Succinic acid production from softwood with genome-edited Corynebacterium glutamicum using the CRISPR-Cpf1 system.

Corynebacterium glutamicum is a useful microbe that can be used for producing succinic acid under anaerobic conditions. In this study, we generated a knock-out mutant of the lactate dehydrogenase 1 gene (ΔldhA-6) and co-expressed the succinic acid transporter (Psod:SucE- ΔldhA) using the CRISPR-Cpf1 genome editing system. The highly efficient HPAC (hydrogen peroxide and acetic acid) pretreatment method was employed for the enzymatic hydrolysis of softwood (Pinus densiflora) and subsequently utilized for production of succinic acid. Upon evaluating a 1%-5% hydrolysate concentration range, optimal succinic acid production with the ΔldhA mutant was achieved at a 4% hydrolysate concentration. This resulted in 14.82 g L-1 succinic acid production over 6 h. No production of acetic acid and lactic acid was detected during the fermentation. The co-expression transformant, [Psod:SucE-ΔldhA] produced 17.70 g L-1 succinic acid in 6 h. In the fed-batch system, 39.67 g L-1 succinic acid was produced over 48 h. During the fermentation, the strain consumed 100% and 73% of glucose and xylose, respectively. The yield of succinic acid from the sugars consumed was approximately 0.77 g succinic acid/g sugars. These results indicate that the production of succinic acid from softwood holds potential applications in alternative biochemical processes.

Optimizing bioconversion processes of rice husk into value-added products: D-psicose, bioethanol, and lactic acid.

Rice husk, rich carbon content, is an agricultural waste produced globally at an amount of 120 million tons annually, and it has high potential as a biorefinery feedstock. Herein, we investigated the feasibility of producing various products as D-psicose, bioethanol and lactic acid from rice husk (RH) through a biorefinery process. Alkali-hydrogen peroxide-acetic acid pretreatment of RH effectively removed lignin and silica, resulting in enzymatic hydrolysis yield of approximately 86.3% under optimal hydrolysis conditions. By using xylose isomerase as well as D-psicose-3-epimerase with borate, glucose present in the RH hydrolysate was converted into D-psicose with a 40.6% conversion yield in the presence of borate. Furthermore, bioethanol (85.4%) and lactic acid (92.5%) were successfully produced from the RH hydrolysate. This study confirmed the high potential of RH as a biorefinery feedstock, and it is expected that various platform chemicals and value-added products can be produced using RH.

Value-Added Products from Coffee Waste: A Review.

Coffee waste is often viewed as a problem, but it can be converted into value-added products if managed with clean technologies and long-term waste management strategies. Several compounds, including lipids, lignin, cellulose and hemicelluloses, tannins, antioxidants, caffeine, polyphenols, carotenoids, flavonoids, and biofuel can be extracted or produced through recycling, recovery, or energy valorization. In this review, we will discuss the potential uses of by-products generated from the waste derived from coffee production, including coffee leaves and flowers from cultivation; coffee pulps, husks, and silverskin from coffee processing; and spent coffee grounds (SCGs) from post-consumption. The full utilization of these coffee by-products can be achieved by establishing suitable infrastructure and building networks between scientists, business organizations, and policymakers, thus reducing the economic and environmental burdens of coffee processing in a sustainable manner.

Predicting the severity of COVID-19 patients using the CD24-CSF1R index in whole blood samples.

