Regioselective Enzymatic Synthesis of Non-Steroidal Anti-Inflammatory Drugs Containing Glucose in Organic Media

Enzymatic transesterification of glucose with the vinyl ester of non-steroidal anti-inflammatory drugs (NSAIDs) was in organic media performed for synthesis of novel NSAIDs-glucose conjugates. Glucose was regioselectively acylated at the 6-hydroxyl group. The indomethacin-glucose conjugate and ketoprofen-glucose conjugate were produced by the catalysis of alkaline protease from Bacillus subtilis in the respective yields of 42% (over 48 h) and 63% (over 40 h). The etodolac-glucose conjugate was obtained in 26% yield (over 144 h) by lipase from Candida antarctica.     

Cellulase and xylanase activity in relation to biofilm formation by two intertidal filamentous fungi in a novel polymethylmethacrylate conico-cylindrical flask

A polymethylmethacrylate (PMMA) conico-cylindrical flask (CCF) with an inner arrangement consisting of eight equidistantly spaced rectangular strips mounted radially on a circular disk to provide additional surface area for fungal attachment was employed for production of cellulase by Chaetomium crispatum and xylanase by Gliocladium viride. The design allowed comparison of production between CCFs with hydrophobic surface (PMMA-CCF), hydrophilic glass surface (GS-CCF) and 500-ml Erlenmeyer flask (EF). Compared with the EF, endo-β-1,4-glucanase and FPase (filter paper degradation) activities increased from 0.044 to 0.156 and from 0.008 to 0.021 IU/ml, respectively, in the PMMA-CCF, while growth of C. crispatum was higher by at most 1.38-fold compared with the other vessels. Xylanase production in the EF was at most 5.08-fold higher and growth of G. viride was at most 1.52-fold higher compared with the other vessels. Temporal pattern of biofilm development based on two-channel fluorescence detection of extracellular polymeric substances (EPSs) and whole cells in a confocal laser scanning microscope demonstrated increase by 100% in biovolume, 25% in thickness and 62.5% both in substratum coverage and total spreading of C. crispatum biofilm in PMMA-CCF over 6 days. Biovolume of G. viride biofilm in GS-CCF increased by 150% over 4 days while that in PMMA-CCF enhanced by 200% over 2 days. Biofilm thickness in PMMA-CCF was 44% higher compared with GS-CCF and increased by 175% over 2 days. Substratum coverage was 38% higher in GS-CCF compared with PMMA-CCF. Thus, reactor surface area and property, shear forces and biofilm formation influenced enzyme production.     

Quantitative Data on Four Species of Eimeria in Eastern Chipmunks and Red Squirrels*

SYNOPSIS. Total counts of Eimeria vilasi and E. wisconsinensis oocysts in 0.05 g fecal samples from the eastern chipmunk over a 2-year period indicated that the 2 species had respective occurrences of 100% and 11% and an actual abundance ratio of 35 to 1. In the red squirrel similar counts of E. tamiasciuri and E. toddi over a 3-year period indicated occurrences of 98% and 6% and an actual abundance ratio of 26 to 1, respectively. These results parallel a similar dichotomy in percent occurrence of 7 Eimeria in the sciurid genera Cynomys and Marmota.     

Self-Organizing Maps in Revealing Variation in Non-obligatory Riverine Fish in Long-term Data

Sensitiveness to the overall influence of the river channel regulation, impoundment and pollution was studied for 12 non-obligatory riverine (NOR) fish species in the Warta River, Poland, over the period of 1963–1998. Their total abundance has not considerably changed unlike the structure of their aggregation, which was revealed by a self-organizing map (SOM, Kohonen unsupervised artificial neural network) almost perfectly separating 1960s samples from 1990s ones. The greatest changes in proportion were recorded for the dominant roach Rutilus rutilus (from 28.6 to 60.0%), perch Perca fluviatilis (1.9 to 9.5%), pike Esox lucius (2.9 to 4.9%), and the former codominant bleak Alburbus alburnus (22.9 to 1.8%). They resulted from highly significant (p < 0.001), reaching even one order of magnitude, differences in the abundance of these species. Such remarkable alternations in dominant NOR ichthyofauna obviously influenced the whole fish assemblage structure, which was best illustrated by a clear vertical gradient over SOM in Simpson and Shannon indices. Thus, NOR species, both studied together and separately, can supply valuable information on the fish assemblage dynamics and should not be neglected in temporal analyses, often focused only on vulnerable obligatory riverine, migratory and/or endemic species.     

