Low-cost cultivation of Nannochloropsis oceanica in newly designed photobioreactors and its productivity trends in semi-continuous cultivation under inland outdoor conditions.

Most marine microalgae are typically cultivated in coastal areas due to challenges in inland cultivation. In this 185 days experiment, Nannochloropsis oceanica was semi-continuously cultivated inland using different photobioreactors (PBRs). The newly designed 700-liter (L) PBR exhibited tolerance to seasonal changes compared to the 150-L PBRs. The innovative in-situ oxygen release rate (ORR) measurement method results indicated that ORR was influenced by light intensity and temperature. The optimal temperature range for N. oceanica growth was 14-25 â„ƒ, demonstrated cold tolerance and lipid accumulation at low temperatures. The maximum lipid content in 700-L and 150-L PBRs was 29 % and 28 %, respectively. Based on the average biomass productivity, the price of N. oceanica was $11.89 kg-1 (or $3.35 kg-1 based on maximum biomass productivity), which is cheaper than the current market price of $20.19 kg-1. From results, smaller PBRs at the same hydro electricity price are more cost-effective.

Optimization and purification of bioproducts from Bacillus velezensis PhCL fermentation and their potential on industrial application and bioremediation.

Bioproduction is considered a promising alternative way of obtaining useful and green chemicals. However, the downstream process of biomolecules has been one of the major difficulties in upscaling the application of bioproducts due to the high purification cost. Acid precipitation is the most common method for purifying biosurfactants from the fermentation broth with high purity. However, the use of strong acids and organic solvents in solvent extraction has limited its application. Hence, in this study, a new strain of Bacillus velezensis PhCL was isolated from phenolic waste, and its production of amylase had been optimized via response surface methodology. After that, amylase and biosurfactant were purified by sequential ammonium sulfate precipitation and the result suggested that even though the purified crude biosurfactant had a lower purification fold compared to the acid precipitation, the yield was higher and both enzymes and biosurfactant also could be recovered for lowering the purification cost. Moreover, the purified amylase and crude biosurfactant were characterized and the results suggested that the purified crude biosurfactant would have a higher emulsion activity and petroleum hydrocarbon removal rate compared to traditional surfactants. This study provided another approach for purifying bioactive compounds including enzymes and biosurfactants from the same fermentation broth and further explored the potential of the crude purified biosurfactant in the bioremediation of polycyclic aromatic hydrocarbons and petroleum hydrocarbons.

Characterization of Potential Virulence, Resistance to Antibiotics and Heavy Metals, and Biofilm-Forming Capabilities of Soil Lignocellulolytic Bacteria.

Soil bacteria participate in self-immobilization processes for survival, persistence, and production of virulence factors in some niches or hosts through their capacities for autoaggregation, cell surface hydrophobicity, biofilm formation, and antibiotic and heavy metal resistance. This study investigated potential virulence, antibiotic and heavy metal resistance, solvent adhesion, and biofilm-forming capabilities of six cellulolytic bacteria isolated from soil samples: Paenarthrobacter sp. MKAL1, Hymenobacter sp. MKAL2, Mycobacterium sp. MKAL3, Stenotrophomonas sp. MKAL4, Chryseobacterium sp. MKAL5, and Bacillus sp. MKAL6. Strains were subjected to phenotypic methods, including heavy metal and antibiotic susceptibility and virulence factors (protease, lipase, capsule production, autoaggregation, hydrophobicity, and biofilm formation). The effect of ciprofloxacin was also investigated on bacterial susceptibility over time, cell membrane, and biofilm formation. Strains MKAL2, MKAL5, and MKAL6 exhibited protease and lipase activities, while only MKAL6 produced capsules. All strains were capable of aggregating, forming biofilm, and adhering to solvents. Strains tolerated high amounts of chromium, lead, zinc, nickel, and manganese and were resistant to lincomycin. Ciprofloxacin exhibited bactericidal activity against these strains. Although the phenotypic evaluation of virulence factors of bacteria can indicate their pathogenic nature, an in-depth genetic study of virulence, antibiotic and heavy metal resistance genes is required.

A comparative study of Cellulomonas sp. and Bacillus sp. in utilizing lignocellulosic biomass as feedstocks for enzyme production.

