High-yield expression,purification, characterization,and structure determination of tag-free <Emphasis Type="Italic">Candida utilis</Emphasis> uricase

We report the successful high-yield expression of Candida utilis uricase in Escherichia coli and the establishment of an efficient three-step protein purification protocol. The purity of the recombinant protein, which was confirmed to be C. utilis uricase by sodium dodecyl sulfate–polyacrylamide gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight mass spectrometer analysis, was >98% and the specific activity was 38.4 IU/mg. Crystals of C. utilis uricase were grown at 18°C using 25% polyethylene glycol 3350 as precipitant. Diffraction by the crystals extends to 1.93 Å resolution, and the crystals belong to the space group P2₁2₁2₁ with unit cell parameters a?=?69.16 Å, b?=?139.31 Å, c?=?256.33 Å, and α?=?β?=?γ?=?90°. The crystal structure of C. utilis uricase shares a high similarity with other reported structures of the homologous uricases from other species in protein database, demonstrating that the three-dimensional structure of the protein defines critically to the catalytic activities.     

Cold-Set Gelation of Commercial Soy Protein Isolate: Effects of the Incorporation of Locust Bean Gum and Solid Lipid Microparticles on the Properties of Gels

This study aimed to evaluate the ability of commercial soy protein isolate (SPI) to form cold-set gels under different pHs (5–11), pre-heating temperatures (60 °C, 80 °C), CaCl₂ (0–15 mM) and SPI (5–15%, w/v) concentrations, and also select a formulation for the investigation of the effects of incorporating locust bean gum (LBG) (0–0.3%, w/v) and solid lipid microparticles (SLM) on gels rheological and microstructural properties. Gels were evaluated in terms of visual aspect, water-holding capacity, microstructure (using confocal laser scanning microscopy and cryo-scanning electronic microscopy) and rheological properties. SPI showed higher solubilities at pHs 7 (32.0%), 9 (51.6%) and 11 (100%). Self-supported gels were obtained under several conditions at alkaline pHs. At pH 7, only systems pre-heated to 80 °C with 15% (w/v) SPI and 10 or 15 mM CaCl₂ gave self-supported gels. At neutral pH, samples showed relative structural instability, which was minimized with LBG incorporation. Formulations G_SPI (pH 7, preheated to 80 °C, 15% (w/v) SPI, 10 mM CaCl₂) and G_MIX (pH 7, preheated to 80 °C, 15% (w/v) SPI, 0.2% (w/v) LBG, 15 mM CaCl₂) were selected for emulsion-filled gels (EFG) production. Power law parameters (K′, K″), calculated from frequency sweep results, revealed that non-filled G_MIX (K′: 472.1; K″: 77.6) was stronger than G_SPI (K′: 170.4; K″: 33.6). Besides, G_MIX showed microphase separation. SLM stabilized with Tween 80-Span 80 were active fillers in EFG, altering microstructures and increasing G’, G” and the Young’s modulus (1.8 to 2.1 kPa for G_SPI and 1.4 to 2.2 kPa for G_MIX).     

Substrate Temperature Effect on Microstructure,Optical, and Glucose Sensing Characteristics of Pulsed Laser Deposited Silver Nanoparticles

This work reports the substrate temperature-influenced change in the structural, morphological, optical, and glucose sensing properties of silver (Ag) nanoparticles (NPs) deposited on p-type Si (100) wafers. AgNP films grown at temperatures ranging from RT to 600 °C clearly show a dependence of orientation texture and surface morphology on substrate temperature (T _s). As T _s increases from RT towards 600 °C, the preferred orientation of AgNP film changes from (111) to (200). The AgNPs size, that is T _s-dependent, reaches the maximum value at T _s = 300 °C. This result is attributed to restructuring of AgNPs texture. Moreover, the AgNP shape also changes from ellipsoid to sphere as T _s increases from RT to 600 °C. Surface plasmon enhancement in photoluminescence intensity is observed with increase in T _s. It is found also that the AgNP film deposited at 300 °C has considerable reflectance reduction relative to the silicon substrate, in wavelength range of 300–800 nm and a progressive red shift of localized surface plasmon resonances caused by the adding of increasing quantities of glucose has been observed. As a proof of concept, we also demonstrate the capability of grown AgNP substrates for glucose detection based on surface enhanced Raman spectroscopy in physiological concentration range with short integration time 10 s, varying with T _s.     

