Molecular and Biochemical Detection of Insecticide Resistance in the Leishmania Vector, Phlebotomus papatasi (Diptera: Psychodidae) to Dichlorodiphenyltrichloroethane and Pyrethroids, in Central Iran.

The aim of the present study was to explore resistance markers and possible biochemical resistance mechanisms in the Phlebotomine sand fly Phlebotomus papatasi in Esfahan Province, central Iran. Homogenous resistant strains of sand flies were obtained by exposing P. papatasi collected from Esfahan to a single diagnostic dose of DDT. The adults from the colony were tested with papers impregnated with four pyrethroid insecticides: Permethrin 0.75%, Deltamethrin 0.05%, Cyfluthrin 0.15%, and Lambdacyhalothrin 0.05% to determine levels of cross-resistance. To discover the presence of mutations, a 440 base pair fragment of the voltage gated sodium channel (VGSC) gene was amplified and sequenced in both directions for the susceptible and resistant colonies. We also assayed the amount of four enzymes that play a key role in insecticide detoxification in the resistant colonies. A resistance ratio (RR) of 2.52 folds was achieved during the selection of resistant strains. Sequence analysis revealed no knockdown resistance (kdr) mutations in the VGSC gene. Enzyme activity ratio of the resistant candidate and susceptible colonies were calculated for α-esterases (3.78), ß-esterases (3.72), mixed function oxidases (MFO) (3.21), and glutathione-S-transferases (GST) (1.59). No cross-resistance to the four pyrethroids insecticides was observed in the DDT resistant colony. The absence of kdr mutations in the VGSC gene suggests that alterations in esterase and MFO enzymes are responsible for the resistant of P. papatasi to DDT in central Iran. This information could have significant predictive utility in managing insecticide resistant in this Leishmania vector.

Bioecological Study on the Sand Flies (Diptera: Psychodidae, Phlebotominae) in Sari County, North of Iran.

Background: Phlebotomine sand flies are vectors of Leishmania species, the causative agents of leishmaniasis in the world. Present study aimed to evaluate the bioecological aspects of sand flies in different ecotopes in Sari County, north of Iran. Methods: Sand flies were collected from four villages in mountainous, forest, plain and peri-urban areas monthly using sticky traps in May-October 2016. Mounted specimens were identified using valid identification keys under optical microscope. The Arc GIS 10.5 software was applied for showing the distribution of sand flies. Shannon-Weiner, Simpson and Evenness species diversity indices were calculated. Results: Generally, 334 specimens were captured and identified, namelly Phlebotomus kandelakii, Ph. papatasi, Ph. major, Ph. sergenti, Ph. longiductus, Ph. halepensis, Ph. tobbi, Sergentomyia dentata, Se. theodori, Se. sintoni, Se. antennata and Se. sumbarica. The most common species was Ph. kandelakii (n= 128, 38.32 %). The highest Simpson index (0.81) and abundance (N= 141) were recorded in the mountaineous area. Shannon diversity index was higher in the forest (H'= 1.53) and the highest evenness index was in the plain area (J'= 0.93). The highest richness (S= 9) and Shannon indices (H'= 1.57) were observed in June. Conclusions: Phlebotomus kandelakii, Ph. sergenti, Ph. tobbi, Ph. longiductus, Se. theodori, Se. antennata and Se. sumbarica were recorded for the first time in the study area. Since some species are incriminated for leishmaniasis transmission, further studies are required in the northern regions of Iran to timely control measures planning.

Ecology of sand flies (Diptera: Psychodidae, Phlebotominae) in Jajarm County, an area with high risk of cutaneous leishmaniasis, in North Khorasan, Iran.

