Lactic acid fermentation by Lactobacillus casei in free cell form and immobilised on gluten pellets

A comparative study of the fermentation of a range of carbohydrate substrates, at various temperatures, was carried out using a commercial Lactobacillus casei strain in a free cell form and immobilised on gluten pellets. This strain required yeast extract, l-cysteine HCl and Mn²⁺ at 5, 0.5 and 0.1 g l^–1, respectively, for maximum growth and lactic acid production. Sugar fermentation using free cells showed preference in the order glucose, sucrose, fructose while lactose was poorly utilised. Optimum temperature for growth and lactic acid production over (18–30 h) was 43 °C. L. casei was successfully immobilised on gluten pellets and fermented glucose and sucrose in a shorter time (18 h) with increased lactic acid production (42 and 41 g l^–1 on glucose and sucrose, respectively).     

Bacterial diversity and community structure during fermentation of Chinese sauerkraut with Lactobacillus casei 11MZ‐5‐1 by Illumina Miseq sequencing

The bacterial diversity and community structure involved in Chinese sauerkraut is one of the most important factors shaping the final characteristics of traditional foods. In this research, Lactobacillus casei 11MZ‐5‐1 was applied in Chinese sauerkraut fermentation as a starter culture. Illumina Miseq sequencing analysis was used to reveal the bacterial diversity and community structure during Chinese sauerkraut fermentation. A total of 177 283 high‐quality reads of 16S rRNA V4 regions were obtained. The inoculation of L. casei 11MZ‐5‐1 decreased considerably the bacterial richness and bacterial diversity. This inoculum led to the replacement of Lactococcus by Lactobacillus. The levels of Pseudomonas and Enterobacter bacteria decreased. These findings reveal the evolution of important bacterial groups that are involved in fermentation and will facilitate improvements in the Chinese sauerkraut fermentation process.

Significance and Impact of the Study

This research thoroughly revealed the effects of Lactobacillus casei 11MZ‐5‐1 starter cultures on bacterial communities during Chinese sauerkraut fermentation. Illumina Miseq sequencing was effective technique to monitor the bacterial diversity and community structure. The inoculation of L. casei 11MZ‐5‐1 led to the decline of bacterial richness and diversity together with a consistent predominance of Lactobacillus during spontaneous fermentation. The result collectively suggested L. casei 11MZ‐5‐1 is a promising starter in Chinese sauerkraut manufacturing.     

Exploring the application of biostimulation strategy for bacteria in the bioremediation of industrial effluent

The purpose of this study was to isolate and characterise toxic element-resistant bacteria from acid mine drainage water and to apply them in the bioremediation of industrial effluent, as well as to identify optimal effluent:nutrient concentration for onsite biostimulation strategy. Wastewater samples were collected from acid mine drainage and industry. Inductively coupled plasma optical emission spectroscopy (ICP-OES) was employed for elemental composition analysis. Isolated bacterial strains were characterised using molecular methods. Bioremediation assays were employed to determine the extent of bacterial tolerance and removal of toxic elements using a biostimulation strategy employing minimal salt medium (MSM) at varied concentrations and positive and negative controls of only MSM and industrial effluent, respectively. Two bacterial strains demonstrated resistance to toxic elements, Bacillus sp. MGI101 and Lysinibacillus sp. MGI102 both isolated from the AMD sites. However, no observable growth of toxic metal-resistant bacteria was obtained from the industrial effluents. Bacterial strains MGI101 and MGI102 demonstrated high resistance to target toxic elements during the screening and tolerance assays. Remarkably, Bacillus sp. MGI101 demonstrated greater ability to remove toxic elements including arsenic, chromium, zinc, copper and aluminium in undiluted solutions of the industrial effluent, with its highest removal capacity observed at > 60% for arsenic and aluminium. Both Bacillus sp.MGI101 and Lysinibacillus sp. MGI102 demonstrated varied abilities for the removal of toxic elements from dilution concentration of effluent mixed with MSM. However, the optimal dilution ratio observed in this experiment was 5:15 (effluent:MSM). Overall results demonstrated that isolated bacterial strains have the potential to be employed in bioremediation programmes of acid mine drainages and multi-element contaminated water.

