The fall armyworm (FAW) Spodoptera frugiperda (J.E. Smith) is a highly damaging invasive omnivorous pest that has developed varying degrees of resistance to commonly used insecticides. To investigate the molecular mechanisms of tolerance to tetraniliprole, spinetoram, and emamectin benzoate, the enzyme activity, synergistic effect, and RNA interference were implemented in S. frugiperda. The functions of cytochrome P450 monooxygenase (P450) in the tolerance to tetraniliprole, spinetoram, and emamectin benzoate in S. frugiperda was determined by analysing changes in detoxification metabolic enzyme activity and the effects of enzyme inhibitors on susceptibility to the three insecticides. 102 P450 genes were screened via transcriptome and genome, of which 67 P450 genes were differentially expressed in response to tetraniliprole, spinetoram, and emamectin benzoate and validated by quantitative real-time PCR. The expression patterns of CYP9A75, CYP340AA4, CYP340AX8v2, CYP340L16, CYP341B15v2, and CYP341B17v2 were analysed in different tissues and at different developmental stages in S. frugiperda. Silencing CYP340L16 significantly increased the susceptibility of S. frugiperda to tetraniliprole, spinetoram, and emamectin benzoate. Furthermore, knockdown of CYP340AX8v2, CYP9A75, and CYP341B17v2 significantly increased the sensitivity of S. frugiperda to tetraniliprole. Knockdown of CYP340AX8v2 and CYP340AA4 significantly increased mortality of S. frugiperda to spinetoram. Knockdown of CYP9A75 and CYP341B15v2 significantly increased the susceptibility of S. frugiperda to emamectin benzoate. These results may help to elucidate the mechanisms of tolerance to tetraniliprole, spinetoram and emamectin benzoate in S. frugiperda.

Kazal-type serine protease inhibitors (KaSPI) play important roles in insect growth, development, digestion, metabolism and immune defence. In this study, based on the transcriptome of Mythimna separata, the cDNA sequence of MsKaSPI with Kazal domain was uploaded to GenBank (MN931651). Spatial and temporal expression analysis showed that MsKaSPI was expressed at different developmental stages and different tissues, and it was induced by 20-hydroxyecdysone in third-instar larvae of M. separata. After 24 h infection by Beauveria bassiana, the expression level of MsKaSPI and the corresponding MsKaSPI content were significantly up-regulated, being 6.42-fold and 1.91-fold to the control group, respectively, while the activities of serine protease, trypsin and chymotrypsin were inhibited. After RNA interference interfered with MsKaSPI for 6 h, the expression decreased by 73.44%, the corresponding content of MsKaSPI protein decreased by 55.66% after 12 h, and the activities of serine protease and trypsin were significantly enhanced. Meanwhile, both the larval and pupal stages of M. separata were prolonged, the weights were reduced and the number of eggs per female decreased by 181. Beauveria bassiana infection also increased the mortality of MsKaSPI-silenced M. separata by 18.96%. These prove MsKaSPI can not only result in slow growth and low fecundity of M. separata by regulating the activity of related protease, but also participate in the resistance to pathogenic fungi by regulating the serine protease inhibitor content and the activities of related serine protease.

Leishmania spp. are parasitic protozoa that cause leishmaniasis, a disease endemic in 98 countries. Leishmania promastigotes are transmitted by the vector and differentiate into amastigotes within phagocytic cells of the vertebrate host. To survive in multiple and hostile environments, the parasite has several virulence factors. Oligopeptidase B (OPB) is a serine peptidase present in prokaryotes, some eukaryotes and some higher plants. It has been considered a virulence factor in trypanosomatids, but only a few studies, performed with Old World species, analysed its role in Leishmania virulence or infectivity.

