scholarly journals Understanding the Role of Prevotella Genus in the Digestion of Lignocellulose and Other Substrates in Vietnamese Native Goats’ Rumen by Metagenomic Deep Sequencing

Animals ◽  
2021 ◽  
Vol 11 (11) ◽  
pp. 3257
Author(s):  
Trong-Khoa Dao ◽  
Thi-Huyen Do ◽  
Ngoc-Giang Le ◽  
Hong-Duong Nguyen ◽  
Thi-Quy Nguyen ◽  
...  
Keyword(s):  
Deep Sequencing ◽  
Gene Encoding ◽  
Amylolytic Enzymes ◽  
E Coli ◽  
Lignocellulose Conversion ◽  
Genes Encoding ◽  
Abundant Genus ◽  
First Time ◽  
Goat Rumen

Bacteria in rumen play pivotal roles in the digestion of nutrients to support energy for the host. In this study, metagenomic deep sequencing of bacterial metagenome extracted from the goats’ rumen generated 48.66 GB of data with 3,411,867 contigs and 5,367,270 genes. The genes were mainly functionally annotated by Kyoto Encyclopedia of Genes and Genomes (KEGG) Carbohydrate-Active enZYmes (CAZy), and HMMER database, and taxonomically classified by MEGAN. As a result, 65,554 genes encoding for 30 enzymes/proteins related to lignocellulose conversion were exploited, in which nine enzymes were seen for the first time in goat rumen. Prevotella was the most abundant genus, contributing 30% hemicellulases and 36% enzymes/proteins for lignocellulose pretreatment, and supporting 98.8% of feruloyl esterases and 71.7% acetylxylan esterases. In addition, 18 of the 22 most lignocellulose digesting- potential contigs belonged to Prevotella. Besides, Prevotella possessed many genes coding for amylolytic enzymes. One gene encoding for endoxylanase was successfully expressed in E. coli. The recombinant enzyme had high Vmax, was tolerant to some salts and detergents, worked better at pH 5.5–6.5, temperature 40–50 °C, and was capable to be used in practices. Based on these findings, we confirm that Prevotella plays a pivotal role for hemicellulose digestion and significantly participates in starch, cellulose, hemicellulose, and pectin digestion in the goat rumen.

scholarly journals Detection and Quantification of Superoxide Formed within the Periplasm of Escherichia coli

2006 ◽  
Vol 188 (17) ◽  
pp. 6326-6334 ◽  
Author(s):  
Sergei Korshunov ◽  
James A. Imlay
Keyword(s):  
Escherichia Coli ◽  
Pathogenic Bacteria ◽  
Phagocytic Cells ◽  
Free Living ◽  
E Coli ◽  
Superoxide Formation ◽  
Genes Encoding ◽  
Copper Zinc ◽  
Primary Substrate

ABSTRACT Many gram-negative bacteria harbor a copper/zinc-containing superoxide dismutase (CuZnSOD) in their periplasms. In pathogenic bacteria, one role of this enzyme may be to protect periplasmic biomolecules from superoxide that is released by host phagocytic cells. However, the enzyme is also present in many nonpathogens and/or free-living bacteria, including Escherichia coli. In this study we were able to detect superoxide being released into the medium from growing cultures of E. coli. Exponential-phase cells do not normally synthesize CuZnSOD, which is specifically induced in stationary phase. However, the engineered expression of CuZnSOD in growing cells eliminated superoxide release, confirming that this superoxide was formed within the periplasm. The rate of periplasmic superoxide production was surprisingly high and approximated the estimated rate of cytoplasmic superoxide formation when both were normalized to the volume of the compartment. The rate increased in proportion to oxygen concentration, suggesting that the superoxide is generated by the adventitious oxidation of an electron carrier. Mutations that eliminated menaquinone synthesis eradicated the superoxide formation, while mutations in genes encoding respiratory complexes affected it only insofar as they are likely to affect the redox state of menaquinone. We infer that the adventitious autoxidation of dihydromenaquinone in the cytoplasmic membrane releases a steady flux of superoxide into the periplasm of E. coli. This endogenous superoxide may create oxidative stress in that compartment and be a primary substrate of CuZnSOD.

scholarly journals Selective expression of KCNA5 and KCNB1 genes in gastric and colorectal carcinoma

