Cometabolic biodegradation of quizalofop-p-ethyl by Methylobacterium populi YC-XJ1 and identification of QPEH1 esterase

BACKGROUND: Quizalofop-p-ethyl (QPE), a unitary R configuration aromatic oxyphenoxypropionic acid ester (AOPP) herbicide, was widely used and had led to detrimental environmental effects. For finding the QPEdegrading bacteria and promoting the biodegradation of QPE, a series of studies were carried out. RESULTS: A QPE-degrading bacterial strain YC-XJ1 was isolated from desert soil and identified as Methylobacterium populi, which could degrade QPE with methanol by cometabolism. Ninety-seven percent of QPE (50 mg/L) could be degraded within 72 h under optimum biodegradation condition of 35°C and pH 8.0. The maximum degradation rate of QPE was 1.4 mg/L/h, and the strain YC-XJ1 exhibited some certain salinity tolerance. Two novel metabolites, 2-hydroxy-6-chloroquinoxaline and quinoxaline, were found by high-performance liquid chromatography/mass spectroscopy analysis. The metabolic pathway of QPE was predicted. The catalytic efficiency of strain YC-XJ1 toward different AOPPs herbicides in descending order was as follows: haloxyfop-pmethyl ≈ diclofop-methyl ≈ fluazifop-p-butyl N clodinafop-propargyl N cyhalofop-butyl N quizalofop-p-ethyl N fenoxaprop-p-ethyl N propaquizafop N quizalofop-p-tefuryl. The genome of strain YC-XJ1 was sequenced using a combination of PacBio RS II and Illumina platforms. According to the annotation result, one α/ß hydrolase gene was selected and named qpeh1, for which QPE-degrading function has obtained validation. Based on the phylogenetic analysis and multiple sequence alignment with other QPE-degrading esterases reported previously, the QPEH1 was clustered with esterase family V. CONCLUSION: M. populi YC-XJ1 could degrade QPE with a novel pathway, and the qpeh1 gene was identified as one of QPE-degrading esterase gene.

The improvement of competitive saprophytic capabilities of Trichoderma species through the use of chemical mutagens

Abstract The antagonistic potential of Trichoderma strains was assayed by studying the effect of their culture filtrate on the radial growth of Sclerotium rolfsii, the causal agent of chickpea collar rot. Trichoderma harzianum-1432 (42.2%) and Trichoderma atroviride (40.3%) were found to be strong antagonists. To enhance their antagonistic potential, mutagenesis of these two selected strains was performed. Two mutants, Th-m1 and T. atroviride m1, were found to be more effective than their parent strains. The enzymatic activities of the selected parent and mutant strains were assayed, and although both mutants were found to have enhanced enzymatic activities compared to their respective parent strains, Th-m1 possessed the maximum cellulase (5.69 U/mL) and β-1,3-glucanase activity (61.9 U/mL). Th-m1 also showed high competitive saprophytic ability (CSA) among all of the selected parent and mutant strains, and during field experiments, Th-m1 was found to successfully possess enhanced disease control (82.9%).

Halotolerant bacteria in the São Paulo Zoo composting process and their hydrolases and bioproducts

Halophilic microorganisms are able to grow in the presence of salt and are also excellent source of enzymes and biotechnological products, such as exopolysaccharides (EPSs) and polyhydroxyalkanoates (PHAs). Salt-tolerant bacteria were screened in the Organic Composting Production Unit (OCPU) of São Paulo Zoological Park Foundation, which processes 4 ton/day of organic residues including plant matter from the Atlantic Rain Forest, animal manure and carcasses and mud from water treatment. Among the screened microorganisms, eight halotolerant bacteria grew at NaCl concentrations up to 4 M. These cultures were classified based on phylogenetic characteristics and comparative partial 16S rRNA gene sequence analysis as belonging to the genera Staphylococcus, Bacillus and Brevibacterium. The results of this study describe the ability of these halotolerant bacteria to produce some classes of hydrolases, namely, lipases, proteases, amylases and cellulases, and biopolymers. The strain characterized as of Brevibacterium avium presented cellulase and amylase activities up to 4 M NaCl and also produced EPSs and PHAs. These results indicate the biotechnological potential of certain microorganisms recovered from the composting process, including halotolerant species, which have the ability to produce enzymes and biopolymers, offering new perspectives for environmental and industrial applications.

