High Protease Activity Research Articles

Virgibacillus halodenitrificans MSK-10P, a Potential Protease-producing Starter Culture for Fermented Shrimp Paste (kapi) Production

ABSTRACTFermented shrimp paste (Kapi) is traditionally produced using a high content of salt and mainly consumed as a food condiment by the Thai people. In order to select potential functional autochthonous starter culture, the protease-producing bacteria from Kapi were isolated, screened, and evaluated for technological and safety aspects. Among 195 isolates, five bacterial strains (MSK-3P, MSK-4P, MSK-5P, MSK-7P, and MSK-10P) exhibited the highest activity of protease using Anson method. All of them were identified as Virgibacillus halodenitrificans using 16S rRNA sequence analysis. All selected strains exhibited growth in the presence of NaCl up to 25%. Neither strain showed hemolytic activity nor biogenic amine formation. Among all selected strains, only strain MSK-10P had no ability to form a biofilm, while four other strains showed a weak ability. Strain MSK-7P exhibited multiple resistances to tetracycline and vancomycin, while strains MSK-3P, MSK-4P, and MSK-10P showed susceptibility to all antibiotics tested. According to the obtained results, this protease-producing V. halodenitrificans MSK-10P is a good candidate for further investigation for use in Kapi fermentation to assess its technological performance as a autochthonous starter culture.

The Application of Crude Ginger Protease as an Inhibitive Assay for Heavy Metals

In this work, a novel source of protease for the bioassay of heavy metals has been developed using proteases extracted from ginger. The result shows that the optimum protease activity was reached at 1 mg/mL protein concentration of the ginger protease. The optimum casein concentration toward crude ginger protease activity was 1.75 mg/mL. The most suitable pH for protease from crude ginger protease was within the range from pH 5.0 to 7.0. The proteases exhibited high protease activity in a broad range of temperature from 20 to 60 oC. The optimum incubation time for the enzyme occurred at minute 30. Among the six heavy metals tested, only three heavy metals inhibited proteolytic activity of ginger crude with an inhibition more than 30% at 1 mg/L. The calculated LC50 for mercury, copper and silver were, 0.182, (95%, confidence interval (C.I.) 0.134 to 0.283), 0.071, (95% C.I. of 0.056 to 0.096), 0.054, (95% C.I. of 0.039 to 0.085), respectively. Data on the sensitivity of various proteases to heavy metals shows that the crude ginger protease is comparable in sensitivity for mercury to the achromopeptidase, bromelain, papain assays. The crude ginger protease assay for copper is comparable in sensitivity to the papain assay and appears to be more sensitive than the rest of the assays. For silver, the ginger protease was the most sensitive while other assay methods are either unable or be able to detect higher concentration of silver. The crude proteases extracted from ginger showed a good potential for the development of a rapid, sensitive, and economic inhibitive assay for the biomonitoring of mercury, copper, and silver in the environment.

Effects of dietary protein level on growth performance, digestive enzyme activity, and gene expressions of the TOR signaling pathway in fingerling Pelteobagrus fulvidraco.

An 8-week feeding trial was conducted to investigate effects of dietary protein levels (37, 40, and 43%) on the growth performance, feed utilization, digestive enzyme activity, and gene expressions of target of rapamycin (TOR) signaling pathway in fingerling yellow catfish. One hundred and eighty fingerlings (average weight 0.77 ± 0.03g) were equally distributed across four replicate tanks for each of the three treatments, with 15 fish per tank. No difference (P > 0.05) was observed in initial body weight, survival rate (SR), hepatosomatic index (HSI), viscera index (VSI), dressing percentage (DP), and condition factor (CF) among all the treatments. The diet containing 40% protein increased significantly (P < 0.05) final body weight, weight gain rate (WGR), specific growth rate (SGR), protein efficiency ratio (PER), nitrogen retention (NRE), and energy retention (ERE) in fish. The highest protease activity in the stomach and intestine was observed in the P40 group (P < 0.05), while amylase and lipase were not significantly different (P > 0.05). The transcriptional levels of IGF-1, IGF-1R, and Akt were significantly (P < 0.05) higher in fish fed P40 or P43 than those of fish fed P37. TOR and S6K1 mRNA expressions were significantly (P < 0.05) increased in the P40 groups. Hence, the diet containing 40% protein would be suitable for the optimum growth and effective protein utilization of fingerling Pelteobagrus fulvidraco. In vitro, the transcriptional levels of IGF-1, IGF-1R, Akt, TOR, and S6K1 in hepatocyte supplemented with a 40-μM mixed amino acids were significantly (P < 0.05) higher compared to other treatments. No difference (P > 0.05) was observed in eukaryotic translation initiation factor 4E-binding protein 1 in vivo and in vitro among all the treatments. Effects of dietary protein level on growth performance likely are involved in the activation of TOR signaling pathway in fingerling Pelteobagrus fulvidraco.

