Modification of culture conditions for production of the anti-tubercular hirsutellones by the insect pathogenic fungus Hirsutella nivea BCC 2594

Aims:  This work aimed to improve the production of anti-tubercular hirsutellones by the insect pathogenic fungus Hirsutella nivea BCC 2594.
Methods and Results:  The fungus was cultivated under different carbon/nitrogen sources and aerations (shake vs static flasks) to improve the production of the anti-tubercular alkaloids, hirsutellones A–D. Under the basal conditions, static cultivation at 25°C in minimum salt medium, only hirsutellone B and C were detected with maximum concentrations of 139·00 and 18·27 mg l⁻¹. Substitution of fructose for glucose and peptone for yeast extract increased the titres of hirsutellones A, B and C about two- to threefold. However, hirsutellone D was not detected in this medium. Culture agitation induced the production of hirsutellone D. As a result, the significant amounts of hirsutellones A–D were obtained with the concentration of 29·93, 169·63, 22·65 and 15·71 mg l⁻¹ within 15 days.
Conclusions:  Improved titres of hirsutellones in H. nivea BCC 2549 were achieved with an agitated (200 rev min⁻¹) fructose–peptone medium at 25°C.
Significance and Impact of the Study:  Improved yields of hirsutellones B–D will enable medicinal chemistry modifications leading to a development of a potential candidate for tuberculosis therapy.     

Compatibility of a natural enemy, Coleomegilla maculata lengi (Col., Coccinellidae) and four insecticides used against the Colorado potato beetle (Col., Chrysomelidae)

Abstract:   The toxicity of four insecticides used to control the Colorado potato beetle, Leptinotarsa decemlineata (Say) (Coleoptera: Chrysomelidae), imidacloprid (Admire^®), cryolite (Kryocide^®), cyromazine (Trigard^®), and Bacillus thuringiensis var. tenebrionis (Novodor^®), to one of its natural enemies, the 12-spotted lady beetle, Coleomegilla maculata lengi Timberlake (Coleoptera: Coccinellidae) was determined in the laboratory. Toxicity assays against C. maculata adults and larvae consisted of (1) topical applications and (2) exposures to treated foliage and prey, using concentrations up to 10 times the manufacturer's recommendations. Over a 6-day period, cyromazine (insect growth regulator) and B. t . var. tenebrionis (microbial insecticide) had no lethal effects on first and third instars C. maculata . For both larval and adult stages, cryolite (inorganic insecticide) caused very low predator mortality when topically applied and moderate mortality when ingested through contaminated eggs of Colorado potato beetles. Imidacloprid (systemic organic insecticide) was highly toxic to adult and larval C. maculata . Its estimated LD₅₀ at 6 days following treatment, corresponded to 0.02–0.09 times the recommended field concentration, depending on the developmental stage and mode of contamination. These results indicate that integrated pest management programmes for Colorado potato beetles using imidacloprid or, to a lesser degree, cryolite, would be detrimental to C. maculata . Cyromazine and B. t . var. tenebrionis seem to present a better compatibility with the protection of C. maculata populations.     

Acid phosphatase production by Aspergillus niger N402A in continuous flow culture

The production of acid phosphatases (E.C.3.1.3.2, ACPs) by Aspergillus niger N402A is regulated by specific growth rate, as well as phosphate availability and pH, as demonstrated by studies in continuous flow culture. Specific ACP activity was highest when A. niger was grown at pH 6.3 (64±8 U g⁻¹) or pH 2.8 (99±11 U g⁻¹), at a dilution rate of 0.07 h⁻¹ and phosphate concentrations below 0.46 mM. ACP production was growth correlated for specific growth rates between 0.07 and 0.13 h⁻¹. Four different ACPs, including two phytases, were produced by A. niger N402A. The ACP and the phytase with maximal activities at pH 5.5 were differentially expressed at different culture pH values, with greater production at low pH.     

