Widespread resistance of insect pests to insecticides has been widely reported in China and there is consequently an urgent need to adjust pest management strategies appropriately. This requires detailed information on the extent and causes of resistance. The aim of the present study was to investigate levels of resistance to 5 insecticides among 12 strains of Culex tritaeniorhynchus, a major vector of Japanese encephalitis in China. Resistance to deltamethrin, beta-cypermethrin, permethrin, dichlorvos, and propoxur were measured using larval bioassays. The allelic frequency of knockdown resistance (kdr) and acetylcholinesterase (AChE) mutations were determined in all strains. Larval bioassay results indicated that the field strains collected from different sites were resistant to deltamethrin, beta-cypermethrin, permethrin, dichlorvos, and propoxur, with resistance ratio values ranging from 1.70- to 71.98-fold, 7.83- to 43.07-fold, 3.54- to 40.03-fold, 291.85- to 530.89-fold, and 51.32- to 108.83-fold, respectively. A polymerase chain reaction amplification of specific alleles method for individual was developed to detect genotypes of the AChE gene mutation F455W in Cx. tritaeniorhynchus. The frequency of the AChE gene mutation F455W was 100.00% in all strains, making this mutation of no value as a marker of resistance to organophosphorous and carbamate pesticides in Cx. tritaeniorhynchus in China. The kdr allele was present in all strains at frequencies of 10.00-29.55%. Regression analysis indicated a significant correlation between kdr allele frequencies and levels of resistance to deltamethrin, beta-cypermethrin, and permethrin. These results highlight the need to monitor and map insecticide resistance in Cx. tritaeniorhynchus and to adjust pesticide use to minimize the development of resistance in these mosquitoes.
Title: Thermophilic esterase from Thermomyces lanuginosus: molecular cloning, functional expression and biochemical characterization Li XJ, Zheng RC, Wu ZM, Ding X, Zheng YG Ref: Protein Expr Purif, 101:1, 2014 : PubMed
A novel esterase encoding gene, tle, was cloned from the thermophilic fungus Thermomyces lanuginosus DSM 10635. The tle had an open reading frame of 945bp encoding TLE of 314 amino acids with a theoretical molecular mass of 34.5kDa. The putative catalytic triad of TLE was consisted of Ser151, His279, and Asp249. TLE was heterologously expressed in Escherichia coli in biologically active form and purified to homogeneity. Several biochemical properties of TLE were studied: Among the tested p-nitrophenol esters, TLE showed the highest hydrolytic activity with p-nitrophenyl butyrate (C4) and exhibited the maximum activity at 60 degrees C and pH 8.5. The enzyme was stable at temperatures below 60 degrees C and retained 53% of the maximum activity after treatment at 70 degrees C for 60min. Esterase activity was notably enhanced by addition of Ca(2+) and Ba(2+), respectively. Furthermore, TLE showed high enantioselectivity (E=95) in the kinetic resolution of 2-carboxyethyl-3-cyano-5-methylhexanoic acid ethyl ester (CNDE), which produce a valuable chiral intermediate-(3S)-2-carboxyethyl-3-cyano-5-methylhexanoic acid for Pregabalin. These unique properties of the esterase indicate that TLE is a potential candidate for industrial application.
