3 Biotech

The objective of this study was to isolate the lactic acid bacteria from fermented silage sample and analyze their antibacterial activities, probiotic properties, and fermentation potential in silage. Eleven lactic acid bacteria (LAB) were selected based on distinct morphologies and preliminary studies. Cell-free supernatant (CFS) was then prepared from the selected strains for antibacterial analysis. L-30 strain and its CFS showed highest inhibition (> 10 mm) against tested foodborne pathogens as compared to other strains. Hereafter, the strain L-30 was named as KCC-30 and used for further studies. KCC-30 can survive in the harsh conditions of GIT such as low pH ( 2) and bile salt environment (oxgal) than standard L. plantarum KACC-91016 (pH 2: 27.2% vs 20.5%; oxgal: 72.3% vs 57.7%, both p < 0.05). In addition, KCC-30 exhibited strong auto-aggregation (68.3% vs 51.5%) and co-aggregation (33% vs 23.9%) properties. For silage experiment, KCC-30 treatment did not alter the nutrient profiles of silage. At the same time, KCC-30 treatment increased the lactic acid content of silage as compared to untreated silage (5.55 DM% vs 3.11 DM%). An increase of lactic acid content in the silage is due to higher lactic acid bacteria population in KCC-30 treated silage (15.33 × 107 CFU/g vs 7.66 × 107 CFU/g) than untreated silage (p < 0.05). Overall data suggested that KCC-30 exhibited strong probiotic potential and improved the quality of Lolium multiflorum silage by increasing the lactic acid level. Therefore, KCC-30 could be considered as potential strain to improve the fermentation quality of L. multiflorum silage.

The accumulation of selenium (Se) by Lactobacillus delbrueckii ssp. bulgaricus (Lb) and Streptococcus thermophilus (St) at the different cultivation conditions, including initial pH, inoculum dose (%), and temperature (°C), was investigated in this work. Se enrichment efficiency was optimized using the Design-Expert software for response surface methodology on a basis of single-factor experiment. The antioxidant activities of Se-enriched Lactic acid bacteria (LAB) were also investigated. The qualitative analysis of Se-enriched LAB was performed by FT-IR spectra. The cell morphology and chemical element components were measured by a scanning electron microscope (SEM) equipped with energy-dispersive X-ray spectroscopy. The results indicated that the optimum initial pH, inoculum doses, and temperatures of Lb and St were 5.96, 6.73%, 33.24 °C, and 6.37, 6%, 40 °C, respectively. Under the optimal conditions, the ratios of Se enrichment reached 94.34% for Lb and 97.05% for St. Furthermore, Se-enriched LAB enhanced scavenging rates on DPPH, ABTS free radical, and also heightened reducing activity. The FT-IR results showed that the two Se-enriched strains had similar characteristic absorption peaks, which were further demonstrated that both Se biomasses had the same carbonyl, carboxyl, and hydroxyl groups. Elemental selenium nanoparticles were verified around cell surfaces of Se-enriched LAB, which implied that both strains had detoxification ability when grown in liquid media containing selenite.

This study was aimed at developing an easy to use and inexpensive biosensor for the detection of typhoidal Salmonella. The technique was designed to be used without expensive instrumentation if necessary. Bacteriophages specifically infecting three typhoidal Salmonella serovars were isolated and purified. Log-phase cultures were mixed with a high titre of a single phage (109 PFUs) in separate wells of a microtitre plate and incubated at room temperature (30 °C) for 1 h. After incubation, resazurin was added and the plates were incubated further for 1 h. Absorbance at 570 nm of each test well was measured using a commercial microplate reader and compared with that of the control well. A significant difference (p < 0.05) between the absorbance of test and control wells indicated the presence of target bacteria. With visual inspection, a delay in colour change from blue to pink was considered a positive result. The system could detect 5 × 104 CFUs in 120 min without pre-enrichment and 10 CFUs with a pre-enrichment of 6 h.