Effect of Temperature on Development and Reproduction of <Emphasis Type="Italic">Epilachna dodecastigma</Emphasis> (Wied.) (Coleoptera: Coccinellidae)

The effect of five constant temperatures of 21, 24, 27, 30 and 33 °C on adult life span, reproduction, oviposition behavior and larval developmental time of a bitter gourd inhabited coleopteran insect Epilachna dodecastigma (Wied.) (Coccinellidae) was determined in laboratory conditions under 70 ± 5 % relative humidity and a photoperiod of 12 L : 12 D. Larval developmental time of E. dodecastigma decreased as temperature increased from 21 to 33 °C. Life table data revealed that overall mortality was lowest at 27 °C and highest at 21 °C. Females lived longer than males at all temperatures; but longevity decreased with increase in temperature. Pre-oviposition period decreased significantly with increase in temperature up to 27 °C and thereafter increased at a slower rate; whereas oviposition period decreased significantly with increase in temperature. Fecundity and egg viability increased significantly with an increase in temperature up to 27 °C and thereafter decreased at a slower rate. The intrinsic rate of increase (r _m ) was 0.1703, 0.1984, 0.2235, 0.2227 and 0.2181 day^?1 at 21, 24, 27, 30 and 33 °C, respectively. The net reproductive rate and finite rate of increase was highest at 27 °C (R _o  = 112.05; λ = 1.4233) and lowest at 21 °C (R _o  = 51.23; λ = 1.2581).     

Biology and Thermal Requirements of <Emphasis Type="Italic">Fopius arisanus</Emphasis> (Sonan, 1932) (Hymenoptera: Braconidae) Reared on <Emphasis Type="Italic">Ceratitis capitata</Emphasis> Eggs (Wiedemann) (Diptera: Tephritidae)

Fopius arisanus (Sonan) is a solitary parasitoid of eggs and the first instar larvae of Tephritidae. Due to the occurrence of Ceratitis capitata (Wiedemann) in various regions and under several climatic conditions, this study aimed to evaluate the effect of different temperatures on the embryonic development (egg–adult) and determine thermal requirements and the number of annual generations F. arisanus on eggs of C. capitata. In the laboratory, eggs of C. capitata (24 h) were submitted to parasitism of F. arisanus during 6 h. Later, the eggs were placed in plastic containers (50 mL) (50 eggs/container) on a layer of artificial diet and packed in chambers at temperatures 15, 18, 20, 22, 25, 28, 30, and 32 ± 1°C, RH 70 ± 10%, and a photophase of 12 h. The largest number of offspring, emergence rate, and weight of adults of F. arisanus were observed at 25°C. The highest sex ratios (sr > 0.75) were recorded at 15 and 18°C, being statistically higher than the temperatures 20°C (0.65), 22°C (0.64), 25°C (0.65), 28°C (0.49), and 30°C (0.47). At 32°C, there was no embryonic development of F. arisanus. The egg–adult period was inversely proportional to temperature. Based on the development of the biological cycle (egg–adult), the temperature threshold (T _t) was 10.3°C and thermal constant (K) of 488.34 degree-days, being the number of generations/year directly proportional to the temperature increase. The data show the ability of F. arisanus to adapt to different thermal conditions, which is important for biological control programs of C. capitata.     