ACCUMULATION OF SULFURIC ACID IN DICTYOTALES (PHAEOPHYCEAE): TAXONOMIC DISTRIBUTION AND ION CHROMATOGRAPHY OF CELL EXTRACTS

Four species in the order Dictyotales ( Dictyopteris latiuscula (Okamura) Okamura, D. prolifera (Okamura) Okamura, D. repens (Okamura) Børgesen, and Spatoglossum crassum J. Tanaka) were found to be highly acidic as in some species of the order Desmarestiales (Phaeophyceae). The pH within their cells, presumably that of the vacuole, was estimated to be 0.5 to 0.9 by pH measurements of their cell extracts in distilled water. However, other species of these genera ( D. divaricata (Okamura) Okamura, D. undulata Holmes, and S. pacificum Yendo) did not show high acidity. Ion chromatography of the cell extracts showed that those species contained high concentrations of SO     within their cells, up to 10 times that in seawater but relatively low Cl⁻. The sum of cations examined (Na⁺, NH     , K⁺, Mg²⁺, Ca²⁺) was significantly lower than that of anions (Cl⁻, Br⁻, NO     , SO     ), and the difference is presumed to represent protons (H⁺), causing the extremely low cell sap pH. Estimated cellular proton concentrations calculated from the pH data roughly agreed with those calculated from differences between the sum of cations and anions and that of anions. Although certain other, nonacidic, dictyotalean species also contained high concentrations of SO     , these species contained high concentrations of Mg²⁺, and the sums of cations and anions were balanced.     

Intrinsic rate of population increase of Amitus fuscipennis MacGown and Nebeker (Hym., Platygasteridae) according to climatic conditions and bean cultivar

Data on adult longevity, reproduction and intrinsic rate of population increase of insect parasitoid Amitus fuscipennis MacGown and Nebeker (Hym., Platygasteridae) were obtained under different climatic conditions on two bean cultivars. The insect host was the greenhouse whitefly Trialeurodes vaporariorum (Westwood) (Hom., Aleyrodidae). On cv. Chocho, at 19°C, 75 ± 5% RH and at 22°C, 70 ± 5% RH, mean longevity of adult Amitus females on the host insect was 9.3 and 5.7 days; mean fertility was 64.1 eggs and 52.7 eggs; and r_m values were 0.102 day^–1 and 0.144 day^–1 for the respective conditions. On cv. ICA-Pijao, at 19°C, 75 ± 5% RH, mean longevity was 15.2 days; mean fertility was highest (162.8 eggs); and r_m value was 0.117 day^–1. The r_m values of Amitus were higher than those of its host under the same test conditions. This parasitoid may therefore be a suitable candidate for use in inoculative or seasonal inoculative control programs against T. vaporariorum .     

α-Synuclein modulation of Ca2+ signaling in human neuroblastoma (SH-SY5Y) cells

Parkinson's disease (PD) is characterized in part by the presence of α-synuclein (α-syn) rich intracellular inclusions (Lewy bodies). Mutations and multiplication of the α-synuclein gene ( SNCA ) are associated with familial PD. Since Ca²⁺ dyshomeostasis may play an important role in the pathogenesis of PD, we used fluorimetry in fura-2 loaded SH-SY5Y cells to monitor Ca²⁺ homeostasis in cells stably transfected with either wild-type α-syn, the A53T mutant form, the S129D phosphomimetic mutant or with empty vector (which served as control). Voltage-gated Ca²⁺ influx evoked by exposure of cells to 50 mM K⁺ was enhanced in cells expressing all three forms of α-syn, an effect which was due specifically to increased Ca²⁺ entry via L-type Ca²⁺ channels. Mobilization of Ca²⁺ by muscarine was not strikingly modified by any of the α-syn forms, but they all reduced capacitative Ca²⁺ entry following store depletion caused either by muscarine or thapsigargin. Emptying of stores with cyclopiazonic acid caused similar rises of [Ca²⁺]_i in all cells tested (with the exception of the S129D mutant), and mitochondrial Ca²⁺ content was unaffected by any form of α-synuclein. However, only WT α-syn transfected cells displayed significantly impaired viability. Our findings suggest that α-syn regulates Ca²⁺ entry pathways and, consequently, that abnormal α-syn levels may promote neuronal damage through dysregulation of Ca²⁺ homeostasis.     