A cellulase encoding gene, Cel PRII, was identified from Mehsani buffalo rumen metagenome, and cloned and expressed in Escherichia coli BL21(DE3)pLysS. The 1170 bp full length gene encodes a 389 residue polypeptide (Cel PRII) containing a catalytic domain belonging to glycosyl hydrolase (GH) 5 family. The fusion protein consisting of the Cel PRII, thioredoxin tag and 6x Histidine tag with predicted molecular weight of 63 kDa when recovered from inclusion bodies under denaturing conditions, exhibited cellulolytic activity against carboxymethyl cellulose (CMC). Recombinant Cel PRII was stable in the pH range 4.0–10.0 with pH optima 6.0. The optimal reaction temperature of Cel PRII was 30 °C with more than 50% of its activity retained at the temperatures ranging from 0 to 50 °C. Cel PRII exhibited enhanced enzymatic activity in the presence of Mn2+ ions and was inhibited in the presence of chelating agent EDTA. The K m and V max values for CMC were found to be 166 mg/mL and 1292 IU/mg, respectively. Cel PRII identified in the present study may act as an excellent candidate for industrial applications, and may aid in lignocellulosic biomass conversion because of its potential cellulolytic activity, thermostability, and excellent pH stability.

Plant–parasitic root-knot nematode Meloidogyne incognita uses an array of effector proteins to establish successful plant infections. Mi-msp-1 and Mi-msp-20 are two known effectors secreted from nematode subventral oesophageal glands; Mi-msp-1 being a putative secretory venom allergen AG5-like protein, whereas Mi-msp-20 is a pioneer gene with a coiled-coil motif. Expression of specific effector is known to cause disturbances in the expression of other effectors. Here, we used RNA-Seq to investigate the pleiotropic effects of silencing Mi-msp-1 and Mi-msp-20. A total of 25.1–51.9 million HQ reads generated from Mi-msp-1 and Mi-msp-20 silenced second-stage juveniles (J2s) along with freshly hatched J2s were mapped to an already annotated M. incognita proteome to understand the impact on various nematode pathways. As compared to control, silencing of Mi-msp-1 caused differential expression of 29 transcripts, while Mi-msp-20 silencing resulted in differential expression of a broader set of 409 transcripts. In the Mi-msp-1 silenced J2s, cytoplasm (GO:0005737) was the most enriched gene ontology (GO) term, whereas in the Mi-msp-20 silenced worms, embryo development (GO:0009792), reproduction (GO:0000003) and nematode larval development (GO:0002119) were the most enriched terms. Limited crosstalk was observed between these two effectors as a sheer 5.9% of the up-regulated transcripts were common between Mi-msp-1 and Mi-msp-20 silenced nematodes. Our results suggest that in addition to the direct knock-down caused by silencing of Mi-msp-1 and Mi-msp-20, the cascading effect on other genes might also be contributing to a reduction in nematode's parasitic abilities.

The potassium transporter high-affinity K+ transporter/K+ uptake permease/K+ transporter (HAK/KUP/KT) family plays a vital role in potassium uptake, and potassium ion (K+)-mediated environmental stress. In the present study, we identified 22 IbHAK/KUP/KT (HAK) genes in sweet potato [Ipomoea batata (L.) Lam] and the same number of HAK genes from sweet potato wild relative Ipomoea trifida. Phylogeny analysis indicated that the HAKs can be divided into five clades. Chromosomal distribution and genome synteny analyses revealed two tandem-duplicated gene pairs IbHAK16/17 and IbHAK17/18 on chromosomes 13 and eight segmental-duplicated gene pairs on chromosomes 1, 3, 5, 8, 10, 12, 14 among the IbHAK gene family. Eleven orthologous HAK gene pairs between I. batata and I. trifida were involved in the duplication of genomic blocks based on comparative genomic analysis. The Ka/Ks ratios of these IbHAK genes ranged from 0.02 to 0.55(< 1), further indicated that purifying selection was the primary force driving the evolution of HAKs in Ipomoea. A heat map based on RNA-seq data showed that 13 HAKs in Xushu32 (a K+-tolerant sweet potato genotype) and 10 HAKs in Ningzi1 (a K+-sensitive sweet potato genotype) in response to K+ deficiency stress. Quantitative real-time PCR (qRT-PCR) analysis revealed IbHAK2, -3, -8, -10, -11, -18, -19, and -21 were induced in both Xushu32 and Ningzi1 under low K+ stress. Compared with other IbHAK genes, IbHAK8 showed more strongly upregulation after exposure to drought and salt stress. Furthermore, co-expression analysis showed that only IbHAK8 of 22 IbHAK genes involved in network interactions with 30 genes related to abiotic and biotic stresses. Taken together, these results are helpful for further functional studies on IbHAK and molecular breeding of sweet potato.

