0.5 mL of FAB were injected into the polypropylene tube and the model system was centrifuged at 168 g. An inoculum of Fusobacterium nucleatum in FAB was placed in each coronal chamber at 7-day intervals and daily observations were made for bacterial growth in the apical reservoir for 12 weeks. All positive control teeth showed bacterial leakage within a week, while the negative control teeth remained uncontaminated throughout the test period. All the experimental teeth exhibited leakage of bacterial metabolites within 12 weeks, ranging from 1 to 12 weeks. The mean time for complete leakage in the AH26 and the TubliSeal EWT groups was 8.4 and 8.2 weeks respectively. There was no statistically significant difference (P > 0.05) in leakage between the AH26 and the TubliSeal EWT groups. [ABSTRACT FROM AUTHOR]

Accurate estimates of DNA quantity are likely to become increasingly important for successful genomic screening of insect populations via recently developed, highly multiplexed genotyping assays and high-throughput sequencing methods. Here we show that genomic DNA extractions from single Anopheles gambiae Giles using a standard commercial kit-based methodology yield extracts with concentrations below the linear range of spectrophotometric absorbance at 260 nm. Concentrations determined by spectrophotometry were not reproducible, and are therefore neither accurate nor reliable. However, DNA quantification using a fluorescent nucleic acid stain (PicoGreen®) gave highly reproducible concentration estimates, and indicated that, on average, single mosquitoes yielded approximately 300 ng of DNA. Such a total yield is currently insufficient for many high-throughput genome screening applications, necessitating whole genome amplification of all or most individuals in a population prior to genotyping. [ABSTRACT FROM AUTHOR]

The Chinese mitten crab, Eriocheir sinensis is an important human food source in Asia and causes considerable ecological and economical damage as a recent invader of North America and Europe. Here we report the isolation and characterization of 12 highly polymorphic microsatellite loci for E. sinensis. The number of observed alleles per locus ranged from 7 to 40 and the observed heterozygosity from 0.50 to 0.95. These markers should prove a useful tool to investigate the colonization process and its evolutionary implications. [ABSTRACT FROM AUTHOR]

The freshwater prosobranch snail Potamopyrgus antipodarum is an important model system for studying the maintenance of sexual reproduction in its native New Zealand. Since introduction to the UK in the 1850s, the species has spread throughout Europe and recently the USA. We present the first microsatellites for P. antipodarum. Using two isolation protocols, which are qualitatively compared, we have developed PCR primers for three di- and four trinucleotide microsatellites. All loci proved to be polymorphic, when screened for variability in several UK populations. These markers should be of considerable utility in future population and ecological genetics studies of this species. [ABSTRACT FROM AUTHOR]

Nonamplified (null) alleles are a common feature of microsatellite genotyping and can bias estimates of allele and genotype frequencies, thereby hindering population genetic analyses. The frequency of microsatellite null alleles in diploid populations can be estimated for populations that are in Hardy–Weinberg equilibrium. However, many microsatellite data sets are from nonequilibrium populations, often with known inbreeding coefficients ( F) or fixation indices ( FIS or FST). Here, we propose a novel null allele estimator that can be used to estimate the null allele frequency and adjust visible allele frequencies in populations for which independent estimates of F, FIS or FST are available. The algorithm is currently available as an Excel macro that can be downloaded at no cost from and will be incorporated into the softwaremicro-checker. [ABSTRACT FROM AUTHOR]