     

Studies on cephalosporin-C production in an air lift reactor using different growth modes of Cephalosporium acremonium

This paper deals with the studies on Cephalosporin-C production in a lab-scale airlift reactor using Cephalosporium acremonium. Various growth modes, viz. pellets and Siran supported bioparticles were used to improve the process over conventional free mycelial fermentation. Cephalosporin-C production was significantly improved by using bioparticles over the free mycelial culture, probably due to the enhanced mass transfer in the fermentation broth. However, the biofilm of the bioparticles became unstable as the fermentation proceeded, and increase in the free cells in the broth occurs. The maximum specific growth rate of free cells, pellets and Siran carrier were observed to be 0·037, 0·033 and 0·045 h⁻¹, respectively. The oxygen transfer coefficient also improved for the immobilised modes (100 h⁻¹, 70 h⁻¹ for Siran carrier and pellets) and thereby enhanced specific antibiotic productivity, 18–28% were observed.     

Influences of yeast extract on specific cellular yield of Ovine growth hormone during fed-batch fermentation of E. coli

In most cases of E. coli high cell density fermentation process, maximizing cell concentration helps in increasing the volumetric productivity of recombinant proteins usually at the cost of lower specific cellular protein yield. In this report, we describe a process for maintaining the specific cellular yield of Ovine growth hormone (oGH) from E. coli by optimal feeding of yeast extract during high cell density fermentation process. Recombinant oGH was produced as inclusion bodies in Escherichia coli. Specific cellular yield of recombinant oGH was maintained by feeding yeast extract along with glucose during fed-batch fermentation. Glucose to yeast extract ratio of 0.75 was found to be optimum for maintaining the specific cellular oGH yield of 66 mg/g of E. coli cells. Continuous feeding of yeast extract along with glucose helped in reducing acetic acid secretion and promoted higher cell growth during fed-batch fermentation. High cell growth of E. coli and high specific yield of recombinant oGH thus helped in achieving high volumetric productivity of the expressed protein. A maximum of 2 g/l of ovine growth hormone was expressed as inclusion bodies in 12 h of fed-batch fermentation.     

Batch fermentation with entrapped growing cells ofLactobacillus casei

Summary Growing cells ofLactobacillus casei were entrapped in-carrageenan/locust bean gum (LBG) (2:1 or 2.75%:0.25% w/w respectively) mixed gel beads (two ranges of diameter: 0.5–1.0 and 1.0–2.0 mm) to fermentLactobacillus Selection (LBS) medium and produce biomass. The results showed significant influence of initial cell loading of the beads and bead size on the fermentation rate. The highest cell release rates were obtained with 2.75%:0.25%-carrageenan/LBG small diameter gel beads. However, 17 h fermentation of LBS medium with immobilized cells resulted in substantial softening of the gel matrix, prohibiting reuse of immobilized biocatalysts as inoculum in subsequent batch fermentation. A dynamic shear rheological study showed that the gel weakness was related to chemical interactions with the medium. Results indicated that part of the matrix-stabilizing K⁺ ions diffused back to the medium. Stabilization of the gel was obtained by adding potassium ions to the LBS medium;L. casei growth was not altered by this supplementation. Fermentation of LBS medium supplemented with KCl byL. casei showed higher cell counts in the broth medium with immobilized cells than with free cells, reaching 10¹⁰ cells/ml after about 10 h with entrapped cells in 0.5–1.0 mm diameter beads and 17 h with free cells. Counts in the gel beads after fermentation were higher than 10¹¹ cells/ml and bead integrity was maintained throughout fermentation.     

The effects of light intensity, inoculum size, and cell immobilisation on the treatment of sago effluent withRhodopseudomonas palustris strain B1

A study was carried out to determine a suitable light intensity and inoculum size for the growth ofRhodopseudomonas palustris strain B1. The pollution reduction of sago effluent using free and immobilisedR. palustris cells was also evaluated. The growth rate in glutamatemalate medium was highest at 4 klux compared to 2.5 and 3 klux. The optimal inoculum size was 10% (v/v). Both the COD and BOD of the sago effluent were reduced by 67% after three days of treatment. The difference in biomass production or BOD and COD removal with higher inoculum sizes of 15 and 20% was minimal. This could be attributed to limited nutrient availabillity in the substrate. The use of immobilised cells ofR. palustris reduced the pollution load 10% less compared to pollution reduction by free cells. Hence, there was no significant difference in using free or immobilised cells for the treatment of sago effluent.     

