Hafiz Azhar Ali Khan

Genetics, cross-resistance and mechanism of resistance to spinosad in a field strain of Musca domestica L. (Diptera: Muscidae)

The house fly, Musca domestica L., is a cosmopolitan insect with the ability to develop resistance to insecticides used for their management. In the present study, we investigated the genetics of spinosad resistance, and cross-resistance potential to other insecticides by selecting a field strain with a commercial spinosad formulation. Bioassays with the field strain, before selection with spinosad, gave resistance ratios (RRs) of 4, 5, 66, 21 and 5 fold for spinosad, indoxacarb, abamectin, imidacloprid and deltamethrin, respectively, in comparison to a laboratory susceptible (Lab-susceptible) strain. After continuous selection of the field strain (Spin-SEL) with spinosad, the RR was increased up to 155 fold; however, the resistance was unstable (RR decreased 1.43 fold) when this strain was not exposed to spinosad for five generations. The Spin-SEL strain did not show cross-resistance to abamectin, indoxacarb or deltamethrin, but showed negative cross-resistance to imidacloprid. Crosses between the Spin-SEL and Lab-susceptible strains revealed an autosomal and incomplete dominant mode of resistance to spinosad. A direct test using a monogenic inheritance model based on Chi-square analysis revealed that the resistance was governed by more than one gene. Moreover, the resistance was neither overcome with the insecticide synergist piperonyl butoxide nor with S,S,S-tributylphosphorotrithioate. Lack of cross-resistance and instability of resistance suggest that rotation with spinosad could be an effective resistance management strategy.

Resistance to conventional insecticides in Pakistani populations of Musca domestica L. (Diptera: Muscidae): a potential ectoparasite of dairy animals.

The house fly, Musca domestica L., is an important hygienic pest of humans and dairy animals with the potential to develop resistance to most chemical classes of insecticides. Six adult house fly strains from dairy farms in Punjab, Pakistan were evaluated for resistance to selected insecticides from organochlorine, organophosphate, carbamate and pyrethroid classes. For a chlorocyclodiene and two organophosphates tested, the resistance ratios (RR) at LC50 were in the range of 5.60–22.02 fold for endosulfan, 7.66–23.24 fold for profenofos and 2.47–7.44 fold for chlorpyrifos. For two pyrethroids and one carbamate, the RR values at LC50 were 30.22–70.02 for cypermethrin, 5.73–18.31 for deltamethrin, and 4.39–15.50 for methomyl. This is the first report of resistance to different classes of insecticides in Pakistani dairy populations of house flies. Regular insecticide resistance monitoring programs on dairy farms are needed to prevent field control failures. Moreover, integrated approaches including the judicious use of insecticides are needed to delay the development of insecticide resistance in house flies.

The Effect of Temperature on the Toxicity of Insecticides against Musca domestica L.: Implications for the Effective Management of Diarrhea

Background Diarrhea is an important cause of childhood mortality in developing countries like Pakistan because of unhygienic conditions, lack of awareness, and unwise use of preventive measures. Mechanical transmission of diarrheal pathogens by house flies, Musca domestica, is believed as the most effective route of diarrhea transmission. Although the use of insecticides as a preventive measure is common worldwide for the management of house flies, success of the measure could be compromised by the prevailing environmental temperature since it significantly affects toxicity of insecticides and thus their efficacy. Peaks of the house fly density and diarrheal cases are usually coincided and season specific, yet little is known about the season specific use of insecticides. Methodology/Principal Findings To determine the temperature-toxicity relationship in house flies, the effect of post-bioassays temperature (range, 20–34°C) on the toxicity of seven insecticides from organophosphate (chlorpyrifos, profenofos), pyrethroid (cypermethrin, deltamethrin) and new chemical (emamectin benzoate, fipronil, spinosad) classes was evaluated by using a feeding bioassay method. From 20–34°C, the toxicities of chlorpyrifos, profenofos, emamectin and fipronil increased 2.10, 2.93, 2.40 and 3.82 fold (i.e. positive temperature coefficient), respectively. Whereas, the toxicities of cypermethrin, deltamethrin and spinosad decreased 2.21, 2.42 and 3.16 fold (i.e. negative temperature coefficient), respectively. Conclusion/Significance These findings suggest that for the reduction in diarrheal cases, house flies should be controlled with insecticides according to the prevailing environmental temperature. Insecticides with a positive temperature coefficient may serve as potential candidates in controlling house flies and diarrhea epidemics in hot season and vice versa.