Pre-plant bacterisation: a strategy for delivery of beneficial endophytic bacteria and production of disease-free plantlets of black pepper (Pipernigrum L.)

Naturally occurring endophytic bacteria from black pepper vines were found to exhibit strong antagonistic activities against Phytophthora capsici and Radopholus similis. In order to deliver these bacterial strains, as well as to produce disease-free plantlets of black pepper, a pre-plant stem and root bacterisation was standardised. Stem bacterisation with endophytic Pseudomonas spp. was found to suppress P. capsici infection (over 90% reduction in lesion length) on cut shoots. Pre-plant root bacterisation with Pseudomonas aeruginosa, Pseudomonas putida and Bacillus megaterium yielded over 60% of plantlets free from P. capsici infection on roots. Curtobacterium luteum and B. megaterium recorded over 70% reduction of nematode population in soil with concomitant production of over 65% of nematode-free plantlets. Besides protecting the plants from the pathogens, the bacteria were also found to enhance the growth of rooted cuttings. The biocontrol potential of the above endophytic bacteria and their exploitation for disease management in the black pepper nursery are discussed.     

Deoxyribonucleic acid reassociation in the classification of coagulase-positive staphylococci

DNA-DNA reassociation studies were performed with coagulase-positive staphylococci belonging to the biotypes A, B, C, D, E and F. These studies present genetic evidence for the existence of at least two distinct species within this group of organisms. The common Staphylococcus aureus strains were represented by organisms from biotypes A to D, and their DNA revealed over 80% nucleotide sequence homology under restrictive conditions. Less than 15% DNA homology was detected between strains from biotypes A to D (S. aureus) and those from biotypes E and F. The DNA of organisms from either the biotypes E or F displayed over 70% homology. Together, both biotypes are considered to represent the species S. intermedius. However, DNA homology values dropped to 50–65% between strains from different biotypes. This may justify the separation of S. intermedius biotypes E and F on a subspecies level.Abbreviations O.D. optical density - SSC standard saline citrate buffer (0.15 M NaCl, 0.015 M sodium citrate, pH 7.0) This work was supported by Deutsche Forschungsgemeinschaft     

Decline in native ladybirds in response to the arrival of Harmonia axyridis: early evidence from England

1. Harmonia axyridis (Coleoptera: Coccinellidae) is an invasive non‐native ladybird in Europe, where it was introduced as a biological control agent of aphids and coccids. 2. This study assesses changes to ladybird species assemblages, in arboreal habitats, over a 3‐year period encompassing the invasion phase of H. axyridis in eastern England. The effects of H. axyridis and other factors (weather and prey availability) on native ladybirds are assessed. 3. Harmonia axyridis increased from 0.1% to 40% of total ladybirds sampled, whilst native aphidophagous species declined from 84% to 41% of total ladybirds. The actual number of native aphidophagous ladybirds per survey decreased from a mean of 19.7 in year 1, to 10.2 in year 3. 4. Three ladybird species in particular experienced declines: Adalia bipunctata, Coccinella septempunctata, and Propylea quattuordecimpunctata. Harmonia axyridis was the most abundant species by the end of the study. 5. The decline in native aphidophagous ladybirds could be attributed to competition for prey and intraguild predation of eggs, larvae, and pupae by H. axyridis. Physiological and behavioural traits of H. axyridis are likely to confer an advantage over native ladybird species.     