Technological and safety characterization of coagulase-negative staphylococci with high protease activity isolated from Traditional Chinese fermented sausages

The aim of this study was to determine the technological and safety properties of coagulase-negative staphylococci (CNS) with high protease activity isolated from traditional Chinese fermented sausages in 10 different regions. According to the criteria of meat starter cultures, eight strains were obtained from 76 Gram-positive and catalase-positive isolates of CNS. These strains could hydrolyze both sarcoplasmic and myofibrillar proteins, tolerate 10% of salts and 150 mg/kg nitrites, possessed good nitrate reductase activity and lipolytic activity, and exhibited non-decarboxylase activity. Furthermore, all the eight strains exhibited antibiotic susceptibilities and negative hemolytic activity, DNase activity, and biofilm formation ability. Whole genome sequencing demonstrated that there were no pathogenic genes in the eight CNS strains, including 18 enterotoxin genes, panton-valentine leukocidin, toxic shock syndrome toxin-1, exfoliative toxin genes, and biofilm genes. Based on the results of API and 16S rRNA sequence analysis, seven of the eight CNS strains were identified as Staphylococcus simulans and the remaining one was S. saprophyticus.

Modulation of coffee flavor precursors by Aspergillus oryzae serine carboxypeptidases

Aspergillus oryzae was isolated from coffee pulp, screened for the production of carboxypeptidase and confirmed by molecular studies. Coffee pulp posses high protease activity (1.4 × 103 nkat/mL) among the agro-substrates used for the production of proteases but wheat bran was used for further studies considering its easy availability in large volumes. Proximal composition, pre-treatment, auxiliary carbon and nitrogen with growth studies indicated carboxypeptidase activity (2.6 × 103 nkat/g). The RSM was employed to generate a predictive model of the combined effects of independent variables and indicated a significant impact on acid carboxypeptidase with 35% moisture, 30 °C temperature, 108 CFU mL−1 inoculum and 100 h duration with 4.8 × 103 nkat/g activity. The partially purified peptidase had a specific activity of 6.1 × 104 nkat/mg with a molecular weight of 50 kDa, optimal pH 3.6, temperature 35 °C and feature of a serine protease. Enzyme stability was maintained with 50% glycerol and 2 mg/mL BSA. The protease application to develop flavor profiles of coffee was noteworthy with an organoleptic score of 7.5 compared to 4.5 of the untreated sample. GC-MS highlighted enhanced flavor annotations of pyrazines contributing sweet fruit notes. Carboxypeptidase augmented the coffee bean with specialized flavor notes and has been reported for the first time.

Processed soybean meal as an alternative protein source for yellow perch ( Perca flavescens ) feed

