To develop an alternative quorum quenching therapy against multidrug-resistant Acinetobacter baum... more To develop an alternative quorum quenching therapy against multidrug-resistant Acinetobacter baumannii. Activity-guided partially purified fraction (F1) from Glycyrrhiza glabra significantly (p < 0.05) reduced quorum sensing regulated virulence factors of A. baumannii viz. motility, biofilm formation and production of antioxidant enzymes. Mechanistically, F1 downregulated the expression of autoinducer synthase gene, abaI, and consequently reduced (92%) the production of 3-OH-C12-HSL as determined by ESI-MS. Q-TOF and Q-TRAP analyses suggested the presence of flavonoids viz. licoricone, glycyrin and glyzarin as the active ingredients. This is the first report on quorum quenching activity of G. glabra linked to its flavonoids that downregulated the expression of abaI and attenuated quorum sensing regulated virulence of A. baumannii.
Quorum Sensing vs Quorum Quenching: A Battle with No End in Sight, 2014
Indiscriminate use of antibiotics has led to the emergence of multiple drug-resistant pathogens. ... more Indiscriminate use of antibiotics has led to the emergence of multiple drug-resistant pathogens. Quorum sensing (QS) is an intercellular communication system which is widely present in pathogens regulating the virulence genes expression and pathogenesis in host. Interruption of QS regulatory system is an attractive strategy to attenuate pathogens’ virulence. Several synthetic and natural compounds with potential QS inhibitory activity have been identified and tested in vitro for their inhibitory activity. To study the in vivo efficacy of the compounds, a model organism is required. Unlike other vertebrate model organisms like mouse or rabbit, C. elegans has emerged as a simple, amenable, genetically tractable worm for studying QS inhibitory activity of compounds. This chapter focuses on the advantages and recent development in using C. elegans as a non-mammalian in vivo model system.
Journal of Environmental Pathology, Toxicology and Oncology, 2003
We evaluated for mutagenicity 14 commercial textile dyes used extensively in the northern part of... more We evaluated for mutagenicity 14 commercial textile dyes used extensively in the northern part of India using both the Ames Salmonella typhimurium microsome reversion test as well as the recombination-repair (rec)-assay. The Ames test revealed that 57.14% of dyes were mutagenic and acting directly. The rec-assay detected 50% of dyes to be mutagenic; of these, 71.43% were direct acting, whereas 28.57% required Aroclor-induced exogenous metabolic activation. Used together, the two tests detected 78.57% of the dyes to be mutagenic, and a 50% correlation was found between these two tests. Groupwise, three out of four azo dyes and all five anthraquinone dyes were found to be mutagenic by the Ames assay; the rec-assay detected methine/polymethine (1 out of 3), an oxazine, and a triphenylmethane dye to be mutagenic, besides the azo (1 out of 4) and the anthraquinone (3 out of 5) dyes. The structure-activity analysis attributed the mutagenicity of dyes to the structural alerts such as phenylenediamine, amino and nitro-groups, methylation, CH=CH, and chloro groups; whereas deamination, bulkier groups, and sulfonation may be responsible for diminishing mutagenicity.
World Journal of Microbiology and Biotechnology, 2014
Pseudomonas aeruginosa possesses an arcade of both cell-associated and extracellular cytotoxic vi... more Pseudomonas aeruginosa possesses an arcade of both cell-associated and extracellular cytotoxic virulence factors which are regulated by a multi-component quorum sensing system. Many research studies report success of lactonase in combating the pathogenicity of P. aeruginosa but delivery of lactonase remains a challenge. The present study aims at developing a delivery vehicle for lactonase. Lactobacillus plantarum NC8 was used as host for aiiA (Bacillus thuringiensis 4A3 lactonase gene) using pSIP409 expression vector. pSIP409: aiiA construct was stably maintained in L. plantarum NC8. Co-culturing of multi-drug resistant (MDR) clinical isolates of P. aeruginosa and PAO1 with recombinant L. plantarum NC8 led to significant reduction (p &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.001) in extracellular virulence factors like pyocyanin, protease, elastase and rhamnolipids in P. aeruginosa and also showed significant reduction in adhesion of P. aeruginosa strains to uroepithelial cells in vitro. This study shows the heterologous expression of AiiA lactonase in L. plantarum NC8. Co-culturing of lactonase expressing L. plantarum NC8 with MDR P. aeruginosa strains led to attenuation of their virulence significantly. These results underscore the potential application of recombinant L. plantarum NC8 with anti-quorum sensing properties to control infections caused by multidrug resistant P. aeruginosa.