Breeding biology and seasonal abundance of Lappet-faced Vultures Torgos tracheliotus in western Saudi Arabia

Lappet-faced Vultures Torgos tracheliotus utilizing a 2200-km², fenced, protected area in western Saudi Arabia were studied over 4 years. Numbers fluctuated seasonally from minima of c. 30 individuals in the spring to maxima of 160 birds in the autumn. All birds foraged predominantly on domestic livestock carrion outside the Reserve. In each year, up to 17 pairs attempted to breed, with 81% of pairs laying eggs, typically in large Maerua crassifolia trees. In all, 81% of the eggs hatched and 85% of nestlings fledged. Most eggs were laid in December, when mean daily air temperatures were lowest, and young usually fledged c. 180 days later. Overall, 56% of nesting attempts fledged young. Juvenile mortality over the first 3 months after fledging was at least 17%. The numbers and proportion of breeders appeared to be stable, but it is not known whether the establishment of the Reserve concentrated the nesting and roosting of vultures in the area or has attracted birds from elsewhere. Breeding success was higher than in most well-studied African populations and probably those nesting elsewhere in Arabia. Four pairs reared 52% of the fledglings observed in the Reserve over 4 years. The Saudi Arabian Lappet-faced Vultures probably belong to the subspecies negevensis and, because of the extinction of this subspecies in the wild in Israel, the well-being of the Saudi Arabian population is critical to the subspecies' conservation.     

The potential impact of a krill fishery upon pelagic fish in the Prydz Bay area of Antarctica

Summary RMT hauls taken during FIBEX in the Prydz Bay region showed a relatively high incidence of fish compared with most previous reports from the Atlantic sector. Overall, fish comprised 3.8% by number and 8.1% by weight, and these figures rose to 26.8 and 46.1%, respectively in aimed hauls over the continental shelf. Even when only hauls with>50% E. superba are considered, fish accounted for almost 1% by weight of the catch overall, and more in aimed hauls or those taken over the continental shelf. Species composition differed markedly between the continental shelf and deep water zones. Over the shelf the fish catch comprised mainly Pleuragramma antarcticum and channichthyid juveniles, while over deep water it consisted mainly of myctophids and Notolepis coatsi. These data indicate that the potentially large krill fishery in this region could affect the populations of some shelf dwelling demersal species by increasing the mortality among their pelagic juvenile stages.     

Reproduction of Lumbricillus rivalis (Levinsen) in laboratory cultures and in decaying seaweed

The cocoon production of 144 Lumbricillus rivalis cultured in pairs at 10 ± 1 °C was high over the first 2 weeks of breeding activity and then declined, chiefly because of high mortality. Cocoon deposition lasted for between 1 and 16 weeks, eight pairs of worms producing cocoons for 9 weeks and one pair for 16 weeks. During the total period of cocoon deposition over 9000 eggs (mean 17·4 per cocoon) were deposited. Two decaying wrack bed populations of L. rivalis showed a low level of cocoon and egg production in autumn, rising to an annual maximum in late winter/early spring. In these populations the mean egg content varied seasonally from 17·1 to 47·8 eggs per cocoon. When cocoons in the laboratory were transferred from the site of deposition to incubation dishes 31% hatched, but those left in the substrate showed a 92% hatch. In the naturally occurring populations 19% of the cocoons detached from seaweed fronds hatched, but 62% of those left in situ. Eggs and worm embryos developed to relatively late stages in most cocoons, whatever the rate of hatching; development often continued for up to 2 months after deposition without hatching. Over 50% of the fertile eggs in cocoons from decaying wrack hatched and developed to 5 mm worms.     

Allozyme and morphological variability,outcrossing rate and core collection formation in lentil germplasm

Summary A survey of qualitative genetic variation at 3 morphological trait loci, 17 isozyme loci and a putative isozyme locus (amylase) was made for 105 lentil (Lens culinaris Medikus) germplasm accessions from Chile, Greece and Turkey. New alleles were found for Lap-1, Me-2, Pgm-c, Pgm-p and 6-Pgd-c. The average proportion of polymorphic loci per population was 0.19, with a range of 0 to 0.42 over populations. Germplasm from Chile was equally variable to that from Greece and Turkey on the basis of individual loci and in a multilocus sense, despite its post-Columbus introduction to the New World. Evidence was found from associations between allelic states at different loci of a complex multilocus structure of lentil populations. A single multilocus genotype represented 10.2% of all plants sampled. The rate of outcrossing varied from 2.2% and 2.9% in Turkish and Greek landraces to 6.6% among Chilean populations. Using the survey data, a random sampling strategy for core collection formation was compared with two stratified sampling methods. The advantage of stratified sampling over random sampling was only significant at P=0.28.     