BMC Cancer ◽  
2020 ◽  
Vol 20 (1) ◽  
Author(s):  
Azer Farah ◽  
Maria Kabbage ◽  
Salsabil Atafi ◽  
Amira Jaballah Gabteni ◽  
Mouadh Barbirou ◽  
...  
Keyword(s):  
Potassium Channels ◽  
Cancer Progression ◽  
Clinical Stage ◽  
Inflammatory Cells ◽  
Tumour Cells ◽  
Malignant Tumours ◽  
Genes Encoding ◽  
First Time ◽  
Formalin Fixed

Abstract Background Gastric and colorectal cancers are the most common malignant tumours, leading to a significant number of cancer-related deaths worldwide. Recently, increasing evidence has demonstrated that cancer cells exhibit a differential expression of potassium channels and this can contribute to cancer progression. However, their expression and localisation at the somatic level remains uncertain. In this study, we have investigated the expression levels of KCNB1 and KCNA5 genes encoding ubiquitous Kv2.1 and Kv1.5 potassium channels in gastric and colorectal tumours. Methods Gastric and colorectal tumoral and peritumoral tissues were collected to evaluate the expression of KCNB1 and KCNA5 mRNA by quantitative PCR. Moreover, the immunohistochemical staining profile of Kv2.1 and Kv1.5 was assessed on 40 Formalin-Fixed and Paraffin-Embedded (FFPE) gastric carcinoma tissues. Differences in gene expression between tumoral and peritumoral tissues were compared statistically with the Mann-Whitney U test. The association between the clinicopathological features of the GC patients and the expression of both Kv proteins was investigated with χ2 and Fisher’s exact tests. Results The mRNA fold expression of KCNB1 and KCNA5 genes showed a lower mean in the tumoral tissues (0.06 ± 0.17, 0.006 ± 0.009) compared to peritumoral tissues (0.08 ± 0.16, 0.16 ± 0.48, respectively) without reaching the significance rate (p = 0.861, p = 0.152, respectively). Interestingly, Kv2.1 and Kv1.5 immunostaining was detectable and characterised by a large distribution in peritumoral and tumoral epithelial cells. More interestingly, inflammatory cells were also stained. Surprisingly, Kv2.1 and Kv1.5 staining was undoubtedly and predominantly detected in the cytoplasm compartment of tumour cells. Indeed, the expression of Kv2.1 in tumour cells revealed a significant association with the early gastric cancer clinical stage (p = 0.026). Conclusion The data highlight, for the first time, the potential role of Kv1.5 and Kv2.1 in gastrointestinal-related cancers and suggests they may be promising prognostic markers for these tumours.

Molecular Cloning of the gyrA andgyrB Genes of Bacteroides fragilis Encoding DNA Gyrase

1999 ◽  
Vol 43 (10) ◽  
pp. 2423-2429 ◽  
Author(s):  
Yoshikuni Onodera ◽  
Kenichi Sato
Keyword(s):  
Hot Spot ◽  
Bacteroides Fragilis ◽  
Dna Gyrase ◽  
E Coli ◽  
Important Target ◽  
Mutant Strains ◽  
Genes Encoding ◽  
Gyrase A ◽  
Selection For

ABSTRACT The genes encoding the DNA gyrase A and B subunits ofBacteroides fragilis were cloned and sequenced. ThegyrA and gyrB genes code for proteins of 845 and 653 amino acids, respectively. These proteins were expressed inEscherichia coli, and the combination of GyrA and GyrB exhibited ATP-dependent supercoiling activity. To analyze the role of DNA gyrase in quinolone resistance of B. fragilis, we isolated mutant strains by stepwise selection for resistance to increasing concentrations of levofloxacin. We analyzed the resistant mutants and showed that Ser-82 of GyrA, equivalent to resistance hot spot Ser-83 of GyrA in E. coli, was in each case replaced with Phe. These results suggest that DNA gyrase is an important target for quinolones in B. fragilis.

scholarly journals Identification of a Streptococcus pneumoniae Gene Locus Encoding Proteins of an ABC Phosphate Transporter and a Two-Component Regulatory System

1999 ◽  
Vol 181 (4) ◽  
pp. 1126-1133 ◽  
Author(s):  
Rodger Novak ◽  
Anje Cauwels ◽  
Emmanuelle Charpentier ◽  
Elaine Tuomanen
Keyword(s):  
Response Regulator ◽  
Regulatory System ◽  
Phosphate Limitation ◽  
Pho Regulon ◽  
Loss Of Function ◽  
Gene Encoding ◽  
E Coli ◽  
Two Component ◽  
Pst System