The present study was conducted to investigate the ecological aspects of sand flies in southwestern North Khorasan, in which cutaneous leishmaniasis caused by Leishmania major has been reported with the highest annual incidence in Iran. Sampling was carried out in four localities including: Khorasha (natural), Ghamiteh (natural), Jorbat (semi urban) and Brick kilns (urban), twice a month using 105 sticky paper traps from indoors and outdoors dwellings during May-December 2017. Specimens were removed from sticky papers, washed in acetone, preserved in 80% ethanol, mounted on microscopic slides by Puri's medium, and identified using valid morphological keys. Simpson (D), richness (S), Menhinick (DMg), Margalef (DMn), Shannon-Weiner (H'), evenness (J') were calculated for species diversity. The synanthropic index was determined for the first time in the area. Totally 517 specimens were collected, 47% in outdoors and 30.4% in human indoor dwellings and 22.6% in animal. Eight species of sand flies including 5 species of the genus Phlebotomus and 3 species of the genus Sergentomyia were identified. Phlebotomus papatasi and Sergentomyia sintoni were the most common and Eudominant species, active in all months, collected in the maximum number and percentage in September and August, respectively, and showed the highest abundance in outdoors. The synanthropic index ranged from 6.25 to 38.9 in the study area. The Shannon-Wiener index was estimated to be up to 1.4 and 1.37 in Khorasha and November, respectively, which showed the highest diversity due to maximal richness and evenness compared to other areas. High abundance of Ph. papatasi, as the main vector of cutaneous leishmaniasis, can enhance the potential risk of emerging CL in new areas, the data can be equally important when vector control measures are considered.

Reemergence of zoonotic cutaneous leishmaniasis in an endemic focus, northeastern Iran.

Zoonotic cutaneous leishmaniasis (ZCL) is a form of disease, which is mostly caused by Leishmania major (L.major) in the Old World with high incidence through early life. Recently, the high incidence of CL has been reported in Jajarm County, northeastern Iran. The aim of the present study was to investigate the epidemiological aspects of CL and to identify Leishmania spp. in the area. Between 2012 and 2018, patients with active lesions suspected of CL were examined. Diagnosis was performed by methods of clinical examination and direct smears. Conventional kDNA polymerase chain reaction (PCR) was used to identify species of Leishmania. During the seven-year study period, 629 subjects were diagnosed as CL cases by clinical and microscopic examination. Considering the population of the study area, the average incidence rate was 237.8 per 100,000 population. Most (29.4%) of the subjects with ZCL were aged <10 years. Most of the patients (58.7%) were male and the rest were female. Most ulcers were observed on the face (32.2%) and hands (23.7%), respectively. Most patients (90%) had two or more ulcers on their body. The highest incidence was in the autumn in October (32%). Our data revealed that an ongoing reemergence of ZCL focus caused by L.major has arisen in the study area. Moreover, direct PCR on the smears for identifying Leishmania showed 100% specificity, and the species was identified as L. major using species -specific kDNA PCR. The use of conventional PCR on skin smears seems to provide a valuable method for confirming the clinical diagnosis of ZCL, which is very specific and sensitive particularly for clinical correlative studies. Further investigations are needed to improve planning strategies of vectors and reservoirs populations.

Anaplasma Infection in Ticks in Southeastern Region of Iran.

BACKGROUND: Anaplasmosis and Ehrlichiosis are the most important tick-borne diseases. This study was conducted in three cities of Kerman Province in Iran to investigate the circulation of the bacteria in ticks collected from sheep. METHODS: Ticks were collected from animals using Srkj forceps and transferred to the Entomology lab in cold chain. After specimen's identification, they kept at -70 °C. Tick DNA was extracted using Bioneers DNA extraction kits followed by Nested PCR technique to amplify ribosomal 16S rRNA gene to detect Anaplasma infection in ticks. RESULTS: 472 sheep were examined from which 349 ticks were collected and identified in laboratory using valid keys. Tick specimens belonged to two genera and four species; Hyalomma marginatum (62.47%) was the most frequent and Hylomma asiaticum (5.73%) showed the least abundance. The infestation rate to different tick species was different in three regions of Kerman Province. Observation revealed that 24 specimens (58.3%) were positive for Anaplasma. There is a significant difference between male and female infection rate. However, there is no significant difference between these variables in each of these cities. CONCLUSION: This study shows high infection rates to Anaplasma in hard ticks. It is essential for health and veterinary authorities and farmers to use appropriate strategies to control ticks to reduce the infestation.

Monitoring of Laboratory Reared of Phlebotomus papatasi (Diptera: Psychodidae), Main Vector of Zoonotic Cutaneous Leishmaniasis to Different Imagicides in Hyper endemic Areas, Esfahan Province, Iran.

