Unveiling the Health Benefits of Prebiotics: A Comprehensive Review.

Prebiotics play a pivotal role in fostering probiotics, essential contributors to the creation and maintenance of a conducive environment for beneficial microbiota within the human gut. To qualify as a prebiotic, a substance must demonstrate resilience to stomach enzymes, acidic pH levels, and intestinal bacteria, remaining unabsorbed in the digestive system while remaining accessible to gut microflora. The integration of prebiotics and probiotics into our daily diet establishes a cornerstone for optimal health, a priority for health-conscious consumers emphasizing nutrition that supports a balanced gut flora. Prebiotics offer diverse biological functions in humans, exhibiting antiobesity, antimicrobial, anticancer, anti-inflammatory, antidiabetic, and cholesterol-lowering properties, along with preventing digestive disorders. Numerous dietary fibers possessing prebiotic attributes are inadvertently present in our diets, emphasizing the broader significance of prebiotics. It is crucial to recognize that, while all dietary fibers are prebiotics, not all prebiotics fall under the category of dietary fibers. The versatile applications of prebiotics extend across various industries, such as dairy, bakery, beverages, cosmetics, pharmaceuticals, and other food products. This comprehensive review provides insights into different prebiotics, encompassing their sources, chemical compositions, and applications within the food industry.

Insertion of a FLAG-tag sequence at the end of exon 9 of the TBX5 gene in three induced pluripotent stem cell lines (DHMi004-A-4, DHMi004-A-5, DHMi004-A-6) by CRISPR/Cas9 technology.

The identification of TBX5-related regulatory sequences in genes essential for heart development is hampered by the absence of antibodies which allow precipitation of TBX5:DNA complexes. Employing CRISPR/Cas9 technology, we have inserted a FLAG-tag sequence at the end of exon 9 of the TBX5 gene prior to the stop codon by homologous recombination. The translated TBX5-FLAG fusion protein of the three iPSC lines can effectively be precipitated by anti-FLAG antibodies and, thus, allow the detection of specific TBX5-binding sites and their associated genes.

Characterization and genome sequencing of three Aeromonas hydrophila-specific phages, CF8, PS1, and PS2.

Aeromonas hydrophila is an important finfish pathogen, besides being an opportunistic human pathogen. In the present study, the genomes of three A. hydrophila-specific phages, CF8, PS1, and PS2, were isolated, characterized and sequenced. Transmission electron microscopy showed that all three phages had typical Myoviridae morphology. The linear dsDNA genomes of CF8, PS1, and PS2 were 238,150 bp, 237,367 bp, and 240,447 bp in length, with a GC content of 42.2%, 38.8%, and 38.8%, respectively. The low sequence similarity (67.6% - 69.8% identity with 27.0% - 29.0% query coverage) to other phage genomes in the NCBI database indicated the novel nature of the CF8, PS1, and PS2 genomes. A total of 244, 247, and 250 open reading frames (ORFs) were predicted in the CF8, PS1, and PS2 genome, respectively. During the annotation process, functional predictions were made for 28-31 ORFs, while the rest were classified as "hypothetical proteins" with yet unknown functions. Genes for tRNAs were also detected in all phage genomes. As all three phages in the present study had a very narrow host range with lytic activity against only one strain of A. hydrophila, these phages could be good candidates for phage typing applications. Moreover, the endolysin- and lytic-transglycosylase-encoding genes could be used for recombinant cloning and expression of anti-microbial proteins.

Correction to: Characterization and genome sequencing of three Aeromonas hydrophila-specific phages, CF8, PS1, and PS2.

Authors would like to correct the incorrect version.

Shotgun metagenomics offers novel insights into taxonomic compositions, metabolic pathways and antibiotic resistance genes in fish gut microbiome.

Gut microbiota of freshwater carp (Labeo rohita) was investigated by shotgun metagenomics to understand its taxonomic composition and functional capabilities. With the presence of 36 phyla, 326 families and 985 genera, the fish gut microbiota was found to be quite diverse in nature. However, at the phylum level, more than three-fourths of gut microbes belonged to Proteobacteria. Very low prevalence of commonly used probiotic bacteria (Bacillus, Lactobacillus, Streptococcus, and Lactococcus) in fish gut suggested the need to search for alternative probiotics for aquaculture use. Biosynthesis pathways were found to be the most dominant (51%) followed by degradation (39%), energy metabolism (4%) and fermentation (2%). In conformity with herbivorous feeding habit of L. rohita, gut microbiome also had pathways for the degradation of cellulose, hemicellulose, chitin, pectin, starch, and other complex carbohydrates. High prevalence of Actinobacteria and antibiotic biosynthesis pathways in the fish gut microbiome indicated its potential for bioprospecting of potentially novel natural antibiotics. Fifty-one different types of antibiotic resistance genes (ARGs) belonging to 15 antimicrobial resistance (AMR) gene families and conferring resistance against 24 antibiotic types were detected in fish gut. Some of the ARGs for multi-drug resistance were also found to be located on sequences of plasmid origin. The presence of pathogenic bacteria and ARGs on plasmid sequences suggested the potential risk due to horizontal gene transfer in the confined gut environment. The role of ARGs in fish gut microbiome needs further investigations.

