SHORT-TERM IMPACTS OF FORMULATIONS OF BACILLUS THURINGIENSIS VAR. ISRAELENSIS DE BARJAC AND THE ORGANOPHOSPHATE TEMEPHOS,USED IN BLACKFLY (DIPTERA: SIMULIIDAE) CONTROL,ON RHEOPHILIC BENTHIC MACROINVERTEBRATES IN THE MIDDLE ORANGE RIVER,SOUTH AFRICA

Summary

The impacts of larvicides used in the control of blackflies (Diptera: Simuliidae) on macroinvertebrates in the stones-in-current biotope were assessed during 8 field trials in the middle Orange River, South Africa. Two Bacillus thuringiensis var. israelensis (B.t.i.) products (Vectobac^R 12AS and Teknar^R HP-D) and the organophosphate temephos (Abate^R 200EC) were applied at recommended and high dosages to simulate “operational” and “worst-possible” scenario's respectively. Mortality was evaluated either by direct counting of invertebrates on stones before and after application, or by ranking invertebrates on a 4-point relative abundance scale before and after application. In addition, the re-appearance of benthic invertebrate population densities after temephos application was examined.

At the recommended dosage (1.2 ppm/10 min), B.t.i. significantly reduced blackfly larval numbers (P<0.001) and those of the chironomid Rheotanytarsus fuscus Freeman (P<0.05). At high dosage (20 ppm/10 min), numbers of the filter-feeding mayfly Tricorythus discolor (Burmeister) (P<0.01) and the chironomid Cardiocladius sp. (P<0.05) were also significantly reduced. No Simulium predators were directly affected by B.t.i., but there were indications of food shortage amongst Hydropsychidae and Hirudinea.

Temephos caused significant reductions in the relative abundance of 5 taxa at 0.05 ppm, 3 to 6 taxa at 0.1 ppm, and 9 taxa at 1.0 ppm (P<0.05). “Non-target” organisms which were most affected included the chironomid R. fuscus, the mayflies Baetis glaucus Agnew and Choroterpes elegans Barnard, and the caddisflies Cheumatopsyche thomasseti Ulmer and Amphipsyche scottae Kimmins. The mayfly T. discolor was tolerant of temephos, even at high dosage (1.0 ppm/10 min). In winter, most taxa re-appeared within 19 days, and population densities were back to pre-treatment levels within 35 days.

It is concluded that good reduction of blackfly populations may be obtained with minimal direct impact on the “non-target” fauna, provided recommended dosages of temephos are not exceeded. Overdosing with temephos may result in high mortality of “non-target” organisms, including blackfly predators, and should be avoided.     

The naturally derived insecticide spinosad is highly toxic to Aedes and Anopheles mosquito larvae

Spinosad is a naturally derived biorational insecticide with an environmentally favourable toxicity profile, so we investigated its potency against mosquito larvae (Diptera: Culicidae). By laboratory bioassays of a suspension concentrate formulation of spinosad (Tracer), the 24 h lethal concentration (LC50) against Aedes aegypti (L.) third and fourth instars was estimated at 0.025 p.p.m. following logit regression. The concentration-mortality response of third- and fourth-instar Anopheles albimanus Weidemann did not conform to a logit model. The LC50 value of spinosad in Anopheles albimanus was 0.024 p.p.m. by quadratic linear regression. A field trial in southern Mexico demonstrated that spinosad 1 p.p.m. compared with the standard temephos (Abate) 1% granules 100 g/m3 water prevented Ae. aegypti breeding in plastic containers of water for 8 weeks; at 10 p.p.m. spinosad prevented breeding for > 22 weeks. In another field trial, spinosad at 5 p.p.m. and temephos both completely eliminated reproduction of Ae. aegypti for 13 weeks. In contrast, the bacterial insecticide Bacillus thuringiensis var. israelensis (Bti, Vectobac) AS) performed poorly with just 2 weeks of complete inhibition of Ae. aegypti breeding. Spinosad also effectively prevented breeding of Culex mosquitoes and chironomids in both trials to a degree similar to that of temephos. We conclude that spinosad merits evaluation as a replacement for organophosphate or Bti treatment of domestic water tanks in Mesoamerica. We also predict that spinosad is likely to be an effective larvicide for treatment of mosquito breeding sites.     