Coronavirus disease 2019 (COVID-19), caused by SARS-CoV-2, has become one of the most serious public health crises worldwide. Most infected people are asymptomatic but are still able to spread the virus. People with mild or moderate illnesses are likely to recover without hospitalization, while critically ill patients face a higher risk of organ injury or even death. In this study, we aimed to identify a novel biomarker that can predict the severity of COVID-19 patients. Clinical information and RNA-seq data of leukocytes from whole blood samples with and without a COVID-19 diagnosis (n = 100 and 26, respectively) were retrieved from the National Center for Biotechnology Information Gene Expression Omnibus database. Raw data were processed using the Transcripts Per Million (TPM) method and then transformed using log2 (TPM+1) for normalization. The CD24-CSF1R index was established. Violin plots, Kaplan-Meier curves, ROC curves, and multivariate Cox proportional hazards regression analyses were performed to evaluate the prognostic value of the established index. The CD24-CSF1R index was significantly associated with ICU admission (n = 50 ICU, 50 non-ICU) and ventilatory status (n = 42 ventilation, 58 non-ventilation) with p = 4.186e-11 and p = 1.278e-07, respectively. The ROC curve produced a relatively accurate prediction of ICU admission with an AUC of 0.8524. Additionally, patients with a high index had significantly fewer mechanical ventilation-free days than patients with a low index (p = 6.07e-07). Furthermore, the established index showed a strong prognostic ability for the risk of using a ventilator in the multivariate Cox regression model (p < 0.001). The CD24-CSF1R index was significantly associated with COVID-19 severity. The established index could have potential implications for prognosis, disease severity stratification, and clinical management.

Converting textile waste into value-added chemicals: An integrated bio-refinery process.

The rate of textile waste generation worldwide has increased dramatically due to a rise in clothing consumption and production. Here, conversion of cotton-based, colored cotton-based, and blended cotton-polyethylene terephthalate (PET) textile waste materials into value-added chemicals (bioethanol, sorbitol, lactic acid, terephthalic acid (TPA), and ethylene glycol (EG)) via enzymatic hydrolysis and fermentation was investigated. In order to enhance the efficiency of enzymatic saccharification, effective pretreatment methods for each type of textile waste were developed, respectively. A high glucose yield of 99.1% was obtained from white cotton-based textile waste after NaOH pretreatment. Furthermore, the digestibility of the cellulose in colored cotton-based textile wastes was increased 1.38-1.75 times because of the removal of dye materials by HPAC-NaOH pretreatment. The blended cotton-PET samples showed good hydrolysis efficiency following PET removal via NaOH-ethanol pretreatment, with a glucose yield of 92.49%. The sugar content produced via enzymatic hydrolysis was then converted into key platform chemicals (bioethanol, sorbitol, and lactic acid) via fermentation or hydrogenation. The maximum ethanol yield was achieved with the white T-shirt sample (537 mL/kg substrate), which was 3.2, 2.1, and 2.6 times higher than those obtained with rice straw, pine wood, and oak wood, respectively. Glucose was selectively converted into sorbitol and LA at a yield of 70% and 83.67%, respectively. TPA and EG were produced from blended cotton-PET via NaOH-ethanol pretreatment. The integrated biorefinery process proposed here demonstrates significant potential for valorization of textile waste.

Utilization of bamboo as biorefinery feedstock: Co-production of xylo-oligosaccharide with succinic acid and lactic acid.

Herein, we investigated the possibility of co-producing xylo-oligosaccharides (XOSs) from bamboo, as value-added products, along with succinic and lactic acids, as platform chemicals. Xylan was extracted from bamboo using the alkali method under mild conditions. From xylan, XOSs were produced by partial enzymatic hydrolysis at a conversion rate of 83.9%, and all reaction conditions resulted in similar degrees of polymerization. Hydrogen peroxide-acetic acid (HPAC) pretreatment effectively removed lignin from NaOH-treated bamboo, and the enzymatic hydrolytic yield of NaOH and HPAC-treated bamboo was 84.3% of the theoretical yield. The production of succinic and lactic acids from the hydrolysate resulted in conversion rates of approximately 63.2% and 91.3% of the theoretical yield using Corynebacterium glutamicum Δldh and Actinobacillus succinogenes, respectively, under facultative anaerobic conditions. This study demonstrates that bamboo has a high potential to produce value-added products using a biorefinery process and is an alternative resource for compounds typically derived from petroleum.

An integrated process for conversion of spent coffee grounds into value-added materials.