Comparative Study of Cr(VI) Removal by Exiguobacterium sp. in Free and Immobilized Forms

Cr(VI) is a toxic environmental pollutant. To determine the potential role of microbes towards chromate bioremediation, two bacterial strains, E1 and E4, that could tolerate Cr(VI) at levels up to 2250 μg ml^?1 were isolated from the soil of a tannery. They were identified as Exiguobacterium sp. To estimate the removal of Cr(VI) using immobilized bacterial cells, 2% sodium alginate and 2.5% agar were used as immobilizing matrices. In the case of sodium alginate, 89% and 93% of Cr(VI) removal by E1 and E4, respectively, were observed. When agar beads were used as an immobilizing matrix, removal was recorded as 39% and 48% for E1 and E4, respectively. Removal of Cr(VI) was also estimated in sterile and nonsterile tannery effluent. More Cr(VI) removal was noted in the nonsterile effluent than in the sterile effluent. The maximum uptake of Cr(VI) of bound cells of E1 and E4 was found to be 17.54 and 20.04 μg ml^?1, respectively. Fourier transform infrared (FTIR) spectra of cells of E4 with Cr(VI), without Cr(VI), and immobilized cells depicted several absorption peaks, mainly for P?OH group, C?H bending, C?O bond, and amide II groups, reflecting the complex nature of the bacterial cells and the contribution of these functional groups to the Cr(VI) binding process.     

Isolation and Growth Characteristics of Chromium(VI) and Pentachlorophenol Tolerant Bacterial Isolate from Treated Tannery Effluent for its Possible Use in Simultaneous Bioremediation

The bacterial strains resistant to pentachlorophenol (PCP) and hexavalent chromium [Cr(VI)] were isolated from treated tannery effluent of a common effluent treatment plant. Most of the physico-chemical parameters analyzed were above permissible limits. Thirty-eight and four bacterial isolates, respectively were found resistant to >50 μg/ml concentration of [Cr(VI)] and the same level of PCP. Out of the above 42 isolates, only one was found simultaneously tolerant to higher levels of both PCP (500 μg/ml) and Cr(VI) (200 μg/ml), and hence was selected for further studies. To the best of our knowledge, this is the first report in which a native bacterial isolate simultaneously tolerant to such a high concentrations of Cr(VI) and PCP has been reported. The culture growth was best at 0.4% (w/v) glucose as an additional carbon source and 0.2% (w/v) ammonium chloride as a nitrogen source. The growth results with cow urine as a nitrogen source were comparable with the best nitrogen source ammonium chloride. The isolate exhibited resistance to multiple heavy metals (Pb, As, Hg, Zn, Co & Ni) and to antibiotics nalidixic acid and polymixin-B. The efficacy of bacterial isolate for growth, PCP degradation (56.5%) and Cr(VI) bioremediation (74.5%) was best at 48 h incubation. The isolate was identified as Bacillus sp. by morphological and biochemical tests. The 16S rDNA sequence analysis revealed 98% homology with Bacillus cereus. However, further molecular analysis is underway to ascertain its likelyhood of a novel species.     

Extracellular secretion of levansucrase from Zymomonas mobilis in Escherichia coli

Secretion of levansucrase from Zymomonas mobilis in Escherichiacoli by glycine supplement was investigated. A significant amount of levansucrase (about 25% of total activity) was found in intact whole-cells. Cell fractionation experiments showed that levansucrase was found both in the periplasmic space and in the cytoplasmic fraction of E. coli. None or only trace amounts of levansucrase was detected in the extracellular culture broth at 24 h of cultivation and it accrued with the increasing concentration of glycine in the culture medium and duration of the culture period. Optimal glycine concentration for the maximum secretion of levansucrase was in the range of 0.8-1%, in which approximately 20-50% of levansucrase was released into the extracellular fraction at 24 h of cultivation, although glycine retarded the bacterial growth.     

Reduction of Toxic Chromium and Partial Localization of Chromium Reductase Activity in Bacterial Isolate DM1

Summary With advances in biotechnology, bioremediation has become one of the most rapidly developing fields in environmental restoration, utilizing microorganism to reduce the concentration and toxicity of heavy metals. Hexavalent chromium reducing bacterial culture (DM1) was isolated from the contaminated sites of chemical industries and its ability to reduce hexavalent chromium to trivalent chromium, a detoxification process in cell suspension and cell extract was examined. Based on the biochemical analysis DM1 was identified as Ochrobactrum sp. It could tolerate chromium upto a maximum concentration of 300 ppm, optimum temperature and pH being 35 °C and 7 respectively for maximum chromium reduction. Assay with the permeabilized cells (treated with toluene and Triton X-100) and cell free extract demonstrated that the hexavalent chromium reduction is mainly associated with the soluble fraction of the cell. The chromium reducing activity is inducible. The presence of an induced protein having molecular weight around 30 kDa in the presence of chromium and absence in cells without chromium points out a possible role of this protein in chromium reduction. The bacterial isolate DM1 can be exploited for bioremediation of hexavalent chromium containing wastes, since it seems to have the potential to reduce the toxic hexavalent form of chromium to its nontoxic trivalent form.     