Obesity and Cardiovascular Disease Risk Factors in Firefighters: A Prospective Cohort Study

Objective: Obesity, despite being a significant determinant of fitness for duty, is reaching epidemic levels in the workplace. Firefighters’ fitness is important to their health and to public safety. Research Methods and Procedures: We examined the distribution of BMI and its association with major cardiovascular disease (CVD) risk factors in Massachusetts firefighters who underwent baseline (1996) and annual medical examinations through a statewide medical surveillance program over 5 years of follow‐up. We also evaluated firefighters’ weight change over time. Results: The mean BMI among 332 firefighters increased from 29 at baseline to 30 at the follow‐up examination (2001), and the prevalence of obesity increased from 35% to 40%, respectively (p < 0.0001). In addition, the proportion of firefighters with extreme obesity increased 4‐fold at follow‐up (from 0.6% to 2.4%, p < 0.0001). Obese firefighters were more likely to have hypertension (p = 0.03) and low high‐density lipoprotein‐cholesterol (p = 0.01) at follow‐up. Firefighters with extreme obesity had an average of 2.1 CVD risk factors (excluding obesity) in contrast to 1.5 CVD risk factors for normal‐weight firefighters (p = 0.02). Finally, on average, normal‐weight firefighters gained 1.1 pounds, whereas firefighters with BMI ≥ 35 gained 1.9 pounds per year of active duty over 5 years of follow‐up. Discussion: Obesity is a major concern among firefighters and shows worsening trends over time. Periodic medical evaluations coupled with exercise and dietary guidelines are needed to address this problem, which threatens firefighters’ health and may jeopardize public safety.     

Occurrence of mycotoxins in cereals and animal feedstuffs in Natal,South Africa 1994

During the year of 1994, 417 samples of agricultural commodities, comprising: maize, compound animal feeds, oil seeds, soya bean, fish meal and forage were examined for fungi and over 20 mycotoxins using a multi-screen augmented with individual assays. Trichothecenes had the highest incidence of over 19% in all samples received, followed by aflatoxin at 6% and then zearalenone at 3%. Selected samples (73) were analyzed for fumonisin B₁ and of these, 69 (94%) were found to be positive. Because of this result and high incidence ofFusarium spp. (over 70%) in maize and maize containing feeds, which was higher than eitherAspergillus spp. (19%) orPenicillium spp. (33%), attention is drawn to the actual and potential presence of fumonisin in the food chain.     

Microspore development during <Emphasis Type="Italic">in vitro</Emphasis> androgenesis in triticale

Microspore division was monitored in three triticale (× Triticosecale Wittmack) genotypes over 21 d of in vitro anther culture, on two media differing in their 2,4-dichlorophenoxyacetic acid content. After low temperature (4 °C) pre-treatment for two weeks, all the microspores were still alive, but they began to die from day one of culture. Both genotype and culture medium affected the number of microspores that aborted over time (82 – 97 % by day 21), the number of microspores that underwent the first symmetrical division (> 82 % over all), the number of microspores that attained four or more nuclei, and the number of divisions per 100 alive microspores after 21 d of culture.     

Nematodes associated with dipterous stem-boring larvae (Chloropidae and Opomyzidae) in ryegrass

Associations between different species of nematodes and stem-boring larvae were studied over two years. Howardula oscinellae was rarely found in stem-boring larvae. Usually mermithids were also uncommon, but in one Italian ryegrass (Lolium multiforum) variety trial over 12.5% of stem-boring larvae were attacked, with different levels of parasitism in larvae from the different ryegrass varieties. Panagrolaimus rigidus nematodes were also identified from stem-boring larvae and Oscinella frit larvae were the most common host. Greater infestation levels of P. rigidus were found in larvae from grazed swards than from cut swards. P. rigidus may cause the death of some stem-boring larvae.     