ABSTRACT The Escherichia coli Pst system belongs to the family of ABC transporters. It is part of a phosphate (PHO) regulon which is regulated by extracellular phosphate. Under conditions of phosphate limitation, the response regulator PhoB is phosphorylated by the histidine kinase PhoR and binds to promoters that share a consensus PHO box. Under conditions of phosphate excess, PhoR, Pst, and PhoU downregulate the PHO regulon. Screening of a library of pneumococcal mutants with defects in exported proteins revealed a putative two-component regulatory system, PnpR-PnpS, and a downstream ABC transporter, similar to the Pst system in E. coli including a gene encoding a PhoU protein. Similar to E. coli, mutagenesis of the ATP-binding cassette gene, pstB, resulted in decreased uptake of phosphate. The effects of the loss of the pneumococcal Pst system extended to decreased transformation and lysis. Withdrawal of phosphate led to transformation deficiency in the parent strain R6x but not to penicillin tolerance, suggesting that reduced bacterial death was independent of phosphate. None of these phenotypes was observed in the pneumococcal loss-of-function mutantphoU. By using a lacZ reporter construct, it was demonstrated that expression of the two-component regulatory system PnpR-PnpS was not influenced by different concentrations of phosphate. These results suggest a more complex role of the Pst system in pneumococcal physiology than in that of E. coli.

scholarly journals CTX-M-Type Extended-Spectrum β-Lactamases in Italy: Molecular Epidemiology of an Emerging Countrywide Problem

2006 ◽  
Vol 50 (8) ◽  
pp. 2700-2706 ◽  
Author(s):  
Claudia Mugnaioli ◽  
Francesco Luzzaro ◽  
Filomena De Luca ◽  
Gioconda Brigante ◽  
Mariagrazia Perilli ◽  
...  
Keyword(s):  
Molecular Epidemiology ◽  
Nationwide Survey ◽  
E Coli ◽  
Extended Spectrum ◽  
Random Amplification ◽  
Genes Encoding ◽  
Italian Territory ◽  
Esbl Producers ◽  
Group 1

ABSTRACT A nationwide survey of extended-spectrum β-lactamase (ESBL) production among Enterobacteriaceae, carried out in 2003, showed that CTX-M-type enzymes have achieved a sizeable prevalence among ESBL producers in Italy, mostly in Escherichia coli and, to a lesser extent, in Klebsiella pneumoniae. In this work, we report on the molecular epidemiology of the CTX-M-producing isolates from that survey and on the mechanisms of dissemination of these emerging resistance determinants. The CTX-M-producing isolates were detected in 10 of the 11 participating centers distributed across the Italian national territory, although at remarkably variable rates in different centers (1.2 to 49.5% of the ESBL producers). All CTX-M determinants were of group 1, with CTX-M-15 and CTX-M-1 being the most prevalent variants (60% and 35%, respectively) and CTX-M-32 carried by a minority (5%) of isolates. Each variant was detected both in E. coli and in K. pneumoniae. Genotyping of the CTX-M-producing isolates by random amplification of polymorphic DNA revealed a notable diversity, especially among those producing CTX-M-1, while clonal expansion was evident with some CTX-M-15-producing strains. Mating experiments revealed a higher overall transferability of bla CTX-M-1 and bla CTX-M-32 than of bla CTX-M-15. Coresistance to quinolones and aminoglycosides was overall higher with the CTX-M-15-producing isolates. The present results indicate that CTX-M-producing strains are now widespread across the Italian territory and underscore the emerging role of these ESBL determinants in the European setting. They also reveal notable differences in the dissemination mechanisms of genes encoding different CTX-M variants of the same lineage.

scholarly journals Toxin, toxin-coregulated pili, and the toxR regulon are essential for Vibrio cholerae pathogenesis in humans.

