Industrial applications of fungal lipases: a review.

Fungal lipases (triacylglycerol acyl hydrolases EC 3.1.1.3) are significant industrial enzymes and have several applications in a number of industries and fields. Fungal lipases are found in several species of fungi and yeast. These enzymes are carboxylic acid esterases, categorized under the serine hydrolase family, and do not require any cofactor during the catalyzing of the reactions. It was also noticed that processes including the extraction and purification of lipases from fungi are comparatively easier and cheaper than other sources of lipases. In addition, fungal lipases have been classified into three chief classes, namely, GX, GGGX, and Y. Fungal lipases have applications not only in the hydrolysis of fats and oils (triglycerides) but are also involved in synthetic reactions such as esterification, acidolysis, alcoholysis, interesterification, and aminolysis. The production and activity of fungal lipases are highly affected by the carbon source, nitrogen source, temperature, pH, metal ions, surfactants, and moisture content. Therefore, fungal lipases have several industrial and biotechnological applications in many fields such as biodiesel production, ester synthesis, production of biodegradable biopolymers, formulations of cosmetics and personal care products, detergent manufacturing, degreasing of leather, pulp and paper production, textile industry, biosensor development, and drug formulations and as a diagnostic tool in the medical sector, biodegradation of esters, and bioremediation of wastewater. The immobilization of fungal lipases onto different carriers also helps in improving the catalytic activities and efficiencies of lipases by increasing thermal and ionic stability (in organic solvents, high pH, and temperature), being easy to recycle, and inducing the volume-specific loading of the enzyme onto the support, and thus, these features have proved to be appropriate for use as biocatalysts in different sectors.

Bioengineering of fungal endophytes through the CRISPR/Cas9 system.

The CRISPR/Cas9 system is a genome-editing tool that allows for precise and efficient modifications to the DNA of a cell. This technology can be used in endophytic fungi, which live within plants and can have beneficial effects on their host, making them important for agriculture. Using CRISPR/Cas9, researchers can introduce specific genetic changes into endophytic fungal genomes, allowing them to study the function of genes, improve their plant-growth-promoting properties, and create new, more beneficial endophytes. This system works by using the Cas9 protein, which acts as a pair of molecular scissors, to cut DNA at specific locations determined by a guide RNA. Once the DNA is cut, the cell's natural repair mechanisms can be used to insert or delete specific genes, allowing for precise editing of the fungal genome. This article discusses the mechanism and applications of CRISPR/Cas9 to fungal endophytes.

CRISPR-Cas system: from diagnostic tool to potential antiviral treatment.

This mini review focuses on the diagnosis and treatment of virus diseases using Crisper-Cas technology. The present paper describes various strategies involved in diagnosing diseases using Crispr-Cas-based assays. Additionally, CRISPR-Cas systems offer great potential as new therapeutic tools for treating viral infections including HIV, Influenza, and SARS-CoV-2. There are several major challenges to be overcome before this technology can be applied routinely in clinical settings, such as finding a suitable delivery tool, toxicity, and immunogenicity, as well as off-target effects. This review also discusses ways to deal with the challenges associated with Crisper-Cas technology. KEY POINTS: • Crisper technology is being applied to diagnose infectious and non-infectious diseases. • A new generation of CRISPR-Cas-based assays has been developed which detect pathogens within minutes, providing rapid diagnosis of diseases. • Crispr-Cas tools can be used to combat viral infections, specifically HIV, influenza, and SARS-CoV-2.

Draft genome sequence data of a 4-nitrophenol- degrading bacterium, Pseudomonas alloputida strain PNP.

A 4-nitrophenol-degrading bacterial strain PNP was isolated from pesticide-contaminated soil collected from Lucknow. Strain PNP utilized 0.5 mM 4-nitrophenol as its carbon source and degraded it completely within 24 h with stoichiometric release of nitrite ions. Strain PNP was associated with the genus Pseudomonas in a phylogentic tree and exhibited highest 16S rRNA gene sequence similarity to Pseudomonas juntendi BML3 (99.79%) and Pseudomonas inefficax JV551A3 (99.79%). Based on values of average nucleotide identity and digital DNA-DNA hybridization among strain PNP and its closely related type strains, it concluded that strain PNP belongs to Pseudomonas alloputida. The Illumina HiSeq platform was used to sequence the PNP genome. The draft genome sequence of Pseudomonas alloputida PNP was presented here. The total size of the draft assembly was 6,087,340 bp, distributed into 87 contigs with N50 value of 139502. The genome has an average GC content of 61.7% and contains 5461 coding sequences and 77 putative RNA genes. This Whole Genome Shotgun project has been submitted at DDBJ/ENA/GenBank under the accession JAGKJH000000000.