L. (L.) amazonensis is an important agent of cutaneous leishmaniasis in Brazil. The L. (L.) amazonensis OPB encoding gene has been sequenced and analysed in silico but has never been expressed. In this work, we produced recombinant L. (L.) amazonensis OPB and showed that its pH preferences, Km and inhibition patterns are similar to those reported for L. (L.) major and L. (L.) donovani OPBs. Since Leishmania is known to secrete OPB, we performed in vitro infection assays using the recombinant enzyme. Our results showed that active OPB increased in vitro infection by L. (L.) amazonensis when present before and throughout infection. Our findings suggest that OPB is relevant to L. (L.) amazonensis infection, and that potential drugs acting through OPB will probably be effective for Old and New World Leishmania species. OPB inhibitors may eventually be explored for leishmaniasis chemotherapy.

The accumulation of nutrients during diapause preparation is crucial because any lack of nutrition will reduce the likelihood of insects completing diapause, thereby decreasing their chances of survival and reproduction. The fall webworm, Hyphantria cunea, diapause as overwintering pupae and their diapause incidence and diapause intensity are regulated by the photoperiod. In this study, we test the hypothesis that photoperiod influences energy reserve accumulation during diapause preparation in fall webworm. We found that the body size and mass, lipid and carbohydrate content of pupae with a short photoperiod during the diapause induction phase were significantly greater than those of pupae with a relatively short photoperiod, and the efficiency of converting digested food and ingested food into body matter was greater in the short-photoperiod diapause-destined larvae than the relatively short-photoperiod diapause-destined larvae. We also observed higher lipase and amylase activities in short-photoperiod diapause-destined larvae relative to the counterparts. However, no obvious difference was found in protein and protease in the pupae with a short photoperiod during the diapause induction phase and short-photoperiod diapause-destined larvae compared with the counterparts. Therefore, we conclude that the energy reserve patterns of diapausing fall webworm pupae are plastic and that short-photoperiod diapause-destined larvae increase their energy reserves by improving their feeding efficiency and increase their lipid and carbohydrate stores by increasing the lipase and amylase activities in the midgut.

Increasing crop diversity has been highly recommended because of its environmental and economic benefits. However, the impacts of crop diversity on soil properties are not well documented. Thus, the present study was conducted to assess the impacts of crop diversity on selected soil quality indicators. The cropping systems investigated here included wheat (Triticum aestivum L.) grown continuously for 5 years as mono-cropping (MC), and a 5-year cropping sequence [(wheat–cover crop (CC)–corn (Zea mays L.)–pea (Pisum sativum L.) and barley (Hordeum vulgare L.)–sunflower (Helianthus annuus L.)]. Each crop was present every year. This study was conducted in the northern Great Plains of North America, and soil quality data were collected for 2016 and 2017. Selected soil quality indicators that include: soil pH, organic carbon (SOC), cold water-extractable C (CWC) and N (CWN), hot water-extractable C (HWC) and N (HWN), microbial biomass carbon (MBC), bulk density (BD), water retention (SWR), wet soil aggregate stability (WAS), and urease and β-glucoside enzyme activity were measured after the completion of 5-year rotation cycle and the following year. Crop diversity did not affect soil pH, CWC, CWN, HWC, HWN and SWR. Cropping systems that contained CC increased SOC at shallow depths compared to the systems that did not have CC. Crop diversity increased WAS, MBC, and urease and β-glucoside enzyme activity compared with the MC. Comparison of electrical conductivity (EC) measured in this study to the baseline values at the research site prior to the establishment of treatments revealed that crop rotation decreased EC over time. Results indicate that crop diversity can improve soil quality, thus promoting sustainable agriculture.

The present study characterized the biological function of the asparaginyl peptidase legumain-1 (LEG-1) of the bovine lungworm Dictyocaulus viviparus and its suitability as a recombinant vaccine against dictyocaulosis. Quantitative real-time PCR and immunoblot analysis revealed LEG-1 to be almost exclusively transcribed and expressed in parasitic lungworm stages. Immunohistochemistry localized the enzyme in the parasite's gut, which was confirmed by immunoblots detecting LEG-1 in the gut as well as male testes. LEG-1 was recombinantly (rLEG-1) expressed in the yeast Pichia pastoris and subsequently analysed in activity assays for its enzyme functions and substrate specificity. For sufficient functionality, rLEG-1 needed trans-activation through D. viviparus cathepsin L-2, indicating a novel mechanism of legumain activation. After trans-activation, rLEG-1 worked best at pH 5·5 and 35–39 °C and cleaved a legumain-specific artificial substrate as well as the natural substrates bovine collagen types I and II. In a clinical vaccination trial, rLEG-1 did not protect against challenge infection. Results of in vitro characterization, transcription pattern and localization enhance the presumption that LEG-1 participates in digestion processes of D. viviparus. Since rLEG-1 needs trans-activation through a cathepsin, it is probably involved in an enzyme cascade and therefore remains interesting as a candidate in a multi-component vaccine.

