Esterase Profiles Research Articles

Study of vigor and esterase activity under NaCl stress deciphering overlap between gametophyte and sporophyte in annual Medicago species

To study the overlapping of gene expression between sporophyte and gametophyte phases, three annual species of the Medicago genus; M. truncatula, M. ciliaris and M. aculeata were used. First, we measured the germination rate of seeds and pollen grains as well as the length of young seedlings and the length of the pollen tubes under different levels of NaCl concentrations (0 mM, 68 mM, 102 mM and 136 mM). Afterwards, we measured the biochemical activity of plants based on electrophoresis of esterase enzymatic activity under salt stress treatments. The results showed that genotype M.acl4 of M. aculeate species is the most tolerant in the two stages, while the genotypes M.cil 1 and M.trur1 are the most sensitive of M. ciliaris and M. truncatula species, respectively. The separation of the esterase activities by electrophoresis revealed four bands with quantitative and qualitative changes in relation to stress in the same sensitive genotype M.cil1 of M. ciliaris species. The presence of a correlation between measured traits like vigor, as well as the connection between esterase profiles during periods of salt stress in both the sporophyte and gametophyte stages, may unveil a shared pattern of gene expression.

Virulence and Reproductive Fitness of Meloidogyne paranaensis Field Populations in Coffee Genotypes

The aim of this study was to characterize Meloidogyne paranaensis populations collected from infested coffee crops. Methodologies used to characterize the 11 studied populations from municipalities in Paraná and Minas Gerais States involved the morphological analysis of perineal patterns, biochemical analysis by isozyme electrophoresis, sequencing of internal transcribes spacer 1 (ITS-1) and D2/D3 ribosomal DNA (rDNA) regions, reproductive fitness, and virulence characterization in coffee genotypes. Morphological evaluations showed the existence of variation between populations, although the majority of them showed typical perineal patterns. The biochemical identification was based on α-esterase isozyme analyses and resulted in the appearance of three distinct profiles: P1 (typical), P2 (atypical), and a nondescribed profile, P2b. BLAST of the ITS-1 and D2/D3 rDNA regions indicated homology (>95%) with other sequences deposited in GenBank. For reproductive fitness and virulence characterization, 13 coffee genotypes (5 Coffea arabica and 8 C. canephora) were inoculated with 11 M. paranaensis populations. Variation in the reproductive fitness of populations was observed for cultivar Mundo Novo, a genotype without resistance genes, and variation in the virulence of populations was observed in genotypes carrying resistance genes. Three populations exhibited virulence combined with high reproductive fitness, while one showed virulence with low reproductive fitness. Some hosts were resistant to 11 populations, while one of the hosts was resistant to only one population, indicating the presence of different resistance genes. Nevertheless, no relationship was observed between the origin of population and their variations in perineal patterns, esterase profiles, phylogeny, or reproductive fitness in coffee genotypes, or between the different characterizations, although differences were observed within each characteristic.

Identification of Two Novel Pathovars of Pantoea stewartii subsp. indologenes Affecting Allium sp. and Millets.

Pantoea stewartii subsp. indologenes is a causative agent of leafspot of foxtail millet and pearl millet; however, novel strains were recently identified that are pathogenic on onion. We phenotypically and genotypically characterized 17 P. stewartii subsp. indologenes strains from onion and other sources (pearl millet, foxtail millet, guar pulse, verbena, and corn). Based on the host range evaluation, we propose two pathovars: P. stewartii subsp. indologenes pv. cepacicola pv. nov. and P. stewartii subsp. indologenes pv. setariae pv. nov. P. stewartii subsp. indologenes pv. cepacicola pv. nov. causes symptoms on Allium spp. (leek, onion, chive, and Japanese bunching onion) and on foxtail millet, pearl millet, and oat. However, P. stewartii subsp. indologenes pv. setariae pv. nov. can only infect the members of Poaceae family (foxtail millet, pearl millet, and oat). We also propose that the type strain of P. stewartii subsp. indologenes (LMG 2632T) should be designated as a pathotype strain of P. stewartii subsp. indologenes pv. setariae and recommend that the strain PNA 14-12 be designated as the pathotype strain of P. stewartii subsp. indologenes pv. cepacicola. The digital DNA-DNA hybridization (dDDH), average nucleotide identity (ANI), and multilocus sequence analysis study showed that the two pathovars are genotypically closely related. Our study also showed that P. stewartii subsp. indologenes pathovars and P. stewartii subsp. stewartii share high genotypic relatedness and cannot be differentiated by dDDH and ANI values. Although the newly proposed pathovars are not clearly distinguishable by their fatty acid and methyl esterase profiles and substrate use patterns, a fatty acid (unknown with retention time: 10.9525) and a few metabolites (3-methyl glucose, Na butyrate, and fusidic acid) can be potentially used to distinguish them. We also report the distribution of previously known pathogenicity (HiVir, hrcC) and virulence (alt) factors of Pantoea spp. in the new pathovars. The impact of these new pathovars in the center rot pathosystem of onion is yet to be determined.

