Investigation on Fermented Milk Quality after the Addition of Flaxseed Mucilage and the Use of Lactobacillus delbrueckii subsp. bulgaricus and Lactiplantibacillus plantarum AG9.

BACKGROUND: Flaxseed mucilage (FSM) is one of the healthy components of flaxseed. FSM is an example of a material that can be used in the food, cosmetic, and pharmaceutical industries due to its rheological properties. FSM consists mainly of two polysaccharides, arabinoxylan, and rhamnogalacturonan I, and it also contains protein components and minerals. The prospect of using FSM in food is due to its gelling, water binding, emulsifying, and foaming properties. In addition, valuable natural sources of phenolic compounds such as lignans, phenolic acids, flavonoids, phenylpropanoids, and tannins are partially extracted from flaxseed in FSM. These antioxidant components have pharmacological properties, including anti-diabetic, anti-hypertensive, immunomodulatory, anti-inflammatory and neuroprotective properties. A combination of FSM and lactobacilli in dairy foods can improve their functional properties. This study aimed to develop dairy products by adding of FSM and using two lactic acid bacteria (LAB). FSM (0.2%) was used as an ingredient to improve both the texture and antioxidant properties of the product. METHODS: Skim milk was fermented with 0.2% flaxseed mucilage using Lactobacillus delbrueckii subs. bulgaricus and the probiotic Lactiplantibacillus plantarum AG9. The finished fermented milk products were stored at 4 °C for 14 days. Quantitative chemical, textural, and antioxidant analyses were carried out. RESULTS: Adding 0.2% FSM to the dairy product stimulated the synthesis of lactic acid. FSM increased the viscosity and water-holding capacity of L. bulgaricus or L. bulgaricus/L. plantarum AG9 fermented milk products. Combining these starter strains with FSM promoted the formation of a hard, elastic, resilient casein matrix in the product. When only L. plantarum AG9 was used for the fermentation, the dairy product had a high syneresis and a low viscosity and firmness; such a product is inferior in textural characteristics to the variant with commercial L. bulgaricus. The addition of FSM improved the textural properties of this variant. The use of L. plantarum AG9 and FSM makes it possible to obtain a fermented milk product with the highest content of polyphenolic compounds, which have the highest antioxidant properties and stimulate lipase and α-glucosidase inhibitor synthesis. Combining of L. bulgaricus and L. plantarum AG9 in the starter (20% of the total mass of the starter) and adding of 0.2% FSM is the optimal combination for obtaining a dairy product with high textural and antioxidant properties. CONCLUSIONS: The physicochemical properties (viscosity, syneresis, water holding capacity, texture) and antioxidant properties of fermented milk were improved. In the future, as part of the work to investigate the functional properties of dairy products with FSM, studies will be conducted using in in vivo models.

A Cybertaxonomic Revision of the "Crocidura pergrisea" Species Complex with a Special Focus on Endemic Rocky Shrews: Crocidura armenica and Crocidura arispa (Soricidae).

The extraction of museum DNA from a unique collection of samples of the "Crocidura pergrisea" species complex, which comprises local endemics of Central and West Asia, allowed us to determine their inter- and intragroup relationships. The first step of this study was the re-evaluation of heavily damaged type specimens of C. armenica via a microcomputed-tomography-based cybertaxonomic approach (CTtax), which enabled a precise description of the species' morphology; three-dimensional models of the cybertypes were made available through the MorphoBank Repository. We developed the "AProMaDesU" pipeline on the basis of five requirements for micro-CT-based cyber-datasets in relation to mammalian collections. Our second step was a combination of several meticulous approaches to morphological investigation against a background of a cytb-based phylogeny, which helped us to make a taxonomic decision about the status of species of the "pergrisea" group, e.g., C. arispa, C. armenica, and C. serezkyensis, when the morphological results were partly incongruent with the molecular phylogeny. Nevertheless, under two assumptions, our findings preserved a separate species-level status of C. serezkyensis and C. arispa. In addition, we restored the species-level status of C. armenica. This taxonomic decision is based on our morphospace analysis, which revealed unique craniomandibular shape transformations within the rocky shrews that helped them with the transition to a new area of morphospace/trophic niches and consequently separated them from the other analyzed Crocidura groups.

