The present study investigated effects of antifungal and carboxylesterase inoculant on rumen fermentation with different rumen pH. Corn silage was treated without inoculant (CON) and with a mixed Lactobacillus brevis 5M2 and L. buchneri 6M1 (MIX). Rumen fluid was collected from two cannulated Hanwoo heifers before morning feeding (high rumen pH at 6.70) and 3 h after feeding (low rumen pH at 6.20). Dried corn silage was incubated in the rumen buffer (rumen fluid + anaerobic culture medium at 1:2 ratio) for 48 h at 39oC. Eight replications for each treatment were used along with two blanks. Both in a high and a low rumen pH, MIX silages presented higher (p<0.05) the immediately degradable fraction, the potentially degradable fraction, total degradable fraction, and total volatile fatty acid (VFA) than those of CON silages. Incubated corn silages in a low rumen pH presented lower (p<0.05) total degradable fraction, ammonia-N, total VFA (p=0.061), and other VFA profiles except acetate and propionate, than those in a high rumen pH. The present study concluded that application of antifungal and carboxylesterase inoculant on corn silage could improve degradation kinetics and fermentation indices in the rumen with high and low pH conditions.

To identify whether higher expression of carboxylesterase (CbE) E4 in Myzus persicae is due to gene duplication, gene copy number was determined by quantitative real-time PCR. In addition, to determine the actual protein concentration of CbE E4 and it activity, Western blotting and activity staining were conducted. CbE gene copy number was highly correlated with carbamate resistance ratio (r2=0.934). However, CbE E4 expression level was little correlated with insecticide resistance ratio (r2<0.046) and no apparent correlation was observed among the gene copy number, protein quantity and total activity of CbE E4. Therefore, it was assumed that not only quantitative changing but also qualitative alteration of CbE E4 occurred in M. persicae. To investigate any potential alteration of CbE E4, mutation survey was conducted by sequencing of CbE E4 from various local strains of M. persicae. G137D and W251L mutations have been known as the main mutations associated with structural change leading to resistance. Interestingly, a new G134C mutation, which is in proximity of G137D mutation, was identified in the oxyanion hole of CbE E4. To predict the functional role of this mutation in resistance, 3-dimensional structure modeling was conducted. In summary, CbE E4 appears to be involved in resistance to both pyrethroids and carbamates as a nonspecific hydrolase or sequestration protein in M. persicae.

The gene encoding an esterase enzyme was cloned from a metagenomic library of cow rumen bacteria. The esterase gene (est2R) was 2,120 bp in length, encoding a protein of 516 amino acid residues with a calculated molecular weight of 57,286 Da. The molecular weight of the enzyme was estimated to be 57,000 Da by SDS-PAGE. Est2R shared 35.6% amino acid identity with esterase (CAH19079) of uncultured prokaryote. The Est2R was most active at 20-40°C, and showed optimum at 30°C and pH 8.0. The most activity of Est2R for the different chain length of p-nitrophenyl ester group as substrate was p-nitrophenyl acetate. Moreover, the enzyme was found to be most active without organic solvent, followed by 98% active with ethanol, and the enzyme activity was highly affected by the acetonitrile. The enzyme was significantly inhibited by Zn2+ but stimulated by Ca2+. So, novel esterase gene est2R is likely to obtain from cow rumen metagenome and supposed to use for industrial purpose.

The hemipteran whitefly Bemisia tabaci (Gennadius) is one of the most destructive pests damaging more than 600 agricultural crop species worldwide. The B and Q biotypes are most widely spread in Korea but they are not distinguishable based on morphological characters. Carboxylesterase 2 (Coe2) was determined to be 5.9 times more expressed in B biotype compared to Q biotype. Comparison of deduced amino acid sequences of Coe2 (595 a.a.) showed a total of 3.85% polymorphisms between B and Q types but no differences in major active sites. Quantitative real-time PCR revealed that both B and Q biotypes possess a single copy of coe2, suggesting that the overexpression of Coe2 in B biotype is likely due to overtranscription. To determine the putative role of Coe2 in insecticide tolerance, esterases were separated by native isoelectric focusing (IEF) and inhibited by various insecticides. The putative Coe2 band was apparently inhibited by pyrethroid and organophosphate insecticides, but not by imidacloprid. These findings suggest that overexpression of Coe2 confers chemical defense against pyrethroid and organophosphate insecticides, perhaps by sequestration.

포장의 약제저항성 변동요인을 조사하기 위하여 복수아혹진딧물의 Carboxyl esterase(CE) 활서측정에 의한 저항성도의 변동상황을 조사하였다. 비닐하우스에서 고추육묘중 발생한 복수아혹진딧물의 CE활성 측정 결과 약제저항성도는 무처리구가 40이였으나 아세페이트를 처리한구는 78로 약제처리로 인한 저항성도가 38이 증가하였다. 또한 노천망실에서 약제처리를 하지 않고 재배한 케일에 발생한 복숭아혹진딧물의 CE활성에 의한 저항성도는 7월이 24이었으나, 8, 9월의 활성이 계속 증가하여 10월이 83으로서 최고에 달하였다가 11월에는 다시 81에서 79로 약천 떨어지고 있어 약제이외에 저항성도의 자연변동요인이 있음을 확인하였다. 전국적으로 18개장소에서 채집한 진딧물의 CE활성을 측정한 결과 저항성도의 평균이 여름은 5014였으며 늦가을인 11월은 8210으로서 여름보다 늦가을인 11월이 32가 높았으며 그중에도 약제를 살포사지 않은 곳 보다 약제살포 회수가 많은 곳일수록 높았다.