Spent coffee grounds (SCG) are inexpensive materials with a complex composition that makes them promising feedstocks for a biorefinery.Here, conversion of SCG into a wide range of high value-added products (coffee oil, bio-ethanol, D-mannose, manno-oligosaccharide (MOS), cafestol and kahweol) using a novel integrated system was evaluated. The process involves oil extraction, MOS production by mannanase obtained from Penicillium purpurogenum, NaOH (Na) and hydrogen peroxide (HP) pretreatment for the degradation of lignin and phenolic compounds, diterpenes extraction, enzymatic hydrolysis, and fermentation, which can be performed using environmentally friendly technologies. Approximately 97 mL of coffee oil, 164 g of D-mannose, 102 g of MOS, 99 g of bioethanol and a dash of cafestol/kahweol were produced from 1 kg of dry SCG. Producing high-value co-products from SCG using an integrated approach as demonstrated here may be an efficient strategy to reduce waste generation, while improving the economics of the biorefinery production process.

A novel signature predicts recurrence risk and therapeutic response in breast cancer patients.

Acetylserotonin O-methyltransferase (ASMT) is a key enzyme in the synthesis of melatonin. Although melatonin has been shown to exhibit anticancer activity and prevents endocrine resistance in breast cancer, the role of ASMT in breast cancer progression remains unclear. In this retrospective study, we analyzed gene expression profiles in 27 data sets on 7244 patients from 11 countries. We found that ASMT expression was significantly reduced in breast cancer tumors relative to healthy tissue. Among breast cancer patients, those with higher levels of ASMT expression had better relapse-free survival outcomes and longer metastasis-free survival times. Following treatment with tamoxifen, patients with greater ASMT expression experienced longer periods before relapse or distance recurrence. Motivated by these results, we devised an ASMT gene signature that can correctly identify low-risk cases with a sensitivity and specificity of 0.997 and 0.916, respectively. This signature was robustly validated using 23 independent breast cancer mRNA array data sets from different platforms (consisting of 5800 patients) and an RNAseq data set from TCGA (comprising 1096 patients). Intriguingly, patients who are classified as high-risk by the signature benefit from adjuvant chemotherapy, and those with grade II tumors who are classified as low-risk exhibit improved overall survival and distance relapse-free outcomes following endocrine therapy. Together, our findings more clearly elucidate the roles of ASMT, provide strategies for improving the efficacy of tamoxifen treatment and help to identify those patients who may maximally benefit from adjuvant or endocrine therapies.

Pluronic F127-Folate Coated Super Paramagenic Iron Oxide Nanoparticles as Contrast Agent for Cancer Diagnosis in Magnetic Resonance Imaging.

Contrast agents have been widely used in medicine to enhance contrast in magnetic resonance imaging (MRI). Among them, super paramagnetic iron oxide nanoparticles (SPION) have been reported to have low risk in clinical use. In our study, F127-Folate coated SPION was fabricated in order to efficiently target tumors and provide imaging contrast in MRI. SPION alone have an average core size of 15 nm. After stabilizing with Pluronic F127, the nanoparticles reached a hydrodynamic size of 180 nm and dispersed well in various kinds of media. The F127-Folate coated SPION were shown to specifically target folate receptor expressing cancer cells by flow cytometry analysis, confocal laser scanning microscope, as well as in vitro MRI. Furthermore, in vivo MRI images have shown the enhanced negative contrast from the F127-Folate coated SPION in tumor-bearing mice. In conclusion, our F127-Folate coated SPION have shown great potential as a contrast agent in MRI, as well as in the combination with drug delivery for cancer therapy.

Silver and gold nanoparticles biosynthesized by aqueous extract of burdock root, Arctium lappa as antimicrobial agent and catalyst for degradation of pollutants.

