Thauera sp. for efficient nitrate removal in continuous denitrifying moving bed biofilm reactor.

Thauera is the most widely found dominant denitrifying genus in wastewater. In earlier study, MBBR augmented with a specially developed denitrifying five-membered bacterial consortium (DC5) where Thauera was found to be the most abundant and persistent genus. Therefore, to check the functional potential of Thauera in the removal of nitrate-containing wastewater in the present study Thauera sp.V14 one of the member of the consortium DC5 was used as the model organism. Thauera sp.V14 exhibited strong hydrophobicity, auto-aggregation ability, biofilm formation and denitrification ability, which indicated its robust adaptability short colonization and nitrate removal efficiency. Continuous reactor studies with Thauera sp.V14 in 10 L dMBBR showed 91% of denitrification efficiency with an initial nitrate concentration of 620 mg L-1 within 3 h of HRT. Thus, it revealed that Thauera can be employed as an effective microorganism for nitrate removal from wastewater based on its performance in the present studies.

Moving bed biofilm reactor developed with special microbial seed for denitrification of high nitrate containing wastewater.

Biological denitrification is the most promising alternative approach for the removal of nitrate from wastewater. MBBR inoculated with activated sludge is a widely studied approach, but very few studies have focused on the bioaugmentation of biofilm forming bacteria in MBBR. Our study revealed that the use of special microbial seed of biofilm forming denitrifying bacteria Diaphorobacter sp. R4, Pannonibacter sp. V5, Thauera sp. V9, Pseudomonas sp.V11, and Thauera sp.V14 to form biofilm on carriers enhanced nitrate removal performance of developed MBBR. Various process parameters C/N ratio 0.3, HRT 3 h at Nitrate loading 2400 mg L-1, Filling ratio 20%, operated with Pall ring carrier were optimized to achieve highest nitrate removal. After 300 days of continuous operation results of whole genome metagenomic studies showed that Thauera spp. were the most dominant and key contributor to the denitrification of nitrate containing wastewater and the reactor was totally conditioned for denitrification. Overall, findings suggest that bench-scale MBBR developed with biofilm forming denitrifying microbial seed accelerated the denitrification process; therefore in conclusion it is suggested as one of the best suitable and effective approach for removal of nitrate from wastewater.

Disrupting the quorum sensing mediated virulence in soft rot causing Pectobacterium carotovorum by marine sponge associated Bacillus sp. OA10.

Strains of genus Pectobacterium are major cause of soft rot diseases in fruits and vegetables worldwide. Traditional control methods have not been very successful in combating the pathogenesis. As a result there has been an emerging need for developing an alternative ecofriendly and economical strategy. The pathogenesis of Pectobacterium sp. is mediated by quorum sensing (QS) and approaches based on inhibition of QS system to shut down the virulence genes without affecting growth of the pathogen may serve the purpose. Bacillus sp. OA10 associated with purple sponge Haliclona sp. was found to possess extracellular quorum quenching activity. The OA10 extract inhibited QS dependent virulence of Pectobacterium carotovorum subsp. carotovorum BR1 (PccBR1) at low concentrations (0.2 mg) as evident from 77.56 ± 6.17% reduction in potato maceration with complete inhibition by 0.8 mg. Inhibition of plant cell wall degrading enzymes (PCWDE) and carbapenem production by PccBR1 in presence of OA10 extract indicated disruption of the two QS pathways ExpI/ExpR and CarI/CarR in PccBR1. Bacillus sp. OA10 was not found to degrade acyl homoserine lactone (AHL), instead exhibited QSI activity by probably inhibiting AHL synthesis in PccBR1. Absence of enzymatic principle in quorum sensing inhibitor (QSI) is beneficial as enzymes may get inhibited by various factors during their application. OA10 extract did not affect growth of PccBR1, thereby reducing the chance of developing resistance against the QSI. Thus, Bacillus sp. OA10 can prove to be a good prospective candidate for QSI based novel biocontrol formulations.

Bacteria associated with marine macroorganisms as potential source of quorum-sensing antagonists.