Biological effects and phytochemical study of the underground part of Iris scariosa Willd. ex Link extract: A new source of bioactive constituents.

The expected toxicity and resistance of chemotherapeutic agents necessitate and encourage for the use of natural chemotherapeutic sources of plant origin in the clinical stage of cancer therapy. Plants of the genus Iris (Iridaceae) used by local populations for the treatment of cancer, bacterial and viral infections. In this study, an ethanol extract of rhizomes of I. scariosa was prepared and tested for the cytotoxicity using the MTT assay. The extract exhibited the most potent cytotoxicity against the breast cancer cell line MCF7 (IC50 = 9.28 ± 0.49 µg/ml, selectively index ˃5), and induced apoptosis in MCF7 lines. Notably, the extract significantly inhibited the colony formation of MCF7 and HepG2 cancer cells at a concentration range from 10.6 to 85.0 µg/ml, including non-toxic concentrations for HepG2 cells. The ethanol extract was analyzed by HPLC, revealed the identification of 5 secondary metabolites (quercetin, rutin, myricetin, apigenin, artemisetin), the content of which was shown to reach around 15% of the extract. The petroleum ether (PE) part of the extract (yield 2.62%) was analyzed by GC-MS. The composition of tert-butyl methyl ether (TBME) part of the extract (yield 23.72%) was studied. Total of 15 individual compounds: two benzophenones, eight isoflavones, four flavones and a (2R)-flavanone were isolated. The pentamethoxyflavone artemisetin and flavanone pinocembrin were isolated for the first from Iris sp. The readily available isoflavones from the TBME part of extract (irilone, iriflogenin, irigenin and tectorigenin) may serve as new leads for the discovery of anticancer drugs.

The Complete Mitochondrial Genome of Dendrogale murina (Tupaiidae) and Phylogeny of Scandentia.

In this paper, we report the complete mitochondrial genome of the northern smooth-tailed treeshrew Dendrogale murina, which was sequenced for the first time using the Illumina next-generation sequencing (NGS) technology. The total length of the mitochondrial genome is 16,844-16,850 bp and encodes 37 genes, including two ribosomal RNAs (rRNAs) 12S and 16S, 22 transfer RNAs (tRNAs), 13 protein-coding genes (PCGs), and a D-loop in the characteristic arrangement of family Tupaiidae (Mammalia: Scandentia). The overall base composition of the complete mitochondrial DNA is A (33.5%), C (25.5%), G (13.9%), and T (27.1%). Phylogenetic analysis of Scandentia mitochondrial genomes showed a classic pattern, which was revealed previously while using individual phylogenetic markers. The result of the current study is consistent with one based on the latest morphological studies, with the basal position of Ptilocercus and Dendrogale sister to the rest of the Tupaiidae genera. The divergence time of the Dendrogale genus is estimated as Eocene-Oligocene, with the mean value of 35.8 MYA, and the Ptilocercus genus probably separated at about 46.3 MYA. We observe an increase in the age of all nodes within the Scandentia, except for a decrease in the age of separation of Ptilocercus. This result can be explained both by the addition of new mitochondrial genome data in the analysis and the usage of new calibration points from recently published data.

Tomato-made edible COVID-19 vaccine TOMAVAC induces neutralizing IgGs in the blood sera of mice and humans.