Antioxidant activity and associated structural attributes of Halomicronema phycoerythrin.

In the present study, blue light absorbing pigment protein phycoerythrin (PE) is purified up to molecular grade purity from marine Halomicronema sp. R31DM. The purification method is based on the use of non-ionic detergent Triton-X 100 in ammonium sulphate precipitation. The purified PE is characterized for its antioxidant activity in vitro and in vivo. PE is noted to show substantial in vitro antioxidant activity probed by various biochemical assays. The PE moderated rise in the intracellular-ROS (reactive oxygen species) in wild type Caenorhabditis elegans upon heat and oxidative stress. Further, the antioxidant asset of PE is noted an expedient in averting the ROS associated abnormalities, i.e. impaired physiological behaviour (health span) and aging in C. elegans. The structural attributes of PE contributing to its antioxidant virtue are analysed; the presence of ample residues having antioxidant activity and chromophore-PEB in PE are identified as a source of its antioxidant activity. Furthermore, the stability of PE is assessed under three physico-chemical stresses, temperature, pH and oxidative stress.

In Operando Impedance Spectroscopic Analysis on NiO-WO3 Nanorod Heterojunction Random Networks for Room-Temperature H2S Detection.

The use of metal-oxide sensors for effectively detecting hydrogen sulfide (H2S) gas at room temperature is currently hindered by their inadequate sensitivity and selectivity. Using a lucid fabrication strategy, we report a room-temperature, highly sensitive, and selective H2S gas sensor using NiO-modified WO3 nanorod (one-dimensional-one-dimensional) random networks. The observed improvements in gas-sensing sensitivity stem from the synergistic effects of various contributions inside the sensing heterostructure, such as bulk nanorod, p-n heterojunction at the interface of these two dissimilar oxides, and gas-induced conducting species due to sulfurization (WS2-x and NiS1-x ). An in situ impedance measurement during gas exposure was used to investigate the influence of these effects. The analysis revealed that these contributing factors can be either cooperating or competing and lead to either increased or decreased sensitivity, respectively. The presence of semimetallic species (NiS, WS2) was further confirmed by in situ X-ray diffraction analysis of the heterostructure nanorod sample with H2S gas exposure. The related sensing mechanism in the heterostructures is presented with a conduction pathway model. The room-temperature-operated nanorod heterostructure sensors showed a lower detection limit of H2S at ∼0.5 ppm, which is significantly lower than its toxicity limiting value ∼10 ppm, per the Environmental Protection Agency. The nanorod heterostructure sensors can be used for real-time, low-cost, room-temperature alarms in an H2S monitoring system.

Purification and antioxidant activity of phycocyanin from Synechococcus sp. R42DM isolated from industrially polluted site.

The cyanobacterium Synechococcus sp. R42DM, isolated from an industrially polluted site Vatva, Gujarat, India was recognized to produce phycocyanin (PC) as major phycobiliprotein. In present study, the combinatorial approach of chemical and physical methods i.e. Triton-X 100 treatment and ultra-sonication was designed for extraction of PC. From cell extract, the intact and functional-PC was purified up to purity 4.03 by ammonium sulphate fractionation and ion-exchange chromatography. The PC displayed considerable in vitro antioxidant and radical-scavenging activity. This PC was further noticed to scavenge intracellular-ROS and to increase tolerance against thermal and oxidative stress in Caenorhabditis elegans. Moreover, the PC was noticed to improve the physiological behaviour and longevity of C. elegans. In addition, the PC showed remarkable stability under physico-chemical stressors, which is desirable for their use in biomedical applications. In conclusion, present paper added up evidence in support of the prospective use of PC as an antioxidant nutraceutical.

Genome dynamics and evolution of codon usage patterns in shrimp viruses.