Ba mươi bốn chủng Xanthomonas citri pv. malvacearum (Xcm) được thu thập từ ba khu vực trồng bông ở Ấn Độ đã được nghiên cứu về độ gây hại và tài liệu chủng loại, và sau đó được so sánh với sự đa dạng di truyền như được tiết lộ bởi các yếu tố lặp lại [palindromic ngoại gen lặp lại (REP), đồng thuận lặp lại giữa các gen vi khuẩn đường ruột (ERIC) và yếu tố BOX] cũng như phân tích PCR của các trình tự lặp lại giữa đơn giản (ISSR). Trong số 34 chủng được thử nghiệm về độ gây hại trên giống dễ bị tổn thương LRA 5166, có 7 chủng được ghi nhận là có độ gây hại cao (HV), 16 chủng có độ gây hại trung bình (MV) và 11 chủng có độ gây hại thấp (LV). Tám chủng khác nhau đã được ghi nhận bằng cách sử dụng mười dòng giống bông khác nhau. Hai mươi hai chủng (65%) thuộc về chủng 18. Mười hai chủng (35%) thuộc về các chủng 3, 5, 6, 7, 8, 11 và 13. Phân tích REP, ERIC, BOX, các yếu tố lặp lại kết hợp và ISSR cho thấy sự hiện diện của 7, 10, 9, 11 và 8 cụm, tương ứng, với hệ số tương đồng 0.70 trong các biểu đồ. Phân tích tọa độ chính (PCoA) cho thấy sự biến thiên tích lũy 76.4% và 77.5% cho các yếu tố lặp lại kết hợp và phân tích ISSR. ERIC tạo ra giá trị thông tin đa hình (PIC) cao nhất (0.928). Có rất nhiều biến động nội bộ trong chủng bệnh về độ gây hại và dấu vân tay gen giữa các chủng Xcm. Nhiều chủng được nhóm lại dựa trên nguồn gốc địa lý mà không phụ thuộc vào độ gây hại hay chủng loại. Sự lây lan của các chủng bệnh ở Ấn Độ có thể là do việc vận chuyển các dòng giống cây trồng và vật tư hạt giống từ nơi này sang nơi khác.

The present study investigated the effect of arylsulfonyl indoline-benzamide (ASIB) on neovascular glaucoma in the mice model in vivo. In the mice model of glaucoma, ASIB treatment significantly (P < 0.05) increased PDGF-B-positive cell count in the corneal tissues. ASIB treatment at 5, 10, 15 and 20 mg/kg doses raised the level of PDGF-B mRNA in the mice cornea by 2.3-, 3.8-, 5.4- and 5.5-fold, respectively. Pre-treatment of the glaucoma mice with ASIB leads to inhibition of TNF-α and IL-6 production. In the glaucoma mice, treatment with ASIB leads to a marked decrease in the level of NOD2 mRNA and protein. ASIB treatment caused a significant decrease in the glaucoma-induced up-regulation of NF-κB p65 activation. The phosphorylation of NF-κB p65 was almost completely inhibited in the glaucoma mice on treatment with 15 mg/kg dose of ASIB. ASIB exhibited inhibitory effect on glaucoma-induced inflammatory cytokine and oxidative factor damage in the mice. It caused up-regulation of PDGF expression and down-regulated NF-κB activation. Therefore, ASIB can be of therapeutic significance for neovascular glaucoma treatment. However, more studies need to be performed to fully understand the molecular mechanism of ASIB in glaucoma treatment.

Plant growth-promoting fungi play an important role in development of sustainable agriculture. In the current study, 13 fungal strains were isolated from the rhizosphere of healthy Triticum aestivum (wheat) plant and screened for their indolic auxin production potential. Aspergillus flavus strain PGFW, Aspergillus niger strain BFW and Aspergillus caespitosus strain DGFW were amongst the most efficient indolic auxin-producing strains. Indolic auxins such as indole 3 acetate (IAA), indole 3 butyrate (IBA) and indole 3 propionate (IPA) are produced by fungi. The conventional method to assess the IAA production is through a spectrophotometric assay using Salkowski’s reagent, which quantifies all indolic auxins and not individual auxins. Moreover, it was also observed that the absorption maxima (λmax) of the samples, when compared to that of standard indole-3-acetic acid, showed deviation from the latter, indicative of production of a mixture of indolic derivatives by the fungi. Hence, for further profiling of these indolic compounds, high-performance thin layer chromatography (HPTLC) based protocol was standardized to precisely detect and quantify individual indolic auxins like IAA, IBA and IPA in the range of 100–1000 ng per spot. HPTLC analysis also showed that the fungal strains produce different indolic auxins in media with and without fortification of tryptophan, with the production of indolic auxins being enhanced in presence of tryptophan. Thus, this standardized HPTLC protocol is an efficient and sensitive methodology to separate and quantify the indolic derivatives.