Strain B1 Research Articles

Differential Expression of Genes Related to Growth and Aflatoxin Synthesis in Aspergillus flavus When Inhibited by Bacillus velezensis Strain B2.

Aspergillus flavus is a saprophytic soil fungus that infects and contaminates seed crops with the highly carcinogenic aflatoxin, which brings health hazards to animals and humans. In this study, bacterial strains B1 and B2 isolated from the rhizosphere soil of camellia sinensis had significant antagonistic activities against A. flavus. Based on the phylogenetic analysis of 16SrDNA gene sequence, bacterial strains B1 and B2 were identified as Bacillus tequilensis and Bacillus velezensis, respectively. In addition, the transcriptome analysis showed that some genes related to A. flavus growth and aflatoxin synthesis were differential expressed and 16 genes in the aflatoxin synthesis gene cluster showed down-regulation trends when inhibited by Bacillus velezensis strain B2. We guessed that the Bacillus velezensis strain B2 may secrete some secondary metabolites, which regulate the related gene transcription of A. flavus to inhibit growth and aflatoxin production. In summary, this work provided the foundation for the more effective biocontrol of A. flavus infection and aflatoxin contamination by the determination of differential expression of genes related to growth and aflatoxin synthesis in A. flavus when inhibited by B. velezensis strain B2.

Reduction of Chromium (VI) by indigenous bacteria isolated from industrial effluents of Pakistan

The present study was aimed to isolate and examine the reduction potential of chromium resistant bacteria. For this purpose, 10 chromium resistant bacterial strains were isolated from three different samples, collected from effluents of industries. These isolated strains were designated as A, B1, B2, C, L1, L2, L3, M, I1, and I2. All strains were able to resist up to 3500 μg/ml of potassium chromate on Lagar. At optimum pH 7 and temperature 37 ℃, all isolated strains showed optimum reduction potential. The strain B1 and L3 showed 97% reduction potential in DE broth while B1 in L-broth and I2 in acetate minimal broth showed 98% and 97% reduction potential respectively. In the case of artificial sewage water, strain M and L2 showed 94% reduction potential. In sample-1 of domestic sewage water (sterile), B2 showed 94% while I2 (non-sterile) had maximum reduction potential. Sample-2 (A, C, L2, I1) from domestic sewage water (non-sterile) had 96% and A, C, L2, I1 (sterile) also showed a 96% maximum reduction potential.

Antagonist Effect of Bacillus Species against Alternaria Rot of Tomato

The PGPR possesses antifungal activity against a broad range of pathogens. In this research study, the LPs of Bacillus strain B1 and B2 against Alternaria alternata were assessed. In vitro and in Planta assay on tomato fruit was made to check the post-harvest quality of fruits. The lipopeptides produced by both strains suppress the growth of the fungal pathogen Alternaria alternata and lessen the disease severity of the Alternaria rot on tomato fruits. The result revealed the quality of the Bacillus-treated fruits was also maintained and satisfactory. The lipopeptides of bacillus strain B1 and B2 represent a new innovative method to lessen the use of fungicides and chemical approaches to reduce the post-harvest loss caused by Alternaria alternata.

Phylogenetic Tree 16S rRNA Gene of Acinetobacter soli Isolated from the Prepuce of Aceh Cattle