Alkali production in the mouth and its relationship with certain patient's characteristics

Objectives To assess the relationships among alkali production, diet, oral health behaviors, and oral hygiene. Methods Data from 52 subjects including demographics, diet, and oral hygiene scores were analyzed against the level of arginine and urea enzymes in plaque and saliva samples. An oral habit survey was completed that included: use of tobacco (TB), alcohol (AH), sugary drinks (SD), and diet. Alkali production through arginine deiminase (ADS) and urease activities were measured in smooth-surface supragingival dental plaque and un stimulated saliva samples from all subjects. ADS and urease activities were measured by quantification of the ammonia generated from the incubation of plaque or saliva samples. Spearman correlations were used to compute all associations. Results Participants in the lowest SES (Socio-economic status) group had the habit of consuming sugary drinks the most and had the highest rate of tobacco use. Males consumed significantly more alcohol than females. No significant relationship was found between age or gender and alkali production. Higher rates of sugary drink consumption and tobacco use were significantly related to lower alkali production. Conclusion The study showed a relationship between alkali production and oral hygiene, diet, and certain oral health behaviors. Poor oral hygiene was significantly associated with age, lower SES, tobacco use, and alcohol, and sugary drinks consumption. Clinical relevance Certain oral health behaviors have an impact on oral hygiene and on alkali production; it is important to address these factors with patients as a strategy for caries control. .

Caries-free subjects have high levels of urease and arginine deiminase activity

Objectives: This study investigated the relationship between urease and arginine deiminase system (ADS) activities and dental caries through a cross-sectional study. Material and Methods: Urease and ADS activities were measured in saliva and plaque samples from 10 caries-free subjects and 13 caries-active. Urease activity was obtained from the ammonia produced by incubation of plaque and saliva samples in urea. ADS activity was obtained from the ammonia generated by the arginine-HCl and Tris-maleate buffer. Specific activity was defined as micromoles of ammonia per minute per milligram of protein. Shapiro-Wilk statistical test was used to analyze the distribution of the data, and Mann-Whitney test was used to determine the significance of the data. Results: The specific urease activity in saliva and plaque was significantly higher in individuals with low DMFT scores. ADS activity in saliva (6.050 vs 1.350, p=0.0154) and plaque (8.830 vs 1.210, p=0.025) was also higher in individuals with low DMFT scores. Conclusions: Caries-free subjects had a higher ammonia generation activity by urease and arginine deiminase system for both saliva and plaque samples than low caries-active subjects. High levels of alkali production in oral environment were related to caries-free subjects. .

Screening and isolation of halophilic bacteria producing industrially important enzymes

Halophiles are excellent sources of enzymes that are not only salt stable but also can withstand and carry out reactions efficiently under extreme conditions. The aim of the study was to isolate and study the diversity among halophilic bacteria producing enzymes of industrial value. Screening of halophiles from various saline habitats of India led to isolation of 108 halophilic bacteria producing industrially important hydrolases (amylases, lipases and proteases). Characterization of 21 potential isolates by morphological, biochemical and 16S rRNA gene analysis found them related to Marinobacter, Virgibacillus, Halobacillus, Geomicrobium, Chromohalobacter, Oceanobacillus, Bacillus, Halomonas and Staphylococcus genera. They belonged to moderately halophilic group of bacteria exhibiting salt requirement in the range of 3-20%. There is significant diversity among halophiles from saline habitats of India. Preliminary characterization of crude hydrolases established them to be active and stable under more than one extreme condition of high salt, pH, temperature and presence of organic solvents. It is concluded that these halophilic isolates are not only diverse in phylogeny but also in their enzyme characteristics. Their enzymes may be potentially useful for catalysis under harsh operational conditions encountered in industrial processes. The solvent stability among halophilic enzymes seems a generic novel feature making them potentially useful in non-aqueous enzymology.