We analysed the genomes and codon usage patterns of seven small (DNA and RNA) shrimp viruses. Effective number of codon (ENC) values indicated moderate (35 < ENC < 50) codon usage bias in shrimp viruses. Correlation analysis between GC compositions at non-synonymous codon and synonymous codon positions (GC1, 2 and GC3) as well as GC3 versus ENC curves indicated varying influences of mutational pressure on codon usage. The presence of deoptimized codons and host-antagonistic codon usage trends in shrimp viruses suggested the adaptation of a slow replication strategy by these viruses to avoid host defences. Low CpG frequencies indicated that shrimp viruses have evolved with underrepresentation of CpGs to avoid the host's immune response.

A detailed analysis of codon usage patterns and influencing factors in Zika virus.

Recent outbreaks of Zika virus (ZIKV) in Africa, Latin America, Europe, and Southeast Asia have resulted in serious health concerns. To understand more about evolution and transmission of ZIKV, detailed codon usage analysis was performed for all available strains. A high effective number of codons (ENC) value indicated the presence of low codon usage bias in ZIKV. The effect of mutational pressure on codon usage bias was confirmed by significant correlations between nucleotide compositions at third codon positions and ENCs. Correlation analysis between Gravy values, Aroma values and nucleotide compositions at third codon positions also indicated some influence of natural selection. However, the low codon adaptation index (CAI) value of ZIKV with reference to human and mosquito indicated poor adaptation of ZIKV codon usage towards its hosts, signifying that natural selection has a weaker influence than mutational pressure. Additionally, relative dinucleotide frequencies, geographical distribution, and evolutionary processes also influenced the codon usage pattern to some extent.

EvaGreen-based Multiplex Real-time PCR Assay for Rapid Differentiation of Wild-Type and Glycoprotein E-Deleted Bovine Herpesvirus-1 Strains.

Bovine herpesvirus-1 (BoHV-1) is an important viral pathogen causing significant economic losses to the cattle industry. Glycoprotein E-deleted marker vaccines form the basis for BoHV-1 control programs widely, wherein detection and differentiation of wild-type and gE-deleted vaccine strains is of crucial importance for proper disease management. In the present study, we report an EvaGreen-based multiplex real-time polymerase chain reaction (EGRT-PCR) assay for rapid differentiation of wild-type and glycoprotein E-deleted strains of BoHV-1. The EGRT-PCR assay could simultaneously detect two viral genes (glycoprotein B and E) and an internal positive control gene (bovine growth hormone- bGH), in a single-tube reaction. The analytical sensitivity of the EGRT-PCR assay was as little as 10 copies of the BoHV-1 DNA per reaction. The modified real-time PCR assay could successfully differentiate wild-type and gE-deleted BoHV-1 strains based on gene specific melting temperatures (Tm) peaks. Our results have shown that the EGRT-PCR developed in this study might prove to be a promising tool in disease management by enabling rapid differentiation of wild-type and gE-deleted strains of BoHV-1.

Phycoerythrin averts intracellular ROS generation and physiological functional decline in eukaryotes under oxidative stress.

In vitro antioxidant virtue and life-prolonging effect of phycoerythrin (PE; a pigment protein isolated from Phormidium sp. A09DM) have been revealed in our previous reports (Sonani et al. in Age 36:9717, 2014a; Sonani et al. in Process Biochem 49:1757-1766, 2014b). It has been hypothesized that the PE expands life span of Caenorhabditis elegans (bears large resemblance with human aging pathways) due to its antioxidant virtue. This hypothesis is tested in present study by checking the effect of PE on intracellular reactive oxygen species (ROS) generation and associated physiological deformities using mouse and human skin fibroblasts, C. elegans, and Drosophila melanogaster Oregon R + and by divulging PE's structural attributes responsible for its antioxidant asset. PE treatment displayed noteworthy decrease of 67, 48, and 77 % in ROS level in mouse fibroblast (3T3-L1), human fibroblast, and C. elegans N2, respectively, arisen under chemical-induced oxidative stress. PE treatment delayed the development of paraquat-induced Alzheimer phenotype by 14.5 % in C. elegans CL4176. Furthermore, PE improved the locomotion of D. melanogaster Oregon R + under oxidative stress with simultaneous up-regulation in super-oxide dismutase and catalase activities. The existence of 52 Glu + Asp + His + Thr residues (having metal ion sequestration capacity), 5 phycoerythrobilin chromophores (potential electron exchangers) in PE's primary structure, and significant hydrophobic patches on the surface of its α- and ß-subunits are supposed to collectively contribute in the antioxidant virtues of PE. Altogether, results support the hypothesis that it is the PE's antioxidant asset, which is responsible for its life-prolonging effect and thus could be exploited in the therapeutics of ROS-associated abnormalities including aging and neurodegeneration in eukaryotes.