A biotype of johnsongrass cross resistant to clethodim, sethoxydim, quizalofop-P, and fluazifop-P was identified in several fields in Washington County, MS. Absorption, translocation, and metabolism studies using 14C-clethodim and acetyl-coenzyme A carboxylase (ACCase) activity assays were conducted to determine the resistance mechanism. Absorption of 14C-clethodim was higher in the resistant than the susceptible biotype 4 hours after treatment (HAT), but at 24, 48, and 72 HAT, similar levels of radioactivity were detected in both johnsongrass biotypes. Consequently, resistant plants had more radioactivity present in the treated leaves at 4 and 24 HAT. However, there was no difference between resistant and susceptible biotypes in the translocation of 14C out of the treated leaf at 4, 8, 24, 48, and 72 HAT as a percentage of total absorbed. Metabolism of clethodim was similar in the resistant and susceptible biotypes. There was no difference in the specific activity of ACCase from the susceptible and resistant johnsongrass biotypes (means of 0.221 and 0.223 nmol mg−1 protein min−1, respectively). ACCase from the susceptible biotype was sensitive to clethodim, with an I 50 value of 0.29 μM clethodim. The ACCase enzyme from the resistant biotype was less sensitive, with an I 50 value of 1.32 μM clethodim. The resultant R/S ratio for clethodim was 4.5. These results indicate that resistance to clethodim in this johnsongrass biotype resulted from an altered ACCase enzyme that confers resistance to clethodim.

This is the first study of the metabolic enzyme activity and energy state of the dwarf oyster Ostrea stentina (Payraudeau, 1826) in Tunisia and in the Mediterranean region. The main purpose of this study was to examine the modulation of the physiological status of the oyster O. stentina depending on the season and the presence of parasite Marteilia refringens. The prevalence of bonamiosis and marteiliosis were established by PCR. Bonamia exitiosa was detected only in 2.91% of oysters while the prevalence of M. refringens was 100% (30 Ind./30) in February and 93.93% (28 Ind./30) in March. A 42–87% mortality rate was reported during the study period. Biochemical analyses were carried out to evaluate the management of the energy resources in regard to the biochemical changes of lipids and carbohydrates. The enzyme activity of pyruvate kinase (PK), citrate synthase (CS) and cytochrome c oxidase (CCO) were measured and the mean level of enzyme activity was respectively 20.31 mU mg−1 protein; 12.06 mU mg−1 protein and 3.59 mU mg−1 protein. Carbohydrate contents of O. stentina were very low all year round with an average of 15.18% in dry weight, and lipid contents remain similar (11.77% in dry weight) compared with the values reported for most other temperate bivalves. Enzyme activity significantly decreased over time (P < 0.001). The energy resources of the oyster O. stentina wherein much of the energy was devoted to reproduction seem to affect the defence system.