<Emphasis Type="Italic">Paradevosia shaoguanensis</Emphasis> gen. nov., sp. nov., Isolated from a Coking Wastewater

A Gram staining negative, rod-shaped, aerobic bacterial strain J5-3^T with a single polar flagellum was isolated from coking wastewater collected from Shaoguan, Guangdong, China. It was motile and capable of optimal growth at pH 6–8, 30 °C, and 0–2 % (w/v) NaCl. Its predominant fatty acids were 11-methyl C_18:1 ω7c (29.2 %), C_16:0 (20.6 %), C_19:0 cyclo ω8c (18.2 %), C_18:0 (11.0 %), and C_18:1 ω7c/C_18:1 ω6c (10.9 %) when grown on trypticase soy agar. The major polar lipids were diphosphatidylglycerol, phosphatidylglycerol, two unknown glycolipids (GL1, GL2), and two unknown phospholipid (PL1, PL2). The predominant ubiquinone was Q-10, and the genome DNA G+C content was 61.7 mol %. Phylogenetic analysis based on the 16S rRNA gene sequences indicated that strain J5-3^T belonged to the family Hyphomicrobiaceae in Alphaproteobacteria. It shared the 16S rRNA gene sequence similarities of 93.8–96.1 % with the genus Devosia, 94.5–94.8 % with the genus Pelagibacterium, and <92.0 % with all the other type strains in family Hyphomicrobiaceae. It can be distinguished from the closest phylogenetic neighbors based on several phenotypic and genotypic features, including α-galactosidase activity, tetracycline susceptibility, major fatty acid composition, polar lipid profile, DNA gyrase B subunit (gyrB) gene sequence, and random-amplified polymorphic DNA profile. Therefore, we consider strain J5-3^T to represent a novel species of a novel genus within the family Hyphomicrobiaceae, for which the name Paradevosia shaoguanensis gen. nov., sp. nov. is proposed. The type strain of Paradevosia shaoguanensis is J5-3^T (=CGMCC 1.12430^T =LMG 27409^T).     

Diversity in growth patterns among strains of the lethal fungal pathogen <Emphasis Type="Italic">Batrachochytrium dendrobatidis</Emphasis> across extended thermal optima

The thermal sensitivities of organisms regulate a wide range of ecological interactions, including host–parasite dynamics. The effect of temperature on disease ecology can be remarkably complex in disease systems where the hosts are ectothermic and where thermal conditions constrain pathogen reproductive rates. Amphibian chytridiomycosis, caused by the pathogen Batrachochytrium dendrobatidis (Bd), is a lethal fungal disease that is influenced by temperature. However, recent temperature studies have produced contradictory findings, suggesting that our current understanding of thermal effects on Bd may be incomplete. We investigated how temperature affects three different Bd strains to evaluate diversity in thermal responses. We quantified growth across the entire thermal range of Bd, and beyond the known thermal limits (T _max and T _min). Our results show that all Bd strains remained viable and grew following 24 h freeze (?12 °C) and heat shock (28 °C) treatments. Additionally, we found that two Bd strains had higher logistic growth rates (r) and carrying capacities (K) at the upper and lower extremities of the temperature range, and especially in low temperature conditions (2–3 °C). In contrast, a third strain exhibited relatively lower growth rates and carrying capacities at these same thermal extremes. Overall, our results suggest that there is considerable variation among Bd strains in thermal tolerance, and they establish a new thermal sensitivity profile for Bd. More generally, our findings point toward important questions concerning the mechanisms that dictate fungal thermal tolerances and temperature-dependent pathogenesis in other fungal disease systems.     

Deciphering the mode of action,structural and biochemical analysis of heparinase II/III (<Emphasis Type="Italic">Ps</Emphasis>PL12a) a new member of family 12 polysaccharide lyase from <Emphasis Type="Italic">Pseudopedobacter saltans</Emphasis>