This study presents an efficient and facile method for biosynthesis of silver nanoparticles (AgNPs) and gold nanoparticles (AuNPs) using aqueous extract of burdock root (BR), A. lappa, and their applications. The nanoparticles were characterized by ultraviolet-visible spectrophotometry, X-ray diffraction, transmission electron microscopy, energy dispersive X-ray, thermogravimetry, and differential thermal analysis. AgNPs capped the BR extract (BR-AgNPs) possessed roughly spherical geometry with an average diameter of 21.3 nm while uneven geometry of AuNPs capped the BR extract (BR-AuNPs) showed multi shapes in average size of 24.7 nm. The BR-AgNPs strongly inhibited five tested microorganism strains. In particular, the nanoparticles showed excellent catalytic activity for the conversion of pollutants within wastewater. Pseudo-first-order rate constants for the degradation of 4-nitrophenol, methyl orange, and rhodamine B were respectively found 6.77 × 10-3, 3.70 × 10-3, and 6.07 × 10-3 s-1 for BR-AgNPs and 6.87 × 10-3, 6.07 × 10-3, and 7.07 × 10-3 s-1 for BR-AuNPs. Graphical abstract ᅟ.

CRC-113 gene expression signature for predicting prognosis in patients with colorectal cancer.

Colorectal cancer (CRC) is the third leading cause of global cancer mortality. Recent studies have proposed several gene signatures to predict CRC prognosis, but none of those have proven reliable for predicting prognosis in clinical practice yet due to poor reproducibility and molecular heterogeneity. Here, we have established a prognostic signature of 113 probe sets (CRC-113) that include potential biomarkers and reflect the biological and clinical characteristics. Robustness and accuracy were significantly validated in external data sets from 19 centers in five countries. In multivariate analysis, CRC-113 gene signature showed a stronger prognostic value for survival and disease recurrence in CRC patients than current clinicopathological risk factors and molecular alterations. We also demonstrated that the CRC-113 gene signature reflected both genetic and epigenetic molecular heterogeneity in CRC patients. Furthermore, incorporation of the CRC-113 gene signature into a clinical context and molecular markers further refined the selection of the CRC patients who might benefit from postoperative chemotherapy. Conclusively, CRC-113 gene signature provides new possibilities for improving prognostic models and personalized therapeutic strategies.

Genome-wide analysis of DNA methylation in pigs using reduced representation bisulfite sequencing.

DNA methylation plays a major role in the epigenetic regulation of gene expression. Although a few DNA methylation profiling studies of porcine genome which is one of the important biomedical models for human diseases have been reported, the available data are still limited. We tried to study methylation patterns of diverse pig tissues as a study of the International Swine Methylome Consortium to generate the swine reference methylome map to extensively evaluate the methylation profile of the pig genome at a single base resolution. We generated and analysed the DNA methylome profiles of five different tissues and a cell line originated from pig. On average, 39.85 and 62.1% of cytosine and guanine dinucleotides (CpGs) of CpG islands and 2 kb upstream of transcription start sites were covered, respectively. We detected a low rate (an average of 1.67%) of non-CpG methylation in the six samples except for the neocortex (2.3%). The observed global CpG methylation patterns of pigs indicated high similarity to other mammals including humans. The percentage of CpG methylation associated with gene features was similar among the tissues but not for a 3D4/2 cell line. Our results provide essential information for future studies of the porcine epigenome.

Analysis of cattle olfactory subgenome: the first detail study on the characteristics of the complete olfactory receptor repertoire of a ruminant.

BACKGROUND: Mammalian olfactory receptors (ORs) are encoded by the largest mammalian multigene family. Understanding the OR gene repertoire in the cattle genome could lead to link the effects of genetic differences in these genes to variations in olfaction in cattle. RESULTS: We report here a whole genome analysis of the olfactory receptor genes of Bos taurus using conserved OR gene-specific motifs and known OR protein sequences from diverse species. Our analysis, using the current cattle genome assembly UMD 3.1 covering 99.9% of the cattle genome, shows that the cattle genome contains 1,071 OR-related sequences including 881 functional, 190 pseudo, and 352 partial OR sequences. The OR genes are located in 49 clusters on 26 cattle chromosomes. We classified them into 18 families consisting of 4 Class I and 14 Class II families and these were further grouped into 272 subfamilies. Comparative analyses of the OR genes of cattle, pigs, humans, mice, and dogs showed that 6.0% (n = 53) of functional OR cattle genes were species-specific. We also showed that significant copy number variations are present in the OR repertoire of the cattle from the analysis of 10 selected OR genes. CONCLUSION: Our analysis revealed the almost complete OR gene repertoire from an individual cattle genome. Though the number of OR genes were lower than in pigs, the analysis of the genetic system of cattle ORs showed close similarities to that of the pig.