This study investigated the effects of short-term subclinical Zn deficiency on exocrine pancreatic activity and changes in digestive capacity. A total of forty-eight weaned piglets were fed ad libitum a basal diet (maize and soyabean meal) with adequate Zn supply (88 mg Zn/kg diet) during a 2-week acclimatisation phase. Animals were then assigned to eight dietary treatment groups (n 6) according to a complete randomised block design considering litter, live weight and sex. All pigs were fed restrictively (450 g diet/d) the basal diet but with varying ZnSO4.7H2O additions, resulting in 28·1, 33·6, 38·8, 42·7, 47·5, 58·2, 67·8 and 88·0 mg Zn/kg diet for a total experimental period of 8 d. Pancreatic Zn concentrations and pancreatic activities of trypsin, chymotrypsin, carboxypeptidase A and B, elastase and α-amylase exhibited a broken-line response to stepwise reduction in dietary Zn by declining beneath thresholds of 39·0, 58·0, 58·0, 41·2, 47·5, 57·7 and 58·0 mg Zn/kg diet, respectively. Furthermore, carboxypeptidase B and α-amylase activities were significantly lower in samples with reduced pancreatic Zn contents. Coefficients of faecal digestibility of DM, crude protein, total lipids and crude ash responded similarly to pancreatic enzyme activities by declining below dietary thresholds of 54·7, 45·0, 46·9 and 58·2 mg Zn/kg diet, respectively. In conclusion, (1) subclinical Zn deficiency impaired pancreatic exocrine enzymes, (2) this response was connected to pancreatic Zn metabolism and (3) the decline in catalytic activity impaired faecal digestibility already after 1 week of insufficient alimentary Zn supply and very early before clinical deficiency symptoms arise.

The aphid species Sitobion avenae and Rhopalosiphum padi are the most important pests in wheat growing regions of many countries. In this study, we investigated the sublethal effects of imidacloprid on fecundity, longevity, and enzyme activity in both aphid species by comparing 3-h exposure for one or three generations. Our results indicated that 3-h exposure to sublethal doses of imidacloprid for one generation had no discernible effect on the survival, fecundity, longevity, or enzyme activity levels of aphids. However, when pulse exposures to imidacloprid were sustained over three generations, both fecundity and longevity were significantly decreased in both S. avenae and R. padi. Interestingly, the fecundity of R. padi had almost recovered by the F5 generation, but its longevity was still deleteriously affected. These results indicated that R. padi laid eggs in shorter time lags and has a more fast resilience. The change in reproduction behavior may be a phenomenon of R. padi to compensate its early death. If this is stable for the next generation, it means that the next generation is more competitive than unexposed populations, which could be the reason underlying population outbreaks that occur after longer-term exposure to an insecticide. This laboratory-based study highlights the sublethal effects of imidacloprid on the longevity and fecundity of descendants and provides an empirical basis from which to consider management decisions for chemical control in the field.

Addition of antibiotics to artificial diets of insects is a key component in the rearing of insects in the laboratory. In the present study an antimicrobial agent, streptomycin sulphate was tested for its influence on survival and fitness of Spodoptera litura (Fabricus) (Lepidoptera: Noctuidae) as well as its gut microbial diversity. The antibiotic did not adversely affect the survival of S. litura. Faster growth of larvae was recorded on diet amended with different concentrations of streptomycin sulphate (0.03, 0.07 and 0.15%) as compared to diet without streptomycin sulphate. The overall activity of various digestives enzymes increased on S+ diet while the activity of detoxifying enzymes significantly decreased. In addition, alteration in microbial diversity was found in the gut of S. litura larvae fed on diet supplemented with antibiotic (S+) and without antibiotic (S−).

Introduction. Angiotensin I-Converting Enzyme(ACE) is a key component in regulation of blood pressure by virtue ofthe rennin-angiotensin system. ACE converts the inactive decapeptide,angiotensin I, into the potent vasopressor octapeptide, angiotensinII, and inactivates bradykinin, which has a vasodilating action. So,inhibition of ACE has become a major target in control ofhypertension. It is well known that the consumption of fruits couldprovide health benefits by lowering the risk of chronic diseases suchas metabolic syndrome diseases including type 2 diabetes andcardiovascular disease. Our current study was focused on investigatingthe ACE inhibitory property of a few underutilized minor fruits ofWest Bengal using an in vitro assay. Materialsand methods. The potential antihypertensive activity ofunderutilized indigenous edible fruits of West Bengal, India, wasevaluated by their ability to inhibit Angiotensin-Converting Enzyme(ACE). The ACE inhibitory property was assayed using ACE from rabbitlung and hippuryl-histidyl-leucine as the substrate. Nineteen fruitspecies belonging to 15 families were investigated. The percentage ACEinhibitory activities of these fruits were studied at 20 µg freshweight of fruit extract per mL. The total phenol content of all thesefruits was determined following the standard Folin-Ciocalteu method.Total flavonoid content was also measured. Results. Theaqueous fruit extracts of the red variety of Trapabispinosa, Phoenix sylvestris,Cicca acida, Achras sapota andAverrhoa carambola presented more than 75% ACEinhibition. On the other hand, Punica granatum,Spondias pinnata, Trapabispinosa (green) and Ziziphus mauritianashowed about 50% inhibition. Aeglemarmelos, Annona squamosa, Annona reticulata,Feronia elephantum, Physalis peruviana andSyzygium jambos showed low activity (< 50%inhibition). Discussions. To date there has been noreport on Angiotensin I-Converting Enzyme inhibitory activities ofthese underutilized minor fruits of West Bengal, India. During ourstudy no correlation could be established between the % ACE inhibitionand the total phenol or flavonoid content of these fruit extracts. So,it appears that non-phenolic components may also be responsible forACE inhibitory activity. In our investigation we tried to establishthe fact that the consumption of these underutilized minor fruitsmight have potential in managing cardiovascular diseases.