BACKGROUND: In domestic and per domestic area, insecticides such as DDT, malathion, fenitrothion, propoxur and, more recently, synthetic pyrethroids such as deltamethrin and lambda-cyhalothrin, have been successfully used to control sand flies in many countries. The present study reports the results of time-mortality bioassay to DDT 4%, lambda-cyhalothrin 0.05%, permethrin 0.75%, cyfluthrin 0.15% and deltamethrin 0.05% in recently colonized Phlebotomus papatasi populations in Iran. METHODS: The insecticide susceptibility status of P. papatasi laboratory population was assessed during 2016-2017, following the standard WHO technique for mosquito (WHO, 2013) based on diagnostic dose. Sand flies collected from rural area of Badrood (Matin Abad), Natanz County, Esfahan Province, using aspirator. RESULTS: Susceptibility test to DDT and pyrethroids was assessed on 3534 laboratory-reared P. papatasi (1746 females and 1788 males). The LT 50 and LT 90 values were measured using probit analysis and regression lines. The test results against males of P. papatasi revealed that LT 50 values to DDT 4%, Permethrin 0.75%, Deltamethrin 0.05%, Cyfluthrin 0.15% and Lambdacyhalothrin 0.05% were 439.28, 108.90, 97.75, 5.00 and 57.84 seconds. The figures for females were 641.62, 136.15, 146.44, 8.71 and 72.69 seconds, respectively. CONCLUSION: According to presented results, the reared population of sand flies collected from a hyper-endemic region of Esfahan Province is still susceptible to prethroids and Resistance candidate to DDT 4%.

First Report of Target Site Insensitivity in Pyrethroid Resistant Anopheles gambiae from Southern Guinea Savanna, Northern-Nigeria.

BACKGROUND: Malaria is a major public health problem and life threatening parasitic vector-borne disease. For the first time, we established and report the molecular mechanism responsible for Anopheles gambiae s.l. resistance to pyrethroids and DDT from Yamaltu Deba, Southern Guinea Savanna, Northern-Nigeria. METHODS: The susceptibility profile of An. gambiae s.l. to four insecticides (DDT 4%, bendiocarb 0.1%, malathion 5% and deltamethrin 0.05%) using 2-3 days old females from larvae collected from study area between August and November, 2018 was first established. Genomic DNA was then extracted from 318 mosquitoes using Livak DNA extraction protocol for specie identification and kdr genotyping. The mosquitoes were identified to species level and then 96 genotyped for L1014F and L1014S kdr target site mutations. RESULTS: The mosquitoes were all resistant to DDT, bendiocarb and deltamethrin but fully susceptible to malathion. An. coluzzii was found to be the dominant sibling species (97.8%) followed by An. arabiensis (1.9%) and An. gambiae s.s (0.3%). The frequency of the L1014F kdr mutation was relatively higher (83.3%) than the L1014S (39%) in the three species studied. The L1014F showed a genotypic frequency of 75% resistance (RR), 17% heterozygous (RS) and 8% susceptible (SS) with an allelic frequency of 87% RR and 13% SS while the L1014S showed a genotypic frequency of RR (16%), RS (38%) and SS (46%) with an allelic frequency of 40% RR and 60% SS, respectively. CONCLUSION: This study reveals that both kdr mutations present simultaneously in Northern-Nigeria, however contribution of L1014F which is common in West Africa was more than twice of L1014S mutation found in East Africa.

Co-encapsulation of imidacloprid and lambda-cyhalothrin using biocompatible nanocarriers: Characterization and application.

A well-known strategy for managing pest resistance is application of mixture of pesticides. Conventionally formulated pesticides have several environmental incompatibilities. The use of biocompatible and biodegradable nanocarriers in formulating pesticides could improve environmental protection. In this study, a mixture of imidacloprid and lambda-cyhalothrin was co-encapsulated for the first time using liposomes as nanocarrier to simultaneously deliver these insecticides. Ethanol injection was used to produce self-assembled liposomes. The formed nanoliposomes were coated with different concentrations of chitosan. Nanoparticles were characterized by dynamic light scattering (DLS), scanning electron microscope (SEM) and FT-IR spectroscopy. The encapsulation efficiencies of lambda-cyhalothrin and imidacloprid were about 93% and 51%, respectively. The insecticide carrying liposomes had a size and surface charge of 57 nm and +0.6 mV, respectively. The size and surface charge of the particles produced were increased to 69 nm and +31 mV after being coated with chitosan (0.1%, W/V). In this study, residual activity of technical grade imidacloprid, lambda-cyhalothrin and their mixture and the effect of adjuvants used in commercial and nano formulations of these insecticides on Myzus persicae Sulzer was investigated. The insecticidal effects and duration of residual activity of nano-formulations was correlated with concentration of chitosan in final formulation. In accordance with the life cycle of M. persicae, using the mixture of imidacloprid and lambda-cyhalothrin improves the residual effect over their use alone. The use of lipid nanocarriers makes the improvement even further and can be a better alternative to conventional combination of these insecticides due to their more environmental friendliness.