Geographical variation in AdhF and alcoholic resource utilization in Indian populations of Drosophila melanogaster

Indian geographical populations of Drosophila melanogaster, collected at latitude 10–32.3 ^oN, longitude 76.1–80.1 ^oE and altitude 16–2050 m, were analysed for changes in Adh ¹ frequency and utilization potential of diverse alcoholic resources. Until now, there have been limited data on climatic association of correlated changes on these aspects in continental populations of D. melanogaster. Indian populations demonstrate an increase in Adh ^F frequency by 0.04 per degree latitude and significantly higher utilization of ethanol (9.0–15.2%), acetic acid (8.1–12.5%), n-propanol (2.55-4.25%), 2-propanol (1.5-3.5%), n-butanol (2.15 3.5%) and 2-butanol (0.68-1.2%). Data also revealed significant correlation with latitude as well as altitude. Regression analysis of climatic data from collection sites confirmed the observed clinal variation of Adh ^F as well as alcoholic utilization in D. melanogaster. On the Indian subcontinent, variation in Adh ^F frequency and alcoholic resource utilization along increasing latitude as well as altitude are due to climatic selection factors such as temperature and rainfall.     

Nisin, temperature and pH effects on growth and viability of Pectinatus frisingensis, a Gram-negative, strictly anaerobic beer-spoilage bacterium

Pectinatus frisingensis , a Gram-negative and strictly anaerobic beer spoilage bacterium is sensitive to nisin. An increase in nisin concentration (0 to 1100 IU ml⁻¹) added to the culture medium prolonged the lag phase, and decreased the growth rate of the bacterium. In addition, late exponential cells of P. frisingensis exposed to low concentrations of nisin lost immediately a part of their intracellular K⁺. Presence of Mg²⁺ up to 15 mmol l⁻¹ did not protect P. frisingensis from nisin-induced loss of viability and K⁺ efflux. Potassium leaks were also measured in P. frisingensis late exponential phase cells exposed to combined effects of nisin addition (100–500 IU ml⁻¹), 10 min mild heat-treatment (50 °C) or rapid cooling (2 °C), and pH (4·0 and 6·2). Net K⁺ efflux from both starving and glucose-metabolizing cells, was more important at pH 6·2, whatever the temperature treatment and nisin addition. Reincubation at 30 °C of P. frisingensis glucose-metabolizing cells exposed to a preliminary combination of nisin addition and mild heat or cooling down treatment, showed that cells exposed to rapid cooling reaccumulated more K₊ than heat-treated cells, whatever the pH conditions. A combination of nisin and mild heat-treatment could thus be of interest to prevent P. frisingensis growth in beers.     

Mercury in tigerfish (Hydrocynus vittatus, Castelnau), green happy (Sargochromis codringtonii, Boulenger) and kapenta (Limnothrissa miodon, Boulenger) from Lake Kariba, Zimbabwe

Concentrations of total mercury were determined in Hydrocynus vittatus (Castelnau), Sargochromis codringtonii (Boulenger), and Limnothrissa miodon (Boulenger) from two localities in Lake Kariba, Zimbabwe.
The mean concentrations of total mercury in H. vittatus from Basin 5 and Basin 2 were 0.08 mg kg⁻¹ and 0.094 mg kg⁻¹, respectively. In S. codringtonii, mean concentrations were 0.004 mg kg⁻¹ and 0.026 mg kg⁻¹ for Basins 5 and 2, respectively. No mercury was detected in L. miodon from Basin 2 while samples from Basin 5 had a mean concentration of 0.069 mg kg⁻¹ (wet weight). Total mercury concentrations were also determined on a dry weight basis.
Within each sampling area, total mercury concentrations were significantly different among species ( P  < 0.05). For H. vittatus and S. codringtonii, total mercury concentrations (in the same species) were not significantly different between the two localities ( P  < 0.05).
The factors causing the observed differences in total mercury between similar species from different localities and among different species in the same locality (sampling area) are discussed. From the observed low levels of mercury in all three species, it was concluded that the mercury constituted 'background levels'. These levels are below the maximum concentrations permissible in human fish foods.     