Abstract – Introduction. Mango fruits grown underhigh-density planting show a progressive decline in crop yield after14–15 years, due to overcrowding of canopies, which suggests regularcanopy management is necessary. Hence, the effects of pruningtreatment on fruit yield and quality of ‘Amrapali’ mango were studiedin India over two consecutive years, 2010 and 2011. Materialsand methods. Mango trees were subjected to pruning(removal of 50 cm of shoot from the apex) in the month of September2009 with unpruned trees serving as control. Fruits were harvested atthe commercial maturity stage and quality parameters were assessedboth in fresh fruits and following ripening at room temperature [(35 ±2) °C and (80 ± 5)% RH)]. Results and discussion. Fruityield of pruned trees was found to decrease during the first yearcompared with the fruit yield of unpruned trees; later on, itincreased during the second year. Pruning resulted in significantlyhigher fruit weight, fruit firmness, total carotenoids, antioxidantcapacity and total phenolic content. Early maturity of fruits wasobserved from unpruned trees with faster color change, higher totalsoluble solids and lower titratable acidity. The fruits harvested frompruned trees showed slower ripening, and lower respiration, ethyleneevolution rate and enzyme activity as compared with fruits fromunpruned trees. Both anthracnose and stem-end rot disease percentagewere reduced in ripe fruits from pruned trees.Conclusion. Pruning treatment appears to be analternative strategy to obtain better yield and quality in denselypopulated old mango orchards.

Introduction. Plum cv. SantaRosa shows climacteric behaviour and grows profitably under sub-temperateconditions in India. After ripening, it has a very limited shelflife (3–4 days) in ambient conditions [(38 ± 2) °C and at (68 ± 4)%RH]. Hence, there is an urgent need to extend its marketabilityusing recent techniques. Materials and methods. Plumswere harvested at the pre-climacteric and climacteric stages ofmaturity and packed with newspaper shreds (control), KMnO4-impregnatedchalks, KMnO4-impregnated newspaper shreds or ethylene-absorbent sachets;then they were transported by road to New Delhi. The biochemicaland physiological analyses were carried out at 3-day intervals onplums stored in supermarket conditions [(20 ± 1) °C and at (90 ± 2)%RH] for 15 days. Results and discussion. Untreatedfruits had less firmness than those which were packed with ethyleneabsorbents. The activities of fruit-softening enzymes such as lipoxygenase(LOX), polygalacturonase (PG) and pectin methylesterase (PME) increasedrapidly in the control treatment in comparison with plums packed withvarious ethylene absorbents, the least being either with ethylene-absorbentsachets or with KMnO4-impregnated newspaper shreds. Plums of pre-climacteric andclimacteric maturity attained respiration and ethylene evolutionpeaks on the 9th day and 6th day, respectively. Conclusions. Theplums of pre-climacteric and climacteric stages of maturity packed withethylene-absorbent sachets showed the best results in supermarketconditions in maintaining the firmness, quality and shelf life ofplum up to the 12th day and 9th day of storage, respectively.