Colour removal from bagasse-based pulp mill effluent using a white rot fungus

Colour removal of pulp plant effluent was studied using white rot fungus, Trametes (Coriolus) versicolor. The batch experiments were carried out using fungus in the form of mycelial pellets. In the present investigation, the effect of pH, concentrations of glucose (substrate), initial effluent colour and ammonium chloride (nutrient) on colour removal efficiency were studied. It was found that the maximum colour removal efficiency of 82.5% was obtained with an optimal glucose and ammonium chloride concentrations of 15 g/l and 0.5 g/l respectively at a pH of 4.5 without diluting the effluent.     

Laboratory scale equipment for the determination of k L a in bio-reactors

The study of the oxygen transfer rate (O.T.R) in reactors designed for cell cultivation and enzyme reaction is a difficult task. In this work a bio-reactor for acetic acid fermentation purposes is studied by using the static gassing out method for K_La evaluation. Results obtained prove that K_La shows linear dependence versus operation temperature.     

Remediation of copper-contaminated soil by Kocuria flava CR1, based on microbially induced calcite precipitation

An indigenous calcifying bacterial strain CR1, identified as Kocuria flava, was isolated from soil of a mining area, Urumqi, China. An extensive copper bioremediation capacity of this isolate was studied based on microbially induced calcite precipitation (MICP). K. flava CR1 removed 97% of copper when initial Cu concentration was 1000 mg L⁻¹. The isolate produced significant amount of urease (472 U mL⁻¹), an enzyme that leads to calcite precipitation. The isolate removed 95% of copper from contaminated soil. The MICP process in bioremediation was further confirmed by FTIR and XRD analyses. FTIR analysis showed two different forms of calcium carbonate, i.e., calcite and aragonite, and the results were well supported by XRD. For the first time, the ability of K. flava has been documented in the bioremediation of polluted soil. This study showed that MICP-based bioremediation by K. flava is a viable, environmental friendly technology for cleaning-up the copper-contaminated site.     

Verticillium lecanii spore production in solid-state and liquid-state fermentations

Verticillium lecanii has been recognized as an entomopathogen with high potential in biological control of pests. Two types of cultivation methods, the solid-state fermentation (SSF) and the liquid-state fermentation (LSF), were examined for V. lecanii. In SSF, the substrate types including rice, rice bran, rice husk, and the mixtures of these components were tested. The results showed that both cooked rice with appropriate water addition and rice bran gave significantly higher spore production of 1.5 2 10⁹ spores/g substrate and 1.4 2 10⁹ spores/g substrate, respectively. In LSF, SMAY liquid medium was used as a base, and the effects of environmental conditions on the spore production of V. lecanii were investigated. From the time course study, on the 9th day the spore yield reached 1.2 2 10⁹ spores/ml of broth at 24v°C, 150 rpm for this strain. A series of medium volumes in the shaker-flask have been tested for the requirement of aeration. The largest surface aeration test, one tenth of the medium volume in the shaker-flask for cultivation, gave the highest spore count. The optimal pH value was tested and the initial pH 5 in the SMAY medium produced a high spore density. Finally, V. lecanii spores from SSF and LSF were different in size, shape, and size distribution; while mean spore length from SSF was 6.1 7m, and mean spore length from LSF was 5.0 7m.     

Lactic acid production from whey permeate by immobilized Lactobacillus casei

Two matrices have been assessed for their ability to immobilize Lactobacillus casei cells for lactic acid fermentation in whey permeate medium. Agar at 2% concentration was found to be a better gel than polyacrylamide in its effectiveness to entrap the bacterial cells to carry out batch fermentation up to three repeat runs. Of the various physiological parameters studied, temperature and pH were observed to have no significant influence on the fermentation ability of the immobilized organism. A temperature range of 40–50°C and a pH range of 4.5–6.0 rather than specific values, were found to be optimum when fermentation was carried out under stationary conditions. In batch fermentation ～90% conversion of the substrate (lactose) was achieved in 48 h using immobilized cell gel cubes of 4 × 2 × 2 mm size, containing 400 mg dry bacterial cells per flask and 4.5% w/v (initial) whey lactose content as substrate. However, further increase in substrate levels tested (>4.5% w/v) did not improve the process efficiency. Supplementation of Mg²⁺ (1 mM) and agricultural by-products (mustard oil cake, 6%) in the whey permeate medium further improved the acid production ability of the immobilized cells under study.     