Contribution of fungal biomass to the growth of the shredder, Pycnopsyche gentilis (Trichoptera: Limnephilidae)

1.  1. It has been accepted that aquatic hyphomycetes colonising submerged leaves increase the nutritional value of leaf detritus and suggested that fungal biomass plays a greater role in the growth of shredders than leaf tissue itself. However, it is not clear what proportion of the nutritional needs of shredders is met by fungal biomass.
2.  We fed Pycnopsyche gentilis larvae with tulip poplar ( Liriodendron tulipifera ) leaf discs colonised by the aquatic hyphomycete, Anguillospora filiformis , which had been radiolabelled to quantify the contribution of fungal carbon to the growth of the shredder at different larval developmental stages. Instantaneous growth rates of larvae on this diet were also estimated.
3.  When provided with fungal-colonised leaves (14–16% fungal biomass), the third and the fifth instar larvae of P. gentilis grew at the rates of 0.061 and 0.034 day⁻¹, respectively, but on a diet of sterile leaves, both larval instars lost weight. The incorporation rates of fungal carbon were 31.6 μg C mg⁻¹ AFDM day⁻¹, accounting for 100% of the daily growth rate of the third instar larvae and 8.6 μg C mg⁻¹ AFDM day⁻¹, accounting for 50% of the daily growth rate of the fifth instar larvae.
4.  These results suggest that leaf material colonised by A. filiformis is a high quality food resource for P. gentilis larvae, and that fungal biomass can contribute significantly to the growth of these larvae. Differences in feeding behaviour and digestive physiology may explain the significantly greater assimilation of fungal biomass by the earlier instar than the final instar. To satisfy their nutritional needs the fifth instar larvae would have to assimilate detrital mass that may have been modified by fungal exoenzymes.     

Atmospheric nitric oxide stimulates plant growth and improves the quality of spinach (Spinacia oleracea)

Nitric oxide (NO) is an endogenous signalling molecule implicated in a growing number of plant processes and has been recognised as a plant hormone. The present research employed spinach plant ( Spinacia oleracea cv. Huangjia) and closed growth chambers to investigate the effects of gaseous NO application on vegetable production in greenhouses. Treatment of low concentration of NO gas (ambient atmosphere with 200 nL L⁻¹ NO gas) significantly increased the shoot biomass of the soil-cultivated plants as compared with the control treatment (ambient atmosphere). In addition, the NO treatment also increased the photosynthetic rate of leaves, indicating that the enhancement of photosynthesis is an important reason leading to more biomass accumulation induced by NO gas. Furthermore, the NO treatment decreased nitrate concentration but increased the concentrations of soluble sugar, protein, antioxidants (vitamin C, glutathione and flavonoids), and ferric reducing-antioxidant power (FRAP) in shoots of the plants grown in soil, suggesting that the gaseous NO treatment can not only increase vegetable production but also improve vegetable quality. In addition, the effects of the combined application of NO and CO₂ (NO 200 nL L⁻¹ and CO₂ 800 μL L⁻¹) on shoot biomass was even greater than the effects of elevated CO₂ (CO₂ 800 μL L⁻¹) or the NO treatment alone, implying that gaseous NO treatment can be used in CO₂-elevated greenhouses as an effective strategy in improving vegetable production.     

Cellular and biochemical response to Cr(VI) in Stenotrophomonas sp.

Chromium (Cr)-resistant bacteria isolated from a soil with 6 g kg⁻¹ of Cr were identified based on 16S rRNA gene sequence analysis as a Stenotrophomonas , and designated as JD1. Growth of JD1 was accompanied by transformation of Cr(VI) to Cr(III) in liquid medium initially containing 300 mg L⁻¹ Cr(VI), the maximum concentration allowing growth. JD1 produced the highest levels of a Cr(VI)-binding exopolysaccharide when grown in medium with 100 mg L⁻¹ Cr(VI). The relative exopolysaccharide monosaccharide composition was analysed by HPLC, which showed that rhamnose+galactose was the major component, and that its relative level increased when cells were grown with Cr(VI). JD1 grew as a biofilm on various inert surfaces. Biofilm macromolecular composition analysis indicated that the relative levels of exopolysaccharide and protein were more abundant in biofilms grown in 100 mg L⁻¹ Cr(VI), whereas relative uronic acid levels remained constant. Biofilm cells exposed to Cr(VI) were elongated, grouped in clusters and exopolysaccharide obtained from the biofilm extracellular matrix had an enhanced capacity to bind Cr(VI). Exopolysaccharide production and composition, and biofilm growth are discussed as a mechanism of protection that allows survival during Cr(VI) stress.     