Characterization of codon usage pattern and influencing factors in Japanese encephalitis virus.

Recently, several outbreaks of Japanese encephalitis (JE), caused by Japanese encephalitis virus (JEV), have been reported and it has become cause of concern across the world. In this study, detailed analysis of JEV codon usage pattern was performed. The relative synonymous codon usage (RSCU) values along with mean effective number of codons (ENC) value of 55.30 indicated the presence of low codon usages bias in JEV. The effect of mutational pressure on codon usage bias was confirmed by significant correlations of A3s, U3s, G3s, C3s, GC3s, ENC values, with overall nucleotide contents (A%, U%, G%, C%, and GC%). The correlation analysis of A3s, U3s, G3s, C3s, GC3s, with axis values of correspondence analysis (CoA) further confirmed the role of mutational pressure. However, the correlation analysis of Gravy values and Aroma values with A3s, U3s, G3s, C3s, and GC3s, indicated the presence of natural selection on codon usage bias in addition to mutational pressure. The natural selection was further confirmed by codon adaptation index (CAI) analysis. Additionally, relative dinucleotide frequencies, geographical distribution, and evolutionary processes also influenced the codon usage pattern to some extent.

Bioinformatics analysis of codon usage patterns and influencing factors in Penaeus monodon nudivirus.

Penaeus monodon nudivirus (PmNV) is one of the most important and most commonly reported shrimp viruses. In the present study, codon usage of PmNV was studied in detail. Based on effective number of codons (ENC) values, strong to low codon usage bias was observed in PmNV genes. Nucleotide composition-ENC correlation analysis and the GC3 versus ENC relationship indicated that compositional constraint has a major effect on codon usage of PmNV. At the whole-genome level, relative synonymous codon usage (RSCU) analysis showed almost complete antagonism between the codon usage pattern of PmNV and its host P. monodon. However, codon adaptive index (CAI) values indicated that forces of selective/translational constraints have been able to overcome this antagonism in some genes.

Loop-Mediated Isothermal Amplification for Rapid Detection and Differentiation of Wild-Type Bovine Herpesvirus-1 and Glycoprotein E-Deleted Marker Vaccine Strain.

Bovine herpesvirus-1 (BoHV-1) is an important viral pathogen affecting cattle and causing numerous reproductive disorders leading to significant economic losses to the cattle industry. The control programs for BoHV-1 are widely based on the use of glycoprotein E-deleted marker vaccines, wherein detection and differentiation of wild-type and gE-deleted vaccine strains is of crucial importance for proper disease management. In this study, we report rapid and simple loop-mediated isothermal amplification (LAMP) assays for detection and differentiation of gE-deleted BoHV-1 from wild-type virus under isothermal conditions. The assays could be completed in 90 mintes, including viral DNA isolation, target amplification and visual interpretation of results with naked eye. The analytical sensitivity of the assays was 10 times higher than conventional PCR and could detect as little as 100 fg of viral DNA per reaction. The applicability of LAMP for detection of BoHV-1 in bovine semen was assessed by testing semen samples collected from breeding bulls and compared with TaqMan real-time PCR (as gold standard). The LAMP assays had diagnostic specificity of 100%. The diagnostic sensitivity was 88.2% and 83.3% for gB- and gE-LAMP, respectively, when compared with TaqMan real-time PCR. Our results have shown that the LAMP method developed in this study is a potential tool for rapid, sensitive, specific, cost-effective, and user-friendly detection and differentiation of wild type BoHV-1 from gE-deleted marker vaccine.

Development of a SYBR Green I based duplex real-time PCR for detection of bovine herpesvirus-1 in semen.

Bovine herpesvirus-1 (BoHV-1) is a viral pathogen found in infected bull semen, which is transmitted to inseminated cows by artificial insemination. BoHV-1 infection can cause reproductive disorders leading to significant economic loss to cattle industry. To detect BoHV-1 in semen, in this study, a SYBR Green I based duplex real-time PCR was developed. The assay included primers from BoHV-1 glycoprotein C (gC) and bovine growth hormone (bGH) genes for simultaneous detection in single tube. The result was interpreted by analysing melting temperature (Tm) peaks obtained after melt curve analysis of the amplified products at the end of reaction. The Tm peaks for BoHV-1-gC indicated presence of BoHV-1 while the bGH peak indicated reaction without inhibition. The sensitivity of the assay was to detect ten BoHV-1 genome copies per reaction. The analytical sensitivity was to detect 0.21 TCID50 infectious BoHV-1 in spiked semen. The assay was validated with 80 semen samples collected from breeding bulls. The diagnostic sensitivity and specificity of the assay was 100% with OIE recommended TaqMan probe based real-time PCR.