The demand for enzymes is increasing continuously due to their applications in various avenues. The pectin-hydrolyzing bacteria, Cellulomonas sp. and Bacillus sp., isolated from forest soil have the potential to produce industrially important enzymes (pectinase, PGase, Cellulase, and xylanase). However, these bacteria have different optimal cultural conditions for pectinase production. The optimal cultural conditions for Cellulomonas sp. were room temperature (25-26℃), pH 7, 1% inoculum volume, and 1.5% citrus pectin with 8.82 ± 0.92 U/mL pectinase activity. And Bacillus sp. illustrated the highest pectinase activity (12.35 ± 0.72 U/mL) at room temperature, pH 10, 1% inoculum volume, and 1.5% pectin concentration. Among the different agro-wastes, the orange peel was found to be the best substrate for pectinase, PGase, and cellulase activity whereas barley straw for xylanase activity. Further, Cellulomonas sp. and Bacillus sp. illustrated higher pectinase activity from commercial pectin compared to orange peel showing their preference for commercial citrus pectin. In addition, the optimization by the Box-Behnken design increased pectinase activity for Cellulomonas sp., while a noticeable increase in activity was not observed in Bacillus sp. Besides, all the agro-wastes exploited in this study can be used for pectinase, PGase, and xylanase production but not cellulase. The study revealed that each bacteria has its specific optimal conditions and there is a variation in the capacity of utilizing the various lignocellulosic biomass.

Optimization of cultural conditions for pectinase production by Streptomyces sp. and characterization of partially purified enzymes.

The cultural parameters of Streptomyces sp. for pectinase production were optimized using the Box-Behnken design. The maximum pectinase production was obtained after 58 hours at 35℃ and pH 7 upon submerged fermentation in yeast extract-containing media. The enzymes were partially purified with acetone precipitation and the analysis by SDS-PAGE and zymogram revealed that Streptomyces sp. produced two pectinases with molecular weights of about 25 and 75 kDa. The pectinase activity was detected in a wide range of temperatures (30℃ to 80℃) and pH (3 to 9) with maximum pectinase activities observed at 70℃ and pHs 5 and 9. The enzymes retained about 30 to 40% of their activities even after incubating the enzyme at different temperatures for 120 mins. The pectinase activities of Streptomyces sp. were enhanced in the media containing 1.5% pectin, 1% casein as a nitrogen source, 0.5 mM MgSO4, and 5 mM NaCl. Further, the addition of Tween-20, amino acids, and vitamins to the media also enhanced the pectinase activity. Moreover, the bacterium illustrated the ability to decolorize crystal violet dye efficiently. The decolorization rate ranged from 39.29 to 53.75% showing the highest bacterial decolorization in the media containing 2mg/mL crystal violet at 144 hours. Therefore, the bacterium has the potential in treating wastewater produced by industries like textile industries.

Optimization of multiple enzymes production by fermentation using lipid-producing Bacillus sp.

The present study identified the pectinase-producing bacterium isolated from the contaminated broth as Bacillus sp. on 16S rDNA sequence analysis. The bacterium illustrated water-like droplets on the colony grown on the Sabouraud dextrose agar plate. It also exhibited multi-enzymes activities, such as pectinase, polygalacturonase, xylanase, and cellulase by using various agro-wastes as low-cost substrates. The orange peel was observed to be the best substrate among the agro-wastes used for maximum multi-enzymes (pectinase, polygalacturonase, xylanase, and cellulase). However, the bacterium demonstrated its capability to produce different enzymes according to the different substrates/agro-wastes used. The Plackett-Burman design was used to determine the essential influencing factors, while the Box Behnken design response surface methodology was for optimizing cultural conditions. At their optimal conditions (40°C incubation temperature, 24 h of incubation period, 1% w/v orange peel, and 2% v/v inoculum volume), the bacterium exhibited the maximum pectinase (9.49 ± 1.25 U/ml) and xylanase (16.27 ± 0.52 U/ml) activities. Furthermore, the study explored the ability of the bacterium to produce bacterial lipids and observed about 25% bacterial lipid content on a dry weight basis. Therefore, the bacterium is a good candidate for producing important multi-enzymes and subsequent agro-waste degradation controlling the environment, and facilitating waste management. Also, the bacterium can be a potential feedstock in producing renewable biofuel.

Characterization of Cellulose-Degrading Bacteria Isolated from Soil and the Optimization of Their Culture Conditions for Cellulase Production.

The characterization of bacteria with hydrolytic potential significantly contributes to the industries. Six cellulose-degrading bacteria were isolated from mixture soil samples collected at Kingfisher Lake and the University of Manitoba campus by Congo red method using carboxymethyl cellulose agar medium and identified as Paenarthrobacter sp. MKAL1, Hymenobacter sp. MKAL2, Mycobacterium sp. MKAL3, Stenotrophomonas sp. MKAL4, Chryseobacterium sp. MKAL5, and Bacillus sp. MKAL6. Their cellulase production was optimized by controlling different environmental and nutritional factors such as pH, temperature, incubation period, substrate concentration, nitrogen, and carbon sources using the dinitrosalicylic acid and response surface methods. Except for Paenarthrobacter sp. MKAL1, all strains are motile. Only Bacillus sp. MKAL6 was non-salt-tolerant and showed gelatinase activity. Sucrose enhanced higher cellulase activity of 78.87 ± 4.71 to 190.30 ± 6.42 U/mL in these strains at their optimum pH (5-6) and temperature (35-40 °C). The molecular weights of these cellulases were about 25 kDa. These bacterial strains could be promising biocatalysts for converting cellulose into glucose for industrial purposes.