Heparinases are widely used for production of clinically and therapeutically important bioactive oligosaccharides and in analyzing the polydisperse, heterogeneous, and complex structures of heparin/heparan sulfate. In the present study, the gene (1911 bp) encoding heparinase II/III of family 12 polysaccharide lyase (PsPL12a) from Pseudopedobacter saltans was cloned, expressed, and biochemically and functionally characterized. The purified enzyme PsPL12a of molecular size approximately 76 kDa exhibited maximum activity in the temperature range 45–50 °C and at pH 6.0. PsPL12a gave maximum activity at 1% (w/v) heparin under optimum conditions. The kinetic parameters, K _m and V_max, for PsPL12a were 4.6?±?0.5 mg/ml and 70?±?2 U/mg, respectively. Ten millimolars of each Mg²⁺ and Mn²⁺ ions enhanced PsPL12a activity by 80%, whereas Ni²⁺ inhibited by 75% and Co²⁺ by 10%, and EDTA completely inactivated the enzyme. Protein melting curve of PsPL12a gave a single peak at 55 °C and 10 mM Mg²⁺ ions and shifted the peak to 60 °C. The secondary structure analysis of PsPL12a by CD showed 65.12% α-helix, 11.84% β-strand, and 23.04% random coil. The degradation products of heparin by PsPL12a analyzed by ESI-MS spectra displayed peaks corresponding to heparin di-, tetra-, penta-, and hexa-saccharides revealing the endolytic mode of enzyme action. Heparinase II/III (PsPL12a) from P. saltans can be used for production of low molecular weight heparin oligosaccharides for their utilization as anticoagulants. This is the first report on heparinase cloned from P. saltans.     

Expression and characterization of glutamate decarboxylase from <Emphasis Type="Italic">Lactobacillus brevis</Emphasis> HYE1 isolated from <Emphasis Type="Italic">kimchi</Emphasis>

A putative gene (gadlbhye1) encoding glutamate decarboxylase (GAD) was cloned from Lactobacillus brevis HYE1 isolated from kimchi, a traditional Korean fermented vegetable. The amino acid sequences of GADLbHYE1 showed 48% homology with the GadA family and 99% identity with the GadB family from L. brevis. The cloned GADLbHYE1 was functionally expressed in Escherichia coli using inducible expression vectors. The expressed recombinant GADLbHYE1 was successfully purified by Ni–NTA affinity chromatography, and had a molecular mass of 54 kDa with optimal hydrolysis activity at 55 °C and pH 4.0. Its thermal stability was determined to be higher than that of other GADs from L. brevis, based on its melting temperature (75.18 °C). Kinetic parameters including K_m and V_max values for GADLbHYE1 were 4.99 mmol/L and 0.224 mmol/L/min, respectively. In addition, the production of gamma-aminobutyric acid in E. coli BL21 harboring gadlbhye1/pET28a was increased by adding pyridoxine as a cheaper coenzyme.     

Molecular and cytogenetic analyses provide evidence of the introgression of chromosomal segments from the wild <Emphasis Type="Italic">Cucumis hystrix</Emphasis> into the cultivated cucumber through the bridge of a synthetic allotetraploid

Cucumis × hytivus (2n = 4× = 38) is a synthetic allotetraploid obtained from interspecific hybridization between the cucumber (2n = 2× = 14) and its wild relative C. hystrix (2n = 2× = 24). The synthesis of this species built a bridge for cucumber improvement through gene introgression. Allotriploid and introgression lines (ILs) have previously been produced and characterized with respect to morphology, cytology, and molecular markers. However, no clear evidence of how the chromosomal segments of C. hystrix were introgressed and inherited was found owing to the small size of chromosomes. In the present study, cucumber-C. hystrix introgression lines were developed by backcrossing the allotriploid to North China cucumber breeding line “P01” followed by self-pollination. The introgressed segments of C. hystrix in the ILs were revealed by meiotic pachytene chromosome analysis. Fluorescence in situ hybridization (FISH) was performed on pachytene chromosomes using fosmid clones from cucumber, which confirmed that introgression occurred in the long arm of chromosome 7. Molecular analysis using a set of 53 simple sequence repeats (SSRs) indicated that the chromosomal segments of C. hystrix were introduced into 4 cucumber chromosomes, the short arms of chromosomes 2 and 6, and long arms of chromosomes 3 and 7. The inheritance of alien sequences in the long arm of chromosome 7 was investigated with 21 SSRs in self-pollinated progenies. C. hystrix-specific bands of several SSRs were still present in some individuals, indicating that the introgressed segment was partially preserved. The first unambiguous identification of alien chromosome segments in cucumber ILs using combined molecular cytogenetics could facilitate the determination of effects of wild alleles and promote cucumber improvement.     