Rapid detection of bovine herpesvirus 1 in bovine semen by loop-mediated isothermal amplification (LAMP) assay.

Bovine herpesvirus 1 (BoHV-1) is the most common viral pathogen found in bovine semen, causing numerous reproductive disorders leading to economic losses to the cattle industry. For rapid detection of BoHV-1 in bovine semen, in this study, we applied a loop-mediated isothermal amplification (LAMP) assay. The assay could be completed within 90 min, including total DNA isolation, target amplification, and visual interpretation of positive or negative results with the naked eye. The assay detected as little as 10 fg of BoHV-1 DNA per reaction. The analytical sensitivity of the assay was 0.2 TCID50 BoHV-1 per reaction, which was 100 times more sensitive than conventional PCR and comparable to TaqMan real-time PCR. The applicability of the assay was assessed by analysing 118 semen samples collected from breeding bulls. On comparison with TaqMan real-time PCR, the LAMP assay had a diagnostic sensitivity of 97 %, specificity of 100 %, and accuracy of 99.2 % for detection of BoHV-1 in bovine semen. The LAMP assay developed in this study is a rapid, sensitive, and cost-effective alternative for detection of BoHV-1 in bovine semen.

Protection of mice against lethal rabies virus challenge using short interfering RNAs (siRNAs) delivered through lentiviral vector.

The antiviral potential of small interfering RNAs (siRNAs) targeting rabies virus (RV) polymerase (L) and nucleoprotein (N) genes delivered through lentiviral vector was investigated. For in vitro evaluation, siRNAs expressing BHK-21 cell lines (BHK-L and BHK-N) were developed using transduction with Lenti-L and Lenti-N lentiviruses encoding siRNAs against RV-L and N genes, respectively. When these cell lines were challenged in vitro with RV Pasteur virus-11 (PV-11) strain, there was reduction in number of RV-specific foci and target gene transcripts indicating inhibitory effect on RV multiplication. For in vivo evaluation, mice were treated intracerebrally with lentiviruses and challenged with 20 LD50 of RV challenge virus standard-11 (CVS-11) strain by intramuscular route in masseter muscle. Five out of eight mice treated with Lenti-N survived indicating 62.5 % protection. The control and Lenti-L-treated mice died within 7-10 days indicating lethal nature of challenge virus and no protection. These results demonstrated that siRNA targeting RV-N could not only inhibit RV multiplication, but also conferred protection in mice against lethal RV challenge. These findings have implication on therapeutic use of siRNA targeting RV-N against RV infection.

Evaluation of single and dual siRNAs targeting rabies virus glycoprotein and nucleoprotein genes for inhibition of virus multiplication in vitro.

Small interfering RNAs (siRNAs) targeting rabies virus (RV) glycoprotein (G) and nucleoprotein (N) genes were evaluated as antiviral agents against rabies virus in vitro in BHK-21 cells. To select effective siRNAs targeting RV-G, a plasmid-based transient co-transfection approach was used. In this, siRNAs were expressed as short hairpin RNAs (shRNAs), and their ability to inhibit RV-G gene expression was evaluated in cells transfected with a plasmid expressing RV-G. The nine different siRNAs designed to target RV-G exhibited varying degrees of knockdown of RV-G gene expression. One siRNA (si-G7) with considerable effect in knockdown of RV-G expression also demonstrated significant inhibition of RV multiplication in BHK-21 cells after in vitro challenge with the RV Pasteur virus-11 (PV-11) strain. A decrease in the number of fluorescent foci in siRNA-treated cells and a reduction (86.8 %) in the release of RV into infected cell culture supernatant indicated the anti-rabies potential of siRNA. Similarly, treatment with one siRNA targeting RV-N resulted in a decrease in the number of fluorescent foci and a reduction (85.9 %) in the release of RV. As a dual gene silencing approach where siRNAs targeting RV-G and RV-N genes were expressed from single construct, the anti-rabies-virus effect was observed as an 87.4 % reduction in the release of RV. These results demonstrate that siRNAs targeting RV-G and N, both in single and dual form, have potential as antiviral agent against rabies.

Mechanochemical synthesis of an yttrium based metal-organic framework.

For the first time a metal hydride has been used for the preparation of a metal-organic framework. MIL-78 has been synthesized by the solid-state mechanochemical reaction between yttrium hydride and trimesic acid. The process does not involve solvents and does not generate liquid by-products, thus proving the viability of the solid-state approach to the synthesis of MOFs.