BACKGROUND: In the pre-seed area of healthy Aceh cattle, it is possible to be contaminated with pathogenic bacteria that can interfere with the reproductive system. This study is needed to identify these pathogenic bacteria using a molecular approach, in an effort to prevent infection. AIM: The aim of the present study was to construct phylogenetic tree relationships of Acinetobacter soli identified in the preputial area of Aceh cattle by molecular analysis using 16S rRNA gene sequencing. MATERIALS AND METHODS: A total of 75 preputial specimens were obtained from Indrapuri’s Breeding and Forages Center of Aceh Cattles, Indrapuri district, Banda Aceh, Indonesia. The samples were processed for culture using standard conventional methods. The extraction of genomic DNA and the amplification of the 16S rRNA gene were assayed using polymerase chain reaction. A phylogenetic tree was constructed using distance matrices using the neighbor-joining model of the molecular evolutionary genetic analysis software 6.1 software. RESULTS: The results showed that of 75 preputial swab samples, 18 (24%) were positive for A. soli isolates. There was a 100% sequence similarity to A. soli prototype strain B1 and a 99% similarity to Acinetobacter parvus prototype strain LUH4616, Acinetobacter baylyi strain B2, A. venetianus strain ATCC 31012, as well as a 99% similarity to Acinetobacter baumannii strain DSM 30007, the strain ATCC 19606, and the strain JCM 6841, respectively. We concluded that A. soli-positive presentation in the preparation of Aceh cattle has 100% sequence similarity of 16S rRNA with A. soli strain B1. CONCLUSIONS: The conclusion of this study is that, based on the construction of a phylogenetic tree, it shows that 24% of the bacterial isolate is related to A. soli. It is essential to conduct a regular survey for bacterial contamination and to increase worker awareness and education about hygiene standards.

Live-attenuated YF17D-vectored COVID-19 vaccine protects from lethal yellow fever virus infection in mouse and hamster models.

SummaryBackgroundThe live-attenuated yellow fever vaccine YF17D holds great promise as alternative viral vector vaccine platform, showcased by our previously presented potent severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccine candidate YF-S0. Besides protection from SARS-CoV-2, YF-S0 also induced strong yellow fever virus (YFV)-specific immunity, suggestive for full dual activity. A vaccine concomitantly protecting from SARS-CoV-2 and YFV would be of great benefit for those living in YFV-endemic areas with limited access to current SARS-CoV-2 vaccines. However, for broader applicability, pre-existing vector immunity should not impact the potency of such YF17D-vectored vaccines.MethodsThe immunogenicity and efficacy of YF-S0 against YFV and SARS-CoV-2 in the presence of strong pre-existing YFV immunity were evaluated in mouse and hamster challenge models.FindingsHere, we show that a single dose of YF-S0 is sufficient to induce strong humoral and cellular immunity against YFV as well as SARS-CoV-2 in mice and hamsters; resulting in full protection from vigorous YFV challenge in either model; in mice against lethal intracranial YF17D challenge, and in hamsters against viscerotropic infection and liver disease following challenge with highly pathogenic hamster-adapted YFV-Asibi strain. Importantly, strong pre-existing immunity against the YF17D vector did not interfere with subsequent YF-S0 vaccination in mice or hamsters; nor with protection conferred against SARS-CoV-2 strain B1.1.7 (Alpha variant) infection in hamsters.InterpretationOur findings warrant the development of YF-S0 as dual SARS-CoV-2 and YFV vaccine. Contrary to other viral vaccine platforms, use of YF17D does not suffer from pre-existing vector immunity.FundingStated in the acknowledgments.

Esterase TriH responsible for the hydrolysis of trifloxystrobin in Hyphomicrobium sp. B1

Strobilurin fungicides are widely used in rice and corn planting due to their broad-spectrum and fungal mitochondrial inhibitory activities. However, their environmental residues will cause damage to eukaryotes such as chlorella, Daphnia magna and zebrafish, etc. In this study, a bacterial strain B1 was isolated and identified as Hyphomicrobium sp., which could degrade trifloxystrobin, a representative of strobilurin fungicides. A novel esterase gene triH responsible for the trifloxystrobin detoxification and trifloxystrobin acid generation was cloned. TriH shared low identity with the reported alpha/beta-hydrolase superfamily esterases (only 26–28%). The Km and kcat values of TriH for trifloxystrobin were 2.35 ± 0.24 μM and 48.66 ± 6.43 s−1. Also, TriH could catalyze the de-esterification reaction of azoxystrobin, picoxystrobin, and kresoxim-methyl to generate the corresponding daughter acid and methanol. Strain B1 can effectively remediate the trifloxystrobin-polluted freshwater and protect chlorella, Daphnia magna and zebrafish from the harm of trifloxystrobin. This work offers a potential strain as well as an enzyme for the remediation of strobilurin fungicides-contaminated environments.

Calcium Determines Lactiplantibacillus plantarum Intraspecies Competitive Fitness.