Temephos-resistant larvae of Simulium sanctipauli associated with a distinctive new chromosome inversion in untreated rivers of south-western Ghana

Larvae of the Simulium damnosum Theobald complex (Diptera: Simuliidae) were sampled in June 1996 from two sites in south-west Ghana where larviciding has not been applied: Sutri Rapids on the Tano river (05 degrees 23 minutes N 02 degrees 38 minures W) and Sekyere-Heman on the Pra river (05 degrees 11 minutes N 01 degrees 35 minutes W). All specimens were identified as Simulium sanctipauli Vajime & Dunbar sensu stricto (Diptera: Simuliidae). Bioassays with temephos (organophosphorus larvicide employed by the Onchocerciasis Programme for systematic treatment of most rivers across West Africa since the 1970s) showed about five-fold resistance in the Tano population (LC95 2.37-3.14 mg/L) and slight tolerance to temephos in the Pra population (LC95 0.67-0.76 mg/L), vs. the diagnostic concentration of 0.625 mg/L. Larval salivary polytene chromosomes of S. sanctipauli showed fixed inversions 1S-24/24, standard IIL-6 and a new inversion IL/36 polymorphism at Sutri on the Tano. These karyotype characteristics differ from those of temephos-resistant S. sanctipauli in rivers of C te d'Ivoire and other sites on the Tano in Ghana. Thus, temephos resistance in S. sanctipauli at Sutri is associated with distinct chromosomal configurations, showing that immigration was unlikely. This resistance could have been locally selected by exposure of S. sanctipauli larval populations to agrochemicals run-off from cocoa, coffee and oil plantations flanking the rivers.     

Defence mechanisms against insecticides temephos and diflubenzuron in the mosquito Aedes caspius: the P-glycoprotein efflux pumps

P-glycoproteins (P-gps) are efflux transporters found in cells of a broad range of both procaryotic and eukaryotic taxa, whose action is to relieve the cells of multiple, structurally dissimilar, toxic compounds. The possible role of P-gps in defence against the insecticides temephos and diflubenzuron was investigated in the mosquito Aedes caspius (Pallas), also known as Ochlerotatus (Aedes) caspius (Diptera: Culicidae), and the genomic DNA sequences encoding for P-gp transporters were isolated to provide molecular instruments for future research into the expression and characterization of genes codifying for P-gps in this mosquito species. Mosquito larvae were treated with insecticides alone and in conjunction with a sublethal dose of the P-gp inhibitor verapamil. The inhibition of P-gps reduced the LD(50) values of temephos and diflubenzuron by factors of 3.5 and 16.4, respectively, suggesting the potential involvement of P-gps in insecticide defence. Using a polymerase chain reaction (PCR)-based approach, a 481-bp sequence was isolated. The inferred nucleotide sequence shows high homology with the C-terminal sequence of known P-gps. The isolation and characterization of a putative P-gp sequence from Ae. caspius is the first step towards a better molecular understanding of the role played by multidrug transporters in the defence against insecticides in this species. This knowledge may open the way to a novel control strategy based on the inhibition of pest defences. The beneficial consequences of the inhibition of efflux pumps in improving insecticide performance are discussed.     

Comparative sensitivity of larval mosquitoes to vegetable polyphenols versus conventional insecticides

The larvicidal effects of polyphenols of natural crude decomposed alder leaf litter and commercially available tannic acid were experimentally compared with those of two common conventional insecticides (Bacillus thuringiensis ssp. israelensis: microbial insecticide; temephos: organophosphate insecticide). Comparative standard bioassays using third instar larval Aedes aegypti, A. albopictus, Culex pipiens and Coquillettidia richiardii as references indicated that Aedes and Culex taxa are far more sensitive to alder leaf litter than to tannic acid and conventional insecticides. C. richiardii is far more resistant to conventional insecticides than Aedes and Culex taxa, but its sensitivity to tannic acid is close to that of those taxa. Dietary vegetable polyphenols are thus proposed as new, practical, alternative chemicals for mosquito control when conventional insecticides are difficult and costly to be used (e.g., in the management of Aedes and Culex populations in man-made breeding sites and Coquillettidia control strategy).     

Temephos resistance and esterase activity in the mosquito Aedes aegypti in Havana, Cuba increased dramatically between 2006 and 2008

Aedes aegypti (L.) (Diptera: Culicidae) control programmes in Cuba rely on the application of the organophosphate temephos for larval control. Hence, the monitoring of resistance to this insecticide is an essential component of such programmes. Here, 15 field populations from different municipalities of Havana City were assayed for resistance to temephos. High levels of resistance were detected in all strains and resistance ratios were highly correlated with esterase activity (P = 0.00001). Populations from three municipalities were tested in both 2006 and 2008; resistance and esterase activities both significantly increased during this 2-year period. Synergist studies demonstrated that neither glutathione transferases nor monooxygenases were associated with the increase in resistance to temephos in this period. The duration of the efficacy of commercial formulations of temephos in controlling Ae. aegypti populations in Havana City was reduced by the high level of temephos resistance observed; hence these data are of clear operational significance for the dengue control programme in Cuba. New integrated strategies to avoid further increases in temephos resistance in Cuba are necessary.     