Samples were collected from different undisturbed areas along the coast of Gujarat like Okha, Diu, Veraval, and Somnath. A total of 68 marine isolates were obtained out of which 53 were associated with various marine macroorganisms like sponges, gastropods, and algae, whereas 15 were free living. Quorum-quenching ability of all the isolates was tested against Chromobacterium violaceum MK by co-culture technique as a way to simultaneously detect signal-degrading as well as nondegrading quorum-sensing inhibitors. Nineteen macroorganism-associated bacteria and eight free-living bacteria were found to possess quorum-sensing inhibitory activity against C. violaceum MK without affecting its growth. Isolate OA22 from grape alga and OA10 from purple sponge (Haliclona sp.) were found to possess the highest C6-HSL degradation activity and extracellular non-N-acyl-homoserine lactone degrading QSI activity, respectively. OA22 was also found to degrade 3-oxo-C12 homoserine lactone. Acid recovery of both the C6- and C12-HSL after degradation by OA22 indicated the presence of lactonase enzyme in the isolate. Cell-free supernatant of OA10 was extracted with ethyl acetate to obtain the quorum-quenching compound. Pigment inhibition in C. violaceum MK treated with OA10 extract was demonstrated in various ways and was indicative of QSI activity of the extract without degradation of the quorum-sensing signaling molecule. The isolates OA22 and OA10 were identified as Desemzia incerta and Bacillus sp., respectively, by 16S ribosomal DNA sequence analysis.

Optimization of concurrent production of xylanolytic and pectinolytic enzymes by Bacillus safensis M35 and Bacillus altitudinis J208 using agro-industrial biomass through Response Surface Methodology.

Application of crude xylanolytic and pectinolytic enzymes in diverse industrial processes make these enzymes commercially valuable and demand their production process to be cost-effective. Out of four different agrowaste biomass, wheat bran (WB) and citrus peel (CP), when amended as fermentation substrates, respectively induced the highest xylanolytic enzymes and pectinolytic enzymes from both, B. safensis M35 and B. altitudinis J208. Further, the simultaneous amendment of WB and CP yielded concurrent production of these cellulase free xylanolytic and pectinolytic enzymes. Hence, the quadratic model was developed using the Central Composite Design of Response Surface Method (CCD-RSM). The model gave the concentration values for WB and CP substrates to be amended in one single production medium for obtaining two optimized predicted response values of xylanase activity and pectinase activity units, which were further practically validated for the xylanase and pectinase production responses from the optimized production medium (OPM). These practically obtained response values from OPM were found to be in accordance with a range of 95% predicted intervals (PI) values. These observations verified the validity of the predicted quadratic model from RSM and suggested that both xylanase and pectinase enzymes can be induced concurrently from both of the bacterial strains.

Valorization of sugarcane bagasse by chemical pretreatment and enzyme mediated deconstruction.

After chemical pretreatment, improved amenability of agrowaste biomass for enzymatic saccharification needs an understanding of the effect exerted by pretreatments on biomass for enzymatic deconstruction. In present studies, NaOH, NH4OH and H2SO4 pretreatments effectively changed visible morphology imparting distinct fibrous appearance to sugarcane bagasse (SCB). Filtrate analysis after NaOH, NH4OH and H2SO4 pretreatments yielded release of soluble reducing sugars (SRS) in range of ~0.17-0.44%, ~0.38-0.75% and ~2.9-8.4% respectively. Gravimetric analysis of pretreated SCB (PSCB) biomass also revealed dry weight loss in range of ~25.8-44.8%, ~11.1-16.0% and ~28.3-38.0% by the three pretreatments in the same order. Release of soluble components other than SRS, majorly reported to be soluble lignins, were observed highest for NaOH followed by H2SO4 and NH4OH pretreatments. Decrease or absence of peaks attributed to lignin and loosened fibrous appearance of biomass during FTIR and SEM studies respectively further corroborated with our observations of lignin removal. Application of commercial cellulase increased raw SCB saccharification from 1.93% to 38.84%, 25.56% and 9.61% after NaOH, H2SO4 and NH4OH pretreatments. Structural changes brought by cell wall degrading enzymes were first time shown visually confirming the cell wall disintegration under brightfield, darkfield and fluorescence microscopy. The microscopic evidence and saccharification results proved that the chemical treatment valorized the SCB by making it amenable for enzymatic saccharification.

