Assessment of the Bemisia tabaci C YP6CM l vQ transcript and protein levels in laboratory and field-derived imidacloprid-resistant insects and cross-metabolism potential of the recombinant enzyme

Over-expression of the cytochrome P450 CYP6CM1 gene has been associated with imidacloprid resistance in a number of Q and B biotype Bemisia tabaci laboratory strains from distinct geographical origins worldwide. We recently demonstrated that the Q biotype version of the CYP6CM 1 protein （CYP6CMlvQ） is capable of metabolizing imida- cloprid. Here, we show that the levels of BtCYP6CMlvQ were also elevated in laboratory- resistant strains and field-derived populations, with variable imidacloprid resistance levels, collected in Crete. High levels of CYP6CMlvQ transcripts were also determined in survivors of a heterogeneous field population, after exposure to discriminating imidacloprid dosage. Using peptide antibody-based detection assays, we demonstrated that in line with transcriptional data, the CYP6CMlvQ protein levels were higher in imidacloprid-resistant insects, which further implicates the gene as the causal factor of resistance. Finally, assess- ment of the cross-metabolism potential of CYP6CMlvQ against additional neonicotinoid molecules used for B. tabaci control revealed that clothianidin and thiacloprid, but not acetamiprid or thiamethoxam, are metabolized by the recombinant enzyme in vitro.

Resurgence of the cotton bollworm Helicoverpa armigera in northern Greece associated with insecticide resistance

Helicoverpa armigera has been controlled effectively with chemical insec- ticides in the major cotton crop production areas of northern Greece for many years. However, a resurgence of the pest was observed in 2010, which significantly affected crop production. During a 4-year survey （2007-2010）, we examined the insecticide re- sistance status of 1-1. armigera populations from two major and representative cotton production areas in northern Greece against seven insecticides （chlorpyrifos, diazinon, methomyl, alpha-cypermethrin, cypermethrin, gamma-cyhalothrin and endosulfan）. Full dose-response bioassays on third instar larvae were performed by topical application. Lethal doses at 50% were estimated by probit analysis and resistance factors （RF） were calculated, compared to a susceptible laboratory reference stra^a. Resistance levels were relatively moderate until 2009, with resistance ratios below 10-fold for organophosphates and carbamates and up to 16-fold for the pyrethroid alpha-cypermethrin. However, resis- tance rose to 46- and 81-fold for chlorpyrifos and alpha-cypermethrin, respectively in 2010, when the resurgence of the pest was observed. None of the known pyrethroid resistance mutations were found in the pyrethroid-resistant insects. The possible association between resistance and H. armigera resurgence in Greece is discussed.

Insecticide resistance in Bemisia tabaci from Cyprus

A comprehensive study on the Bemisia tabaci （biotype B） resistance to neonicotinoid insecticides imidacloprid, acetamiprid and thiamethoxam, and pyrethroid bifenthrin was conducted in Cyprus. The resistance level to eight field-collected B. tabaci populations was investigated. The activities of enzymes involved in metabolic detoxification and the frequencies ofpyrethroid and organophosphates target site resistance mutations were determined. Moderate to high levels of resistance were detected for imidacloprid （re- sistance factor [RF] 77-392） and thiamethoxam （RF 50-164） while low resistance levels were observed for acetamiprid （RF 7-12）. Uniform responses by the Cypriot whiteflies could be observed against all neonicotinoid insecticides. No cross-resistance between the neonicotinoids was detected as well as no association with the activity of the P450 microsomal oxidases. Only imidacloprid resistance correlated with carboxylesterase activity. Low to extremely high resistance was observed for insecticide bifenthrin （RF 49 1 243） which was associated with the frequency of the resistant allele in the sodium channel gene but not with the activity of the detoxification enzymes. Finally, the F331W mutation in the acetylcholinesterase enzyme acel gene was fixed in all B. tabaci populations from Cyprus.

Population dynamics, pathogen detection and insecticide resistance of mosquito and sand fly in refugee camps, Greece

Background:As of 2015 thousands of refugees are being hosted in temporary refugee camps in Greece.Displaced populations,travelling and living under poor conditions with limited access to healthcare are at a high risk of exposure to vector borne disease(VBD).This study sought to evaluate the risk for VBD transmission within refugee camps in Greece by analyzing the mosquito and sand fly populations present,in light of designing effective and efficient context specific vector and disease control programs.Methods:A vector/pathogen surveillance network targeting mosquitoes and sand flies was deployed in four temporary refugee camps in Greece.Sample collections were conducted bi-weekly during June-September 2017 with the use of Centers for Disease Control(CDC)light traps and oviposition traps.Using conventional and molecular diagnostic tools we investigated the mosquito/sand fly species composition,population dynamics,pathogen infection rates,and insecticide resistance status in the major vector species.Results:Important disease vectors including Anopheles sacharovi,Culex pipiens,Aedes albopictus and the Leishmania vectors Phlebotomus neglectus,P.perfiliewi and P.tobbi were recorded in the study refugee camps.No mosquito pathogens(Plasmodium parasites,flaviviruses)were detected in the analysed samples yet high sand fly Leishmania infection rates are reported.Culex pipiens mosquitoes displayed relatively high knock down resistance(kdr)mutation allelic frequencies(ranging from 41.0 to 63.3%)while kdr mutations were also detected in Ae.albopictus populations,but not in Anopheles and sand fly specimens.No diflubenzuron(DFB)mutations were detected in any of the mosquito species analysed.Conclusions:Important disease vectors and pathogens in vectors(Leishmania spp.)were recorded in the refugee camps indicating a situational risk factor for disease transmission.The Cx.pipiens and Ae.albopictus kdr mutation frequencies recorded pose a potential threat against the effectiveness of pyrethroid insecticides in these settings.In contrast,pyrethroids appear suitable for the control of Anopheles mosquitoes and sand flies and DFB for Cx.pipiens and Ae.albopictus larvicide applications.Targeted actions ensuring adequate living conditions and the establishment of integrated vector-borne disease surveillance programs in refugee settlements are essential for protecting refugee populations against VBDs.