The effects of five organophosphorus insecticides on thermal stress in tadpoles of the Pacific tree frog, Hyla regilla

Three week old tadpoles of Hyla regilla were exposed to the insecticides temephos, fenthion, methyl parathion, chlorpyrifos, and malathion for 24 h at concentrations less than and at field applied rates as recommended for mosquito control in California. The thermal tolerance was significantly lowered by all chemicals and concentrations tested. Chlorpyrifos and methyl parathion were the most active compounds tested whereas malathion was the least toxic to H regilla tadpoles. Activity was depressed, in comparison to control tadpoles, by the chemicals methyl parathion (100 ppb) and malathion (500 ppb).     

Relative toxicity of different combinations of temephos and fenthion with Solanum xanthocarpum extract against anopheline larvae

The relative toxicity of different concentrations of temephos and fenthion with petroleum ether root extract of Solanum xanthocarpum (Schrader) at 1 : 1, 1 : 2 and 1 : 4 ratios was evaluated against Anopheles stephensi (Liston) larvae. All combinations exhibited antagonism at concentrations lower than LC₅₀ values and synergism at concentrations higher than their LC₉₀ values. A 1 : 1 product ratio was the most effective.     

IMPACTS OF REPEATED APPLICATIONS OF BACILLUS THURINGIENSIS VAR. ISRAELENSIS DE BARJAC AND TEMEPHOS,USED IN BLACKFLY (DIPTERA: SIMULIIDAE) CONTROL,ON MACROINVERTEBRATES IN THE MIDDLE ORANGE RIVER,SOUTH AFRICA

Summary

The impacts of five consecutive treatments of blackfly larvicides on macroinvertebrates in the middle Orange River were assessed. The abundance of the midge Xenochironomus sp. and the limpet Burnupia sp. was lower at sites treated with Bacillus thuringiensis var. israelensis (B.t.i.) compared to control sites (P<0.05). There was no medium-term impact on the abundance of two B.t.i.-sensitive taxa: Simulium spp. and the midge Rheotanytarsus fuscus. Repeated applications of the organophosphate temephos (Abate^R 200EC) significantly reduced the abundance of 25% of the invertebrate taxa in the stones-in-current biotope (P<0.1). The most affected taxa were R. fuscus, certain Baetis spp. mayflies and Coenagrionidae damselflies. Ordination of the species/sample matrix using detrended correspondence analysis showed that changes in invertebrate abundance and composition caused by either larvicide were well within the range of natural seasonal and spatial variation. However, the control programme may be detrimental to non-pest blackfly species, particularly the South African endemic Simulium gariepense, which may be endangered because of river regulation. It is concluded that the medium-term impacts of operational dosages of B.t.i. and temephos are safe provided that the recommended dosages of temephos are adhered to, and provided certain sections of river are left untreated as refugia for sensitive taxa.     

Differential protein expression in the midgut of Culex quinquefasciatus mosquitoes induced by the insecticide temephos

Mosquitoes are vectors for pathogens of malaria, lymphatic filariasis, dengue, chikungunya, yellow fever and Japanese encephalitis. Culex quinquefasciatus Say, 1823 (Diptera: Culicidae) is a known vector of lymphatic filariasis. Its control in Brazil has been managed using the organophosphate temephos. Studies examining the proteins of Cx. quinquefasciatus that are differentially expressed in response to temephos further understanding of the modes of action of the insecticide and may potentially identify resistance factors in the mosquito. In the present study, a comparative proteomic analysis, using 2‐dimensional electrophoresis coupled with matrix‐assisted laser desorption/ionization (MALDI) time of flight (TOF)/TOF mass spectrometry, and bioinformatics analyses were performed to identify midgut proteins in Cx. quinquefasciatus larvae that were differentially expressed in response to exposure to temephos relative to those in untreated controls. A total of 91 protein spots were differentially expressed; 40 were upregulated and 51 were downregulated by temephos. A total of 22 proteins, predominantly upregulated, were identified as known to play a role in the immune response, whereas the downregulated proteins were involved in energy and protein catabolism. This is the first proteome study of the midgut of Cx. quinquefasciatus and it provides insights into the molecular mechanisms of insecticide‐induced responses in the mosquito.