ABSTRACTThe importance of individual nutrients for microbial strain robustness and coexistence in habitats containing different members of the same species is not well understood. To address this for Lactiplantibacillus plantarum in food fermentations, we performed comparative genomics and examined the nutritive requirements and competitive fitness for L. plantarum strains B1.1 and B1.3 isolated from a single sample of teff injera fermentation batter. Compared to B1.1 and other L. plantarum strains, B1.3 has a smaller genome, limited biosynthetic capacities, and large mobilome. Despite these differences, B1.3 was equally competitive with B1.1 in a suspension of teff flour. In commercially sourced, nutrient-replete MRS (cMRS) medium, strain B1.3 reached 3-fold-higher numbers than B1.1 within 2 days of passage. Because B1.3 growth and competitive fitness were poor in mMRS medium (here called mMRS), a modified MRS medium lacking beef extract, we used mMRS to identify nutrients needed for robust B1.3 growth. No improvement was observed when mMRS was supplemented with nucleotides, amino acids, vitamins, or monovalent metals. Remarkably, the addition of divalent metal salts increased the growth rate and cell yields of B1.3 in mMRS. Metal requirements were confirmed by inductively coupled plasma mass spectrometry, showing that total B1.3 intracellular metal concentrations were significantly (up to 2.7-fold) reduced compared to B1.1. Supplemental CaCl2 conferred the greatest effect, resulting in equal growth between B1.1 and B1.3 over five successive passages in mMRS. Moreover, calcium supplementation reversed a B1.3 strain-specific, stationary-phase, flocculation phenotype. These findings show how L. plantarum calcium requirements affect competitive fitness at the strain level.IMPORTANCE Ecological theory states that the struggle for existence is stronger between closely related species. Contrary to this assertion, fermented foods frequently sustain conspecific individuals, in spite of their high levels of phylogenetic relatedness. Therefore, we investigated two isolates of Lactiplantibacillus plantarum, B1.1 and B1.3, randomly selected from a single batch of teff injera batter. These strains spanned the known genomic and phenotypic range of the L. plantarum species, and in laboratory culture medium used for strain screening, B1.3 exhibited poor growth and was outcompeted by the more robust strain B1.1. Nonetheless, B1.1 and B1.3 were equally competitive in teff flour. This result shows how L. plantarum has adapted for coexistence in that environment. The capacity for the single macronutrient calcium to restore B1.3 competitive fitness in laboratory culture medium suggests that L. plantarum intraspecies diversity found in food systems is fine-tuned to nutrient requirements at the strain level.

Biodegradation study of Polyethylene and PVC using naturally occurring plastic degrading microbes.

One of the most serious man-made concerns today is the ever-increasing amount of plastic waste overwhelming the planet. The worldwide interest in using polymers consistently expanded over the years. Because of the plastic wastes thrown into the environment, outrageously the plastic pollution is increasing. In the present study, degradation of PVC and polyethylene-derived synthetic polymers has been carried out. The fungi and bacteria were isolated from the soil of the plastic waste environment and were used for the biodegradation of plastic films. Successful bacterial candidates for biodegradation were identified after screening. The bacterial strain Sb1 was identified as Bacillus licheniformis and Sb2 as Achromobacter xylosoxidans. The fungal strains Sf.1 and Sf.2 were identified as Aspergillus niger and Aspergillus glaucus, respectively. The degraded polymeric films were critically assessed by following the characterization methods like weight loss, FTIR and SEM. The results indicate that the polymers of polyethylene sample showed 32.2% degradation using bacterial strains and 40% using fungal strains in a time duration of just 4weeks. PVC samples degraded 17 and 32% by fungal strains after 4weeks. The changes in surface topography was confirmed by scanning electron microscopy and the changes in functional groups intensity was observed using the FTIR. Different parameters, varying temperature, pH, and inoculum concentration, were also evaluated, which implied that plastic waste treated by fungal and bacterial strains gives significant (p < 0.05) result in polymer degradation. As a result, the current research gave a scientific justification that bacteria and fungus could be further developed as promising candidates for plastic bioremediation.