1988 ◽  
Vol 168 (4) ◽  
pp. 1487-1492 ◽  
Author(s):  
D A Herrington ◽  
R H Hall ◽  
G Losonsky ◽  
J J Mekalanos ◽  
R K Taylor ◽  
...  
Keyword(s):  
Cholera Toxin ◽  
Regulatory Protein ◽  
Deletion Mutation ◽  
Intestinal Colonization ◽  
Gene Encoding ◽  
Disease Symptoms ◽  
Mutant Strains ◽  
A Subunit ◽  
First Time

Isogenic mutant strains of V. cholerae O1 lacking elements of a genetic regulon controlled by toxR and implicated in virulence were tested in volunteers. A deletion mutation in ctxA, the gene encoding the A subunit of cholera toxin, markedly attenuated disease symptoms without affecting intestinal colonization. Deletion of toxR, the gene encoding the cholera toxin-positive regulatory protein resulted in a diminution in colonizing capacity. A deletion mutation in tcpA, encoding the major subunit of the toxin coregulated pilus (regulated by toxR), abolished the colonizing capacity of this strain. These results show for the first time the role of a specific pilus structure in colonization of the human intestine by V. cholerae O1 and exemplify the significance of a genetic regulon in pathogenesis.

scholarly journals Biochemical and Mutational Analysis of Glutamine Synthetase Type III from the Rumen Anaerobe Ruminococcus albus 8

2005 ◽  
Vol 187 (21) ◽  
pp. 7481-7491 ◽  
Author(s):  
Kensey R. Amaya ◽  
Svetlana A. Kocherginskaya ◽  
Roderick I. Mackie ◽  
Isaac K. O. Cann
Keyword(s):  
Glutamine Synthetase ◽  
Mutational Analysis ◽  
Sufficient Conditions ◽  
Kinetic Studies ◽  
Type I ◽  
Gene Encoding ◽  
Genes Encoding ◽  
Subunit Organization ◽  
Glutamyl Transferase ◽  
First Time

ABSTRACT Two different genes encoding glutamine synthetase type I (GSI) and GSIII were identified in the genome sequence of R. albus 8. The identity of the GSIII protein was confirmed by the presence of its associated conserved motifs. The glnN gene, encoding the GSIII, was cloned and expressed in Escherichia coli BL21 cells. The recombinant protein was purified and subjected to biochemical and physical analyses. Subunit organization suggested a protein present in solution as both monomers and oligomers. Kinetic studies using the forward and the γ-glutamyl transferase (γ-GT) assays were carried out. Mutations that changed conserved glutamic acid residues to alanine in the four GSIII motifs resulted in drastic decreases in GS activity using both assays, except for an E380A mutation, which rather resulted in an increase in activity in the forward assay compared to the wild-type protein. Reduced GSIII activity was also exhibited by mutating, individually, two lysines (K308 and K318) located in the putative nucleotide-binding site to alanine. Most importantly, the presence of mRNA transcripts of the glnN gene in R. albus 8 cells grown under ammonia limiting conditions, whereas little or no transcript was detected in cells grown under ammonia sufficient conditions, suggested an important role for the GSIII in the nitrogen metabolism of R. albus 8. Furthermore, the mutational studies on the conserved GSIII motifs demonstrated, for the first time, their importance in the structure and/or function of a GSIII protein.

scholarly journals Role of the Escherichia coli glgX Gene in Glycogen Metabolism

2005 ◽  
Vol 187 (4) ◽  
pp. 1465-1473 ◽  
Author(s):  
David Dauvillée ◽  
Isabelle S. Kinderf ◽  
Zhongyi Li ◽  
Behjat Kosar-Hashemi ◽  
Michael S. Samuel ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Glycogen Synthesis ◽  
High Specificity ◽  
Glycogen Metabolism ◽  
Futile Cycle ◽  
E Coli ◽  
Debranching Enzymes ◽  
Genes Encoding ◽  
Definition Of

ABSTRACT A role for the Escherichia coli glgX gene in bacterial glycogen synthesis and/or degradation has been inferred from the sequence homology between the glgX gene and the genes encoding isoamylase-type debranching enzymes; however, experimental evidence or definition of the role of the gene has been lacking. Construction of E. coli strains with defined deletions in the glgX gene is reported here. The results show that the GlgX gene encodes an isoamylase-type debranching enzyme with high specificity for hydrolysis of chains consisting of three or four glucose residues. This specificity ensures that GlgX does not generate an extensive futile cycle during glycogen synthesis in which chains with more than four glucose residues are transferred by the branching enzyme. Disruption of glgX leads to overproduction of glycogen containing short external chains. These results suggest that the GlgX protein is predominantly involved in glycogen catabolism by selectively debranching the polysaccharide outer chains that were previously recessed by glycogen phosphorylase.

scholarly journals Overexpression of Mannitol-1-Phosphate Dehydrogenase Increases Mannitol Accumulation and Adds Protection Against Chilling Injury in Petunia