The pancreatic enzyme secretory phospholipase A2 group IB (sPLA2 IB) hydrolyses phospholipids at the sn-2 position, resulting in a NEFA and a lyso-phospholipid, which are then absorbed by the enterocytes. The sPLA2 IB is a member of a family of nineteen enzymes sharing the same catalytic ability, of which nine are cytosolic and ten are secretory. Presently, there are no pharmacological tools to separate between the different secretory enzymes when measuring the enzymatic activity. Thus, it is important to support activity data with more precise techniques when isolation of intestinal content is not possible for analysis, as in the case of small teleost larvae, where the whole animal is sometimes analysed. In the present study, we characterise the sPLA2 IB gene in Atlantic cod (Gadus morhua) and describe its ontogeny at the genetic and protein level and compare this to the total sPLA2 activity level. A positive correlation was found between the expression of sPLA2 IB mRNA and protein. Both remained stable and low during the larval stage followed by an increase from day 62 posthatch, coinciding with the development of the pyloric ceaca. Meanwhile, total sPLA2 enzyme activity in cod was stable and relatively high during the early stages when larvae were fed live prey, followed by a decrease in activity when the fish were weaned to a formulated diet. Thus, the expression of sPLA2 IB mRNA and protein did not correlate with total sPLA2 activity.

This work aims at the identification of relevant intermediate metabolism enzymes contributing to improved meat production due to genetic selection. A wild rabbit (WR) breed and a highly meat selected breed (New Zealand (NZ) rabbit) were used. Food restriction was used as an experimental condition so as to enhance differences within the metabolic pathways under study. During a period of 30 days, NZ and WR experimental breeds were subjected to, respectively, 40% and 60% ad libitum food restriction leading to 17.7% and 21.1% initial weight. Hepatic glycolytic, lipidic and protein regulatory enzyme activity, transcriptional and metabolite levels were determined. Insulin-like growth factor (IGF-1), triiodothyronine, and cortisol were also evaluated. In the glycolytic pathways, the NZ control rabbits presented a higher phosphofructokinase and pyruvate kinase activity level when compared to the WR, while the latter group showed a higher expression of glycogen synthase, although with less glycogen content. In the nitrogen metabolism, our results showed a lower activity level of glutamate dehydrogenase in WR when subjected to food restriction. Within the lipid metabolism, results showed that although WR had a significantly higher mRNA hepatic lipase, non-esterified fatty acid levels were similar between the experimental groups. NZ rabbits presented a better glycemia control and greater energy substrate availability leading to enhanced productivities in which triiodothyronine and IGF-1 played a relevant role.

Plant oils in the diet are known to alter milk fat composition owing to changes in the supply of fatty acid precursors and/or activity of lipogenic enzymes in the mammary gland. Thirteen mid-lactating Alpine goats were used in a 3×3 Latin square design with 28-d periods to evaluate possible mechanisms regulating milk fat synthesis and fatty acid composition on grass hay-based diets containing none (H) or 55 g/kg diet dry matter of sunflower-seed oil (HSO) or linseed oil (HLO). Inclusion of oils in the diet had no effect on milk yield but enhanced (P<0·05) milk fat secretion. Compared with the control, HLO and HSO decreased (P<0·05) C10–C16 secretion and increased (P<0·05) C18 output in milk, responses that were accompanied by reductions in milk fat cis-9 14:1/14:0, cis-9 18:1/18:0 and cis-9, trans-11 18:2/cis-9 18:1 concentration ratios. Plant oil supplements decreased (P<0·05) mammary stearoyl-CoA desaturase (SCD) activity but had no effect on SCD mRNA. Treatments had no effect on glucose-6-phosphate dehydrogenase, malic enzyme and glycerol-3-phosphate dehydrogenase activity, or mRNA abundance and/or activity of lipoprotein lipase, acetyl-CoA carboxylase and fatty acid synthase in mammary, hepatic or adipose tissue. The results provided little support for milk fatty acid secretion responses to HLO and HSO being mediated via changes in mammary, hepatic or adipose mRNA abundance or in the activity of key lipogenic enzymes. In conclusion, plant oils in the diet enhance milk fat synthesis, alter milk fatty acid composition and specifically inhibit mammary SCD activity in the goat. Furthermore, the results suggest that the regulation of mammary lipogenesis in response to plant oils appears related to factors other than altered mammary gene expression or potential lipogenic enzyme activity.