Agro-industrial residues and starch for growth and co-production of Polyhydroxyalkanoate copolymer and alfa-amylase by Bacillus. SP CFR67îen

Polyhydroxyalkanoates (PHA) and α-amylase (α-1,4 glucan-4-glucanohydrolase, E.C. 3.2.1.1) were co-produced by Bacillus sp. CFR-67 using unhydrolysed corn starch as a substrate. Bacterial growth and polymer production were enhanced with the supplementation of hydrolysates of wheat bran (WBH) or rice bran (RBH) individually or in combination (5-20 g L-1, based on weight of soluble substrates-SS). In batch cultivation, a mixture of WBH and RBH (1:1, 10 g L-1 of SS) along with ammonium acetate (1.75 g L-1) and corn starch (30 g L-1) produced maximum quantity of biomass (10 g L-1) and PHA (5.9 g L-1). The polymer thus produced was a copolymer of polyhydroxybutyrate-co-hydroxyvalerate of 95:5 to 90:10 mol%. Presence of WBH and corn starch (10-50 g L-1) in the medium enhanced fermentative yield of α-amylase (2-40 U mL-1 min-1). The enzyme was active in a wide range of pH (4-9) and temperature (40-60ºC). This is the first report on simultaneous production of copolymer of bacterial PHA and α-amylase from unhydrolysed corn starch and agro-industrial residues as substrates.

Variability in the production of extracellular enzymes by entomopathogenic fungi grown on different substrates

Entomopathogenic fungi are important controllers of pest-insects populations in agricultural production systems and in natural environment. These fungi have enzymatic machinery which involve since the recognition and adherence of spores in their hosts culminating with infection and death of these insects. The main objective of this study was to analyzed extracellular enzyme production of the fungi strains Beauveria bassiana, Metarhizium anisopliae and Paecilomyces sp when cultured on substrates. These fungi were grown in minimal media containing specific substrates for the analysis of different enzymes such as amylases, cellulases, esterases, lipases, proteases (gelatin and caseinase), pectinases and cuticles of Musca domestica larvae and adults. All the assays were performed with and without the presence of dextrose in the culture media. The quantification of enzyme activity was performed by the ratio of halo / colony (H/C) and the results subjected to variance analysis level of 5% (ANOVA) followed by post-Tukey test. All strains were positive for lipase and also they showed a high significant enzyme production for gelatin at concentrations of 4 and 1%. B. bassiana and Paecilomyces sp. were positive for amylase, pectinase and caseinase, and only Paecilomyces sp. showed cellulase activity.

Evaluation of sorghum straw hemicellulosic hydrolysate for biotechnological production of xylitol by Candida guilliermondii

A preliminary study on xylitol production by Candida guilliermondii in sorghum straw hemicellulosic hydrolysate was performed. Hydrolysate had high xylose content and inhibitors concentrations did not exceed the commonly found values in other hemicellulosic hydrolysates. The highest xylitol yield (0.44 g/g) and productivity (0.19 g/Lh) were verified after 72 hours.

Tannase production by Penicillium atramentosum KM under SSF and its applications in wine clarification and tea cream solubilization

Tannin acyl hydrolase commonly known as tannase is an industrially important enzyme having a wide range of applications, so there is always a scope for novel tannase with better characteristics. A newly isolated tannase-yielding fungal strain identified as Penicillium atramentosum KM was used for tannase production under solid-state fermentation (SSF) using different agro residues like amla (Phyllanthus emblica), ber (Zyzyphus mauritiana), jamun (Syzygium cumini), Jamoa (Eugenia cuspidate) and keekar (Acacia nilotica) leaves. Among these substrates, maximal extracellular tannase production i.e. 170.75 U/gds and 165.56 U/gds was obtained with jamun and keekar leaves respectively at 28ºC after 96 h. A substrate to distilled water ratio of 1:2 (w/v) was found to be the best for tannase production. Supplementation of sodium nitrate (NaNO3) as nitrogen source had enhanced tannase production both in jamun and keekar leaves. Applications of the enzyme were studied in wine clarification and tea cream solubilization. It resulted in 38.05 percent reduction of tannic acid content in case of jamun wine, 43.59 percent reduction in case of grape wine and 74 percent reduction in the tea extract after 3 h at 35ºC.

Xylitol production from wheat straw hemicellulosic hydrolysate: hydrolysate detoxification and carbon source used for inoculum preparation / Produção de xilitol em hidrolisado hemicelulósico de palha de trigo: destoxificação do hidrolisado e fonte de carbono utilizada para o preparo do inóculo

Wheat straw hemicellulosic hydrolysate was used for xylitol bioproduction. The use of a xylose-containing medium to grow the inoculum did not favor the production of xylitol in the hydrolysate, which was submitted to a previous detoxification treatment with 2.5 percent activated charcoal for optimized removal of inhibitory compounds.