A cross sectional survey of knowledge, attitude and practices related to house flies among dairy farmers in Punjab, Pakistan

BackgroundHouse flies are of major public health concerns in areas with poor sanitation and hygienic conditions. Unfortunately, sanitation and hygiene have always been ignored in dairy farms particularly in the developing or low-income countries. Management of these flies mostly depends on the awareness regarding associated hazards and protective measures taken by the people to minimize risks associated with flies. The present study therefore explores the knowledge, attitude and practices taken by dairy farmers in Punjab, Pakistan against house flies.MethodsThe present study was based on a cross sectional self administered survey to a convenience sample of 173 small scale dairy farmers in four localities – Multan, Lahore, Shorkot and Faisalabad – of Pakistan. The relationships between socio-demographics, knowledge and preventive practices were investigated through logistic regression analysis and chi-square test of association.ResultsConsiderable number of dairy farmers 71/173 (41.04%) had no idea about the problems associated with house flies. Although 77/173 (44.51%) dairy farmers reported house flies as disease transmitters, only 23 (29.87%) farmers were familiar with diseases and 22 (28.57%) had somewhat idea of the mode of disease transmission. We found a positive association between dairy farmer’s education level and overall knowledge of house flies in multivariate analysis. Farmer’s education level and knowledge of the house flies breeding sites had a positive association with the adoption of house fly prevention practices by the respondents. However, knowledge of the problems associated with house flies and preventive measures had no association with house fly prevention practices.ConclusionThe present ethnoentomological survey provides information about knowledge, attitude and practices of dairy farmers related to house flies in Punjab, Pakistan. We conclude that the farmers’ education level and knowledge of the breeding sites had a positive association with the adoption of prevention practices against house flies. The study also highlights the need of targeting the lack of knowledge of dairy farmers for the successful management of house flies.

Thiamethoxam Resistance in the House Fly, Musca domestica L.: Current Status, Resistance Selection, Cross-Resistance Potential and Possible Biochemical Mechanisms

The house fly, Musca domestica L., is an important ectoparasite with the ability to develop resistance to insecticides used for their control. Thiamethoxam, a neonicotinoid, is a relatively new insecticide and effectively used against house flies with a few reports of resistance around the globe. To understand the status of resistance to thiamethoxam, eight adult house fly strains were evaluated under laboratory conditions. In addition, to assess the risks of resistance development, cross-resistance potential and possible biochemical mechanisms, a field strain of house flies was selected with thiamethoxam in the laboratory. The results revealed that the field strains showed varying level of resistance to thiamethoxam with resistance ratios (RR) at LC50 ranged from 7.66-20.13 folds. Continuous selection of the field strain (Thia-SEL) for five generations increased the RR from initial 7.66 fold to 33.59 fold. However, resistance declined significantly when the Thia-SEL strain reared for the next five generations without exposure to thiamethoxam. Compared to the laboratory susceptible reference strain (Lab-susceptible), the Thia-SEL strain showed cross-resistance to imidacloprid. Synergism tests revealed that S,S,S-tributylphosphorotrithioate (DEF) and piperonyl butoxide (PBO) produced synergism of thiamethoxam effects in the Thia-SEL strain (2.94 and 5.00 fold, respectively). In addition, biochemical analyses revealed that the activities of carboxylesterase (CarE) and mixed function oxidase (MFO) in the Thia-SEL strain were significantly higher than the Lab-susceptible strain. It seems that metabolic detoxification by CarE and MFO was a major mechanism for thiamethoxam resistance in the Thia-SEL strain of house flies. The results could be helpful in the future to develop an improved control strategy against house flies.