Bio-molecular characterization of stress enzyme profile on esterase in selected silkworm races of Bombyx mori (L.) for biomarker selection

Silkworm Bombyx mori exhibits for eliminating pathogenic defenses by their secretory component of haemolymph and gut component. This study aimed to investigate the stress enzyme role of carboxyl esterase, plays a major function in toxic component detoxification by their lipid assembly. Hence, the study has planned to identify their carboxyl esterase role in haemolymph of six silkworm races using innovative tools. The six silkworm races are characterized based on their esterase profiles and their morphological origins. It was concluded that Nistari, Kollegal Jawan and Hosa Mysore are considered as tolerant races and remaining races are susceptible by eliminating the toxic component from their body. Therefore, the present study could be an index for identifying the esterase role and used as a biomarker for the screening of a wide range of silkworm races for identifying genetic hardiness.

Determination of intracellular pH in phytoplankton using the fluorescent probe, SNARF, with detection by fluorescence spectroscopy

The maintenance of pH homeostasis is critical for a variety of cellular metabolic processes. Although ocean acidification is likely to influence cellular metabolism and energy balance, the degree to which intracellular pH in phytoplankton differs from the external environment under varying environmental pH levels is not well characterized. While there are numerous existing methods for the determination of intracellular pH in the form of single peak emission (e.g., BCECF) and radioisotopic (e.g., 14C-DMO) indicators for use with phytoplankton, the fluorescent pH indicator seminaphtharhodafluor (SNARF) has not been established as a robust method for measuring in vivo pH in phytoplankton. SNARF has superior accuracy and sensitivity since it exhibits dual emission peaks from a single excitation wavelength and the ratio of the two are related to pH. The use of a ratio limiting variations in fluorescence due to dye loading, photobleaching, and instrument variation; moreover, like other fluorescence-based assays, it does not require the specialized equipment and permits that radioisotopic methods do. As a first step, we tested the performance of SNARF for measuring intracellular pH in vivo in a number of phytoplankton taxa. SNARF detection was accomplished using fluorescence spectroscopy (FS) and laser scanning microscopy (LSM). Since SNARF fluorescence is activated by cleavage of an ester group from the core fluorophore by non-specific esterases, we measured esterase activity using fluorescein diacetate (FDA) to characterize variability in esterase activity among phytoplankton taxa, with a view towards its influence on assay performance. Esterase activity cell volume; however, there was no indication that enzyme specificity and differences in individual esterase profiles adversely affected SNARF performance in phytoplankton. Assays of intracellular pH using SNARF were comparable to those made with 14C-labeled DMO, an accepted standard method. Thus, SNARF provides robust measurements of intracellular pH in phytoplankton, constituting a useful tool in investigations of the effects of ocean acidification and fluctuations in environmental pH on cellular physiology.