Hidrolisado hemicelulósico de palha de trigo foi utilizado para a bioprodução de xilitol. O uso de meio contendo xilose para crescer o inóculo não favoreceu a produção de xilitol no hidrolisado, que foi submetido a um tratamento prévio de destoxificação com 2.5 por cento de carvão ativo para remoção otimizada de compostos inibitórios.

Alteraciones clínicas y bioquímicas en ratas tratadas con dosis altas de vitamina A / Clinical and biochemical alterations in rats treated with high doses of vitamin A

En el presente trabajo se estudió el efecto de la administración intramuscular de 30.000, 50.000 y 100.000 UI de palmitato de vitamina A/día, durante 7 días, respectivamente, sobre la actividad enzimática hepática en 45 ratas Wistar machos, de 12 semanas de edad, con pesos entre 180 y 200 gramos. El grupo control estuvo integrado por 15 ratas Wistar sanas, con género, edad y peso similares a los animales tratados. El consumo de alimentos y de agua, y el peso de las ratas se determinó al finalizar el período experimental. Las ratas se examinaron en busca de manifestaciones clínicas de toxicidad. Al final el estudio, las ratas se sacrificaron bajo anestesia con éter y se tomaron muestras de tejido hepático para la determinación de la actividad enzimática. La administración de vitamina A en exceso incrementó de manera significativa (p menor que 0,05) el contenido hepático del retinol, determinó diversos y variados signos clínicos (tales como: anorexia, pérdida de peso, alopecia, conjuntivitis, hemorragias internas y externas, alteraciones cutáneas y muerte de los animales) e incrementó (p menor que 0,05) la actividad de las siguientes enzimas: alanina aminotransferasa, aspartato aminotransferasa, maltasa ácida (alfa-1,4-glucosidasa ácida), proteasas ácidas, lactato dehidrogenasa y fosfatasa alcalina mientras que las actividades de la glucosa-6-fosfatasa, glucógeno fosforilasa, alfa-amilasa, colinesterasa y arginasa disminuyeron (p menor que 0,05) al comparar con los controles no tratados. Estos cambios son proporcionales a las dosis inyectadas de vitamina A. En conclusión, nuestros resultados proporcionan evidencias que la administración de dosis altas de vitamina A a corto plazo determina diversos y variados signos clínicos y produce una marcada alteración de la actividad enzimática hepática.

In the present work the effect of intramuscular administration of 30.000, 50.000 and 100.000 IU of vitamin A palmitate daily for seven days, respectively, on the liver enzyme activity in 45 white male Wistar rats, aged 12 weeks and weighing 180-200 g, have been studied. The group control was integrated by 15 healthy rats with similar characteristics (strain, gender, age and weight) to treated animals. Food and water consumption and body weights were recorded at the end of the experimental period. Rats were observed for clinical signs of toxicity. At the end of the study, rats were sacrificed under ether anesthesia. Liver samples were taken for the determination of enzyme activity. Administration of excess of vitamin A produced a significant (p menor 0.05) increase in the content of liver vitamin A, determined diverse and variable clinical signs (such as, anorexia, loss of body weight, alopecia, conjunctivitis, external and internal hemorrhages, skin abnormalities and death) and increased (p menor que 0.05) the activity of the following enzymes: alanine aminotransferase, aspartate aminotransferase, acid maltase (acid alfa-1,4-glucosidase), acid proteases, lactate dehydrogenase and alkaline phosphatase while glucose-6-phosphatase, glycogen phosphorylase, alfa-amylase, cholinesterase and arginase decreased (p menor que 0.05) as compared with untreated controls. These changes depend on the doses given of vitamin A. In conclusion, our results provide evidence that short-term administration of high doses of vitamin A determined diverse and variable clinical signs and produces a marked alteration of activity of liver enzymes.