A review of the history of research and control of <Emphasis Type="Italic">Rhipicephalus</Emphasis> (<Emphasis Type="Italic">Boophilus</Emphasis>) <Emphasis Type="Italic">microplus</Emphasis>, babesiosis and anaplasmosis in Uruguay

The effect of five constant temperatures (16, 20, 24, 28 and 32 °C) on the development, survival and reproduction of Tetranychus cinnabarinus (Boisduval) [=?Tetranychus urticae Koch (red form)] fed on cassava leaves was examined in the laboratory at 85% relative humidity. Development time of various immature stages decreased with increasing temperature, with total egg-to-adult development time varying from 27.7 to 6.7 days. The lower thermal threshold for development was 10.8 °C and the thermal constant from egg to adult was 142.4 degree-days. Pre- and post-oviposition period and female longevity all decreased as temperature increased. The longest oviposition period was observed at 20 °C with 20.4 days. Under different temperatures, mated females laid, on average, 1.0, 2.9, 4.7, 4.7 and 4.9 eggs per day, respectively. The maximum fecundity (81.5 eggs per female) was at 28 °C and the intrinsic rate of increase (r _m ) was highest (0.25) at 32 °C. The results of this study indicate that T. cinnabarinus population could increase rapidly when cassava leaves serve as a food source. At the appropriate temperature T. cinnabarinus could seriously threaten growth of cassava.     

A novel thermostable and organic solvent-tolerant lipase from <Emphasis Type="Italic">Xanthomonas oryzae</Emphasis> pv. <Emphasis Type="Italic">oryzae</Emphasis> YB103: screening,purification and characterization

Thermostable lipases offer major biotechnological advantages over mesophilic lipases. In this study, an intracellular thermostable and organic solvent-tolerant lipase-producing strain YB103 was isolated from soil samples and identified taxonomically as Xanthomonas oryzae pv. oryzae. The lipase from X. oryzae pv. oryzae YB103 (LipXO) was purified 101.1-fold to homogeneity with a specific activity of 373.9 U/mg. The purified lipase showed excellent thermostability, exhibiting 51.1 % of its residual activity after incubation for 3 days at 70 °C. The enzyme showed optimal activity at 70 °C, suggesting it is a thermostable lipase. LipXO retained 75.1–154.1 % of its original activity after incubation in 20 % (v/v) hydrophobic organic solvents at 70 °C for 24 h. Furthermore, LipXO displayed excellent stereoselectivity (e.e._p >99 %) toward (S)-1-phenethyl alcohol in n-hexane. These unique properties of LipXO make it promising as a biocatalyst for industrial processes.     

Genetic population structure and low genetic diversity in the over-exploited sea cucumber <Emphasis Type="Italic">Holothuria edulis</Emphasis> Lesson, 1830 (Echinodermata: Holothuroidea) in Okinawa Island

Understanding genetic connectivity is fundamental for ecosystem-based management of marine resources. Here we investigate the metapopulation structure of the edible sea cucumber Holothuria edulis Lesson, 1830 across Okinawa Island, Japan. This species is of economic and ecological importance and is distributed from the Red Sea to Hawai‘i. We examined sequence variation in fragments of mitochondrial cytochrome oxidase subunit I (COI) and 16S ribosomal RNA (16S), and nuclear histone (H3) at six locations across Okinawa Island. We found higher haplotype diversity for mtDNA (COI: Hd = 0.69 and 16S: Hd = 0.67) and higher heterozygosity of nDNA (H3: H _E = 0.39) in populations from the west coast of Okinawa compared to individuals from populations on the east coast (COI: Hd = 0.40; 16S: Hd = 0.21; H3: H _E = 0.14). Overall population structure was significant (AMOVA results for COI: Φ _ST = 0.49, P < 0.0001; 16S: Φ _ST = 0.34, P < 0.0001; H3: Φ _ST = 0.12, P < 0.0001). One population in the east, Uruma, showed elevated pairwise Φ _ST values in comparisons with all other sites and a marked reduction of genetic diversity (COI: Hd = 0.25 and 16S: Hd = 0.24), possibly as a consequence of a shift to a more dominant asexual reproduction mode. Recent reports have indicated that coastal development in this area influences many marine organisms, and ecosystem degradation in this location could cause the observed decrease of genetic diversity and isolation of H. edulis in Uruma. Our study should provide valuable data to help with the urgently needed management of sea cucumber populations in Okinawa, and indicates particular attention needs to be paid to vulnerable locations.     