Utilization of several plant proteins by gibel carp (Carassius auratus gibelio)

Six isonitrogenous (gross protein content 35%) and isoenergetic (gross energy content 17 kJ g⁻¹) diets were formulated to investigate the effects of inclusion of plant proteins on the gibel carp ( Carassius auratus gibelio L.). The plant proteins tested were: soybean cake (SBC), potato protein concentrate (PPC), peanut cake (PNC), cottonseed cake (CSC) and rapeseed cake (RSC). Fish meal (FM) was used as control. In each diet, 27% of the protein was supplied by fish meal, and the rest supplied by the plant protein tested. Each diet was fed to three groups of gibel carp for 8 weeks in a recirculation system. Specific growth rate (SGR) in fish fed the control diet was significantly higher than those in the other groups, and SGR in fish fed the PPC was significantly lower than in fish fed other plant proteins. There was no significant difference in SGR among the other groups. Feeding rates were ranked in the order: RSC > CSC > FM > PNC > SBC > PPC. Conversion efficiency was highest in groups fed FM, SBC and PNC, followed by groups fed CSC and RSC, and was lowest in the group fed PPC. The fish fed PPC showed lower protein retention than those fed FM and SBC. FM showed highest energy retention while PPC showed lowest. There was no significant relationship between SGR and intake of digestible protein (g g⁻¹ day⁻¹), digestible lysine (g g⁻¹ day⁻¹), digestible methionine (g g⁻¹ day⁻¹) or digestible total essential amino acids (g g⁻¹ day⁻¹), suggesting that the differences in SGR could not alone account for any of these variables.     

Age, growth, mortality and population structure of the oyster, Crassostrea madrasensis, in the Moheskhali Channel (southeastern coast of Bangladesh)

The population structure, age, growth, mortality and harvest intensity of the oyster Crassostrea madrasensis were examined in the Moheskhali Channel, Bangladesh between June 2003 and May 2004. The channel is a representative habitat for the area. C. madrasensis monthly length frequency data were analyzed using FiSAT software for estimating population parameters, including asymptotic length ( L _∝ ), growth co-efficient ( K ) and recruitment pattern to assess the status of the stock. Asymptotic length ( L∝ ) and growth co-efficient ( K ) were 20.88 cm and 0.35 year⁻¹, respectively. The growth performance index (φ') was calculated with 2.18. The growth pattern showed negative allometric growth ( b  < 3), with an asymptotic weight ( W _∝ ) of about 1124.6 g. The oyster attained an average length of 6.17 cm at the end of 1 year. Total mortality ( Z ) by length-converted catch curve was estimated at 1.78 year⁻¹, fishing mortality ( F ) at 0.77 year⁻¹, and natural mortality ( M ) at 1.01 year⁻¹. The exploitation level ( E ) of C. madrasensis was 0.43, while the maximum allowable limit of exploitation ( E _max) was 0.45 for the highest yield. The recruitment pattern was continuous, displaying a single major peak event per year. Habitat temperatures were 25.5–31.0°C (mean ± SD, 29 ± 1.62°C); salinity range was from 12.36 to 26.0 ppt (mean ± SD, 19.6 ± 4.7 ppt). The exploitation level (0.43) indicated that the oyster stock was exploited at almost maximum yield in this channel.     

Age, growth and feeding of the common stingray (Dasyatis pastinaca, L., 1758) in the Cilician coastal basin, northeastern Mediterranean Sea

Growth and feeding of stingray ( Dasyatis pastinaca ) were studied using 346 specimens from the Cilician Basin coastal area (northeastern Mediterranean). Age classes between 0 to XII were found. The total length of all specimens ranged from 14.6 to 100.9 cm, and total weight was between 22.5 and 6800 g. Total length (TL)-weight (W) and disc width (DW)-weight (W) relationships were W = 0.0033*L^3.1429 and W = 0.0039*DW^3.4914, respectively. The age data, derived from central readings, were used to estimate the von Bertalanffy length and weight growth parameters. The results were L_∞ = 294.9 cm, W_∞ = 198690.1 g, K = 0.029 year⁻¹; t₀ = −2.2 year. The D. pastinaca diet was composed mainly of crustaceans.     