The goal of this study was to compare conventional soybean meal (SBM) with modified SBM (MSBM) after chemical and enzyme pretreatment to potentially reduce the antinutritional factors (ANF), as a fishmeal (FM) replacer at 50% or 100% in the diets of yellow perch (Perca flavescens). Half of the SBM and MSBM diets contained an enzyme cocktail (of phytase and carbohydrate-degrading enzyme), and the other half received no enzyme supplementations. Fingerlings (297; initial weight, 11.01 ± 0.19 g) were randomly distributed over nine treatments, in triplicate, and fed these isonitrogenous diets (crude protein 410 g/kg) that included replacing FM with SBM at 50% or 100% without or with enzyme supplementations (S50, S100, S50+E or S100+E, respectively), MSMB at 50% or 100% without or with enzyme supplementations (MS50, MS100, MS50+E or MS100+E, respectively) or a control FM-based diet. After 10 weeks, the growth performance, feeding efficiencies, proximate composition, intestinal/pyloric caeca digestive enzymes and liver metabolic/antioxidant enzymes in P. flavescens were measured. The highest (p < 0.05) growth performance and nutrient utilization parameters (protein efficiency ratio and protein productive value, PPV) were observed for the MS50+E group, which was not statistically different to parameters for the control and MS50 groups, and significantly (p < 0.05) higher than all other groups. The lowest and highest growth performance and feed conversion ratios, respectively, were observed in the S100, S100+E and MS100 groups. The highest protease activity (in both intestine and pyloric caeca) was observed for the control group, but was significantly similar to MS50, MS50+E and S50+E groups. The lowest value was observed for 100% replacement of FM protein by SBM and MSBM fed groups. However, inclusion of exogenous enzymes in feed showed positive effects in MS50+E (compared to MS50) for PPV, lipid productive value and amylase activity in the intestine. Activity of protein metabolism enzymes (i.e., alanine transaminase and aspartate aminotransferase) in the liver was the highest in the control group, which was similar (p < 0.05) to the MS50+E and MS50 groups. Antioxidant enzymes (catalase, superoxide dismutase and glutathione peroxidase) in the liver were the lowest in the control group, which was significantly similar (p < 0.05) to the group given the feed with 50% replacement of FM protein. Complete (100%) replacement of FM protein exhibited the highest antioxidant enzyme activity. Conclusively, performance of the MS50 and MS50+E groups was similar to the FM group and better than SBM group; therefore, MSBM with high protein and low ANFs has considerable potential as an alternative to FM in aquafeed.

Formation of ACE-inhibitory peptides during fermentation of jack bean tempe inoculated by usar Hibiscus tiliaceus leaves starter

Jack bean is one of the potential legumes as a source of ACE-inhibitory peptides in tempe fermentation due to its high protein content and amino acids composition. A traditional inoculum for tempe fermentation is usar from Hibiscus tiliaceus leaves. The aim of this study was to investigate the optimum tempe fermentation time inoculated using usar on the formation of ACE-inhibitory peptides. The study applied the prolonged tempe fermentation (120 h) at room temperature (+30°C). The change of pH, soluble protein, protease activity, and ACE-inhibitory activity during tempe fermentation were determined using standard methods. There were significant differences (α=0.05) in pH, proteolytic activity, soluble protein content, and ACE-inhibitory activity during tempe fermentation. The highest protease activity was produced at 96 h of fermentation while the highest soluble protein content was produced at 120 h of fermentation with the value of 12.71 unit/g and 3.65 mg/mL, respectively. The highest ACE-inhibitory activity was reached at 72 h of fermentation (53.89 %). Conclusion: The optimum fermentation time of the formation of ACE-inhibitory peptide was at 72 h of fermentation.

Drying and rewetting effects on organic matter mineralisation of contrasting soils after 36 years of storage

Soil incubation studies commonly use archived, dry stored soil samples. Numerous studies have detected a profound impact of dry storage on soil microbial community composition and activity. However, comparatively few studies have explored the impact of storage on the short-term mineralisation of soil organic matter (SOM) after moistening dry soils to revive and stabilise microbial activity (“pre-incubation”). We examined whether pre-incubation of dry stored soil of contrasting texture (Alfisol and Vertisol) can recover native soil microbial functions in fresh soil or soil stored for 3 weeks, 3 or 36 years. The oldest samples had a delayed CO2 response after moistening, but matched CO2 production rates of younger samples within hours. The two soil types displayed contrasting CO2 emission responses after moistening; the Alfisols had a rapid but short-lived CO2 emission peak (the so-called “Birch Effect”) while the Vertisols demonstrated a long, sustained release of CO2 reflecting its higher abundance of microsite abundance and microaggregates which may protect SOM and microorganisms during adverse conditions like drying. After 10 days of pre-incubation, older Alfisol samples had higher protease activity and response to amino acid addition demonstrating altered microbial physiological responses relating to nitrogen (N) compared to fresher counterparts. However, such responses in Vertisols were unchanged suggesting high clay soils with high potential microsite abundance may improve preservation of soil N functions through dry storage or even drought. This work highlights both limitations and possibilities of incubating dry archived soils for SOM mineralisation studies.