Development of ESAT-6 Based Immunosensor for the Detection of Mycobacterium tuberculosis.

Early secreted antigenic target of 6 kDa (ESAT-6) has recently been identified as a biomarker for the rapid diagnosis of tuberculosis. We propose a stable and reusable immunosensor for the early diagnosis of tuberculosis based on the detection and quantification of ESAT-6 via cyclic voltammetry (CV). The immunosensor was synthesized by polymerizing aniline dispersed with the reduced graphene oxide (rGO) and Ni nanoparticles, followed by surface modification of the electroconductive polyaniline (PANI) film with anti-ESAT-6 antibody. Physicochemical characterization of the prepared materials was performed by several analytical techniques, including FE-SEM, EDX, XRD, FT-IR, Raman, TGA, TPR, and BET surface area analysis. The antibody-modified Ni-rGO-PANI electrode exhibited an approximately linear response (R2 = 0.988) towards ESAT-6 during CV measurements over the potential range of -1 to +1 V. The lower detection limit for ESAT-6 was approximately 1.0 ng mL-1. The novelty of this study includes the development of the reusable Ni-rGO-PANI-based electrochemical immunosensor for the early diagnosis of tuberculosis. Furthermore, this study successfully demonstrates that electro-conductive PANI may be used as a polymeric substrate for Ni nanoparticles and rGO.

Draft genome sequence data of a chromium reducing bacterium, Bacillus licheniformis strain KNP.

A chromium-reducing bacterium designated as strain KNP was isolated from a sample collected from a tannery effluent of Kanpur, India. Phylogenetic analysis based on the 16S rRNA gene sequences revealed that strain KNP belonged to the Bacillus genus and showed 100% similarity with Bacillus licheniformis. Furthermore, average nucleotide identity and digital DNA-DNA hybridization between strain KNP and its closely related strains confirmed its affiliation with Bacillus licheniformis species. Whole-genome sequencing of Bacillus licheniformis KNP was performed using the Illumina Hiseq platform. Here, we present the draft genome sequence of Bacillus licheniformis KNP. The total size of the draft assembly was 4,280,093 bp, distributed into 21 contigs with an N50 value of 4,186,229. The genome has 45.9% G + C content, 4255 coding sequences and 86 putative RNA genes. This Whole Genome Shotgun project has been deposited at DDBJ/ENA/GenBank under the accession JACDXS000000000. The version described in this paper is version JACDXS010000000.

Bacilli-Mediated Degradation of Xenobiotic Compounds and Heavy Metals.

Xenobiotic compounds are man-made compounds and widely used in dyes, drugs, pesticides, herbicides, insecticides, explosives, and other industrial chemicals. These compounds have been released into our soil and water due to anthropogenic activities and improper waste disposal practices and cause serious damage to aquatic and terrestrial ecosystems due to their toxic nature. The United States Environmental Protection Agency (USEPA) has listed several toxic substances as priority pollutants. Bacterial remediation is identified as an emerging technique to remove these substances from the environment. Many bacterial genera are actively involved in the degradation of toxic substances. Among the bacterial genera, the members of the genus Bacillus have a great potential to degrade or transform various toxic substances. Many Bacilli have been isolated and characterized by their ability to degrade or transform a wide range of compounds including both naturally occurring substances and xenobiotic compounds. This review describes the biodegradation potentials of Bacilli toward various toxic substances, including 4-chloro-2-nitrophenol, insecticides, pesticides, herbicides, explosives, drugs, polycyclic aromatic compounds, heavy metals, azo dyes, and aromatic acids. Besides, the advanced technologies used for bioremediation of environmental pollutants using Bacilli are also briefly described. This review will increase our understanding of Bacilli-mediated degradation of xenobiotic compounds and heavy metals.

Microbial lipases and their industrial applications: a comprehensive review.