Journal of Environmental Pathology, Toxicology and Oncology, 2004
Fourteen textile and biological dyes, belonging to the azo, triphenylmethane, anthraquinone, hete... more Fourteen textile and biological dyes, belonging to the azo, triphenylmethane, anthraquinone, heterocyclic, oxazine, and methine/polymethine groups, were degraded using the PhotoFenton treatment (PFT) and the Phanerochaete chrysosporium crude ligninase enzyme (ED) treatment. The genotoxicity of the dyes and of their degradation products were assessed with the rec-assay. We found that the genotoxicity depended on the dye and on the method of degradation. In general, PFT was better than ED in decreasing the genotoxicity. Basic dyes showed complete or maximum loss of genotoxicity, whereas the vat group was more resistant. The azo group showed varied results. Crystal Violet was the only dye whose genotoxicity increased after PFT. Our results suggest that PFT and ED are two effective treatment methods to reduce the genotoxicity of dyes in waste waters.
Thermophilic Microbes in Environmental and Industrial Biotechnology, 2013
ABSTRACT Restriction endonucleases (REases) are enzymes that recognize and cleave DNA in a sequen... more ABSTRACT Restriction endonucleases (REases) are enzymes that recognize and cleave DNA in a sequence specific manner. The recognition site consists of a sequence of nucleotides in the DNA duplex, typically four to eight base pairs long. Most of the commercially produced REases are isolated from the mesophilic bacteria. But the disadvantage of REases from mesophilic sources is that these enzymes are usually denatured at ambient and high temperature. As temperature produces opposite effects on both enzyme activity and stability, it is therefore a key variable in any biocatalytic process. Also, mesophilic enzymes are unstable, have low reactivity, lose activity during purification, and require refrigerated transport and storage. So, thermostable REases are preferred to circumvent these problems. This chapter deals mainly with thermophilic REases. The increasing interest in this field is reflected by the growing information on the discovery, purification, and characterization of REases from thermophilic sources. The properties associated with these enzymes offer additional advantages over their mesophilic counterparts.
A Gram-negative, alkalotolerant bacterium, isolated from the soil continually drained with indust... more A Gram-negative, alkalotolerant bacterium, isolated from the soil continually drained with industrial wastewater and identified as gamma-proteobacterium by partial 16S rRNA sequence analysis, produced a polyphenol oxidase, which showed laccase but not tyrosinase activity. The organism grew well from pH 6 to 10 and produced laccase maximally at pH 10. The enzyme was stable from pH 3 to 10.6 for at least 24 h
ABSTRACT An alkali-thermostable β-mannanase from Bacillus nealsonii PN-11 was purified 38.96-fold... more ABSTRACT An alkali-thermostable β-mannanase from Bacillus nealsonii PN-11 was purified 38.96-fold to homogeneity with specific activity of 2,288.90 ± 27.80 U mg−1 protein and final recovery of 8.92 ± 0.09 %. The purified β-mannanase was an extracellular monomeric protein with a molecular mass of 50 kDa on SDS–PAG E. The first 20 N-terminal amino acid sequence of mannanase enzyme was MVVKKLSSFILILLLVTSAL. The optimal temperature and pH for enzyme were 65 °C and 8.8, respectively. It was completely stable at 60 °C for 3 h and retained &gt;50 ± 1.0 % activity at 70 °C up to 3 h. The β-mannanase was highly stable between pH 5–10 and retained &gt;85 % of the initial activity for 3 h. The metal ions Ni+2, Co+2, Zn+2 and Mg+2 enhanced the enzyme activity. The enzyme remained stable after 3 h of preincubation with most of the tested organic solvents. According to substrate specificity study, the purified mannanase had high specificity to locust bean gum which was degraded mainly to mannooligosaccharides (MOS) like mannotriose, mannotetraose and mannopentose. These MOS enhanced the growth of Lactobacillus casei but inhibited the growth of Salmonella enterica indicating potential prebiotic properties. The properties of the purified β-mannanase from B. nealsonii PN-11 make this enzyme attractive for biotechnological applications.