Alpha (1,2)-fucosyltransferase M307A polymorphism improves piglet survival.

To confirm the beneficial effects of alpha (1,2)-fucosyltransferase (FUT1) M307 (A) on piglet survival on commercial farms, we performed PCR-RFLP analysis of FUT1 M307 in successfully marketed (n = 245) and disease affected/deceased pigs during weaning (n = 252) at a commercial farm. We also evaluated the FUT1 genotypes of 190 healthy pigs from three different genetic backgrounds. The distribution of genotypes differed between the successfully marketed and disease affected/deceased pig groups. The frequency of the A allele, associated with resistance to edema and post-weaning diarrhea, was higher in the post-weaning survival group (0.21) than in the non-survival group (0.16, P < 0.05). The odds ratio for piglet survival between AA and GG genotypes was 1.98; thus, piglet survival for individuals with the AA genotype was almost two-fold greater than for GG individuals. The FUT1 gene polymorphism can be used as an effective marker for selection programs to improve post-weaning piglet survival.

The complete swine olfactory subgenome: expansion of the olfactory gene repertoire in the pig genome.

BACKGROUND: Insects and animals can recognize surrounding environments by detecting thousands of chemical odorants. Olfaction is a complicated process that begins in the olfactory epithelium with the specific binding of volatile odorant molecules to dedicated olfactory receptors (ORs). OR proteins are encoded by the largest gene superfamily in the mammalian genome. RESULTS: We report here the whole genome analysis of the olfactory receptor genes of S. scrofa using conserved OR gene specific motifs and known OR protein sequences from diverse species. We identified 1,301 OR related sequences from the S. scrofa genome assembly, Sscrofa10.2, including 1,113 functional OR genes and 188 pseudogenes. OR genes were located in 46 different regions on 16 pig chromosomes. We classified the ORs into 17 families, three Class I and 14 Class II families, and further grouped them into 349 subfamilies. We also identified inter- and intra-chromosomal duplications of OR genes residing on 11 chromosomes. A significant number of pig OR genes (n = 212) showed less than 60% amino acid sequence similarity to known OR genes of other species. CONCLUSION: As the genome assembly Sscrofa10.2 covers 99.9% of the pig genome, our analysis represents an almost complete OR gene repertoire from an individual pig genome. We show that S. scrofa has one of the largest OR repertoires, suggesting an expansion of OR genes in the swine genome. A significant number of unique OR genes in the pig genome may suggest the presence of swine specific olfactory stimulation.

Genome-level identification, gene expression, and comparative analysis of porcine ß-defensin genes.

BACKGROUND: Beta-defensins (ß-defensins) are innate immune peptides with evolutionary conservation across a wide range of species and has been suggested to play important roles in innate immune reactions against pathogens. However, the complete ß-defensin repertoire in the pig has not been fully addressed. RESULT: A BLAST analysis was performed against the available pig genomic sequence in the NCBI database to identify ß-defensin-related sequences using previously reported ß-defensin sequences of pigs, humans, and cattle. The porcine ß-defensin gene clusters were mapped to chromosomes 7, 14, 15 and 17. The gene expression analysis of 17 newly annotated porcine ß-defensin genes across 15 tissues using semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) showed differences in their tissue distribution, with the kidney and testis having the largest pBD expression repertoire. We also analyzed single nucleotide polymorphisms (SNPs) in the mature peptide region of pBD genes from 35 pigs of 7 breeds. We found 8 cSNPs in 7 pBDs. CONCLUSION: We identified 29 porcine ß-defensin (pBD) gene-like sequences, including 17 unreported pBDs in the porcine genome. Comparative analysis of ß-defensin genes in the pig genome with those in human and cattle genomes showed structural conservation of ß-defensin syntenic regions among these species.