Lipases are very versatile enzymes, and produced the attention of the several industrial processes. Lipase can be achieved from several sources, animal, vegetable, and microbiological. The uses of microbial lipase market is estimated to be USD 425.0 Million in 2018 and it is projected to reach USD 590.2 Million by 2023, growing at a CAGR of 6.8% from 2018. Microbial lipases (EC 3.1.1.3) catalyze the hydrolysis of long chain triglycerides. The microbial origins of lipase enzymes are logically dynamic and proficient also have an extensive range of industrial uses with the manufacturing of altered molecules. The unique lipase (triacylglycerol acyl hydrolase) enzymes catalyzed the hydrolysis, esterification and alcoholysis reactions. Immobilization has made the use of microbial lipases accomplish its best performance and hence suitable for several reactions and need to enhance aroma to the immobilization processes. Immobilized enzymes depend on the immobilization technique and the carrier type. The choice of the carrier concerns usually the biocompatibility, chemical and thermal stability, and insolubility under reaction conditions, capability of easy rejuvenation and reusability, as well as cost proficiency. Bacillus spp., Achromobacter spp., Alcaligenes spp., Arthrobacter spp., Pseudomonos spp., of bacteria and Penicillium spp., Fusarium spp., Aspergillus spp., of fungi are screened large scale for lipase production. Lipases as multipurpose biological catalyst has given a favorable vision in meeting the needs for several industries such as biodiesel, foods and drinks, leather, textile, detergents, pharmaceuticals and medicals. This review represents a discussion on microbial sources of lipases, immobilization methods increased productivity at market profitability and reduce logistical liability on the environment and user.

Punjabi version of the European Organization for Research and Treatment of Cancer Quality of Life Questionnaire-OES 18 and Quality of Life Questionnaire-OG25 modules for quality of life assessment.

CONTEXT: Health-related quality of life (HRQOL) assessment plays an important role in the decision-making process in oncology. AIMS: The aim of the study was to translate European Organization for Research and Treatment of Cancer (EORTC) quality of life questionnaire (QLQ) OES18 and OG25 in Punjabi language for HRQOL assessment of patients diagnosed with esophagus and esophagogastric malignancies. SUBJECTS AND METHODS: The EORTC translation guidelines were duly followed to translate QLQ-OES18 and OG25 into Punjabi language. Each set of questionnaire was independently translated by two forward translators, followed by backward translation of the reconciled version by two independent translators. The final version was submitted to the EORTC Translation Team and served to the patients for the pilot testing. RESULTS: The questionnaire was administered to ten patients each of esophagus and esophagogastric malignancies who were evaluated and treated at our hospital. Every patient underwent an interview to check if any of the questions was difficult, uncomfortable, or upsetting to answer. Their concerns were recorded as per the template provided by the EORTC team and due changes done if required. CONCLUSIONS: The EORTC QLQ-OES18 and OG25 questionnaire has been translated to Punjabi language and subsequently approved for usage.

Recent advances in degradation of chloronitrophenols.

Chloronitrophenols (CNPs) constitute a group of environmental pollutants that are widely distributed in our surrounding environment due to human based activities. This group of chemicals is highly toxic to living beings due to its mutagenic and carcinogenic nature. Examples include 2-chloro-4-nitrophenol, 4-chloro-2-nitrophenol, 2-chloro-5-nitrophenol, 4-chloro-3-nitrophenol and 2,6-dichloro-4-nitrophenol. Several methods including advanced oxidation processes, adsorption and bacterial degradation have been used for degradation of CNPs. Among, bacterial degradation is an eco-friendly and effective way to degrade CNPs. Several bacterial metabolic pathways have been proposed for degradation of CNPs and their genes and enzymes have been identified in bacteria. These bacteria were able to degrade CNPs in broth culture and soil. Therefore, CNPs-degrading bacteria are suitable candidates for bioremediation of CNPs-contaminated sites. Few CNP-degrading bacteria exhibited chemotaxis towards CNPs to enhance their biodegradation. The present review summarizes recent progress in degradation of CNPs.

Lipase catalysis in organic solvents: advantages and applications.

Lipases are industrial biocatalysts, which are involved in several novel reactions, occurring in aqueous medium as well as non-aqueous medium. Furthermore, they are well-known for their remarkable ability to carry out a wide variety of chemo-, regio- and enantio-selective transformations. Lipases have been gained attention worldwide by organic chemists due to their general ease of handling, broad substrate tolerance, high stability towards temperatures and solvents and convenient commercial availability. Most of the synthetic reactions on industrial scale are carried out in organic solvents because of the easy solubility of non-polar compounds. The effect of organic system on their stability and activity may determine the biocatalysis pace. Because of worldwide use of lipases, there is a need to understand the mechanisms behind the lipase-catalyzed reactions in organic solvents. The unique interfacial activation of lipases has always fascinated enzymologists and recently, biophysicists and crystallographers have made progress in understanding the structure-function relationships of these enzymes. The present review describes the advantages of lipase-catalyzed reactions in organic solvents and various effects of organic solvents on their activity.

Amyand's Hernia: Rare Presentation of a Common Ailment.