Optimization of protease production in indigenous Bacillus species isolated from soil samples in Lagos, Nigeria using response surface methodology

Abstract Proteases catalyse the hydrolysis of peptide bonds in proteins and offer a huge potential for application in industries, including detergent, dairy, leather, baking, pharmaceutical and beverage industries. In this study, indigenous Bacillus species were isolated from soil samples collected from abattoir, refuse and non-refuse sites in Lagos, Nigeria and optimized for protease production. The isolates were purified on Bacillus agar and screened for protease production on casein agar. Three strains showing high potential for protease production were identified as Bacillus cereus ABBA1, Bacillus subtilis RD7 and Bacillus subtilis NRD9 via amplification and analysis of 16S rRNA genes. Protease optimization was done Insilco using Box-Behnken Design (BBD) by response surface methodology (RSM) with Design-Expert software and then validated experimentally. Factors optimized include temperature, pH, carbon and nitrogen source and inoculum density. Statistical analysis was done using ANOVA. The results obtained from the Insilco experimental model revealed high protease activity of 159.43 U/ml, 141.28 U/ml and 138.17 U/ml while experimental validation generated a high protease activity of 200.56 U/ml, 176.00 U/ml and 163.76 U/ml for strains ABBA1, RD7 and NRD9, respectively in optimized medium. This corresponds to 33.54-, 42.21- and 36.64- fold increase in protease production compared to the unoptimized protease production medium. The optimum conditions for extracellular protease production obtained from quadratic model of RSM were 40 °C, pH 8.5, 2.5% (v/v) inoculum density, 1.5 g/L maltose and 2.0 g/L beef extract powder. The model prediction agreed with the experimental data (R2 = 0.98) and was statistically significant (p ≤ 0.05). This results further confirms the need to optimize the production parameters to achieve maximum yield and economical use of available resources during production of industrially important enzymes.

Antifungal susceptibilities, biofilms, phospholipase and proteinase activities in the Candida rugosa complex and Candida pararugosa isolated from tertiary teaching hospitals.

Non-albicansCandida species have emerged as fungal pathogens that cause invasive infections, with many of these species displaying resistance to commonly used antifungal agents. This study was confined to studying the characteristics of clinical isolates of the C. rugosa complex and C. pararugosa species. Seven isolates of the C. rugosa complex and one isolate of C. pararugosa were obtained from two tertiary referral hospitals in Malaysia. Their antifungal susceptibilities, biofilm, proteinase, phospholipase, esterase and haemolysin activities were characterized. Biofilms were quantified using crystal violet (CV) and tetrazolium (XTT) reduction assays at 1.5, 6, 18, 24, 48 and 72 h.Results/Key findings. The E-test antifungal tests showed that both species have elevated MICs compared to C. albicans and C. tropicalis. The highest biomass was observed in one of the C. rugosa isolates (0.237), followed by C. pararugosa (0.206) at 18 h of incubation. However, the highest bioactivity was observed in the C. rugosa ATCC 10571 strain at 24 h (0.075), followed by C. pararugosa at 48 h (0.048) and the same C. rugosa strain at 24 h (0.046), with P<0.05. All isolates exhibited high proteinase activity (+++) whereas six isolates showed very strong esterase activity (++++). All the isolates were alpha haemolytic producers. None of the isolates exhibited phospholipase activity. Elevated MICs were shown for the C. rugosa complex and C. pararugosa for commonly used antifungal drugs. Further studies to identify virulence genes involved in the pathogenesis and genes that confer reduced drug susceptibility in these species are proposed.

Guava (Psidium guajava L.) leaf protease activity enriched by controlled-stress and putrescine application

Plant-derived proteases are widely used in the food and pharmaceutical industries. Some enzyme operations require enzyme activity and stability at high temperatures or extremes of pH. We sought a simple, low-priced, and nature-friendly method to improve protease activity and stability. Guava was chosen as the plant protease source. Plants were treated with saline water (0, 5 and 10 dS /m) and foliar application of putrescine (0, 250 and 500 ppm). Leaf protease activity was evaluated at a range of pH (2–8) and temperature (30–90 °C). Salt, putrescine and their combination significantly influenced protease activity. Putrescine application (250 ppm) increased enzyme activity at high temperatures and acidic pH in leaf extracts. For plants irrigated with high saline, 500 ppm putrescine treatment increased both the low pH and high pH protease activities. The existence of two pH optima of guava leaf protease suggests that at least two major proteases are present in the guava leaf extract.