Cyromazine resistance in a field strain of house flies, Musca domestica L.: Resistance risk assessment and bio-chemical mechanism.

Developing resistance management strategies for eco-friendly insecticides is essential for the management of insect pests without harming the environment. Cyromazine is a biorational insecticide with very low mammalian toxicity. Resistance to cyromazine has recently been reported in house flies from Punjab, Pakistan. In order to propose a resistance management strategy for cyromazine, experiments were planned to study risk for resistance development, possibility of cross-resistance and bio-chemical mechanisms. A field strain of house flies with 8.78 fold resistance ratio (RR) to cyromazine was re-selected under laboratory conditions. After seven rounds of selection (G1-G7), the RR values rapidly increased from 8.8 to 211 fold. However, these values declined to 81fold when the cyromazine selected (CYR-SEL) strain was reared without selection pressure, suggesting an unstable nature of resistance. The CYR-SEL strain showed lack of cross-resistance to pyriproxyfen, diflubenzuron, and methoxyfenozide. Synergism bioassays using enzyme inhibitors: piperonyl butoxide (PBO) and S,S,S-tributylphosphorotrithioate (DEF), and metabolic enzyme analyses revealed increased activity of carboxylesterase (CarE) and mixed-function oxidase (MFO) in the CYR-SEL strain compared to the laboratory susceptible (Lab-susceptible) strain, suggesting the metabolic resistance mechanism responsible for cyromazine resistance in the CYR-SEL strain. In conclusion, risk of rapid development of cyromazine resistance under consistent selection pressure discourages the sole reliance on cyromazine for controlling house flies in the field. The unstable nature of cyromazine resistance provides window for restoring cyromazine susceptibility by uplifting selection pressure in the field. Moreover, lack of cross-resistance between cyromazine and pyriproxyfen, diflubenzuron, or methoxyfenozide in the CYR-SEL strain suggest that cyromazine could be rotated with these insecticides whenever resistance crisis occur in the field.

Toxicity and resistance of field collected Musca domestica (Diptera: Muscidae) against insect growth regulator insecticides.

The house fly, Musca domestica, is a serious pest of public health importance with the ability to develop insecticide resistance. The focus of the present study was to evaluate toxicity and resistance of the field collected house flies from Punjab, Pakistan, against insect growth regulator (IGR) insecticides. House flies collected from six different localities exhibited very low levels of resistance to cyromazine, triflumuron, and methoxyfenozide compared with the Lab-susceptible reference strain, with resistance ratios (RRs) ranging between 3.56- to 8.19-fold, 1.45- to 3.68-fold, and 2.20- to 8.60-fold, respectively. However, very low to low levels of resistance were observed for pyriproxyfen and very low to moderate levels for lufenuron with RRs ranged from 4.13- to 11.63-fold to 8.57- to 22.75-fold, respectively. There was a significant correlation between RRs of cyromazine and triflumuron (r = 0.976, p < 0.001), suggesting the possibility for cross-resistance. Susceptibility status of different IGRs in Pakistani strains of house flies is reported here for the first time. The trend observed in decreasing susceptibility towards different IGR insecticides tested will continue unless resistance management practices are followed.