Comparative Pharmaceutical Evaluation of Candesartan and Candesartan Cilexetil: Physicochemical Properties, <Emphasis Type="Italic">In Vitro</Emphasis> Dissolution and <Emphasis Type="Italic">Ex Vivo In Vivo</Emphasis> Studies

The aim of the present work is to answer the question is it possible to replace the ester prodrug candesartan cilexetil (CC) by its active metabolite candesartan (C) to bypass the in vivo variable effect of esterase enzymes. A comparative physicochemical evaluation was conducted through solubility, dissolution, and stability studies; additionally, ex vivo permeation and in vivo studies were assessed. C demonstrated higher solubility over CC at alkaline pH. Moreover, dissolution testing using the pharmacopeial method showed better release profile of C even in the absence of surfactant in the testing medium. Both drugs demonstrated a slight degradation in acidic pH after short-term stability. Instead, shifting to alkaline pH of 6.5 and 7.4 showed superiority of C solution stability compared to CC solution. The ex vivo permeation results demonstrated that the parent compound C has a significant (P < 0.05) enhanced permeation compared to its prodrug from CC, that agreed with in vivo results in which C suspension reached significantly (P < 0.05) higher C _max of 1.39 ± 0.59 μg/mL at T _max of 0.66 ± 0.11 h, while CC suspension reached C _max of 0.47 ± 0.22 μg/mL at T _max of 2.00 ± 0.27 h, a lag period of 40 min is needed prior to detection of any absorbed CC in plasma. Those findings are not in agreement with the previously reported rationale on the prodrug formation owing to the poor permeability of the parent compound, suggesting the possibility of marketing the parent drug candesartan for clinical use similarly to azilsartan and its prodrug.     

Bacterial metabolites from intra- and inter-species influencing thermotolerance: the case of <Emphasis Type="Italic">Bacillus cereus</Emphasis> and <Emphasis Type="Italic">Geobacillus stearothermophilus</Emphasis>

Bacterial metabolites with communicative functions could provide protection against stress conditions to members of the same species. Yet, information remains limited about protection provided by metabolites in Bacillus cereus and inter-species. This study investigated the effect of extracellular compounds derived from heat shocked (HS) and non-HS cultures of B. cereus and Geobacillus stearothermophilus on the thermotolerance of non-HS vegetative and sporulating B. cereus. Cultures of B. cereus and G. stearothermophilus were subjected to HS (42 or 65 °C respectively for 30 min) or non-HS treatments. Cells and supernatants were separated, mixed in a combined array, and then exposed to 50 °C for 60 min and viable cells determined. For spores, D values (85 and 95 °C) were evaluated after 120 h. In most cases, supernatants from HS B. cereus cultures added to non-HS B. cereus cells caused their thermotolerance to increase (D ₅₀ 12.2–51.9) when compared to supernatants from non-HS cultures (D ₅₀ 7.4–21.7). While the addition of supernatants from HS and non-HS G. stearothermophilus cultures caused the thermotolerance of non-HS cells from B. cereus to decrease initially (D ₅₀ 3.7–7.1), a subsequent increase was detected in most cases (D ₅₀ 18–97.7). In most cases, supernatants from sporulating G. stearothermophilus added to sporulating cells of B. cereus caused the thermotolerance of B. cereus 4810 spores to decline, whereas that of B. cereus 14579 increased. This study clearly shows that metabolites in supernatants from either the same or different species (such as G. stearothermophilus) influence the thermotolerance of B. cereus.     