Efficient and scalable synthesis of 1,5-diamino-2-hydroxy-pentane from l-lysine via cascade catalysis using engineered Escherichia coli

Background1,5-Diamino-2-hydroxy-pentane (2-OH-PDA), as a new type of aliphatic amino alcohol, has potential applications in the pharmaceutical, chemical, and materials industries. Currently, 2-OH-PDA production has only been realized via pure enzyme catalysis from lysine hydroxylation and decarboxylation, which faces great challenges for scale-up production. However, the use of a cell factory is very promising for the production of 2-OH-PDA for industrial applications, but the substrate transport rate, appropriate catalytic environment (pH, temperature, ions) and separation method restrict its efficient synthesis. Here, a strategy was developed to produce 2-OH-PDA via an efficient, green and sustainable biosynthetic method on an industrial scale.ResultsIn this study, an approach was created for efficient 2-OH-PDA production from l-lysine using engineered E. coli BL21 (DE3) cell catalysis by a two-stage hydroxylation and decarboxylation process. In the hydroxylation stage, strain B14 coexpressing l-lysine 3-hydroxylase K3H and the lysine transporter CadB-argT enhanced the biosynthesis of (2S,3S)-3-hydroxylysine (hydroxylysine) compared with strain B1 overexpressing K3H. The titre of hydroxylysine synthesized by B14 was 2.1 times higher than that synthesized by B1. Then, in the decarboxylation stage, CadA showed the highest hydroxylysine activity among the four decarboxylases investigated. Based on the results from three feeding strategies, l-lysine was employed to produce 110.5 g/L hydroxylysine, which was subsequently decarboxylated to generate a 2-OH-PDA titre of 80.5 g/L with 62.6% molar yield in a 5-L fermenter. In addition, 2-OH-PDA with 95.6% purity was obtained by solid-phase extraction. Thus, the proposed two-stage whole-cell biocatalysis approach is a green and effective method for producing 2-OH-PDA on an industrial scale.ConclusionsThe whole-cell catalytic system showed a sufficiently high capability to convert lysine into 2-OH-PDA. Furthermore, the high titre of 2-OH-PDA is conducive to separation and possesses the prospect of industrial scale production by whole-cell catalysis.

Engineering Escherichia coli for the High-Titer Biosynthesis of Lacto-N-tetraose.

Lacto-N-tetraose (LNT), a member of the human milk oligosaccharides family, has received widespread attention because of its importance in infant health. We constructed a whole-cell biotransformation method in Escherichia coli BL21(DE3) for high-titer LNT synthesis. The approach was performed by using a systematic design and metabolic engineering based on the metabolic pathway of LNT. The lgtA (encoding β-1,3-N-acetylglucosaminyltransferase) and wbgO (encoding β-1,3-galactosyltransferase) genes were introduced into the engineered E. coli BL21(DE3) to construct an LNT-producing starting strain B1 (0.22 g/L). Then, the genes related to the LNT metabolic pathway were screened in two vectors to evaluate LNT synthesis. The lgtA-wbgO and galE-galT-galK genes were overexpressed through the two-plasmid system in E. coli BL21(DE3). The titer of LNT (3.42 g/L) had a gain of 14.55 times compared with that of B1. Furthermore, the ugd gene, which was associated with the UDP-Gal bypass pathway, was inactivated to further improve LNT production in shake-flask cultivation (4.14 g/L). The final fed-batch cultivation of the engineered strain produced 31.56 g/L of LNT. This study provided a strategy for the effective production of LNT in E. coli.