Applicability of pectate-entrapped <Emphasis Type="Italic">Lactobacillus casei</Emphasis> cells for <Emphasis Type="SmallCaps">l</Emphasis>(+) lactic acid production from whey

Lactic acid is a versatile organic acid, which finds major application in the food, pharmaceuticals, and chemical industries. Microbial fermentation has the advantage that by choosing a strain of lactic acid bacteria producing only one of the isomers, an optically pure product can be obtained. The production of l(+) lactic acid is of significant importance from nutritional viewpoint and finds greater use in food industry. In view of economic significance of immobilization technology over the free-cell system, immobilized preparation of Lactobacillus casei was employed in the present investigation to produce l(+) lactic acid from whey medium. The process conditions for the immobilization of this bacterium using calcium pectate gel were optimized, and the developed cell system was found stable during whey fermentation to lactic acid. A high lactose conversion (94.37%) to lactic acid (32.95 g/l) was achieved with the developed immobilized system. The long-term viability of the pectate-entrapped bacterial cells was tested by reusing the immobilized bacterial biomass, and the entrapped bacterial cells showed no decrease in lactose conversion to lactic acid up to 16 batches, which proved its high stability and potential for commercial application.     

p LA-PEDV-S1质粒拷贝数与发酵重组乳酸菌S1表达量之间的相关性研究

探究质粒拷贝数以及目标蛋白S1表达量与发酵时间的关系,从而确定放大生产p LA-PEDV-S1/Lactobacillus casei的最佳发酵时间。通过发酵重组干酪乳杆菌,绘制重组干酪乳杆菌的生长曲线,确定其生长的最佳时期。将p LA-PEDV-S1/L. casei分别接种至添加抗生素和不添加抗生素的MRS培养基中传代培养,进行稳定性实验。使用荧光定量PCR方法检测重组干酪乳杆菌中质粒的拷贝数,使用流式细胞术检测乳酸菌表达的目标蛋白。重组菌在7 h达到生长顶点,传至120代时外源质粒并无丢失情况出现。质粒拷贝数在9 h达到峰值29. 34,表达目标蛋白的重组菌在7 h达到峰值97. 98%。结果显示在细菌生长的对数生长期末期,质粒拷贝数最高且S1表达量最多;在平台期随着发酵时间的增加,质粒拷贝数逐渐降低,S1表达量也相应减少。发酵的最佳时间为7～10 h,质粒拷贝数与S1表达量之间存在着正相关性。     

Cold tolerance of Pseudomonas sp. 30-3 isolated from oil-contaminated soil, Antarctica

Pseudomonas sp. 30-3 was enriched from oil-contaminated soil from Wright Valley, Antarctica using JP8 jet fuel as sole carbon source. This isolate exhibited tolerance to temperatures ranging from 0°C to 35°C when cultured in laboratory medium. In a freeze-thaw study, an 89% survival was observed when Pseudomonas sp. 30-3 was exposed to 4°C prior to freezing. PCR amplification of a 248-bp DNA fragment in Pseudomonas sp. 30-3 using capB-gene specific primers showed a 98% amino acid sequence homology with CapB of Pseudomonas fragi and 62% homology with CspA of Escherichia coli. Radiolabeling of total cellular proteins exhibited elevated expression of an 8-kDa protein at 4°C, which suggests that the CapB in Pseudomonas sp. 30-3 may play a pivotal role in survival and tolerance at cold and subzero temperatures. Tolerance to cold temperatures and the ability to degrade hydrocarbons by Pseudomonas sp. 30-3 provide support for the application of bioremediation for petroleum hydrocarbons in Antarctic soils.     

Effect of environmental parameters on decolorization of textile wastewater using Phanerochaete chrysosporium

Effect of various parameters such as size of inoculum, temperature, carbon source on decolorization of textile wastewater by Phanerochaete chrysosporium was investigated. Textile wastewater decolorization occurred during the primary phase of growth and secondary metabolism in carbon and nitrogen limited medium, respectively. It was found that glucose concentration up to 0.3 g/l has considerable effect on decolorisation rate. Further, it was also found that the concentration of the organic nitrogen of the effluent stream was sufficient to furnish the decolorisation process. It was observed that the inoculum size in this case within 10% increased the decolorisation rate rapidly. It was found that the temperature rise from 20 to 38 °C enhanced the rate of decolorization. The optimum temperature for decolorisation was found to be about 35 °C. Effect of pH from 2-4 on decolorization was also investigated. It is concluded that using Phanerochaete chrysosporium, decolorization of the azo dye containing effluent of the textile industry was achieved to about 96% within 28 h of operation.