To develop an alternative quorum quenching therapy against multidrug-resistant Acinetobacter baum... more To develop an alternative quorum quenching therapy against multidrug-resistant Acinetobacter baumannii. Activity-guided partially purified fraction (F1) from Glycyrrhiza glabra significantly (p < 0.05) reduced quorum sensing regulated virulence factors of A. baumannii viz. motility, biofilm formation and production of antioxidant enzymes. Mechanistically, F1 downregulated the expression of autoinducer synthase gene, abaI, and consequently reduced (92%) the production of 3-OH-C12-HSL as determined by ESI-MS. Q-TOF and Q-TRAP analyses suggested the presence of flavonoids viz. licoricone, glycyrin and glyzarin as the active ingredients. This is the first report on quorum quenching activity of G. glabra linked to its flavonoids that downregulated the expression of abaI and attenuated quorum sensing regulated virulence of A. baumannii.
Quorum Sensing vs Quorum Quenching: A Battle with No End in Sight, 2014
Indiscriminate use of antibiotics has led to the emergence of multiple drug-resistant pathogens. ... more Indiscriminate use of antibiotics has led to the emergence of multiple drug-resistant pathogens. Quorum sensing (QS) is an intercellular communication system which is widely present in pathogens regulating the virulence genes expression and pathogenesis in host. Interruption of QS regulatory system is an attractive strategy to attenuate pathogens’ virulence. Several synthetic and natural compounds with potential QS inhibitory activity have been identified and tested in vitro for their inhibitory activity. To study the in vivo efficacy of the compounds, a model organism is required. Unlike other vertebrate model organisms like mouse or rabbit, C. elegans has emerged as a simple, amenable, genetically tractable worm for studying QS inhibitory activity of compounds. This chapter focuses on the advantages and recent development in using C. elegans as a non-mammalian in vivo model system.
Journal of Environmental Pathology, Toxicology and Oncology, 2003
We evaluated for mutagenicity 14 commercial textile dyes used extensively in the northern part of... more We evaluated for mutagenicity 14 commercial textile dyes used extensively in the northern part of India using both the Ames Salmonella typhimurium microsome reversion test as well as the recombination-repair (rec)-assay. The Ames test revealed that 57.14% of dyes were mutagenic and acting directly. The rec-assay detected 50% of dyes to be mutagenic; of these, 71.43% were direct acting, whereas 28.57% required Aroclor-induced exogenous metabolic activation. Used together, the two tests detected 78.57% of the dyes to be mutagenic, and a 50% correlation was found between these two tests. Groupwise, three out of four azo dyes and all five anthraquinone dyes were found to be mutagenic by the Ames assay; the rec-assay detected methine/polymethine (1 out of 3), an oxazine, and a triphenylmethane dye to be mutagenic, besides the azo (1 out of 4) and the anthraquinone (3 out of 5) dyes. The structure-activity analysis attributed the mutagenicity of dyes to the structural alerts such as phenylenediamine, amino and nitro-groups, methylation, CH=CH, and chloro groups; whereas deamination, bulkier groups, and sulfonation may be responsible for diminishing mutagenicity.
World Journal of Microbiology and Biotechnology, 2014
Pseudomonas aeruginosa possesses an arcade of both cell-associated and extracellular cytotoxic vi... more Pseudomonas aeruginosa possesses an arcade of both cell-associated and extracellular cytotoxic virulence factors which are regulated by a multi-component quorum sensing system. Many research studies report success of lactonase in combating the pathogenicity of P. aeruginosa but delivery of lactonase remains a challenge. The present study aims at developing a delivery vehicle for lactonase. Lactobacillus plantarum NC8 was used as host for aiiA (Bacillus thuringiensis 4A3 lactonase gene) using pSIP409 expression vector. pSIP409: aiiA construct was stably maintained in L. plantarum NC8. Co-culturing of multi-drug resistant (MDR) clinical isolates of P. aeruginosa and PAO1 with recombinant L. plantarum NC8 led to significant reduction (p &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.001) in extracellular virulence factors like pyocyanin, protease, elastase and rhamnolipids in P. aeruginosa and also showed significant reduction in adhesion of P. aeruginosa strains to uroepithelial cells in vitro. This study shows the heterologous expression of AiiA lactonase in L. plantarum NC8. Co-culturing of lactonase expressing L. plantarum NC8 with MDR P. aeruginosa strains led to attenuation of their virulence significantly. These results underscore the potential application of recombinant L. plantarum NC8 with anti-quorum sensing properties to control infections caused by multidrug resistant P. aeruginosa.