Histopathological analysis of placental lesions caused by Chlamydia abortus 1B vaccine strain in vaccinated ewes

Background Chlamydia abortus is one of the most diagnosed causes of infectious abortion in small ruminants. Infections can be controlled using the live, attenuated C. abortus strain 1B vaccine, which has been associated with infection and abortion in animals. This study aimed to compare the severity and the distribution of lesions caused by this vaccine strain (vt) with those resulting from a wild-type (wt) infection. Methods Two grossly affected and 1B-positive (by qPCR and RFLP analysis) placentas from a vaccinated sheep flock were analysed. Histopathological lesions and immunohistochemical labelling (IHC) were graded (increasing score from 0 to 5) according to their severity and distribution. Pathology in the vt infected placentas was compared with that in two wt infected placentas. Datasets generated for observed histological and pathological features were analysed using principal component analysis (PCA). Results Histopathologically, the lesions in both vt and wt-placentas presented as typical multifocal necrosuppurative placentitis, associated with vasculitis, mural necrosis, and thrombosis. IHC for C. abortus revealed intense staining with a multifocal distribution in most cotyledons in both vt and wt placentas. Comparison of the pathological lesions between vt and wt by PCA revealed a similar distribution and severity, revealing a strong association with features such as necrosis and inflammatory infiltration between vt and wt placentas. A weaker association with IHC was observed. Conclusion This study shows that the C. abortus attenuated 1B vaccine strain presents no significant differences in severity and distribution of pathological lesions from those typically observed following a wt infection.

A New Biosurfactant/Bioemulsifier from Gordonia alkanivorans Strain 1B: Production and Characterization

Biosurfactants and bioemulsifiers (BS/BE) are naturally synthesized molecules, which can be used as alternatives to traditional detergents. These molecules are commonly produced by microorganisms isolated from hydrocarbon-rich environments. Gordonia alkanivorans strain 1B was originally found in such an environment, however little was known about its abilities as a BS/BE producer. The goal of this work was to access the potential of strain 1B as a BS/BE producer and perform the initial characterization of the produced compounds. It was demonstrated that strain 1B was able to synthesize lipoglycoprotein compounds with BS/BE properties, both extracellularly and adhered to the cells, without the need for a hydrophobic inducer, producing emulsion in several different hydrophobic phases. Using a crude BS/BE powder, the critical micelle concentration was determined (CMC = 16.94 mg/L), and its capacity to reduce the surface tension to a minimum of 35.63 mN/m was demonstrated, surpassing many commercial surfactants. Moreover, after dialysis, emulsification assays revealed an activity similar to that of Triton X-100 in almond and sunflower oils. In benzene, the E24 value attained was 83.45%, which is 30% greater than that of the commercial alternative. The results obtained highlight for the presence of promising novel BS/BE produced by strain 1B.

An Otorhinolaryngology Perspective Into a Hospital COVID-19 Cluster

In this commentary, we share our experience of a COVID-19 cluster which developed within a frontline healthcare facility designated for treating COVID-19 patients. We provide an Otorhinolaryngology perspective into the key challenges, analyses and responses. We discuss how we identified and isolated infected patients and staff, uncovered the responsible COVID-19 variant strain B1.617.2 and instituted various measures to overcome this cluster. The measures include ceasing non-essential services, limiting transfers of patients, a heightened stance of personal protective equipment, ring-fencing of staff and enhanced COVID-19 testing. With rapid hospital wide efforts, there were no new non-isolated cases from our hospital cluster 3 days after the wards were locked down. The cluster which developed on 28-April-2021 was closed on 6-Jun-2021, with 48 cases, ten of whom were healthcare workers. Some of these lessons may be useful for consideration should another healthcare institution face a similar crisis in the future.

Rhino-orbito-Cerebral mucormycosis during COVID 19 pandemic in western Uttar Pradesh India

Mucormycosis (Black fungus) is a designated as a rare, rapidly progressive fatal disease of immunocompromised caused by saprophytic fungus of family mucorales. Early diagnosis with prompt medical and surgical treatment is the only tool available. Rhino-orbito-cerebral is the most common subtype. In India we saw a sudden rise in mucormycosis cases during second wave of COVID 19. This necessitated a systematic review of epidemic of mucormycosis in COVID 19.A Retrospective multi-centric study was conducted comprising of 51 cases of Rhino-orbito-cerebral mucormycosis with present or recent COVID19 in Western Uttar Pradesh positive status presenting to us during 14 April 2021- 31 May 2021.Either Type2 Diabetes Mellitus or history of recent use of steroids in high doses was present in all the patients. Contribution of virulence of the Delta strain B1.617.2 is significant. FESS with sino-nasal debridement contributes significantly towards mortality reduction and cost of total treatment by significantly reducing days of Liposomal Amphotericin B therapy.Early diagnosis with prompt medical and surgical management along with blood sugar control and avoiding use of high dose of steroids remain to key to mortality and morbidity reduction.