Characterization of Solibacillus silvestris strain AM1 that produces amyloid bioemulsifier.

Solibacillus silvestris AM1 was the first strain from the genus to be reported for the production of a functional amyloid and its potential use as a surface active agent, a thermostable glycoprotein amyloid bioemulsifier BE-AM1 capable of influencing environment and biofilm formation. Phylogenetic analysis based on 16S rRNA gene, molecular characterization studies on the basis of DNA-DNA hybridization and chemotaxonomic fatty acid methyl ester (FAME) analysis showed that S. silvestris AM1 as a strain matches with the type strain S. silvestris HR3-23. But strain AM1 differs from the type strain HR3-23 in carbon substrate utilization studies along with amyloid bioemulsifier production ability with potential industrial and environmental applications. S. silvestris AM1 exhibited bioemulsifier production at wide range of factors like pH and NaCl concentrations, while temperature influenced the bioemulsifier production indirectly (since it affected the growth). Bioemulsifier production was observed even at oligotrophic conditions (0.5 mg ml-1 ) seen usually in its native environment. In this study, we have characterized the amyloid producing S. silvestris AM1 taxonomically and also analyzed 16S rDNA of 103 sequences of Solibacillus sp. available, which indicated the possibility of new species in this genus and can be studied for industrially and environmentally important biomolecules.

A Novel Methylotrophic Bacterial Consortium for Treatment of Industrial Effluents.

Considering the importance of methylotrophs in industrial wastewater treatment, focus of the present study was on utilization of a methylotrophic bacterial consortium as a microbial seed for biotreatment of a variety of industrial effluents. For this purpose, a mixed bacterial methylotrophic AC (Ankleshwar CETP) consortium comprising of Bordetella petrii AC1, Bacillus licheniformis AC4, Salmonella subterranea AC5, and Pseudomonas stutzeri AC8 was used. The AC consortium showed efficient biotreatment of four industrial effluents procured from fertilizer, chemical and pesticide industries, and common effluent treatment plant by lowering their chemical oxygen demand (COD) of 950-2000 mg/l to below detection limit in 60-96 h in 6-l batch reactor and 9-15 days in 6-l continuous reactor. The operating variables of wastewater treatment, viz. COD, BOD, pH, MLSS, MLVSS, SVI, and F/M ratio of these effluents, were also maintained in the permissible range in both batch and continuous reactors. Therefore, formation of the AC consortium has led to the development of an efficient microbial seed capable of treating a variety of industrial effluents containing pollutants generated from their respective industries.

Attenuation of Quorum Sensing Regulated Virulence of Pectobacterium carotovorum subsp. carotovorum through an AHL Lactonase Produced by Lysinibacillus sp. Gs50.

Quorum sensing (QS) is a mechanism in which Gram negative bacterial pathogens sense their population density through acyl homoserine lactones (AHLs) and regulate the expression of virulence factors. Enzymatic degradation of AHLs by lactonases, known as quorum quenching (QQ), is thus a potential strategy for attenuating QS regulated bacterial infections. We characterised the QQ activity of soil isolate Lysinibacillus sp. Gs50 and explored its potential for controlling bacterial soft rot of crop plants. Lysinibacillus sp. Gs50 inactivated AHL, which could be restored upon acidification, suggested that inactivation was due to the lactone ring hydrolysis of AHL. Heterologous expression of cloned gene for putative hydrolase (792 bp) designated adeH from Lysinibacillus sp. Gs50 produced a ~29 kDa protein which degraded AHLs of varying chain length. Mass spectrometry analysis of AdeH enzymatic reaction product revealed that AdeH hydrolyses the lactone ring of AHL and hence is an AHL lactonase. Multiple sequence alignment of the amino acid sequence of AdeH showed that it belongs to the metallo- ß- lactamase superfamily, has a conserved "HXHXDH" motif typical of AHL lactonases. KM for AdeH for C6HSL was found to be 3.089 µM and the specific activity was 0.8 picomol min-1µg-1. AdeH has not so far been reported from any Lysinibacillus sp. and has less than 40% identity with known AHL lactonases. Finally we found that Lysinibacillus sp. Gs50 can degrade AHL produced by Pectobacterium carotovorum subsp. carotovorum (Pcc), a common cause of soft rot. This QQ activity causes a decrease in production of plant cell wall degrading enzymes of Pcc and attenuates symptoms of soft rot in experimental infection of potato, carrot and cucumber. Our results demonstrate the potential of Lysinibacillus sp. Gs50 as a preventive and curative biocontrol agent.