Prevalence of pediculosis and its related factors among primary school girls in the north of Iran.

INTRODUCTION: Head lice infestation is one of the main public health problems worldwide and a sanitary, cultural and economic indicator for communities' health. This study aims to estimate the prevalence of head lice infestation among female primary school children in selected districts of Mazandaran Province, Iran. METHODS: In this cross-sectional study, 3033 female school children were recruited using the consensus method. Head lice infestation was defined as visible adult lice, nymph or egg with the naked eye. Data were described by percent frequency and analyzed using the chi-squared test and multivariate logistic regression models. A p-value less than 0.05 was considered significant. RESULTS: Prevalence of head lice infestation among female school children was estimated at 7.9%. The adjusted odds ratios (OR) [95% confidence intervals (CI)] for rural residents, low level educated mothers, family size more than five and not combing their hair were 1.96 (1.41-2.78), 2.46 (1.35-4.49), 2.04 (1.12-3.70) and 3.94 (1.73-8.96), respectively. In addition, considering first grade students as a reference group, the ORs (95% CI) for second and third grades were 1.89 (1.18-3.05) and 1.69 (1.05-2.74), respectively. CONCLUSION: Our study showed that studying in urban schools, low educational level of mothers, grade, family size and no history of regular combing were predictors of head lice infestation.

First Report of Biochemical Mechanisms of Insecticide Resistance in the Field Population of Culex pipiens (Diptera: Culicidae) from Sari, Mazandaran, North of Iran.

BACKGROUND: Culex pipiens play an important role in transmission of infectious diseases. Vector control by chemical pesticides, leads inevitably to resistance development. Understanding the underlying resistance mechanisms can help improve the control programmes and insecticide resistance management. METHODS: The total contents of cytochrome p450s and the activities of glutathione S-transferases, alpha- and beta-esterases and inhibition rates of acetylcholine esterase (by propoxur) were measured in the field population of Cx. pipiens collected from Sari County, North of Iran, in 2016 and the results were compared with those of the laboratory susceptible strain according to the biochemical assay methods of WHO for adult mosquitoes. Independent sample t-test was used to compare the mean values of enzyme activities/contents between filed and laboratory susceptible populations. RESULTS: The enzyme ratio of cytochrome p450s, alpha- and beta-esterases in the field population was 2.07, 3.72 and 1.36 respectively when compared with the results of the laboratory population. Although not statistically significant, the mean GSTs activities in the field population was marginally less than the laboratory population (ER=0.92). Acetylcholinesterase was insensitive to propoxur in 62.82% of the individuals of the tested field population. There was a significant difference (P< 0.05) between all values of the activities/contents of the enzyme in the field population except for GSTs compared with the laboratory susceptible strain. The highest enzyme activity was related to alpha esterase. CONCLUSION: The present study showed a range of metabolic mechanisms, comprising p450s and esterases combined with target site insensitivity of AChE, contributing to organophosphate, carbamate and pyrethroid resistance in the field population of Cx. pipiens.

Biochemical Basis of Cyfluthrin and DDT Resistance in Anopheles stephensi (Diptera: Culicidae) in Malarious Area of Iran.

BACKGROUND: Anopheles stephensi is a key urban malaria vector in the Indian subcontinent and Middle East including south and southeast of Iran. Wide application of insecticides resulted in resistance of this species to various insecticides in these regions. This study was conducted to reveal the role of metabolic mechanisms in the development of resistance in An. stephensi to DDT and cyfluthrin. METHODS: Field mosquito specimens were collected from Chabahar Seaport, southeast corner of Iran, in 2015. Insecticide susceptibility and enzyme assays were conducted as recommended by WHO. RESULTS: Mean enzyme ratios were 3.95 and 3.04 for α- esterases and 2.40 and 1.97 for ß- esterases in the DDT and cyfluthrin- resistant populations correspondingly compared with the susceptible strain. The GSTs enzyme mean activity ratios were 5.07 and 2.55 in the DDT and cyfluthrin- resistant populations compared with the susceptible beech strain. The cytochrome p450s enzyme ratios were 1.11 and 1.28 in the DDT and cyfluthrin- resistant populations respectively compared with the susceptible beech strain. CONCLUSION: Metabolic mechanisms play a crucial role in the development of DDT and cyfluthrin resistance in An. stephensi, therefore, further evaluation of the mechanisms involved as well as implementation of proper insecticide resistance management strategies are recommended.