Flora surrounding rice fields as a source of alternative prey for coccinellids feeding on the pests of rice

Coccinellids are effective predators and a key component of the predator guild in rice ecosystems. In order to enhance their efficacy, a study was undertaken to assess the seasonal movement of coccinellids into rice fields and the role of the surrounding flora on their colonization. The seasonal abundance of coccinellids and their prey was recorded on the rice crop and the surrounding flora at fortnightly intervals from 2012 to 2015. Coccinellid prey range was assessed using PAGE electrophoresis. The herbivorous insects associated with weeds were Aphis gossypii Glover, Aphis craccivora (Koch), Cicadulina bipunctata (Melichar), Schizaphis graminum (Rondani), Sitobion sp., Thaia oryzivora Ghauri and Zygina maculifrons Matsumura. Of the species of coccinellids recorded in rice fields, Harmonia octomaculata (Fabricius), Micraspis discolor (F.), Propylea dissecta (Mulsant), Coccinella transversalis Fabricius, Cheilomenes sexmaculata (Fabricius), Scymnus nubilus Mulsant and Brumoides suturalis (Fabricius) were also recorded on weeds. The esterase profiles indicated that the leafhoppers and aphids on the weeds were the prey of the coccinellids before they colonized the rice fields. The coccinellids recorded on the weeds showed bands corresponding to the insects present on the weeds. Beetles collected from rice fields had different bands, some of which corresponded to the green leafhopper (GLH) Nephotettix virescens Distant, the brown planthopper (BPH) Nilaparvata lugens Stal and white backed planthopper (WBPH) Sogatella furcifera Hovarth infesting rice. In addition, some bands corresponded to hoppers and aphids that were present on the surrounding flora. The results indicate the importance of surrounding flora in the conservation and colonization of rice fields by coccinellids.

Investigation of the Esterase Status as a Complex Biomarker of Exposure to Organophosphorus Compounds

Development of biomarkers of human exposures to organophosphorus compounds OPCs and their quantification is a vital component of a system of prediction and early diagnostics of OPC-induced diseases. Our study was focused on investigation of esterase status as a complex biomarker of exposure to OPCs and an aid in accurate diagnosis. We suggest that this complex biomarker should be more effective and informative than standard assays of plasma butyrylcholinesterase (BChE), erythrocyte acetylcholinesterase (RBC AChE), and lymphocyte neuropathy target esterase (NTE). It will help: 1) to assess an exposure as such and to confirm the nonexposure of individuals suspected to have been exposed; 2) to determine if the exposure was to agents expected to produce acute and/or delayed neurotoxicity; 3) to perform dosimetry of the exposure, which provides valuable information for medical treatment. To confirm this hypothesis, we have examined the changes in activity of blood AChE, NTE, BChE and carboxylesterase (CaE) 1 h after i.p. administration of increasing doses of three OPCs with different esterase profiles: the known neuropathic compound O,O-dipropyl-O-dichlorovinyl phosphate (C3H 7O)2P(O)OCH=CCl2 (diPr-DClVP) as the control compound and two model dialkylphosphates (C2H5O)2P(O)OCH(CF3)2 (diEt-PFP) and (C4H9O)2P(O)OCH(CF3)2 (diBu-PFP). The esterases assay was performed in hemolysed blood by spectrophotometric (AChE, BChE, CaE) and biosensor (NTE) methods. Analysis of the obtained dose-dependences for blood esterases inhibition showed that blood BChE and CaE were the most sensitive biomarkers, allowing detection of low doses. Inhibition of blood NTE and AChE can be used to assess the likelihood that an exposure to OPC would produce cholinergic and/or delayed neuropathic effects.

Phenotypical characterization and molecular identification of clinical isolates of Candida tropicalis

BackgroundCandida tropicalis is an increasingly important human pathogen which usually affects neutropenic oncology patients with common hematogenous seeding to peripheral organs and high mortality rates. Candida pathogenicity is facilitated by several virulence attributes, including secretion of hydrolytic enzymes; however, little is known regarding the C. tropicalis ability to secrete them and their role in the disease. AimsTo confirm by molecular means the identification of 187 clinical isolates (127 from blood, 52 from urine, and 8 from diverse clinical origins) phenotypically identified as C. tropicalis, and to investigate their in vitro aspartyl proteinase, phospholipase, esterase, hemolysin, DNase and coagulase activities. MethodsThe molecular confirmation was performed by ITS sequencing, and the enzymatic determinations were conducted using plate assays with specific substrates, with the exception of coagulase, which was determined by the classical tube test. ResultsThe majority of the strains exhibited a very strong or strong activity of aspartyl proteinase, phospholipase and esterase. A 4.7% of the bloodstream isolates were hemolysin producers, and all were negative for the coagulase and DNase assays. ConclusionsVery strong activities of aspartyl proteinase, phospholipase and esterase profiles were detected, and a statistical association between phospholipase production and blood and urine isolates was found.