Molecular Cloning,Expression and Characterization of Pectin Methylesterase (<Emphasis Type="Italic">Ct</Emphasis>PME) from <Emphasis Type="Italic">Clostridium thermocellum</Emphasis>

Many phytopathogenic micro-organisms such as bacteria and fungi produce pectin methylesterases (PME) during plant invasion. Plants and insects also produce PME to degrade plant cell wall. In the present study, a thermostable pectin methylesterase (CtPME) from Clostridium thermocellum belonging to family 8 carbohydrate esterase (CE8) was cloned, expressed and purified. The amino acid sequence of CtPME exhibited similarity with pectin methylesterase from Erwinia chrysanthemi with 38% identity. The gene encoding CtPME was cloned into pET28a(+) vector and expressed using Escherichia coli BL21(DE3) cells. The recombinant CtPME expressed as a soluble protein and exhibited a single band of molecular mass approximately 35.2 kDa on SDS-PAGE gels. The molecular mass, 35.5 kDa of the enzyme, was also confirmed by MALDI-TOF MS analysis. Notably, highest protein concentration (11.4 mg/mL) of CtPME was achieved in auto-induction medium, as compared with LB medium (1.5 mg/mL). CtPME showed maximum activity (18.1 U/mg) against citrus pectin with >85% methyl esterification. The optimum pH and temperature for activity of CtPME were 8.5 and 50 °C, respectively. The enzyme was stable in pH range 8.0–9.0 and thermostable between 45 and 70 °C. CtPME activity was increased by 40% by 5 mM Ca²⁺ or Mg²⁺ ions. Protein melting curve of CtPME gave a peak at 80 °C. The peak was shifted to 85 °C in the presence of 5 mM Ca²⁺ ions, and the addition of 5 mM EDTA shifted back the melting peak to 80 °C. CtPME can be potentially used in food and textile industry applications.     

Using hydrothermal time concept to describe sesame (<Emphasis Type="Italic">Sesamum indicum</Emphasis> L.) seed germination response to temperature and water potential

To quantify both temperature (T) and water potential (ψ) effects on sesame (Sesamum indicum L.) seed germination (SG) and also to determine the cardinal T _s for this plant, a laboratory experiment was carried out using hydrothermal time model (HTT). For this purpose, four sesame cultivars (‘Asbomahalleh’, ‘Darab’, ‘Dashtestan’ and ‘Yellowhite’) were germinated at seven constant T _s (20, 25, 30, 35, 37, 39 and 43 °C) at each of the following ψ _s (0, ? 0.12, ? 0.24 and ? 0.36 MPa; provided by PEG 8000). Germination rate (GR) and germination percentage (GP) significantly influenced by ψ, T and their interactions in all cultivars (P ≤ 0.01). There was no significant difference, based on the confidence intervals of the model coefficients, between cultivars, so an average of cardinal T _s was 14.7, 35.4 and 47.2 °C for the minimum (T _b), optimum (T _o) and maximum (T _c) T _s, respectively, in the control condition (0 MPa). Hydrotime values in all cultivars decreased when T was increased to T _o and then remained constant at T _s > T _o (15 MPa h^?1). An average value of ψ _b(50) was estimated to be ? 1.23 MPa at T _s ≤ T _o and then increased linearly (0.1041 MPa°Ch^?1, the slope of the relationship between ψ _b(50) and supra-optimal T _s) with T when T _s increased above T _o and finally reached to zero at T _c. The T _b and T _o values were not influenced by ψ, but T _c value decreased (from 47.2 for zero to 43.5 °C for ? 0.36 MPa) at supra-optimal T _s as a result of the effect of ψ on GR. Based on our findings, this model (as a predictive tool) and or the estimated parameter values in this study can easily be used in sesame SG simulation models to quantitatively characterize the physiological status of sesame seed populations at different T _s and ψ _s.     