Characterization of Biosurfactant Produced by Bacillus licheniformis TT42 Having Potential for Enhanced Oil Recovery.

Bacillus licheniformis TT42 produced a low-molecular weight anionic biosurfactant that reduced the surface tension of water from 72 to 27 mN/m and the interfacial tension from 12 to 0.05 mN/m against crude oil. We have earlier reported significant enhancement in oil recovery in laboratory sand pack columns and core flood studies, by biosurfactant-TT42 compared to standard strain, Bacillus mojavensis JF2. In the context of this application of the biosurfactant-TT42, its characterization was deemed important. In the preliminary studies, the biosurfactant-TT42 was found to be functionally stable at under conditions of temperature, pH, and salinity generally prevalent in oil reservoirs. Furthermore, the purified biosurfactant-TT42 was found to have a CMC of 22 mg/l. A newly developed activity staining TLC method was used for the purification of biosurfactant-TT42. Structural characterization of biosurfactant-TT42 using TLC, Fourier transform infrared spectroscopy (FTIR), GC-MS, and matrix-assisted laser desorption ionization time of flight (MALDI-TOF)/TOF suggested that it was a mixture of lipopeptide species, all having a common hydrophilic cyclic heptapeptide head with the sequence, Gln-Leu/Ileu-Leu/Ileu-Val-Asp-Leu/Ileu-Leu/Ileu linked to hydrophobic tails of different lengths of 3ß-OH-fatty acids bearing 1043, 1057 and 1071 Da molecular weight, where 3ß-OH-C19 fatty acid was predominant. This is the longest chain length of fatty acids reported in a lipopeptide.

Interference of Quorum Sensing by Delftia sp. VM4 Depends on the Activity of a Novel N-Acylhomoserine Lactone-Acylase.

BACKGROUND: Turf soil bacterial isolate Delftia sp. VM4 can degrade exogenous N-acyl homoserine lactone (AHL), hence it effectively attenuates the virulence of bacterial soft rot pathogen Pectobacterium carotovorum subsp. carotovorum strain BR1 (Pcc BR1) as a consequence of quorum sensing inhibition. METHODOLOGY/PRINCIPAL FINDINGS: Isolated Delftia sp. VM4 can grow in minimal medium supplemented with AHL as a sole source of carbon and energy. It also possesses the ability to degrade various AHL molecules in a short time interval. Delftia sp. VM4 suppresses AHL accumulation and the production of virulence determinant enzymes by Pcc BR1 without interference of the growth during co-culture cultivation. The quorum quenching activity was lost after the treatment with trypsin and proteinase K. The protein with quorum quenching activity was purified by three step process. Matrix assisted laser desorption/ionization-time of flight (MALDI-TOF) and Mass spectrometry (MS/MS) analysis revealed that the AHL degrading enzyme (82 kDa) demonstrates homology with the NCBI database hypothetical protein (Daci_4366) of D. acidovorans SPH-1. The purified AHL acylase of Delftia sp. VM4 demonstrated optimum activity at 20-40°C and pH 6.2 as well as AHL acylase type mode of action. It possesses similarity with an α/ß-hydrolase fold protein, which makes it unique among the known AHL acylases with domains of the N-terminal nucleophile (Ntn)-hydrolase superfamily. In addition, the kinetic and thermodynamic parameters for hydrolysis of the different AHL substrates by purified AHL-acylase were estimated. Here we present the studies that investigate the mode of action and kinetics of AHL-degradation by purified AHL acylase from Delftia sp. VM4. SIGNIFICANCE: We characterized an AHL-inactivating enzyme from Delftia sp. VM4, identified as AHL acylase showing distinctive similarity with α/ß-hydrolase fold protein, described its biochemical and thermodynamic properties for the first time and revealed its potential application as an anti-virulence agent against bacterial soft rot pathogen Pectobacterium carotovorum subsp. carotovorum based on quorum quenching mechanism.