Long-term survival of marine planktonic diatoms and dinoflagellates in stored sediment samples

Sediment samples from Scottish coastal sites, taken over the last 9 years, were stored in closed containers at 5C. Slurry cultures were used to determine the survival of phytoplankton in these sediments. A range of diatom and dinoflagellate species survived for at least 27 months in these stored samples. A number of species grew for which no resting stage has yet been described: Thalassiosira angulata, T.pacifica, T.punctigera, T.eccentrica, T.minima and T.anguste-lineata. Notable results were survival times of 73 months for Skeletonema costatum, 96 months for Chaetoceros socialis, C.didymus and C.diadema, 109 months for Scrippsiella sp. and 112 months for Lingulodinium polyedrum. A single sample was stored and repeatedly cultured for diatoms over a period of 16 months. The number of species cultured from the sediment declined over this time. Lingulodinium polyedrum cysts isolated from sediments collected at least 18 months previously gave a hatching success of 97%, and cysts isolated from a 9-year-old sample gave a hatching success of 3%. The study indicated the potential importance of coastal sediments as a source of phytoplankton to their overlying waters. The validity of using marine planktonic diatoms and dinoflagellates for modelling geological events is discussed.      

Frugivory and seed dispersal by foxes in relation to mammalian prey abundance in a semiarid thornscrub

Abstract We examine the role of the native fox, Pseudalopex culpaeus, as a frugivore and seed disperser in a semiarid thornscrub of Chile. We quantified the fruit and animal components in its diet versus the availability of fruits and small mammals in the field over a 2‐year period (January 1998 through February 2000). We tested the legitimacy and effectiveness of foxes as dispersers by quantifying the percentages of seed viability and of germination of seeds that passed through fox gut versus those picked from plants. We also studied their efficiency as dispersers, monitoring the fate of seeds in faeces placed in the field. The highest frequencies of fruit consumption by foxes were observed when abundances of small mammal prey were <6 individuals per hectare, regardless of fruit abundance in the field. Thus, foxes consumed fruits as a supplementary food resource. Based on 326 faeces, the total number of fruits consumed was about 34 000 over the 2‐year study period, and fruits from the alien shrub Schinus molle represented 98% of that total, with the native Porlieria chilensis a distant second. Germination and viability of defecated seeds of P. chilensis were reduced by 66% and 48%, respectively, in comparison to controls. In contrast, germination of seeds of S. molle increased by 50% and no effect on viability was observed. With regard to P. chilensis, foxes were legitimate (they defecated viable seeds), but ineffective (seeds in faeces had lower germination than those taken directly from parental plants and there was no seedling establishment in the field) and inefficient dispersers (seeds in faeces were deposited on microhabitats hostile to seed germination and seedling establishment). However, with regard to S. molle, foxes were legitimate, effective (seeds in faeces had higher germination than those taken directly from parental plants; there was germination but no establishment in the field), and efficient dispersers (over 41% of seeds were deposited on safe microsites). Thus, a native fox may be contributing to the spread of an alien shrub, co‐opting existing community processes.     

Growth and reproduction of Antarctic vascular plants in response to warming and UV radiation reductions in the field