High Resistance of Vector of West Nile Virus, Culex pipiens Linnaeus (Diptera: Culicidae) to Different Insecticides Recommended by WHO in Northern Iran.

BACKGROUND: Culex pipiens is a mosquito species distributed in different parts of Iran. It is known as vector of some diseases as well as a nuisance insect. A successful control campaign needs to study the biology, ecology and susceptibility status of the target pest. The aim of this study was to investigate the susceptibility status of Culex pipiens to some insecticides recommended by WHO in north of Iran. METHODS: Larvae were collected from different breeding places in Sari County by standard dipping method. They were reared to adult stage in an under standard condition. World Health Organization (WHO) standard test kits and the diagnostic dose of 12 imagicides were used for adults, while two larvicides were used against larvae. RESULTS: The least and highest mortality rates after exposure to insecticides were 4.7% (Propoxur 0.1%) and 76.4% (Etofenprox 0.5%), respectively. Larvae showed high resistance to temephos (19.19%) and malathion (54.63%). Culex pipiens was found to be resistant to all used insecticides/larvicides at diagnostic dose. CONCLUSION: Intensive use of pesticides against agriculture and urban pests increased the resistance level of this species to different insecticides, although currently there is no control program against it. Study on the mechanisms of resistant in this species to different insecticides is recommended.

First report of target site insensitivity to pyrethroids in human flea, Pulex irritans (Siphonaptera: Pulicidae).

The human flea, Pulex irritans, is the most important ectoparasite of humans. Intensive use of pyrethroids for its control has led to insecticide resistance. Monitoring pyrethroid resistance and its underlying mechanisms is essential for flea control. The aims of this study were to identify the susceptibility status of human flea to permethrin and to detect the presence of knockdown resistance (kdr) mutation and its frequency in populations of P. irritans. Adults of P. irritans were collected from Zanjan Province, northwest of Iran, during 2013-2017. Different populations of this flea were exposed to permethrin 0.75% for one and 8 h and then the mortality rate, as well as KD50 and KD95 times were calculated. Total RNA and gDNA of samples were extracted, and the fragments of cDNA encoding the partial voltage-gated sodium channel (VGSC) peptides were amplified using degenerated primers. Specific PCR and TaqMan real-time assays were conducted to characterize the vgsc gene and to detect the presence of mutation and genotyping of the populations. Mortality rates were in the range from 32% to 67% for one-hour and 73% to 90% for eight-hour exposure to permethrin 0.75%. KD50 and KD95 times varied in a range from 46 to 241 and 177 to 899 min, respectively. Sequencing of 70 amplified fragments of gDNA resulted in a 578-bp product. These fragments contained two introns (92 and 63 bp) and three exons (141, 189, and 92 bp) encoding 138 amino acids that encompassed IIS4-IIS6 and the partial linker between domains II and III of VGSC. All the studied populations showed L1014F mutation, substitution of CTT for TTT at the 1014 allele. The result of TaqMan assay for 624 samples showed 96.6% homogenous and 3.36% heterozygous mutant. The development of permethrin resistance and the presence of the L1014F mutation at high frequency in flea populations indicate that pyrethroids are likely ineffective in controlling human flea. Therefore, novel alternative control methods are needed to combat this human ectoparasite.

Prospective Epidemiological Research Studies in Iran (the PERSIAN Cohort Study): Rationale, Objectives, and Design.

Noncommunicable diseases (NCDs) account for 76% of deaths in Iran, and this number is on the rise, in parallel with global rates. Many risk factors associated with NCDs are preventable; however, it is first necessary to conduct observational studies to identify relevant risk factors and the most appropriate approach to controlling them. Iran is a multiethnic country; therefore, in 2014 the Ministry of Health and Medical Education launched a nationwide cohort study-Prospective Epidemiological Research Studies in Iran (PERSIAN)-in order to identify the most prevalent NCDs among Iran's ethnic groups and to investigate effective methods of prevention. The PERSIAN study consists of 4 population-based cohorts; the adult component (the PERSIAN Cohort Study), described in this article, is a prospective cohort study including 180,000 persons aged 35-70 years from 18 distinct areas of Iran. Upon joining the cohort, participants respond to interviewer-administered questionnaires. Blood, urine, hair, and nail samples are collected and stored. To ensure consistency, centrally purchased equipment is sent to all sites, and the same team trains all personnel. Routine visits and quality assurance/control measures are taken to ensure protocol adherence. Participants are followed for 15 years postenrollment. The PERSIAN study is currently in the enrollment phase; cohort profiles will soon emerge.