Nitrate levels modulate the abundance of Paracoccus sp. in a biofilm community.

Conditions required to enhance a particular species efficient in degradative capabilities is very useful in wastewater treatment processes. Paracoccus sp. is known to efficiently reduce nitrogen oxides (NOx) due to the branched denitrification pathway. Individual-based simulations showed that the relative fitness of Paracoccus sp. to Pseudomonas sp. increased significantly with nitrate levels above 5 mM. Spatial structure of the biofilm showed substantially less nitrite levels in the areas of Paracoccus sp. dominance. The simulation was validated in a laboratory reactor harboring biofilm community by fluorescent in situ hybridization, which showed that increasing nitrate levels enhanced the abundance of Paracoccus sp. Different levels of NOx did not display any significant effect on biofilm formation of Paracoccus sp., unlike several other bacteria. This study shows that the attribute of Paracoccus sp. to tolerate and efficiently reduce NOx is conferring a fitness payoff to the organism at high concentrations of nitrate in a multispecies biofilm community.

Mutualism between autotrophic ammonia-oxidizing bacteria (AOB) and heterotrophs present in an ammonia-oxidizing colony.

Coexistence of an autotrophic ammonia-oxidizing bacterium (Nitrosomonas sp. RA) and heterotrophic bacteria was consistently observed when cultured in an inorganic medium without any external supply of organic carbon. The present study was undertaken to understand the association between autotrophs and the associated heterotrophs for which a system containing active autotrophs and heterotrophs controlled by Hg(2+) addition was developed. The study revealed interdependence of heterotrophs and Nitrosomonas sp. RA for growth under iron-limited condition. Growth of Nitrosomonas sp. RA was supported by siderophores produced by the associated heterotroph, Pusillimonas sp., thereby complementing its high iron requirement while the organics (such as pyruvate) excreted by Nitrosomonas sp. RA during its autotrophic growth supported the survival of heterotrophs in the inorganic medium. The study thus sheds light on the nature of the mutual interactions between heterotrophs and autotrophs that play a role in the ammonia-oxidizing system involved in wastewater treatment.

Development of a simultaneous partial nitrification, anaerobic ammonia oxidation and denitrification (SNAD) bench scale process for removal of ammonia from effluent of a fertilizer industry.

A simultaneous partial nitrification, anammox and denitrification (SNAD) process was developed for the treatment of ammonia laden effluent of a fertilizer industry. Autotrophic aerobic and anaerobic ammonia oxidizing biomass was enriched and their ammonia removal ability was confirmed in synthetic effluent system. Seed consortium developed from these was applied in the treatment of effluent in an oxygen limited bench scale SNAD type (1L) reactor run at ambient temperature (∼30°C). Around 98.9% ammonia removal was achieved with ammonia loading rate 0.35kgNH(4)(+)-N/m(3)day in the presence of 46.6mg/L COD at 2.31days hydraulic retention time. Qualitative and quantitative analysis of the biomass from upper and lower zone of the reactor revealed presence of autotrophic ammonia oxidizing bacteria (AOB), Planctomycetes and denitrifiers as the dominant bacteria carrying out anoxic oxidation of ammonia in the reactor. Physiological and molecular studies strongly indicate presence of anammox bacteria in the anoxic zone of the SNAD reactor.

Occurrence of Biosurfactant Producing Bacillus spp. in Diverse Habitats.