Isolation And Characterization Of Biosurfactant Producing Bacteria From Different Environmental Soil Samples

Isolation And Characterization Of Biosurfactant Producing Bacteria From Different Environmental Soil Samples

Synergistic degradation of phenanthrene by constructed Pseudomonas spp. consortium compared with pure strains

Polycyclic aromatic hydrocarbons (PAHs) are highly persistent compounds as well as they have a toxic impact to all living organisms. Removal of these xenobiotic compounds by mixed bacteria is achieving a maximum degradation rate over the pure strains. So, this study substantiates the application of constructed bacterial consortium for degradation of 400 mg/L phenanthrene within 30 days of incubation at 30 °C. The consortium consisted of two bacterial species Pseudomonas pseudoalcaligenes (B1) and Pseudomonas aeruginosa (B2). It exhibited a significantly higher phenanthrene degradation efficiency (97.5%) compared to pure strains B1 and B2 (52.5 and 47.6%), respectively. The phenanthrene degradation rate by the individual strains was enhanced in the presence of salicylic and phthalic acids as intermediates pathway. On the other side, the degradation rate was reduced in the presence of catechol suggesting a feedback inhibition of catabolic enzymes. The ring-cleavage dioxygenases were induced in all treatments. However, the induction of ortho cleavage dioxygenase (2.44 U/mg protein) was higher than meta cleavage dioxygenase (0.42 U/mg protein). Six main metabolites were detected by GC–MS analysis. Among them, four metabolites were detected in the extract of phenanthrene growing consortium suggesting three main pathways, phthalate, salicylic and benzocoumarin by the constructed consortium. Therefore, this study provides a measuring of phenanthrene degradation efficacy by bacterial consortium and paved the way to increase the potential of using the consortium for successful removal of PAHs from the polluted environment.

Rhino-orbito-cerebral mucormycosis (black fungus) in covid 19 patients in western Uttar pradesh India

Mucormycosis (Black fungus) is a designated as a rare, rapidly progressive fatal disease of immunocompromised caused by saprophytic fungus of family mucorales. Early diagnosis with prompt medical and surgical treatment is the only tool available. Rhino-orbito-cerebral is the most common subtype. In India we saw a sudden rise in mucormycosis cases during second wave of COVID 19. This necessitated a systematic review of epidemic of mucormycosis in COVID 19. A Retrospective multi-centric study was conducted comprising of 51 cases of Rhino-orbito-cerebral mucormycosis with present or recent COVID19 in Western Uttar Pradesh positive status presenting to us during 14th April 2021- 31st May 2021. Either Type2 Diabetes Mellitus or history of recent use of steroids in high doses was present in all the patients. Contribution of virulence of the Delta strain B1.617.2 is significant. FESS with sino-nasal debridement contributes significantly towards mortality reduction and cost of total treatment by significantly reducing days of Liposomal Amphotericin B therapy. Early diagnosis with prompt medical and surgical management along with blood sugar control and avoiding use of high dose of steroids remain to key to mortality and morbidity reduction.

Profiles of Bacillus spp. Isolated from the Rhizosphere of Suaeda glauca and Their Potential to Promote Plant Growth and Suppress Fungal Phytopathogens.