Assessing the insecticide susceptibility status of field population of Phlebotomus papatasi (Diptera: Psychodidae) in a hyperendemic area of zoonotic cutaneous leishmaniasis in Esfahan Province, Central Iran.

Leishmaniasis is a Neglected Tropical Disease (NTD) and emerging parasitic infection that affect mainly poor regions around the world. This study aimed to determine the baseline susceptibility of Phlebotomus papatasi to commonly used insecticides in a hyper endemic area using WHO standard procedure in central Iran. A total of 4-5 replicates containing 120-200 sand flies were used for each insecticide. Baseline susceptibility to DDT and pyrethroids was assessed on 5326 specimens collected from the study area. The LT50 and LT90 values were measured according to the World Health Organisation test using probit analysis and regression lines. The test results against males P. papatasi revealed that LT50 values to DDT 4%,Permethrin 0.75%, Deltamethrin 0.05%, Cyfluthrin 0.15% and Lambdacyhalothrin 0.05% were 564.07, 38.08, 1.95, 0.60 and 9.78s and the figures for females were 584.44, 110.10, 11.64, 1.53 and 16.91s, respectively. Our results indicated that P. papatasi as the main cutaneous leishmaniasis vector was susceptible to Cyfluthrin 0.15%, Lambdacyhalothrin 0.05%, Permethrin 0.75% and Deltamethrin 0.05% and tolerant to DDT 4%. This study was carried out in one out of many Leishmaniasis foci in Iran. We recommend that future studies incorporate other regions and use the same procedure for monitoring and evaluating sand fly resistance. Also, WHO can provide a specific guideline and create a test kit for sand fly resistance monitoring and for applying susceptibility test because the tubes prepared for mosquitoes are not actually fit for sand flies.

The role of midgut symbiotic bacteria in resistance of Anopheles stephensi (Diptera: Culicidae) to organophosphate insecticides.

In the current study, the effects of the presence of symbiotic bacteria on the activity of the enzymes involved in An. stephensi resistance to temephos are evaluated for the first time. Four different strains (I. susceptible strain, II. resistant strain, III. resistant strain + antibiotic, and IV. resistant strain + bacteria) were considered in order to determine the possible effects of the symbiotic bacteria on their hosts' resistance to temephos. The median values of all enzymes of susceptible strain were compared with those of other resistant strains. The results of this study indicated a direct relationship between the presence of bacteria in the symbiotic organs of An. stephensi and resistance to temephos. The profile of enzymatic activities in the resistant strain changed to a susceptible status after adding antibiotic. The resistance of An. stephensi to temephos could be completely broken artificially by removing their bacterial symbionts in a resistant population.

Evaluation of Deltamethrin in Combination of Piperonyl Butoxide (PBO) against Pyrethroid Resistant, Malaria Vector, Anopheles stephensi in IRS Implementation: an Experimental Semi-Filed Trial in Iran.

BACKGROUND: The aim of this study was to evaluate different concentrations of deltamethrin combined with formulated piperonyl butoxide (PBO) synergist on various surfaces against the wild strain of Anopheles stephensi, the main malaria vector in Southern Iran under semi-field condition. METHODS: Four concentrations of deltamethrin WG 25% (Tagros) and PBO 800EC-UV (Endura) were prepared and sprayed on the pre-designed surfaces in accordance with WHO alliance line of the IRS Micronair®. The WHO's recommended bioassay kit and method was used during this study. RESULTS: Comparing the mortality rate of mosquitoes, the results showed a significant difference between months after treatment of IRS (Indoor Residual Spraying) (P< 0.05) but didn't show any significant differences between days during the first and second months (P> 0.05).Statistical test revealed a significance difference between mortality rate of mosquitoes in exposing to concentrations of 1 and 4 (P< 0.05) which demonstrated effect of synergizing PBO on mortality rate. CONCLUSION: This research as the first semi-field trial on deltamethrin added to different concentrations of formulated PBO for IRS, indicates that deltamethrin+10X PBO is more effective than other concentrations. Therefore, using synergists can be suggested as a new tool for prevention of pyrethriod resistance, although more studies are recommended.