Plant-based edible vaccines that provide two-layered protection against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) outweigh the currently used parenteral types of vaccines, which predominantly cause a systemic immune response. Here, we engineered and selected a transgenic tomato genotype (TOMAVAC) that stably synthesized an antigenic S1 protein of SARS-CoV-2. Two-course spaced force-feeding of mice with ≈5.4 µg/ml TOMAVAC increased up to 16-fold the synthesis of RBD-specific NAbs in blood serum and the significant induction of S-IgA in intestinal lavage fluid. In a surrogate virus neutralization test, TOMAVAC-induced NAbs had 15-25% viral neutralizing activity. The results suggested early evidence of the immunogenicity and protectivity of TOMAVAC against the coronavirus disease 2019 (COVID-19) infection. Furthermore, we observed a positive trend of statistically significant 1.2-fold (average of +42.28 BAU/ml) weekly increase in NAbs in the volunteers' serum relative to the initial day. No severe side effects were observed, preliminarily supporting the safety of TOMAVAC. With the completion of future large-scale studies, higher-generation TOMAVAC should be a cost-effective, ecologically friendly, and widely applicable novel-generation COVID-19 vaccine, providing two-layered protection against SARS-CoV-2.

Signatures of Adaptation in Mitochondrial Genomes of Palearctic Subterranean Voles (Arvicolinae, Rodentia).

This study evaluates signatures of selection in the evolution of the mitochondrial DNA of voles, subfamily Arvicolinae, during the colonization of subterranean environments. The comparative sequence analysis of mitochondrial protein-coding genes of eight subterranean vole species (Prometheomys schaposchnikowi, three species of the genus Ellobius: Ellobius talpinus, Ellobius fuscocapillus and Ellobius lutescens, two species of the genus Terricola: Terricola subterraneus and Terricola daghestanicus, Lasiopodomys mandarinus, and Hyperacrius fertilis) and their closest aboveground relatives was applied using codon-substitution models. The highest number of selection signatures was detected in genes ATP8 and CYTB. The relaxation of selection was observed in most mitochondrial DNA protein-coding genes for subterranean species. The largest amount of relaxed genes is discovered in mole voles (genus Ellobius). The number of selection signatures was found to be independent of the evolutionary age of the lineage but fits the degree of specialization to the subterranean niche. The common trends of selective pressures were observed among the evolutionary ancient and highly specialized subterranean rodent families and phylogenetically young lineages of voles. It suggests that the signatures of adaptation in individual mitochondrial protein-coding genes associated with the colonization of the subterranean niche may appear within a rather short evolutionary timespan.

A mitochondrial genome phylogeny of voles and lemmings (Rodentia: Arvicolinae): Evolutionary and taxonomic implications.

Arvicolinae is one of the most impressive placental radiations with over 150 extant and numerous extinct species that emerged since the Miocene in the Northern Hemisphere. The phylogeny of Arvicolinae has been studied intensively for several decades using morphological and genetic methods. Here, we sequenced 30 new mitochondrial genomes to better understand the evolutionary relationships among the major tribes and genera within the subfamily. The phylogenetic and molecular dating analyses based on 11,391 bp concatenated alignment of protein-coding mitochondrial genes confirmed the monophyly of the subfamily. While Bayesian analysis provided a high resolution across the entire tree, Maximum Likelihood tree reconstruction showed weak support for the ordering of divergence and interrelationships of tribal level taxa within the most ancient radiation. Both the interrelationships among tribes Lagurini, Ellobiusini and Arvicolini, comprising the largest radiation and the position of the genus Dinaromys within it also remained unresolved. For the first time complex relationships between genus level taxa within the species-rich tribe Arvicolini received full resolution. Particularly Lemmiscus was robustly placed as sister to the snow voles Chionomys in the tribe Arvicolini in contrast with a long-held belief of its affinity with Lagurini. Molecular dating of the origin of Arvicolinae and early divergences obtained from the mitogenome data were consistent with fossil records. The mtDNA estimates for putative ancestors of the most genera within Arvicolini appeared to be much older than it was previously proposed in paleontological studies.