Members of the genus Bacillus are known to play an important role in promoting plant growth and protecting plants against phytopathogenic microorganisms. In this study, 21 isolates of Bacillus spp. were obtained from the root micro-ecosystem of Suaeda glauca. Analysis of the 16S rRNA genes indicated that the isolates belong to the species Bacillus amyloliquefaciens, Bacillus velezensis, Bacillus subtilis, Bacillus pumilus, Bacillus aryabhattai and Brevibacterium frigoritolerans. One of the interesting findings of this study is that the four strains B1, B5, B16 and B21 are dominant in rhizosphere soil. Based on gyrA, gyrB, and rpoB gene analyses, B1, B5, and B21 were identified as B. amyloliquefaciens and B16 was identified as B. velezensis. Estimation of antifungal activity showed that the isolate B1 had a significant inhibitory effect on Fusarium verticillioides, B5 and B16 on Colletotrichum capsici (syd.) Butl, and B21 on Rhizoctonia cerealis van der Hoeven. The four strains grew well in medium with 1-10% NaCl, a pH value of 5-8, and promoted the growth of Arabidopsis thaliana. Our results indicate that these strains may be promising agents for the biocontrol and promotion of plant growth and further study of the relevant bacteria will provide a useful reference for the development of microbial resources.

Integrative multiomics analysis of the acid stress response of Oenococcus oeni mutants at different growth stages

BackgroundAcid stress is one of the most important environmental stresses that adversely affect the growth of lactic acid bacteria (LAB), such as Oenococcus oeni which was isolated from grape-berries and mainly used in wine fermentation. The aim of this paper is to comprehensively characterize the mechanisms of acid stress regulation in O. oeni and to provide a viable theoretical basis for breed and improvement of existing LAB. MethodFirst, six O. oeni mutants with acid-sensitive (strains b2, a1, c2) and acid-tolerant (strains b1, a3, c1) phenotypes were screened from three wild-type O. oeni, and then their genome (sequencing), transcriptome and metabolome (LC-MS/MS) were examined. ResultsA total of 459 genes were identified with one or more intragenic single nucleotide polymorphisms (SNPs) in these mutants, and were extensively involved in metabolism and cellular functions with a high mutation rates in purine (46%) and pyrimidine (48%) metabolic pathways. There were 210 mutated genes that cause significant changes in expression levels. In addition, 446 differentially accumulated metabolites were detected, and they were consistently detected at relatively high levels in the acid-tolerant O. oeni mutant. The levels of intracellular differentially expressed genes and differential metabolites changed with increasing culture time. ConclusionThe integrative pathways analysis showed that the intracellular response associated with acid regulation differed significantly between acid-sensitive and acid-tolerant O. oeni mutants, and also changed at different growth stages.

Design and validation of an expeditious analytical method to quantify the emulsifying activity during biosurfactants/bioemulsifiers production

Biosurfactants (BS) and bioemulsifiers (BE) are amphiphilic molecules that are produced by a wide range of microorganisms. Although the chemical composition of BS and BE is different, both BS/BE have recognized emulsifying properties, which are the focus of this study. Herein, a rapid and simple analytical method to quantify the emulsifying activity (EA) of a product produced by the actinomycete Gordonia alkanivorans strain 1B (BS/BE), which exhibits emulsifying properties, was developed. The analytical approach was based on the ability of a BS/BE solution to form a stable emulsion when mixed with n-heptane. So, using 4 mL screw cap glass tubes (10 × 75 mm, ND10 caps with PTFE septum), the EA was assessed by adding 1 mL of n-heptane to 1 mL of an aqueous solution containing the test product, mix by vortexing at high speed (2 min) and place the tube in an upright stable position for 10 min before analyzing. A set of emulsification tests with increasing volumes of test product solutions was carried out until 100% emulsion was obtained in the organic phase. One emulsification unit was defined as the minimum volume of product (Volmin of emulsifier/surfactant, up to 1 mL) needed to form and maintain 100% emulsion in the organic phase. The corresponding emulsifying activity value is presented in U/mL, and it is calculated as: EA (product) = 1 U/Volmin (mL). Further validation by testing several synthetic surfactants and industrial/domestic dishwashing detergents, in parallel with the bacterial crude BS/BE, towards emulsifying activity determination (U/mL) was performed demonstrating the wide range of the method applicability. Moreover, the specific emulsifying activity for each product tested was estimated though correlation analysis (linear regression) between volumetric emulsifying activity (U/mL) and product concentration (g/L). Indeed, this new analytical approach to quantify the emulsifying activity is accurate and reproducible, and consequently it can be a promising tool to apply in screening/monitorization studies on BS/BE production enabling reliable comparisons.