Diversity among biosurfactant producing Bacillus spp. from diverse habitats was studied among 77 isolates. Cluster analysis based on phenotypic characteristics using unweighted pair-group method with arithmetic averages (UPGMAs) method was performed. Bacillus isolates possessing high surface tension activity and five reference strains were subjected to amplified 16S rDNA restriction analysis (ARDRA). A correlation between the phenotypic and genotypic characterization of Bacillus spp. is explored. Most of the oil reservoir isolates showing high surface activity clustered with B. licheniformis and B. subtilis, the hot water spring isolates clustered in two ingroups, while the petroleum contaminated soil isolates were randomly distributed in all the three ingroups. Present work revealed that diversity exists in distribution of Bacillus spp. from thermal and hydrocarbon containing habitats where majority of organisms belonged to B. licheniformis and B. subtilis group. Isolate B. licheniformis TT42 produced biosurfactant which reduced the surface tension of water from 72 mNm(-1) to 28 mNm(-1), and 0.05 mNm(-1) interfacial tension against crude oil at 80°C. This isolate clustered with B. subtilis and B. licheniformis group on the basis of ARDRA. These findings increase the possibility of exploiting the Bacillus spp. from different habitats and their possible use in oil recovery.

Carbon sources influence the nitrate removal activity, community structure and biofilm architecture.

Influence of the frequently used carbon sources in nitrate removal processes were evaluated in a lab-scale biofilm reactor. The NO3-N removal efficiency was in the order acetate>glucose>methanol>ethanol. Acetate-fed biofilm reduced nearly 100% NO3-N with negligible amount of NO2-N accumulation. Although 99% NO3-N was reduced in the glucose-fed biofilm, substantial NH3-N and NO2-N accumulated. Methanol-fed biofilm reduced 72% of NO3-N with accumulation of 2.2 mg L(-1) of NO2-N, while biofilm formed in presence of ethanol showed 61% reduction in NO3-N although relatively higher ratio of denitrifiers were observed. Acetate and ethanol-fed biofilm displayed characteristic biofilm architecture with voids, but the former had relatively higher thickness and diffusion distance. In presence of glucose and methanol, a confluent biofilm without characteristic voids was formed. Pseudomonas sp. numerically dominated the acetate and ethanol-fed biofilm, while Enterobacter sp. and Methylobacillus sp., were abundant in glucose and methanol biofilms respectively.

Assessment of denitrifying bacterial composition in activated sludge.

The abundance and structure of denitrifying bacterial community in different activated sludge samples were assessed, where the abundance of denitrifying functional genes showed nirS in the range of 10(4)-10(5), nosZ with 10(4)-10(6) and 16S rRNA gene in the range 10(9)-10(10) copy number per ml of sludge. The culturable approach revealed Pseudomonas sp. and Alcaligenes sp. to be numerically high, whereas culture independent method showed betaproteobacteria to dominate the sludge samples. Comamonas sp. and Pseudomonas fluorescens isolates showed efficient denitrification, while Pseudomonas mendocina, Pseudomonas stutzeri and Brevundimonas diminuta accumulated nitrite during denitrification. Numerically dominant RFLP OTUs of the nosZ gene from the fertilizer factory sludge samples clustered with the known isolates of betaproteobacteria. The data also suggests the presence of different truncated denitrifiers with high numbers in sludge habitat.

Comparison of denitrification between Paracoccus sp. and Diaphorobacter sp.

Denitrification was compared between Paracoccus sp. and Diaphorobacter sp. in this study, both of which were isolated from activated sludge of a denitrifying reactor. Denitrification of both isolates showed contrasting patterns, where Diaphorobacter sp. showed accumulation of nitrite in the medium while Paracoccus sp. showed no accumulation. The nitrate reduction rate was 1.5 times more than the nitrite reduction in Diaphorobacter sp., as analyzed by the resting state denitrification kinetics. Increasing the nitrate concentration in the medium increased the nitrite accumulation in Diaphorobacter sp., but not in Paracoccus sp., indicating a branched electron transfer during denitrification. Diaphorobacter sp. was unable to denitrify efficiently at high nitrate concentrations from 1 M, but Paracoccus sp. could denitrify even up to 2 M nitrate. Paracoccus sp. was found to be an efficient denitrifier with insignificant amounts of nitrite accumulation, and it could also denitrify high amounts of nitrate up to 2 M. Efficient denitrification without accumulation of intermediates like nitrite is desirable in the removal of high nitrates from wastewaters. Paracoccus sp. is shown to suffice this demand and could be a potential organism to remove high nitrates effectively.