Chloroplast genome analysis of Chrysotila dentata.

Chrysotila dentata is an ecologically important marine alga contributing to the coccolith formation. In this study, a complete chloroplast (cp DNA) genome of Chrysotila dentata was sequenced by using Illumina Hiseq and was analyzed with the help of a bioinformatics tool CPGAVAS2. The circular chloroplast genome of Chrysotila dentata has a size of 109,017 bp with two inverted repeats (IRs) regions (4513 bp each) which is a common feature in most land plants and algal species. The Chrysotila dentata cp genome consists of 61 identified protein-coding genes, 30 tRNA genes and 6 rRNAs with 21 microsatellites. The phylogenetic relationship with other select algal species revealed a close phylogeny of Chrysotila dentata with Phaeocystis antarctica. This is the first report of the cp genome analysis of genus Chrysotila and the results from this study will be helpful for understanding the genetic structure and function of chloroplast in other species of Chrysotila.

mtDNA analysis confirms the endangered Kashmir musk deer extends its range to Nepal.

Musk deer Moschus spp. are endemic to the high mountain forests of central Asia. The taxonomic status of musk deer in the central and western Himalayas is poorly understood. We investigated the phylogenetic relationship of musk deer from the central and western Himalayas based on mitochondrial genomic data of Cytochrome b (380 bps) and D-loop (1000 bps). Our results distinguished two divergent lineages using higher bootstrap support (bs) values from the Maximum likelihood and Bayesian posterior probabilities (bpp). Both the Manang and Kaski lineages from central Nepal are confirmed as Himalayan musk deer Moschus leucogaster and represent a species complex widespread throughout the central and eastern Himalayan region. The musk deer Mustang lineage was confirmed as Kashmir musk deer Moschus cupreus and has wide distribution in the western Himalayas (from central Nepal to Afghanistan). Our analysis validates that Kashmir musk deer is a genetically distinct species and it clarifies that Himalayan musk deer and Kashmir musk deer are confirmed instead of Alpine musk deer Moschus chrysogaster which has been previously described from the southern parts of Himalayas of Nepal, India and Pakistan.

Phylogenetic surveys on the newt genus Tylototriton sensu lato (Salamandridae, Caudata) reveal cryptic diversity and novel diversification promoted by historical climatic shifts.

Global climatic transitions and Tibetan Plateau uplifts are hypothesized to have profoundly impacted biodiversity in southeastern Asia. To further test the hypotheses related to the impacts of these incidents, we investigated the diversification patterns of the newt genus Tylototriton sensu lato, distributed across the mountain ranges of southeastern Asia. Gene-tree and species-tree analyses of two mitochondrial genes and two nuclear genes revealed five major clades in the genus, and suggested several cryptic species. Dating estimates suggested that the genus originated in the early-to-middle Miocene. Under different species delimitating scenarios, diversification analyses with birth-death likelihood tests indicated that the genus held a higher diversification rate in the late Miocene-to-Pliocene era than that in the Pleistocene. Ancestral area reconstructions indicated that the genus originated from the northern Indochina Peninsula. Accordingly, we hypothesized that the Miocene Climatic Transition triggered the diversification of the genus, and the reinforcement of East Asian monsoons associated with the stepwise uplifts of the Tibetan Plateau promoted the radiation of the genus in southeastern Asia during the Miocene-to-Pliocene period. Quaternary glacial cycles likely had limited effects on speciation events in the genus, but mainly had contributions on their intraspecific differentiations.

A new species of the genus Microhyla (Anura: Microhylidae) from Eastern Nepal.

A new species of the genus Microhyla is described from Jamun Khadi, Jhapa district of eastern Nepal, based on molecular and morphological comparisons. This species is the sister taxon of Microhyla ornata and can be distinguished by a unique vocalization, morphology and molecular phylogeny. The uncorrected genetic divergences based on rRNA gene between the new species and its closest congeners, M. nilphamariensis, M. ornata and M. rubra were 5.34%, 6.67%, and 8.31%, respectively. The new species, Microhyla taraiensis sp. nov., is distinguished from each other of Microhyla by a combination of the following morphological characters: (1) relatively larger body size (SVL ranges 19.9-20.3 mm, n = 4 in the males and 22.1-24.9 mm, n = 3 in the females); (2) dorsal surface of head and body with light red dots; (3) toes webbing poorly developed or absent; (4) a large round inner metacarpal tubercle; and an (5) elongated outer metacarpal tubercle. In addition, our study also provides a new record of Microhyla nilphamariensis from Nepal.