Journal of Environmental Pathology, Toxicology and Oncology, 2004
Fourteen textile and biological dyes, belonging to the azo, triphenylmethane, anthraquinone, hete... more Fourteen textile and biological dyes, belonging to the azo, triphenylmethane, anthraquinone, heterocyclic, oxazine, and methine/polymethine groups, were degraded using the PhotoFenton treatment (PFT) and the Phanerochaete chrysosporium crude ligninase enzyme (ED) treatment. The genotoxicity of the dyes and of their degradation products were assessed with the rec-assay. We found that the genotoxicity depended on the dye and on the method of degradation. In general, PFT was better than ED in decreasing the genotoxicity. Basic dyes showed complete or maximum loss of genotoxicity, whereas the vat group was more resistant. The azo group showed varied results. Crystal Violet was the only dye whose genotoxicity increased after PFT. Our results suggest that PFT and ED are two effective treatment methods to reduce the genotoxicity of dyes in waste waters.
Thermophilic Microbes in Environmental and Industrial Biotechnology, 2013
ABSTRACT Restriction endonucleases (REases) are enzymes that recognize and cleave DNA in a sequen... more ABSTRACT Restriction endonucleases (REases) are enzymes that recognize and cleave DNA in a sequence specific manner. The recognition site consists of a sequence of nucleotides in the DNA duplex, typically four to eight base pairs long. Most of the commercially produced REases are isolated from the mesophilic bacteria. But the disadvantage of REases from mesophilic sources is that these enzymes are usually denatured at ambient and high temperature. As temperature produces opposite effects on both enzyme activity and stability, it is therefore a key variable in any biocatalytic process. Also, mesophilic enzymes are unstable, have low reactivity, lose activity during purification, and require refrigerated transport and storage. So, thermostable REases are preferred to circumvent these problems. This chapter deals mainly with thermophilic REases. The increasing interest in this field is reflected by the growing information on the discovery, purification, and characterization of REases from thermophilic sources. The properties associated with these enzymes offer additional advantages over their mesophilic counterparts.
A Gram-negative, alkalotolerant bacterium, isolated from the soil continually drained with indust... more A Gram-negative, alkalotolerant bacterium, isolated from the soil continually drained with industrial wastewater and identified as gamma-proteobacterium by partial 16S rRNA sequence analysis, produced a polyphenol oxidase, which showed laccase but not tyrosinase activity. The organism grew well from pH 6 to 10 and produced laccase maximally at pH 10. The enzyme was stable from pH 3 to 10.6 for at least 24 h
ABSTRACT An alkali-thermostable β-mannanase from Bacillus nealsonii PN-11 was purified 38.96-fold... more ABSTRACT An alkali-thermostable β-mannanase from Bacillus nealsonii PN-11 was purified 38.96-fold to homogeneity with specific activity of 2,288.90 ± 27.80 U mg−1 protein and final recovery of 8.92 ± 0.09 %. The purified β-mannanase was an extracellular monomeric protein with a molecular mass of 50 kDa on SDS–PAG E. The first 20 N-terminal amino acid sequence of mannanase enzyme was MVVKKLSSFILILLLVTSAL. The optimal temperature and pH for enzyme were 65 °C and 8.8, respectively. It was completely stable at 60 °C for 3 h and retained &gt;50 ± 1.0 % activity at 70 °C up to 3 h. The β-mannanase was highly stable between pH 5–10 and retained &gt;85 % of the initial activity for 3 h. The metal ions Ni+2, Co+2, Zn+2 and Mg+2 enhanced the enzyme activity. The enzyme remained stable after 3 h of preincubation with most of the tested organic solvents. According to substrate specificity study, the purified mannanase had high specificity to locust bean gum which was degraded mainly to mannooligosaccharides (MOS) like mannotriose, mannotetraose and mannopentose. These MOS enhanced the growth of Lactobacillus casei but inhibited the growth of Salmonella enterica indicating potential prebiotic properties. The properties of the purified β-mannanase from B. nealsonii PN-11 make this enzyme attractive for biotechnological applications.
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