Inguinal hernia with vermiform appendix as content is known as Amyand's hernia. It is a rare entity but we encountered four cases within six months. A 52-year-old female had high grade fever and evidence of inflammatory pathology involving the ileocaecal region. She was initially managed conservatively and subsequently underwent exploratory laparatomy. The appendix was perforated and herniating in the inguinal canal. Appendectomy was done with herniorrhaphy without mesh placement. A 74-year-old male with bilateral inguinal hernia, of which, the right side was more symptomatic, underwent open exploration. Operative findings revealed a lipoma of the sac and a normal appearing appendix as content. Contents were reduced without appendectomy and mesh hernioplasty was performed. A 63-year-old male with an obstructed right sided hernia underwent emergency inguinal exploration which revealed edematous caecum and appendix as content without any inflammation. Contents were reduced without any resection. Herniorrhaphy was performed without mesh placement. A 66-year-old male with an uncomplicated right inguinal hernia underwent elective surgery. The sac revealed an appendix with adhesions at the neck. Contents were reduced after adhesiolysis and hernioplasty was performed with mesh placement. Emphasis is made to the rarity of disease, variation in presentation, and difference in treatment modalities depending upon the state of appendix.

Identification of new members of the MAPK gene family in plants shows diverse conserved domains and novel activation loop variants.

BACKGROUND: Mitogen Activated Protein Kinase (MAPK) signaling is of critical importance in plants and other eukaryotic organisms. The MAPK cascade plays an indispensible role in the growth and development of plants, as well as in biotic and abiotic stress responses. The MAPKs are constitute the most downstream module of the three tier MAPK cascade and are phosphorylated by upstream MAP kinase kinases (MAPKK), which are in turn are phosphorylated by MAP kinase kinase kinase (MAPKKK). The MAPKs play pivotal roles in regulation of many cytoplasmic and nuclear substrates, thus regulating several biological processes. RESULTS: A total of 589 MAPKs genes were identified from the genome wide analysis of 40 species. The sequence analysis has revealed the presence of several N- and C-terminal conserved domains. The MAPKs were previously believed to be characterized by the presence of TEY/TDY activation loop motifs. The present study showed that, in addition to presence of activation loop TEY/TDY motifs, MAPKs are also contain MEY, TEM, TQM, TRM, TVY, TSY, TEC and TQY activation loop motifs. Phylogenetic analysis of all predicted MAPKs were clustered into six different groups (group A, B, C, D, E and F), and all predicted MAPKs were assigned with specific names based on their orthology based evolutionary relationships with Arabidopsis or Oryza MAPKs. CONCLUSION: We conducted global analysis of the MAPK gene family of plants from lower eukaryotes to higher eukaryotes and analyzed their genomic and evolutionary aspects. Our study showed the presence of several new activation loop motifs and diverse conserved domains in MAPKs. Advance study of newly identified activation loop motifs can provide further information regarding the downstream signaling cascade activated in response to a wide array of stress conditions, as well as plant growth and development.

Biodegradation of 4-chloroindole by Exiguobacterium sp. PMA.

Exiguobacterium sp. PMA utilized 4-chloroindole as its sole source of carbon and energy. The effect of initial concentrations of substrate on the 4-chloroindole degradation was studied and observed that strain PMA was capable of degrading 4-chloroindole up to concentration of 0.5mM. The degradation pathway of 4-chloroindole was studied for Exiguobacterium sp. PMA based on metabolites identified by gas chromatography-mass spectrometry. 4-Chloroindole was initially dehalogenated to indole that was further degraded via isatin, anthranilic acid, and salicylic acid. The potential of strain PMA to degrade 4-chloroindole in soil was monitored using soil microcosms, and it was observed that the cells of strain PMA efficiently degraded 4-chloroindole in the soil. The results of microcosm studies show that strain PMA may be used for bioremediation of 4-chloroindole-contaminated sites. This is the first report of the bacterial degradation of 4-chloroindole.

Identification of new metabolites of bacterial transformation of indole by gas chromatography-mass spectrometry and high performance liquid chromatography.

Arthrobacter sp. SPG transformed indole completely in the presence of an additional carbon source. High performance liquid chromatography and gas chromatography-mass spectrometry detected indole-3-acetic acid, indole-3-glyoxylic acid, and indole-3-aldehyde as biotransformation products. This is the first report of the formation of indole-3-acetic acid, indole-3-glyoxylic acid, and indole-3-aldehyde from indole by any bacterium.

Metabolic pathway for degradation of 2-chloro-4-aminophenol by Arthrobacter sp. SPG.