9-Substituted acridine derivatives as acetylcholinesterase and butyrylcholinesterase inhibitors possessing antioxidant activity for Alzheimer's disease treatment

We investigated the inhibitory activity of 4 groups of novel acridine derivatives against acetylcholinesterase (AChE), butyrylcholinesterase (BChE) and carboxylesterase (CaE) using the methods of enzyme kinetics and molecular docking. Antioxidant activity of the compounds was determined using the 2,2′-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS+) radical decolorization assay as their ability to scavenge free radicals. Analysis of the esterase profiles and antiradical activities of the acridine derivatives showed that 9-aryl(heteroaryl)-N-methyl-9,10-dihydroacridines have a high radical-scavenging activity but low potency as AChE and BChE inhibitors, whereas 9-aryl(heteroaryl)-N-methyl-acridinium tetrafluoroborates effectively inhibit cholinesterases but do not exhibit antiradical activity. In contrast, a group of derivatives of 9-heterocyclic amino-N-methyl-9,10-dihydroacridine has been found that combine effective inhibition of AChE and BChE with rather high radical-scavenging activity. The results of molecular docking well explain the observed features in the efficacy, selectivity, and mechanism of cholinesterase inhibition by the acridine derivatives. Thus, in a series of acridine derivatives we have found compounds possessing dual properties of effective and selective cholinesterase inhibition together with free radical scavenging, which makes promising the use of the acridine scaffold to create multifunctional drugs for the therapy of neurodegenerative diseases.

GENDER SPECIFIC CHANGES IN MOUSE BLOOD ESTRERASE PROFILE AT THE ACUTE INTOXICATION BY 2-(О-CRESYL)-4Н-1,3,2-BENZODIOXAPHOSPHORIN-2-OXIDE

A comparative investigation of the esterase profiles of blood of mice of both genders exposed to various doses of 2-(o-crezyl)-4H-1,3,2-benzodioxaphosphorin-2-oxide (CBDP) was made an hour after its percutaneous injection to animals. A lesser amount of esterase in males blood made them more susceptible to CBDP action as compared to females. It was shown that CBDP equally inhibits the activity of carboxyl esterase and butyryl choline esterase in blood serum of both male and female mice. Statistically significant differences in inhibition degree of enzymes between males and females were found out and therefore it is not recommended to use mixed groups of animals when performing testing of serine esterase inhibitors.

Esterase profiles of organophosphorus compounds in vitro predict their behavior in vivo

We studied 4 serine esterases (EOHs) that are associated with the following consequences from their inhibition by organophosphorus compounds (OPCs): acetylcholinesterase (AChE: acute neurotoxicity; cognition enhancement), butyrylcholinesterase (BChE: inhibition of drug metabolism and/or stoichiometric scavenging of EOH inhibitors; cognition enhancement), carboxylesterase (CaE; inhibition of drug metabolism and/or stoichiometric scavenging of EOH inhibitors), and neuropathy target esterase (NTE: delayed neurotoxicity, OPIDN). The relative degree of inhibition of these EOHs constitutes the “esterase profile” of an OPC, which we hypothesize can serve as a predictor of its overall physiological effects. To test this hypothesis, we selected 3 OPCs known from previous work on reference enzymes to span a wide range of esterase profiles, neuropathic potential, and acute cholinergic toxicity. For each compound, we determined in vitro IC50 and in vivo ED50 values for inhibition of AChE, BChE, CaE, and NTE in mouse brain and blood. The results showed good correlations between in vitro and in vivo measures of potency and selectivity except for brain CaE, a tissue-specific isoform of the enzyme that was less sensitive to the test compounds than expected. Thus, this synthesis of new and previously published results indicates that the concept of the esterase profile of OPCs is useful for the prediction of therapeutic and toxic effects in vivo.