Along the west coast of the Antarctic Peninsula springtime ozone depletion events can lead to a two-fold increase in biologically effective UV-B radiation (UV-B_BE) and summer air temperatures have risen ≈1.5°C during the past 50 years. We manipulated levels of UV radiation and temperature around Colobanthus quitensis (a cushion-forming plant, Caryophyllaceae) and Deschampsia antarctica (a tussock grass) along the Peninsula near Palmer Station for two field seasons. Ambient levels of UV were manipulated by placing filters that either transmitted UV (filter control), absorbed UV-B (reducing diurnal levels of UV-B_BE by about 82%), or absorbed both UV-B and UV-A (reducing UV-B_BE and UV-A_BE by about 88 and 78%, respectively) on frames over naturally growing plants from November to March. Half the filters of each material completely surrounded the frames and raised diurnal and diel air temperatures around plants by an average of 2.3°C and 1.3°C, respectively. Reducing UV or warming had no effect on leaf concentrations of soluble UV-B absorbing compounds, UV-B absorbing surface waxes or chlorophylls. Warming had few effects on growth of either species over the first season. However, over the second field season warming improved growth of C. quitensis, leading to a 50% increase in leaf production (P < 0.10), a 26% increase in shoot production, and a 6% increase in foliar cover. In contrast, warming reduced growth of D. antarctica, leading to a 20% decline in leaf length, a 17% decline in leaf production (P < 0.10), and a 5% decline in foliar cover. Warming improved sexual reproduction in both species, primarily through faster development of reproductive structures and greater production of heavier seeds. Over the second field season, the percentage of reproductive structures that had reached the most developed (seed) stage in C. quitensis and D. antarctica was 20% and 15% higher, respectively, under warming. Capsules of C. quitensis produced 45% more seeds under warming and these seeds were 11% heavier. Growth of D. antarctica was improved when UV was reduced and these effects appeared to be cumulative over field seasons. Over the second season, tillers produced 55% more leaves and these leaves were 32% longer when UV-B was reduced. Tillers produced 137% more leaves that were 67% longer when both UV-B and UV-A were reduced. The effects of UV reduction were not as pronounced on C. quitensis, although over the second season cushions tended to be 17% larger and produce 21% more branches when UV-B was reduced, and tended to be 27% larger and produce 38% more branches when both UV-B and UV-A were reduced (P < 0.10). Few interactions were found between UV reduction and warming, although in the absence of warming, reducing UV led to slower development of reproductive structures in both species. The effects of warming and UV reduction were species specific and were often cumulative over the two field seasons, emphasizing the importance of long-term field manipulations in predicting the impacts of climate change. Received: 4 August 1998 / Accepted: 1 December 1998     

Population size,growth and movements of Anguilla australis in a small lake

To study growth rates, movements and estimate population size of shortfin eels Anguilla australis in a small lake (2·5 ha) near Christchurch, New Zealand, 617 A. australis were tagged with PIT tags. Tag retention was high (95%) and over the seven recapture events spread over 2 years, 55% of tagged A. australis were recaptured. Growth of recaptured A. australis averaged 13·1 mm year^?1 and declined slightly with increasing total length. Distance moved from original capture site increased with increasing time at large. Population estimates of A. australis > 400 mm (susceptible to capture by fyke net) from recaptures of individuals averaged 1451 A. australis, with a biomass of 170 kg ha^?1. An average of 6·6% of the estimated total population matured as male silver A. australis each year. Results from radio‐tracking of four A. australis gave an average nightly foraging area of 2780 m², and there was no apparent preference for inshore movement (within 5–6 m of the shoreline) or offshore movement. Fyke‐net efficiency (total catch relative to the estimated total population available to each net) measured over four consecutive nights fishing was 88%. The lack of precision of the shoreline triangulation system used, ±10 m, meant that the positional data were considered too coarse to be used in a proposed novel population estimation technique based on determining population size within foraging areas.     

Translocation and plant availability of radio caesium in an organic soil

¹³⁴Caesium chloride solution was injected into the surface of a peaty podzol at 3 cm depth with 5 cm spacing over a M² at two upland sites, one of which had been fertilized and reseeded. The movement of radio caesium in the soil was subsequently followed by coring and/or taking out 10-cm square blocks at monthly intervals over a period of 2 years. There was very little movement of the caesium down the profile, with more than 95% remaining at the point of application. Lateral movement was also minimal. The caesium did not concentrate in the roots. Samples of herbage collected from the surface during the growing season showed a progressively decreasing concentration of caesium over the period. Total caesium removed in herbage amounted to 3.5 and 0.6% of the original present for the improved and umimproved soils, respectively.The extractability of the caesium from the ¹³⁴Cs-amended peaty soil was compared with that from a low-ash peat which had been treated with the radionuclide for 2 months. Initially 1 M ammonium acetate (pH 7.2) extracted 30 and 56% of the Cs from 0–3 and 3–6 horizous of the improved peaty podzol, respectively. After 5 months only about 12 and 7% of the Cs was extracted by 1 M ammonium acetate and 4 and 1% by 0.1 M sodium pyrophosphate from the 0–3 and 3–6 cm horizons, respectively. After 24 months 1 M ammonium acetate extracted 1.6 and 3.3% of the Cs from the 0–3 and 3–6 cm horizons, respectively. In contrast, the values for the peat after 2 months were 100 and 80% for 1 M ammonium acetate and 0.1 M pyrophosphate, respectively.