A degradation pathway of 2-chloro-4-aminophenol (2C4AP) was studied in an Arthrobacter sp. SPG that utilized 2C4AP as its sole source of carbon and energy. The 2C4AP degradation was initiated by a 2C4AP-deaminase that catalyzed the conversion of 2C4AP into chlorohydroquinone (CHQ) with removal of ammonium ion. In the next step, a CHQ-dehalogenase dehalogenated CHQ to hydroquinone (HQ) that cleaved into γ-hydroxymuconic semialdehyde by a HQ-dioxygenase. The 2C4AP degradation was also investigated in sterile and non-sterile soil microcosms using strain SPG. The results show that the SPG cells degraded 2C4AP more rapidly in sterile soil than non-sterile soil. Our studies showed that strain SPG may be used for bioremediation of 2C4AP-contaminated sites. This is the first report of the 2C4AP degradation by any bacteria.

Biotransformation and chemotaxis of 4-chloro-2-nitrophenol by Pseudomonas sp. JHN.

Pseudomonas sp. JHN decolourized and biotransformed 4-chloro-2-nitrophenol (4C2NP) in the presence of additional carbon source. The effect of the various concentrations of the 4C2NP was studied on the decolourization of 4C2NP by Pseudomonas sp. JHN. It was observed that strain JHN decolourized and biotransformed 4C2NP up to concentration of 0.6 mM. Gas chromatography and gas chromatography-mass spectrometry detected 5-chloro-2-methylbenzoxazole as a major metabolite of the co-metabolism of 4C2NP. Furthermore, strain JHN exhibits positive chemotaxis toward 4C2NP based on the drop plate and capillary assays. This is the first report of the chemotaxis toward 4C2NP by any bacterium.

Integration of bioinformatics to biodegradation.

Bioinformatics and biodegradation are two primary scientific fields in applied microbiology and biotechnology. The present review describes development of various bioinformatics tools that may be applied in the field of biodegradation. Several databases, including the University of Minnesota Biocatalysis/Biodegradation database (UM-BBD), a database of biodegradative oxygenases (OxDBase), Biodegradation Network-Molecular Biology Database (Bionemo) MetaCyc, and BioCyc have been developed to enable access to information related to biochemistry and genetics of microbial degradation. In addition, several bioinformatics tools for predicting toxicity and biodegradation of chemicals have been developed. Furthermore, the whole genomes of several potential degrading bacteria have been sequenced and annotated using bioinformatics tools.

Bacterial degradation of chlorophenols and their derivatives.

Chlorophenols (CPs) and their derivatives are persistent environmental pollutants which are used in the manufacture of dyes, drugs, pesticides and other industrial products. CPs, which include monochlorophenols, polychlorophenols, chloronitrophenols, chloroaminophenols and chloromethylphenols, are highly toxic to living beings due to their carcinogenic, mutagenic and cytotoxic properties. Several physico-chemical and biological methods have been used for removal of CPs from the environment. Bacterial degradation has been considered a cost-effective and eco-friendly method of removing CPs from the environment. Several bacteria that use CPs as their sole carbon and energy sources have been isolated and characterized. Additionally, the metabolic pathways for degradation of CPs have been studied in bacteria and the genes and enzymes involved in the degradation of various CPs have been identified and characterized. This review describes the biochemical and genetic basis of the degradation of CPs and their derivatives.

Degradation of 4-chloro-3-nitrophenol via a novel intermediate, 4-chlororesorcinol by Pseudomonas sp. JHN.

A 4-chloro-3-nitrophenol (4C3NP)-mineralizing bacterium, Pseudomonas sp. JHN was isolated from a waste water sample collected from a chemically-contaminated area, India by an enrichment method. Pseudomonas sp. JHN utilized 4C3NP as a sole carbon and energy source and degraded it with the release of stoichiometric amounts of chloride and nitrite ions. Gas chromatography and gas chromatography-mass spectrometry detected 4-chlororesorcinol as a major metabolite of the 4C3NP degradation pathway. Inhibition studies using 2,2'-dipyridyl showed that 4-chlororesorcinol is a terminal aromatic compound in the degradation pathway of 4C3NP. The activity for 4C3NP-monooxygenase was detected in the crude extracts of the 4C3NP-induced JHN cells that confirmed the formation of 4-chlororesorcinol from 4C3NP. The capillary assay showed that Pseudomonas sp. JHN exhibited chemotaxis toward 4C3NP. The bioremediation capability of Pseudomonas sp. JHN was monitored to carry out the microcosm experiments using sterile and non-sterile soils spiked with 4C3NP. Strain JHN degraded 4C3NP in sterile and non-sterile soil with same degradation rates. This is the first report of (i) bacterial degradation and bioremediation of 4C3NP, (ii) formation of 4-chlororesorcinol in the degradation pathway of 4C3NP, (iii) bacterial chemotaxis toward 4C3NP.