Searching for signatures of positive selection in cytochrome b gene associated with subterranean lifestyle in fast-evolving arvicolines (Arvicolinae, Cricetidae, Rodentia).

BACKGROUND: Mitochondrial genes encode proteins involved in oxidative phosphorylation. Variations in lifestyle and ecological niche can be directly reflected in metabolic performance. Subterranean rodents represent a good model for testing hypotheses on adaptive evolution driven by important ecological shifts. Voles and lemmings of the subfamily Arvicolinae (Rodentia: Cricetidae) provide a good example for studies of adaptive radiation. This is the youngest group within the order Rodentia showing the fastest rates of diversification, including the transition to the subterranean lifestyle in several phylogenetically independent lineages. RESULTS: We evaluated the signatures of selection in the mitochondrial cytochrome b (cytB) gene in 62 Arvicolinae species characterized by either subterranean or surface-dwelling lifestyle by assessing amino acid sequence variation, exploring the functional consequences of the observed variation in the tertiary protein structure, and estimating selection pressure. Our analysis revealed that: (1) three of the convergent amino acid substitutions were found among phylogenetically distant subterranean species and (2) these substitutions may have an influence on the protein complex structure, (3) cytB showed an increased ω and evidence of relaxed selection in subterranean lineages, relative to non-subterranean, and (4) eight protein domains possess increased nonsynonymous substitutions ratio in subterranean species. CONCLUSIONS: Our study provides insights into the adaptive evolution of the cytochrome b gene in the Arvicolinae subfamily and its potential implications in the molecular mechanism of adaptation. We present a framework for future characterizations of the impact of specific mutations on the function, physiology, and interactions of the mtDNA-encoded proteins involved in oxidative phosphorylation.

Phylogenetic relationships and taxonomic position of genus Hyperacrius (Rodentia: Arvicolinae) from Kashmir based on evidences from analysis of mitochondrial genome and study of skull morphology.

In this article, we present the nearly complete mitochondrial genome of the Subalpine Kashmir vole Hyperacrius fertilis (Arvicolinae, Cricetidae, Rodentia), assembled using data from Illumina next-generation sequencing (NGS) of the DNA from a century-old museum specimen. De novo assembly consisted of 16,341 bp and included all mitogenome protein-coding genes as well as 12S and 16S RNAs, tRNAs and D-loop. Using the alignment of protein-coding genes of 14 previously published Arvicolini tribe mitogenomes, seven Clethrionomyini mitogenomes, and also Ondatra and Dicrostonyx outgroups, we conducted phylogenetic reconstructions based on a dataset of 13 protein-coding genes (PCGs) under maximum likelihood and Bayesian inference. Phylogenetic analyses robustly supported the phylogenetic position of this species within the tribe Arvicolini. Among the Arvicolini, Hyperacrius represents one of the early-diverged lineages. This result of phylogenetic analysis altered the conventional view on phylogenetic relatedness between Hyperacrius and Alticola and prompted the revision of morphological characters underlying the former assumption. Morphological analysis performed here confirmed molecular data and provided additional evidence for taxonomic replacement of the genus Hyperacrius from the tribe Clethrionomyini to the tribe Arvicolini.

Influence of Genipin Crosslinking on the Properties of Chitosan-Based Films.

Chitosan is a promising environment friendly active polymer packaging material due to its biodegradability, exceptional film forming capacity, great mechanical strength, appropriate barrier property along with intrinsic antioxidant and antimicrobial features. Bifunctional reagent was used for producing water insoluble chitosan films. Biopolymeric films crosslinked by Genipin (Gp), which is a reagent of natural origin, should have high potential in food packaging. The influence of the ratio of functional groups in the chitosan-Gp system on film absorption in the visible and ultraviolet regions of the spectrum, sorption, physical, and mechanical properties of the films has been studied. The degree of chitosan crosslinking in the films obtained from solutions containing Gp was estimated using the experimental data on film swelling and water vapor sorption isotherms. It is demonstrated that crosslinking with genipin improves swelling, water resistance, and mechanical properties of the films.