Esterase profiles of hexafluoropropan-2-ol-based dialkyl phosphates as a major determinant of their effects in mouse brain in vivo

The relationship between the esterase profiles of two O-phosphorylated hexafluoropropan2-ols and their inhibitory activities and selectivities against mouse brain esterases was studied on brain preparations and in the whole animals. The predictions made from the analysis of esterase profiles of the studied organophosphorus compounds were found to be in complete agreement with their in vivo inhibitory activities against acetylcholinesterase, butyrylcholinesterase, and neuropathy target esterase.

Discrimination of Alicyclobacillus Strains by Lipase and Esterase Fingerprints

Alicyclobacillus spp. have recently become a major issue in the fruit juice industry due to its implication in the spoilage of pasteurized juices/beverages. In this study, lipase and esterase fingerprints of 37 Alicyclobacillus strains and 14 Bacillus strains in apple juice were collected using a five-substrate cocktail including four C8-esters and one C4-diester. Characterization of each strain could be finished in a single assay followed by high-performance liquid chromatography (HPLC) analysis for product separation and quantification. The obtained lipase and esterase fingerprints were highly species-dependent. Differences between the relative peak areas of each product for different species were significant. Coupled to chemometric techniques including principal component analysis (PCA) and stepwise linear discriminant analysis (SLDA), the lipase and esterase profiles led to 100 % of correct species identification for Alicyclobacillus and Bacillus strains. This approach is promising, reliable, and with good repeatability and could be used as an alternative tool to discriminate Alicyclobacillus spp. in fruit juices.

Analysis of genetic diversity within and among four rabbit genotypes using biochemical and molecular genetic markers

Genetic variations were detected in four different rabbit genotypes; Animal Production Research Institute (APRI) line, New-Zealand White (NZW), Baladi Black (BB) and Gabali (GAB) breeds. Ten individuals from the first three genotypes and seven individuals from GAB, were blood sampled. Isozymes (esterase and peroxidase) and sodium dodecyl sulphate (SDS)-protein markers were used to detect the genetic variations within these genotypes, whereas the random amplified polymorphic DNApolymerase chain reaction (RAPD-PCR) analysis using six random arbitrary primers were employed to assess the genetic variations and phylogenetic relationships among the four genotypes. The results show a variation in biochemical activity levels since, esterase profiles showed higher percentage of polymorphism (67.21%) than peroxidase (34.78%) and SDS-protein profiles (39.11 %). Moreover, the mean of the genetic similarity within the genotypes based on overall biochemical markers were 0.81 (APRI), 0.91 (NZW), 0.89 (BB) and 0.86 (GAB), which indicates high homogeneity within each of the four genotypes. The polymorphism percentage based on overall RAPD primers was 35.44% among these genotypes, which presents a kind of genetic diversity. From the dendrogram tree, the genotypes BB and GAB appeared to be closely related, while the APRI genotype was the most different. Keywords : Rabbits, genetic diversity, sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDSPAGE), isozymes, random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) African Journal of Biotechnology Vol. 12(20), pp. 2830-2839

Organophosphorus compound esterase profiles as predictors of therapeutic and toxic effects

Certain organophosphorus compounds (OPCs) inhibit various serine esterases (EOHs) via phosphorylation of their active site serines. We focused on 4 EOHs of particular toxicological interest: acetylcholinesterase (AChE: acute neurotoxicity; cognition enhancement), butyrylcholinesterase (BChE: inhibition of drug metabolism and/or stoichiometric scavenging of EOH inhibitors; cognition enhancement), carboxylesterase (CaE: inhibition of drug metabolism and/or stoichiometric scavenging of EOH inhibitors), and neuropathy target esterase (NTE: delayed neurotoxicity, OPIDN). The relative degree of inhibition of these EOHs constitutes the “esterase profile” of an OPC and serves as a major determinant of its net physiological effects. Thus, understanding and controlling the esterase profile of OPC activity and selectivity toward these 4 target enzymes is a significant undertaking. In the present study, we analyzed the inhibitor properties of 52 OPCs against the 4 EOHs, along with pairwise and multitarget selectivities between them, using 2 QSAR approaches: Hansch modeling and Molecular Field Topology Analysis (MFTA). The general formula of the OPCs was (RO)2P(O)X, where R=alkyl, X=– SCH(Hal)COOEt (Hal=Cl, Br), –SCHCl2, –SCH2Br, –OCH(CF3)R1 (R1=C6H5, CF3, COOEt, COOMe). The Hansch model showed that increasing neuropathic potential correlated with rising R hydrophobicity; moreover, OPC binding to scavenger EOHs (BChE and CaE) had different effects on potential acute and delayed neurotoxicity. Predicted protective roles of BChE and CaE against acute toxicity were enhanced with increasing hydrophobicity, but projected protection against OPIDN was decreased. Next, Molecular Field Topology Analysis (MFTA) models were built, considering atomic descriptors, e.g., effective charge, van der Waals radius of environment, and group lipophilicity. Activity/selectivity maps confirmed predictions from Hansch models and revealed other structural factors affecting activity and selectivity. Virtual screening based on multitarget selectivity MFTA models was used to design libraries of OPCs with favorable esterase profiles for potential application as selective inhibitors of CaE without untoward side effects.