Methanol accumulates in axis tissues of maturing soybean seeds, correlating with preharvest seed deterioration. Accumulation of methanol appears to be associated with the enzymic demethylation of pectin methyl esters by pectinesterase (PE; EC 3.1.1.11). To characterize PE in developing and maturing soybean (Glycine max (L.) Merrill) seeds, enzyme activity was assayed in axis and cotyledon tissues. Activity per g fresh weight was 20–25 times higher in axes than in cotyledons with highest activities between 45 and 60 days after flowering (DAF). Twenty to 33% of the total PE activity was in the ‘soluble’ form (extracted with water, 0.5 M sucrose, 1 M sucrose and water). Soluble and cell-wall-bound PE (subsequently extracted with 1 M NaCI) were purified and characterized from axes of seeds at 45–60 DAF. Purification of PE was achieved through concentration of extracts by ultra-filtration, precipitation with ammonium sulfate (30–80% saturation), dialysis, gel filtration on Sephadex G-75 columns, and ion exchange chromatography on CM Sepharose CL-6B. Further purification of both soluble and bound PE was by isoelectric focusing (IEF) on ultrathin layers of polyacrylamide gel with simultaneous detection of protein and PE activity. It was possible to follow seven bands exhibiting PE activity with pl values between 6.0 and 9.5 in 1 M NaCI-extracts of total homogenates. Differences in the IEF patterns of bound and soluble PE were observed. Whereas the bound enzyme exhibited more basic PE bands (pl 8–9.5), the soluble enzyme had more active bands at pl 6.5, 7.0 and 7.5. The Mr was close to 33 000 and the pH optimum was 7.8 for both soluble and bound PE.

Glutathione transferases (GSTs) are a family of multifunctional enzymes with fundamental roles in cellular detoxication. Here we report the molecular characterization of 3 recombinant GSTs belonging to the mu- and delta-class from the parasitic mite Sarcoptes scabiei. Kinetic constants were determined, and the effect of acaricides, including organothiophosphates, pyrethroid esters, a formamidine, a macrocyclic lactone, an organochlorine as well as a bridged diphenyl acaricide, on the activity of the GSTs were tested using 1-chloro-2,4-dinitrobenzene (CDNB) as model substrate. Our results showed that enzymes from the same class and with high amino acid sequence identity have significantly different kinetic properties. For instance, one mu-class GST lost more than 50% of its activity in the presence of one of the organothiophosphates while the activity of the second mu-class GST was only slightly reduced under identical conditions. Tertiary structure modulations indicated that structural differences were the crucial factor for the different kinetic patterns observed. Genome analysis showed that the two mu-class GSTs are organized in tandem in the S. scabiei genome. Taken together these results show that GSTs might be involved in the metabolism of acaricides in S. scabiei.

Conjugated linoleic acid (CLA) has a range of biological properties, including effects on lipid metabolism, milk and body composition in animals. This study investigated the effects of dietary CLA on lactating rats and development of the suckling pups. Dams were fed either a control diet or the same diet supplemented with 25 g/kg of a fat supplement containing 540 g CLA/kg (final concentration of 13·5 g CLA/kg diet) from parturition to the 15th day post-partum. The CLA mixture used in this study contained the following isomers (per 100 g): cis-9, trans-11 (24 g); cis-10, trans-12 (35 g); cis-8, trans-10 (15 g); cis-11, trans-13 (17 g) and others (9 g). On d 15 post partum, CLA supplementation reduced milk fat content by 33% and pup growth by 21%. The milk fatty acid profile, with decreased content of short and medium chain acids, suggests CLA inhibition was more pronounced for de novo lipid synthesis. Consistent with these results, activities of fatty acid synthase, glucose 6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase were reduced by CLA treatment in the mammary gland and liver. In contrast, the activity of NADP-malate dehydrogenase was unchanged.