High Insecticides Resistance in Culex pipiens (Diptera: Culicidae) from Tehran, Capital of Iran.

BACKGROUND: During recent years transmission of Dirofilaria immitis (dog heart worm) by Culex pipiens and West Nile virus have been reported from Iran. The present study was preformed for evaluating the susceptibility status of Cx. pipiens collected from capital city of Tehran, Iran. METHODS: Four Insecticides including: DDT 4%, Lambdacyhalothrin 0.05%, Deltamethrin 0.05% and Cyfluthrin 0.15 % according to WHO standard methods were used for evaluating the susceptibility status of Cx. pipiens from Tehran moreover For comparison susceptibility status a Laboratory strain also was used. Bioassay data were analyzed using Probit program. The lethal time for 50% and 90% mortality (LT50 and LT90) values were calculated from regression line. RESULTS: The susceptibility status of lab strain of Cx. pipiens revealed that it is susceptible to Lambdacyhalothrin, Deltamethrin, Cyfluthrin and resistant to DDT. Moreover cyfluthrin with LT50=36 seconds and DDT with LT50=3005 seconds had the least and most LT50s. Field population was resistance to all tested insecticides and DDT yielded no mortality. CONCLUSION: Highly resistance level against all WHO recommended imagicides were detected in field populations. We suggest more biochemical and molecular investigations to detect resistance mechanisms in the field population for further decision of vector control.

Current Susceptibility Status of Anopheles stephensi (Diptera: Culicidae) to Different Imagicides in a Malarious Area, Southeastern of Iran.

BACKGROUND: Anopheles mosquitoes are an important group of arthropods due to their role in transmission of malaria. The present study was conducted for determination of susceptibility status of Anopheles stephensi to different imagicides collected from malarious area in Chabahar city, Iran. METHODS: In the present study seven insecticides including: DDT 4%, lambdacyhalothrin 0.05%, deltamethrin 0.05%, permethrin 0.75%, cyfluthrin 0.15% and etofenprox 0.5% were tested based on WHO method. Regression line was plotted for each insecticide using mortality of different exposure times. Bioassay data were analyzed using Probit software and the lethal time for 50% and 90% mortality (LT50 and LT90) values were calculated. RESULTS: The susceptibility levels of field strain of An. stephensi to the discriminative dose of different imagicides were determined 100, 98, 96, 89, 82 and 62% for etofenprox, permethrin, deltamethrin, lambdacyhalothrin, cyfluthrin and DDT, respectively. Our finding indicated that An. stephensi is resistant to DDT, lambdacyhalothrin and cyfluthrin, and susceptible to etofenprox and permethrin and candidate of resistant to deltamethrin based on WHO criteria. CONCLUSION: Our findings indicated that An. stephensi is resistant to DDT and some pyrethroid insecticides which can be developed due to application of insecticides in health and agriculture. These results can provide a clue for future chemical control program in the study area.

Resistance Mechanisms of Anopheles stephensi (Diptera: Culicidae) to Temephos.

BACKGROUND: Anopheles stephensi is a sub-tropical species and has been considered as one of the most important vector of human malaria throughout the Middle East and South Asian region including the malarious areas of southern Iran. Current reports confirmed An. stephensi resistance to temephos in Oman and India. However, there is no comprehensive research on mechanisms of temephos resistance in An. stephensi in the literature. This study was designed in order to clarify the enzymatic and molecular mechanisms of temephos resistance in this species. METHODS: Profile activities of α- and ß-esterases, mixed function oxidase (MFO), glutathione-S-transferase (GST), insensitive acetylcholinesterase, and para-nitrophenyl acetate (PNPA)-esterase enzymes were tested for An. stephensi strain with resistance ratio of 15.82 to temephos in comparison with susceptible strain. RESULTS: Results showed that the mean activity of α-EST, GST and AChE enzymes were classified as altered indicating metabolic mechanisms have considerable role in resistance of An. stephensi to temephos. Molecular study using PCR-RFLP method to trace the G119S mutation in ACE-1 gene showed lack of the mutation responsible for organophosphate insecticide resistance in the temephos-selected strain of An. stephensi. CONCLUSION: This study showed that the altered enzymes but not targets site insensitivity of ACE-1 are responsible for temephos resistance in An. stephensi in south of Iran.