Assessment of Extra Cellular Enzymes of Bacteria Isolated from Mangrove Rhizosphere Soil of Different Places of Gujarat in Monsoon Season

Background/objectives: Mangroves are one of the most diverse ecosystems on the earth and harbours many organisms, including extremophiles which help the ecosystem to grow and survive despite harsh environment. These microbes produce novel enzymes as well as secondary metabolites that find major applications in extreme environments. Methodology: In this study, total 56 bacteria were isolated from rhizospheric soil of mangrove collected from seven sampling sites of Gujarat, India i.e. Kandla, Mundra, Tuna, Guneri, Machhad, Diu, and Madhwad. These isolates were screened for their production of extracellular hydrolytic enzymes i.e. amylase, protease, cellulase, and catalase. All the isolates were also assessed for their ability to survive in different salt concentration ranging from 0.5% to 25% in Zobell marine agar. Findings: After 48 h of incubation, isolates K10–2(11)ZO and M100(5)NASW had the highest amylase activity, K100(1)NASW had the highest cellulase activity, and G10–2(9)ZO had the highest protease activity. Among all sites, Madhwad had the highest catalase positives. Three isolates named as MD10–2C3, MW10–3C2, and MWC4 emerged as possible extreme halotolerant and were able to grow at 25% of salt concentration while Diu10–4C1, MW10–3, and G10–1(5) ZO were also possible halotolerant and were able grow in till 20% of salt concentration. Application/ improvements: Few isolates that had showed higher extracellular enzyme activity and could be potential high producers that can be used in industries such as detergent industry, textile industries, environmental remediation, etc.Keywords: Mangroves, Enzymes, Cellulase, Amylase, Protease, Extreme Halotolerant

Screening of lactic acid bacteria with high protease activity from fermented sausages and antioxidant activity assessment of its fermented sausages

ABSTRACTThis study was conducted to screen the lactic acid bacteria (LAB) with admirable technological properties and high protease activity from traditional Chinese fermented sausages, and to investigate the effects of LAB on the antioxidant activity of peptides of fermented sausages. Seven strains were obtained from 133 LAB isolates. They complied with the criteria of meat starter cultures with high proteolytic activity. These strains were analyzed by API and sequence analysis of 16S rRNA to identify the five strains of Lactobacillus plantarum, one strain of L. fermentum, and one strain of L. pentosus. Furthermore, L. plantarum CD101 was inoculated with fermented sausages. LC-MS/MS was used to analyze the peptide composition. Compared with control group, using L. plantarum CD101 as starter cultures (Group L) led to a significant increase (P < 0.05) in antioxidant activity. The relative content of most peptides in Group L samples was largely higher than that in control samples.

Properties of Tetragenococcus halophilus Strains Isolated from Myeolchi (anchovy)-jeotgal

Halophilic lactic acid bacteria (LAB) were isolated from myeolchi-jeotgal (23% NaCl, w/v) fermented in jangdok (Korean earthenware) located outside a house. They were identified as Tetragenococcus halophilus by 16S rRNA and recA gene sequencing. Four T. halophilus isolates showing high protease activities were selected for further studies. Four strains grew well, reaching OD600 values of 0.75-0.92 at 18% NaCl content (w/v) and 0.28-0.44 at 23% salt. They showed rapid growth, attaining OD600 values of 1.1-1.2 at 20-30℃, but did not grow at 4℃. At 15℃, the highest OD600 values, which exceeded 0.6, were observed at 20 days, and were higher than those of cultures at 37℃ and 42℃ (approximately 0.5). Four isolates grew best in broth where the initial pH was adjusted to 8 and did not grow at pH ≤ 4. T. halophilus BS2-36 showed the highest survival ratio of 18.7% after 2 hours of exposure at pH 3. BS2-36 showed the highest survival ratio (1.29%) in presence of 0.3% bile salts. T. halophilus BS2-36 seems a promising candidate as a starter for jeotgal and other fermented foods with high salinities.

Mass spectrometry-based peptidome profiling of human serous ovarian cancer tissues.