2005 ◽  
Vol 130 (4) ◽  
pp. 605-610 ◽  
Author(s):  
Yu-Jen Chiang ◽  
C. Stushnoff ◽  
A.E. McSay ◽  
M.L. Jones ◽  
H.J. Bohnert
Keyword(s):  
Chilling Stress ◽  
Chilling Injury ◽  
Dry Weight ◽  
Wild Type ◽  
Gene Encoding ◽  
E Coli ◽  
Horticultural Crops ◽  
Transgenic Lines ◽  
And Control

Petunia ×hybrida (Hook) Vilm. cv. Mitchell was transformed with an E. coli gene encoding mannitol-1-phosphate dehydrogenase (mtlD). Four plant lines that grew on kanamycin and contained the mtlD transgene were identified. Two of these lines contained high levels of mannitol [high-mannitol lines M3 and M8; mean mannitol = 3.39 μmol·g-1 dry weight (DW)] compared to nontransformed wild-type plants (0.86 μmol·g-1 DW), while two lines had mannitol levels similar to wild-type plants (low-mannitol lines M2 and M9; mean mannitol = 1.05 μmol·g-1 DW). Transgenic and control plants were subjected to chilling stress (3 ± 0.5 °C day/0 ± 0.5 °C night, 12-hour photoperiod and 75% relative humidity) to evaluate the role of mannitol in chilling tolerance. Based upon foliage symptoms and membrane leakage after a 3-week chilling treatment, the high-mannitol containing lines, M3 and M8, were more tolerant of chilling stress than the low-mannitol containing transgenic lines, M2 and M9, and wild-type. Under nonchilling conditions mannitol was the only carbohydrate that differed among transgenic lines, but all carbohydrates were present. When subjected to chilling stress, mannitol levels dropped by 75%, sucrose by 52%, and inositol by 54% in the low-mannitol lines (M2 and M9). In M3 and M8, the high-mannitol lines, mannitol levels decreased by 36%, sucrose by 25%, and inositol by 56%, respectively. Raffinose increased 2- to 3-fold in all lines following exposure to low-temperature chilling stress. In the higher mannitol lines only 0.04% to 0.06% of the total osmotic potential generated from all solutes could be attributed to mannitol, thus its action is more like that of an osmoprotectant rather than an osmoregulator. This study demonstrates that metabolic engineering of osmoprotectant synthesis pathways can be used to improve stress tolerance in horticultural crops.

scholarly journals Toxicity of Recombinant Necrosis and Ethylene-Inducing Proteins (NLPs) from Neofusicoccum parvum

Toxins ◽  
2020 ◽  
Vol 12 (4) ◽  
pp. 235
Author(s):  
Forough Nazar Pour ◽  
Rebeca Cobos ◽  
Juan José Rubio Coque ◽  
João Serôdio ◽  
Artur Alves ◽  
...  
Keyword(s):  
Recombinant Proteins ◽  
Fungal Pathogen ◽  
Vero Cells ◽  
Cytotoxic Effects ◽  
Neofusicoccum Parvum ◽  
Induce Cell Death ◽  
E Coli ◽  
Wide Range ◽  
Genes Encoding ◽  
First Time

Neofusicoccum parvum is a fungal pathogen associated with a wide range of plant hosts. Despite being widely studied, the molecular mechanism of infection of N. parvum is still far from being understood. Analysis of N. parvum genome lead to the identification of six putative genes encoding necrosis and ethylene-inducing proteins (NLPs). The sequence of NLPs genes (NprvNep 1-6) were analyzed and four of the six NLP genes were successfully cloned, expressed in E. coli and purified by affinity chromatography. Pure recombinant proteins were characterized according to their phytotoxic and cytotoxic effects to tomato leaves and to mammalian Vero cells, respectively. These assays revealed that all NprvNeps tested are cytotoxic to Vero cells and also induce cell death in tomato leaves. NprvNep2 was the most toxic to Vero cells, followed by NprvNep1 and 3. NprvNep4 induced weaker, but, nevertheless, still significant toxic effects to Vero cells. A similar trend of toxicity was observed in tomato leaves: the most toxic was NprvNep 2 and the least toxic NprvNep 4. This study describes for the first time an overview of the NLP gene family of N. parvum and provides additional insights into its pathogenicity mechanism.

Sign in / Sign up

Export Citation Format

Share Document