Localização histoquímica de algumas hidrolases no epidídimo do gerbilo / Histochemical localization of some hydrolases in the epididymis of the gerbil

Neste estudo verificou-se, principalmente, que a expressão tissular da FoAl esteve presente nas regiões proximais do epidídimo de gerbilo. As reatividades da FoAc e da ATPase foram intensas (fortes), na maioria das regiões do epidídimo caracterizadas, marcadamente ao nível do citoplasma apical das células do epitélio tubular, exceto no corpo epididimal um delgado ístmo de conexão da cabeça à cauda epidiclimárias nesta espécie, com notória hiporeatividade enzimática a praticamente todas as enzimas aqui estudadas. A SDH mostrou também fraca atividade em todas as regiões, e estruturas do ducto epidiclimário do gerbilo, exceto no compartimento intraluminal onde se verificou atividade SDH-intensa nos espermatozóides estocados no lúmen. As reações enzimáticas caracterizadas foram correlacionadas com alguns papéis histofisiológicos tais como a mediação enzimática das fosfatases e ATPase em processos como endocitose, secreção, absorção e processo de transporte ativo das células do epitélio tubular. Uma possível marcação específica de mitocôndrias da peça média dos espermatozóides intraluminais pela SDH foi sugerida, com embasamento em processos metabólicos das mitocôndrias.

This study mainly showed that alkaline phosphatase expression had been present in the proximal regions of the epididymidis ductus of the gerbil which comprised the initial segment and proximal caput. The reactivities of acid phosphatase and ATPase were strong in the proximal and clistal regions of the epididymidis ductus at the level of the apical cytoplasm and epithelium, except at the corpus level, a very thin isthmus located between the caput and cauda epididymidis, and as a general rule a low enzymatic reactive region of the epididymidis of gerbil. SDH revealed also low activities in ali the regions and regional structures of the duct, except into the luminal content formed by storaged spermatozoa, prior on the cauda leveI. The enzymes presented in the epididymidis were correlated to some histophysiological roles such as the enzymatic mecliation of endocytosis, secretion, absorption and active transport concerning to phosphatases and A TPase and a possible mitochondrial role of SDH could occur at the spermatozoa level in which the middle pieces were formed by a great amount of mitochondria.

Hydrolytic enzymes in Paracoccidioides brasiliensis--ecological aspects

Paracoccidioides brasiliensis is a thermally dimorphic fungus that causes paracoccidioidomycosis. The yeast form of this pathogen is found in the animal host whereas the mycelial form is recovered from living and non-living organic material. The sole carbon source available in these habitats is represented by polysaccharides from the plant cell wall. Hydrolytic enzymes are necessary to convert these polymers into simple sugars for fungal metabolism. We report on the presence of ortholog genes of hydrolytic enzymes identified in the P. brasiliensis transcriptome and on hydrolytic activities in supernatants of induced P. brasiliensis cultures of mycelium and yeast cells. Enzymatic assays have shown cellulase and xylanase activities, both being higher in mycelium than in the yeast form. Amylase and chitinase activities were detected only in mycelium. Data so far reinforce the idea that mycelial P. brasiliensis is a saprobe.

Efeito das vastatinas sobre a hidrólise hepática do colesterol esterificado, em células de hepatoblastoma humano HEP G2 / Effects of vastatins in the hepatic hidrolase of esterified cholesterol in human HEP G2 hepatoblastoma cells

O presente trabalho se valeu de células de hepatoma humano Hep G2 para estudar se existe algum efeito das vastatinas sobre a atividade das enzimas que governam a hidrólise hepática do coleterol: colesterol éster hidrolases lisossomal, citosólica e microssomal. As enzimas foram incubadas com ou sem amostras de vastatina, dissovidas em dimetilsulfóxido. A adiçao das vastatinas ofereceu distintos resultados, conforme a maneira que se apresentou o substrato às enzimas. No caso da colesterol éster hidrolase lisossomal, obteve-se inibiçao em substrato Hajjar, com sais biliares, e estimulaçao em substrato Brecher, desprovido de sais biliares. A hidrolase citosólica teve sua atividade inibida em substrato Hajjar e constante em substrato Brecher.

Hydrolytic enzymes in macrophages from leprosy patients in presence of Mycobacterium leprae.

Presence of Mycobacterium leprae in association with in vitro cultured macrophages, from bacillary negative long term treated lepromatous leprosy patients, induces reduced level of protein and lowering of hydrolytic enzymes like p-glucuronidase, Lysozyme and Lactic dehydrogenase. Alkaline phosphatase, on the other hand is increased. In the macrophages from normal healthy individuals or tuberculoid leprosy patients, presence of M.leprae increases both protein and levels of all the above enzymes. This observation shows that macrophages from lepromatous leprosy patients are unable to manifest in presence of M. leprae, the key enzymes involved in degradation of complex biological entities phagocytosed by the cells.