Bioactive peptides existing in vivo have been considered as an important class of natural medicines for the treatment of diseases. Peptidome analysis of tissues and biofluids had provided important information about the differentially expressed bioactive peptides in vivo. Here, we analyzed the peptidome of serous ovarian cancer tissue samples and normal ovarian epithelial tissue samples by mass spectrometry and further investigated the possible bioactive peptides that were differentially expressed. We identified 634 differentially expressed peptides, 508 of these peptides were highly abundant in serous ovarian cancer tissues, a result consistent with higher protease activity in ovarian cancer patients. The difference in preferred cleavage sites between the serous ovarian cancer tissues and normal ovarian epithelium indicated the characteristic peptidome of ovarian cancer and the nature of cancer-associated protease activity. Interestingly, KEGG pathway analysis of the peptide precursors indicated that the differentially regulated pathways in ovarian cancer are highly consistent with the pathways discovered in other cancers. Besides, we found that a proportion of the differentially expressed peptides are similar to the known immune-regulatory peptides and anti-bacterial peptides. Then we further investigated the function of the two down-regulated peptides in ovarian cancer cells and found that peptide P1DS significantly inhibited the invasion and migration of OVCAR3 and SKOV3 ovarian cancer cells. Our results are the first to identify the differentially expressed peptides between the serous ovarian cancer tissue and the normal ovarian epithelium. Our results indicate that bioactive peptides involved in tumorigenesis are existed in vivo.

Pathogenic significance of hemorrhagic pneumonia in hematologic malignancy patients with Stenotrophomonas maltophilia bacteremia: clinical and microbiological analysis

Hemorrhagic pneumonia (HP) is known as the clinical manifestation of Stenotrophomonas maltophilia infection, while catheter-related blood stream infection (CRBSI) is the common clinical presentation of S. maltophilia bacteremia (SMB). The purpose of this study is to evaluate the risk factors for mortality in hematologic malignancy patients with SMB and to analyze clinical and microbiological characteristics of HP associated with SMB and CRBSI. SMB cases of patients with a hematologic malignancy were collected from 2006 through 2016. The overall 30-day mortality rate and mortality risk factors were assessed. The expression of major virulence-associated genes from S. maltophilia isolates, which included genes encoding type-1 fimbriae (smf-1), proteases (StmPr1 and StmPr2), and esterase (Smlt3773), from the blood of patients with HP and CRBSI was investigated. The phenotypic and genotypic traits were also compared. A total of 118 cases of SMB were included. The overall 30-day mortality rate was 61.0%. A multivariable analysis showed that HP was the most important risk factor for mortality (adjusted OR = 106.41; 95% CI = 5.18-2184.55). Although no statistical significance was observed in microbiological analysis, isolates from HP have a trend toward a higher protease activity (93.8% vs. 73.3%, P = 0.172). Clinical analysis showed that thrombocytopenia (P = 0.037) and prolonged neutropenia (P = 0.043) were significant factors associated with HP. Our data, which includes hematologic malignancy patients with SMB, suggest that HP is the significant risk factor for mortality and that the unique characteristics of patients and microbes contribute to the pathogenesis.

Strategical isolation of efficient chicken feather-degrading bacterial strains from tea plantation soil sample.

Chicken feather waste is generally insufficiently utilized despite its high content of protein, constituting an environmental issue. Biodegradation of the waste with enabling microbes provides an advantageous option among the available solutions. In this study, an efficient whole feather-degrading strain was strategically isolated from a soil sample taken from a local tea plantation that has little or nothing to do with feathers. The strain was identified as Bacillus thuringiensis (designated as FDB-10) according to the cloned complete 16S rRNA sequence. The FDB-10 could efficiently degrade briefly heat-treated whole feather (102°C, 5min; up to 90% of a maximum concentration of 30g/L) in a salt medium supplemented with 0.1g/L yeast extract within 24h (37°C, 150rpm). Addition of carbon sources (glycerol, glucose, starch, Tween 20, Tween 80, 1.25g/L as glycerol) to the fermentation medium could improve the degradation. However, significant inhibition could be observed when the added carbon source reached the amount usually adopted in the investigation of carbon source preference (1%). Nitrogen source (NH4Cl, (NH4)2SO4, peptone) adversely influenced the performance of the strain. When the molar concentrations of NH4+ were equal for the two salt, the inhibitory effect on degradation of whole feathers was similar. Entirely different from other reported feather-degrading strains showing a preference to melanin-free feather substrates, the strain isolated in this study could degrade melanin-containing feather equally efficiently, and higher protease activity could be detected in the digest mix. As a plus, the strain could degrade feathers in rice wash produced in daily cooking, indicating its potential use in the simultaneous treatment of rice cooker wastewater produced by a rice processing plant. All these results imply that the FDB-10 is a strain with great potential in the biodegradation of feather waste.