Genetics and mechanism of resistance to deltamethrin in the house fly, Musca domestica L., from Pakistan

Deltamethrin (a pyrethroid insecticide) has widely been used against the house fly, Musca domestica, a pest found in livestock facilities worldwide. Although, cases of both metabolic and physiological resistance to deltamethrin have been reported in different parts of the world, no studies have been reported to characterize this resistance in house flies from Pakistan. In the present study, we investigated a field strain of house flies for potential to develop resistance to deltamethrin. Also, its stability, possible mechanisms and cross-resistance potential to other insecticides. Before the selection experiments, the field strain showed 8.41-, 3.65-, 8.39-, 2.68-, 19.17- and 5.96-fold resistance to deltamethrin, bifenthrin, lambda-cyhalothrin, chlorpyrifos, profenofos and spinosad, respectively, compared with the reference strain (Lab-susceptible). Continuous selection of the field strain (Delta-SEL) with deltamethrin for six generations (G1–G6) in the laboratory increased the resistance ratio to 176.34 after bioassay at G7. The Delta-SEL strain was reared for the next four generations without exposure to deltamethrin and bioassayed at G11 which revealed that the resistance was stable. The Delta-SEL strain at G7 showed cross-resistance to all other insecticides except spinosad, when compared to the bioassays before the selection experiment (G1). Crosses between Delta-SEL and Lab-susceptible strains revealed an autosomal and incomplete dominant mode of resistance to deltamethrin. A direct test using a monogenic inheritance model revealed that the resistance was governed by more than one factor. Moreover, synergism studies with the enzyme inhibitors PBO and DEF reduced the resistance to deltamethrin in the selected strain up to 2.51- and 2.19-fold, respectively, which revealed that the resistance was possibly due to microsomal oxidase and esterase activity. It is concluded that the resistance to deltamethrin was autosomal and incompletely dominant. The high cross-resistance of bifenthrin, lambda-cyhalothrin, chlorpyrifos and profenofos in the Delta-SEL strain suggests that other insecticides would be necessary to counter the resistance. These results are therefore suggestive for implications in the management of insecticide resistance in house flies.

Combination of Phagostimulant and Visual Lure as an Effective Tool in Designing House Fly Toxic Baits: A Laboratory Evaluation

House flies (Diptera: Muscidae), potential vectors of a variety of pathogens, characteristically search and feed on sugar sources just after emergence for their survival. Phagostimulants like sugars, and visual characteristics of feeding materials play an important role in foraging success in house flies. Therefore, development of toxic baits by using the combination of phagostimulant and visual lure may prove effective in localized control of house flies. In the present study, visual attraction of house flies to different fabric colors was studied in choice and no choice experiments. Dark blue was the most preferred color in both experiments. In toxicity experiments, insecticide solutions were prepared in 20% sugar solution. Dark blue fabric strips were prepared by moistening with 20% sugar water solution containing median lethal concentrations of one of the four insecticides viz., fipronil, Imidacloprid, indoxacarb and Spinosad. The fabric strips treated with fipronil and Imidacloprid took minimum time (7.66 and 7.81 h, respectively) to cause 50% mortality, while those treated with Spinosad and indoxacarb took relatively more time (13.62 and 17.91 h, respectively) to cause 50% mortality. In conclusion, the combination of phagostimulant and visual lure could be used in designing toxic baits for house flies.

Impact of copper toxicity on stone-head cabbage (Brassica oleracea var. capitata) in hydroponics

Arable soils are frequently subjected to contamination with copper as the consequence of imbalanced fertilization with manure and organic fertilizers and/or extensive use of copper-containing fungicides. In the present study, the exposure of stone-head cabbage (Brassica oleracea var. capitata) to elevated Cu2+ levels resulted in leaf chlorosis and lesser biomass yield at ≥2 µ M. Root nitrate content was not statistically affected by Cu2+ levels, although it was substantially decreased at ≥5 µ M Cu2+ in the shoot. The decrease in nitrate contents can be related to lower nitrate uptake rates because of growth inhibition by Cu-toxicity. Shoot sulfate content increased strongly at ≥2 µ M Cu2+ indicating an increase in demand for sulfur under Cu stress. Furthermore, at ≥2 µM concentration, concentration of water-soluble non-protein thiol increased markedly in the roots and to a smaller level in the shoot. When exposed to elevated concentrations of Cu2+ the improved sulfate and water-soluble non-protein thiols need further studies for the evaluation of their direct relation with the synthesis of metal-chelating compounds (i.e., phytochelatins).