Inactivation or removal of the budded particles of a nuclear polyhedrosis virus of a silkworm, Bombyx mori L. (Lep., Bombycidae)

Effective methods to inactivate or remove budded particles of a nuclear polyhedrosis virus of Bombyx mori (BmNPV) from a cell-cultured media or from host haemolymph that is infected by this virus have been developed. Two types of suspensions containing BmNPV budded virus particles, TC-100 media that cultured BmN4 cells infected by this virus and haemolymph of B. mori larvae infected by this virus, were treated by 6% (w/v) polyethylene glycol (PEG), 0.01% (w/v) chitosan, 0.05% (v/v) linoleic acid (an emulsion), and/or diethylether. Treatment by linoleic acid followed by PEG-precipitation and treatment by diethylether followed by PEG-precipitation were so effective that these treatments suppressed the viral titre of BmNPV-infected larval haemolymph from an original titre (> 10⁹ TCID₅₀ units/ml) to below a detectable limit. These methods are suggested as being potentially useful in an insect factory system; that is, a protein production system utilizing a baculovirus vector and its insect host or cultured cells on a large scale.     

Author idex volume 36 (1986)

Abstract Nitrate reduction to ammonia by marine Vibrio species was studied in batch and continuous culture. In pH-controlled batch cultures (pH 7.4; 50 mM glucose, 20 mM KNO₃), the nitrate consumed accumulated to more than 90% as nitrite. Under these conditions, the nitrite reductase (NO⁻₂→ NH₃) was severely repressed. In pH-controlled continuous cultures of V. alginolyticus with glucose or glycerol as substrates ( D = 0.045 h⁻¹) and limiting N-source (nitrate or nitrite), nitrite reductase was significantly derepressed with cellular activities in the range of 0.7–1.2 μmol min⁻¹ (mg protein)⁻¹. The enzyme was purified close to electrophoretic homogeneity with catalytic activity concentrations of about 1800 nkat/mg protein. It catalyzed the reduction of nitrite to ammonia with dithionite-reduced viologen dyes or flavins as electron donors, had an M _r of about 50 000 (determined by gel filtration) and contained c-type heme groups (probably 4–6 per molecule).     

Ecology and exploitation pattern of a landlocked population of sand smelt, Atherina boyeri (Risso 1810), in Trichonis Lake (western Greece)

Age, growth, mortality and exploitation pattern of sand smelt, Atherina boyeri (Risso 1810), in Trichonis Lake (western Greece) were studied from samples taken from catches of local fishermen. Individuals ranged between 44 and 109.53 mm in total length (TL). Age determinations based on scale readings show that the population has a 4-year life cycle. Sand smelt grows allometrically (b=3.18) and relatively rapidly, achieving 52.3% of the growth during the first year; thereafter the annual growth rate drops quickly. The von Bertalanffy growth parameters for all individuals are: L_∞=112.40 mm, k= 0.42 years⁻¹και t_o=−0.40 years. The total mortality rate was Z=1.65 years⁻¹ and the natural mortality rate M=1.07 years⁻¹. The exploitation rate indicates that the population is rather underexploited (E=0.35).     

Evaluation of an entomopathogenic fungus, Paecilomyces fumosoroseus (Wize) Brown and Smith (Deuteromycota: Hyphomycetes) obtained from Formosan subterranean termites (Isop., Rhinotermitidae)

Abstract:  An isolate of Paecilomyces fumosoroseus was obtained from Coptotermes formosanus collected in Hong Kong, and a commercially available isolate of Metarhizium anisopliae , were both tested against C. formosanus shipped live from China. Survivorship of termites treated with a suspension of 5 × 10⁵ M. anisopliae conidia/ml and kept alone declined more rapidly than for those treated at the same concentration of P. fumosoroseus conidia. At a 5 × 10⁶ conidia/ml concentration, no significant differences in terms of termite survivorship were observed between the two fungal species. However, among termites kept in groups of 10 after treatment, those sprayed with P. fumosoroseus conidia at either 5 × 10⁵ or 5 × 10⁶ conidia/ml had significantly lower survivorship than those sprayed with M. anisopliae conidia. All the cadavers of termites treated with P. fumosoroseus and kept alone sporulated and among grouped termites 29% of the cadavers sporulated. By comparison, 53% of the cadavers of termites treated with M. anisopliae and kept alone sporulated, and only 4% of the cadavers of treated termites kept in groups sporulated.