<Emphasis Type="Italic">Talaromyces heiheensis</Emphasis> and <Emphasis Type="Italic">T. mangshanicus</Emphasis>, two new species from China

Two new species, Talaromyces heiheensis from rotten wood and T. mangshanicus isolated from soil, are illustrated and described as new to science in sections Trachyspermi and Talaromyces. The phylogenetic positions of the two new species inferred from the internal transcribed spacer, beta-tubulin, calmodulin and RNA polymerase II second largest subunit regions were carried out. Talaromyces heiheensis is phylogenetically closely related to T. albobiverticillius, T. rubrifaciens, T. solicola and T. erythromellis, and characterised by slow growth on Czapek yeast autolysate agar at 25 °C, orange conidia en masse on malt extract agar at 25 °C, biverticillate and terverticillate conidiophores, acerose phialides and subglobose to ellipsoidal, smooth-walled conidia. Talaromyces mangshanicus is related to T. kendrickii, T. qii and T. thailandensis, and characterised by slow-growing colonies with absent or sparse sporulation on CYA agar at 25 °C, conidia en masse greyish purple, purplish red soluble pigment on yeast extract agar (YES) at 25 °C, biverticillate conidiophores, ampulliform phialides and subglobose to ellipsoidal conidia with echinulate walls. They are distinguished from the known species in culture characteristics on four standard media, microscopic features and sequence data.     

Impact of Preservation Conditions on Fatty Acids,Xanthan Gum Production and Other Characteristics of <Emphasis Type="Italic">Xanthomonas campestris</Emphasis> pv. <Emphasis Type="Italic">mangiferaeindicae</Emphasis> IBSBF 2103

The conditions of storage, cultivation and maintenance of microbial cultures should preserve the microbiological homogeneity, phenotypic and genotypic characteristics to ensure better reproducibility of metabolic production. To evaluate the influence of the storage condition on the composition of cell fatty acids, genetic profile and biochemical characteristics of Xanthomonas campestris pv. mangiferaeindicae IBSBF 2103, as well as, to identify its relationship with the yielding and viscosity of the xanthan gum produced, this study monitored the strain preserved in two simple and widely used conditions, ultra-freezer (?80 °C) and refrigeration (3–8 °C) during 5 months. Were identified and quantified 13 fatty acids. The cells preserved at ?80 °C showed more stable concentration of all fatty acids, producing more xanthan gum and with higher viscosity. The chromosomal analysis obtained with the enzyme XbaI revealed 17 distinct fragments with maximum size of 485 kilobases, without variations among the subcultures maintained in both storage conditions. The X. campestris pv. mangiferaeindicae subcultures preserved at ?80 °C showed less pronounced phenotypic variations, which had positive influence in the qualitative and quantitative characteristics of the xanthan gum produced.     

Nisin Z produced by <Emphasis Type="Italic">Lactococcus lactis</Emphasis> from bullfrog hatchery is active against <Emphasis Type="Italic">Citrobacter freundii</Emphasis>, a red-leg syndrome related pathogen

Lactococcus lactis subsp. lactis CRL 1584 isolated from a bullfrog hatchery produces a bacteriocin that inhibits both indigenous Citrobacter freundii (a Red-Leg Syndrome related pathogen) and Lactobacillus plantarum, and Listeria monocytogenes as well. Considering that probiotics requires high cell densities and/or bacteriocin concentrations, the effect of the temperature on L. lactis growth and bacteriocin production was evaluated to find the optimal conditions. Thus, the growth rate was maximal at 36 °C, whereas the highest biomass and bacteriocin activity was achieved between 20 and 30 °C and 20–25 °C, respectively. The bacteriocin synthesis was closely growth associated reaching the maximal values at the end of the exponential phase. Since bacteriocins co-production has been evidenced in bacterial genera, a purification of the bacteriocin/s from L. lactis culture supernatants was carried out. The active fraction was purified by cationic-exchange chromatography and then, a RP-HPLC was carried out. The purified sample was a peptide with a 3353.05 Da, a molecular mass that matches nisin Z, which turned out to be the only bacteriocin produced by L. lactis CRL 1584. Nisin Z showed bactericidal effect on C. freundii and L. monocytogenes, which increased in the presence l-lactic acid?+?H₂O₂. This is the first report on nisin Z production by L. lactis from a bullfrog hatchery that resulted active on a Gram-negative pathogen. This peptide has potential probiotic for raniculture and as food biopreservative for bullfrog meat.