Combined QSAR studies of inhibitor properties of O-phosphorylated oximes toward serine esterases involved in neurotoxicity, drug metabolism and Alzheimer's disease

Oxime reactivation of serine esterases (EOHs) inhibited by organophosphorus (OP) compounds can produce O-phosphorylated oximes (POXs). Such oxime derivatives are of interest, because some of them can have greater anti-EOH potencies than the OP inhibitors from which they were derived. Accordingly, inhibitor properties of 58 POXs against four EOHs, along with pair-wise selectivities between them, have been analysed using different QSAR approaches. EOHs (with their abbreviations and consequences of inhibition in parentheses) comprised acetylcholinesterase (AChE: acute neurotoxicity; cognition enhancement), butyrylcholinesterase (BChE: inhibition of drug metabolism or stoichiometric scavenging of EOH inhibitors; cognition enhancement), carboxylesterase (CaE: inhibition of drug metabolism or stoichiometric scavenging of EOH inhibitors), and neuropathy target esterase (NTE: delayed neurotoxicity). QSAR techniques encompassed linear regression and backpropagation neural networks in conjunction with fragmental descriptors containing labelled atoms, Molecular Field Topology Analysis (MFTA), Comparative Molecular Similarity Index Analysis (CoMSIA), and molecular modelling. All methods provided mostly consistent and complementary information, and they revealed structural features controlling the ‘esterase profiles’, i.e. patterns of anti-EOH activities and selectivities of the compounds of interest. In addition, MFTA models were used to design a library of compounds having a cognition-enhancement esterase profile suitable for potential application to the treatment of Alzheimer's disease.

Biological characterization of two Bacillus thuringiensis strains toxic against Spodoptera frugiperda

Two Bacillus thuringiensis strains isolated from diseased Spodoptera frugiperda larvae collected in the northwest of Argentina were molecularly and phenotypically characterized. Insecticidal activity against Spodoptera frugiperda larvae was also determined. Both strains were highly toxic against first instar larvae. One strain (Bacillus thuringiensis LSM) was found to be even more toxic than the reference strain Bacillus thuringiensis var. kurstaki 4D1. This strong biological effect was represented by both a higher mortality which reached 90%, and a shorter LT50. Molecular characterization showed that Bacillus thuringiensis LSM carried a cry gene profile identical to that of Bacillus thuringiensis var. kurstaki 4D1. Evaluation of length polymorphism of the intergenic transcribed spacers between the 16S and 23S rDNA genes revealed an identical pattern between native strains and Bacillus thuringiensis var. kurstaki 4D1. In contrast, phenotypic characterization allowed differentiation among the isolates by means of their extracellular esterase profiles. Lytic activity that would contribute to Bacillus thuringiensis effectiveness was also studied in both strains. Analyses like those presented in the current study are essential to identify the most toxic strains and to allow the exploitation of local biodiversity for its application in biological control programmes.