Proteogenomic Analyses of Cellular Lysates Using a Phenol-Guanidinium Thiocyanate Reagent.

Protein and RNA profiles are highly informative when generating a comprehensive understanding of the dynamics of complex biological systems. Quantitative correlations between protein and RNA profiles are not always high, yet simultaneous acquisition of both profiles remains challenging, in part because of the limited availability of samples and the inconvenience of separately preparing protein and RNA fractions. In a previous study, protein, DNA, and RNA fractions were simultaneously prepared from the same sample using phenol-guanidinium isothiocyanate reagent (P/GTC), although the performance of P/GTC-extracts in proteogenomic analyses remains poorly understood. We therefore evaluated the performance of the P/GTC-extraction method in proteogenomic analyses using standard HEK293-F cells and human peripheral blood neutrophils. The latter cell type is renowned for its extreme vulnerability to protein/RNA degradation, reflecting high protease and RNase activities. Our data indicate that the P/GTC extraction method provides superior protein profiles from neutrophil and HEK293-F cell samples for simultaneous preparation of RNA and protein, as compared with those from conventional protein extraction methods. The P/GTC extraction method therefore provides a powerful and robust tool for a broad range of proteogenomic studies.

Genetic Analysis and In Vitro Enzymatic Determination of Bacterial Community in Compost Teas from Different Sources

Compost tea has been used throughout the world to control diseases and promote the growth of plants. The microbial community is the main factor involved both in the antagonistic effect against relevant phytopathogens and as a stimulator of plant growth. The objective of this research is to determine the bacterial diversity present in four types of compost as well as some of the mechanisms that may be involved in the positive effects of compost tea on crops. Aerated (ACT) and non-aerated (NCT) compost teas were obtained from four different composts: spent mushroom substrate compost (SMC), grape marc compost (GMC), greenhouse horticultural crop residues compost (CRC), and vermicompost (CRV). 16S rDNA-based DGGE profiles were obtained for each compost tea and their respective bacterial communities were analyzed. Of the 100 clones obtained, those typical for being dominant for all profiles were chosen. In total, 20 different clones were sequenced. The results showed that the bacterial communities of most compost teas had high richness, diversity, and evenness values, with relative abundance of species belonging to Bacteroidetes and Proteobacteria. The NCTs obtained from all the composts, especially CRC and CRV, showed high levels of siderophore production, while teas from GMC manifested high and consistent cellulase activity. The ACTs from all the composts, especially SMC, had high protease activity.

Characteristics of herring marinated in reused brines after microfiltration

Brines after herring marinating pose a serious financial problem to the industry and natural environment. Paradoxically, the brine waste containing biological active compounds like proteases and peptides being responsible for marinade quality is discarded with sewage. Results show that the reuse of brine without filtration is not possible because of increase in the bacterial count and lipid oxidation in marinated herring. The desired parameters of marinades were achieved using brine permeate-50 µm free of the suspension and lipids. The best quality and sensory parameters of marinated herring meat were obtained using permeate-0.22 brine, which not contain microorganisms and lipids, and also a high activity of proteases. Reuse of brine allows reducing by half the losses of protein hydrolysis products (PHP) from meat to brine and for reverse diffusion of PHP and peptidases to meat. The marinades produced with the newly-developed method had up to 25% more PHP, up to 20% lower hardness, 10–20% higher activity of proteases, 40–97% lower indices of lipid oxidation, and 5% higher scores in sensory assessment, compared to the marinades produced with fresh brine. The inexpensive and easy to perform microfiltration of brine affords the possibility of increasing the quality and nutritional value of marinades and minimizing both waste volume and production costs.