Biochemical and thermal stabilization parameters of polygalacturonase from Erwinia carotovora subsp. carotovora BR1.

With emphasis on thermal behavior in presence of different pH conditions and salts, the kinetic and thermodynamic parameters of purified polygalacturonase (PG) of E. carotovora subsp. carotovora (Ecc) BR1 were studied since characterization of an enzyme is significant in the context of burgeoning biotechnological applications. Thermodynamic parameters for polygalacturonic acid hydrolysis by purified PG were, deltaH* = 7.98 kJ/mol, deltaG* = 68.86 kJ/mol, deltaS*= -194.48 J/mol/K, deltaG(E-S) = -1.04 kJ/mol and deltaG(E-T) = -8.96 kJ/mol. Its turnover number (k(cat)) was 21/sec. Purified PG was stable in 20-50 degrees C temperature range and was deactivated at 60 degrees C and 70 degrees C. Thermodynamic parameters (deltaH*, deltaG*, deltaS*) for irreversible inactivation of PG at different temperatures (30-60 degrees C) were determined, where effectiveness of various salts and different pH (4-8) individually for thermal stability of PG were characterized. The efficacy of various salts for thermal stability of PG was in the following order: MgCl2 >BaCl2 >KCl >CaCl2 >NaCl. Present work projects biochemical, thermodynamics of substrate hydrolysis as well as thermal stabilization parameters of PG from Ecc.

Structural and molecular characteristics of lichenysin and its relationship with surface activity.

Lichenysins are most potent anionic cyclic lipoheptapeptide biosurfactants produced by Bacillus licheniformis on hydrocarbonless medium with mainly glucose as carbon source. They have the capacity to lower the surface tension of water from 72 to 27 mN/m. Based on species specific variations they are named lichenysin A, B, C, D, G and surfactant BL86. The lowest ever interfacial tension against decane of 0.006 mN/m is obtained with acid precipitated lichenysin B. Surfactant BL86 and lichenysin B have recorded lowest ever CMC of 10 mg/L by any surfactant under optimal conditions. Surface and interfacial tension lowering ability bears significance in the context of oil recovery from oil reservoir. Similarity exists between structure and biosynthesis of surfactin and lichenysin. Surfactin being the most studied of the two, understanding its structure and biosynthesis gives an insight into the structure and biosynthesis of lichenysin. Lichenysin is synthesized by a multienzyme complex, lichenysin synthetase (LchA/Lic) encoded by 32.4 (26.6 kb) lichenysin operon lchA (lic). The structure of lichenysin and its operon indicate the nonribosomal biosynthesis with the same multifunctional modular arrangement as seen in surfactin synthetase SrfA. The lchA operon consists of lchAA-AC (lic A-C) and lchA TE (licTE) genes encoding the proteins LchAA, LchAB, LchAC and thioesterase LchA-TE. The licA (lchAA) gene is 10,746 bp and codes for a 3,582 amino acids protein, licB (lchAB) gene is 10,764 bp and codes for a similar sized protein, while licC (lchAC) gene is 3,864 bp and codes for protein containing 1,288 amino acid. The biotechnological potential of lichenysin in MEOR has triggered research on structure-activity relationship. Both the nature of peptide and fatty acid dictate the activity of the biosurfactant. Tailormade biosurfactant with desired attributes can be obtained from engineered synthetases. Basic studies are lacking on mechanism of biosynthesis by lichenysin synthetase however, studies on various aspects of lichenysin including regulation are expected to swell in coming years.