Genetics and Mechanisms of Permethrin Resistance in the Santa Luiza Strain of <I>Boophilus microplus</I> (Acari: Ixodidae)

The Santa Luiza strain of the southern cattle tick, Boophilus microplus (Canestrini) (Acari: Ixodidae), is resistant to both permethrin and amitraz. A study was conducted at the USDA Cattle Fever Tick Research Laboratory in Texas to investigate the genetic basis of permethrin resistance with cross-mating experiments, and to determine the mechanisms of permethrin resistance through synergist bioassays and biochemical analysis of esterase profiles. The Muñoz strain, an acaricide-susceptible reference strain, was used as the susceptible parent and the Santa Luiza strain, originating in Brazil, was used as the resistant parent. The Food and Agriculture Organization larval packet test was used to measure the levels of susceptibility of larvae of the parental strains, F1, backcross, F2, and F3 generations to permethrin. Results of reciprocal crossing experiments suggested that permethrin resistance was inherited as an incomplete recessive trait. There was no significant maternal effect on larval progeny's susceptibility to permethrin in the F1 and subsequent generations. The values of the degree of dominance were estimated at -0.700 and -0.522 for the F1 larvae with resistant and susceptible female parents, respectively. Results of bioassays on larval progeny of the F1 backcrossed with the resistant parent strain and of the F2 generations suggested that one major gene was responsible for permethrin resistance in the Santa Luiza strain. Selection of F3 larvae with either permethrin or amitraz led to significantly increased resistance to both permethrin and amitraz, indicating a close linkage between genes responsible for permethrin and amitraz resistance. The possible involvement of metabolic enzymes in permethrin resistance in the Santa Luiza strain of B. microplus was dismissed by the lack of enhanced synergism by TPP or PBO, as observed in synergist bioassays, as well as by the lack of enhanced esterase activity in the Santa Luiza strain relative to the susceptible strain. The results of this study suggest that other mechanisms, including a possible new sodium channel mutation that is different from the one currently known, may be responsible for permethrin resistance in the Santa Luiza strain of B. microplus.

A novel phenotype based on esterase electrophoretic polymorphism for the differentiation of Lactococcus lactis ssp. lactis and cremoris

To evaluate the esterase phenotype in Lactococcus lactis strains isolated from traditional Tunisian dairy products. A collection of 55 L. lactis strains isolated from traditional fermented milk products and three reference strains were identified at species and subspecies level using molecular methods targeted to the 16S rRNA gene and to the histidine operon. The genotypic data obtained allowed the identification of the strains as L. lactis ssp. lactis and L. lactis ssp. cremoris with the prevalence of the ssp. lactis. The phenotypic identification based on arginine hydrolysis, the growth at 40 degrees C and in presence of 4% NaCl showed several discrepancy with the identification data based on genotypic analysis. Additional experiments carried out evaluating the esterase electrophoretic patterns revealed four classes of esterases identified on the basis of their electrophoretic mobility and specific activity on alpha- and beta-naphthyl ester of acetate and propionate. Esterase profiles discriminated the strains in two main groups corresponding to the subspecies cremoris and lactis according to a DNA-based identification. The evaluation of esterase activity represents a novel phenotype for the taxonomic discrimination of the L. lactis ssp. lactis and cremoris. Besides the DNA-based techniques that allow the rapid and accurate species/subspecies identification, the electrophoretic esterase profiles of L. lactis strains represents: (i) a new phenotypic tool to understand the physiology and the ecology of this species; and (ii) a new test for the potential selection of flavour producing strains.

Detection of Gonatocerus ashmeadi (Hymenoptera: Mymaridae) parasitism of Homalodisca coagulata (Homoptera: Cicadellidae) eggs by polyacrylamide gel electrophoresis of esterases

Abstract Gonatocerus ashmeadi is an important egg parasitoid of the glassy-winged sharpshooter, Homalodisca coagulata , in California. Although adults of this parasitoid are easily identified from morphological characteristics, their identification as immatures within eggs is more difficult, particularly during the early stages of development. In this study, we show that esterase profiles determined by polyacrylamide gel electrophoresis (PAGE) can be used to detect this species in parasitized eggs within two days of parasitization. The expression of esterase E 0.17 occurred at a time when mymarid parasitoids would be expected to hatch within the host egg. The intensity of this band increased as the parasitoid developed within the sharpshooter egg. After five days, a second parasitoid esterase, E 0.13 , was detected. These two esterases were clearly distinct from all sharpshooter esterases. The electrophoretic technique was also sensitive enough to detect a polymorphic esterase locus in adult insects originating from San Juan Capistrano in California.