Chromones and coumarins from Saposhnikovia divaricata (Turcz.) Schischk. Growing in Buryatia and Mongolia and their cytotoxicity.

ETHNOPHARMACOLOGICAL RELEVANCE: Saposhnikovia divaricata (family Apiaceae) a traditional medicinal plant distributed in many provinces of China, is well known for the pharmaceutical value and has been used for rheumatic arthritis, and anxiety in children. Antiviral, antioxidant and antiproliferative activities were also mentioned. The application of this plant are recorded in the Chinese Medicine (CM) classical text the Shen Nong's Materia Medica (Shen Nong Ben Cao Jing). In this monograph S. divaricata (syn Radix Ledebouriella divaricata) is graded as a premium-grade herb, with their broad-spectrum of therapeutic applications for the treatment of cough, common cold, arthralgia, as well as in rheumatic disorders. AIM OF THE STUDY: To isolate and identify chemical constituents (chromones and coumarins) from S. divaricata, collected in Buryatia and Mongolia and to study their in vitro anticancer activity against MEL-8, U-937, DU-145, MDA-MB-231 and ВТ-474 cell lines. MATERIALS AND METHODS: An 40% aqueous ethanol extract of the roots of S. divaricata was prepared and further successively fractionated by extraction with petroleum ether, diethyl ether, tert-butyl methyl ether and ethyl acetate. The obtained extracts were subjected to a series of chromatographic separations on silica gel for isolation of individual compounds. Isolated compounds were tested for their cytotoxicity with respect to model cancer cell lines using the conventional MTT assays. RESULTS: Total of 15 individual compounds: coumarins scopoletin 2, bergapten 3, isoimperatorin 4, marmesin 5, (+)-decursinol 9, (-)-praeruptorin B 10, oxypeucedanin hydrate 11, chromones: hamaudol 6, cimifugin 7, 5-О-methylvisamminol 8, chromone glycosides: prim-O-glucosylcimifugin 12, sec-O-glucosylhamaudol 13, 4'-O-ß-D-glucopyranosyl-5-О-methylvisamminol 14, 4'-O-ß-D-glucopyranosylvisamminol (15) and also polyyne compound panaxinol 1 were isolated and characterized. The structure of dihydropyranocoumarin 10 was confirmed by X-ray diffraction analyses. HPLC-UV method was used for determination of the content of most abundant chromones 7, 12 and 14 in the roots of S. divaricata, collected in Mongolia. Compounds 3-11 and 13, 14 were evaluated for their cytotoxicity with respect to model cancer cell lines. All the compounds were non-toxic in the hemolysis test. CONCLUSION: This report about the phytochemical profiles of S. divaricata growing in Mongolia and Buryatia led to the identification of 14 compounds including coumarins and chromones. The available coumarins and chromones may serve as new leads for the discovery of anticancer drugs.

The complete mitochondrial genomes of three Ellobius mole vole species (Rodentia: Arvicolinae).

The subterranean voles of the genus Ellobius are species of subfamily Arvicolinae well adapted to underground life. In this paper, we report the assemblies of complete mitochondrial genomes for three mole voles from genus Ellobius - northern mole vole Ellobius talpinus (16,376 bp), transcaucasian mole vole E. lutescens (16,540 bp), and southern mole vole E. fuscocapillus (16,388 bp). Each of three mitogenomes encode for 12S and 16S rRNAs, 22 tRNAs, 13 protein-coding genes, and D-loop in the characteristic arrangement of subfamily Arvicolinae (Rodentia: Cricetidae). This study verifies the evolutionary status of subgenera Bramus and Ellobius within the genus Ellobius at the molecular level. The mitochondrial genome would be a significant supplement for the Ellobius genetic background. The three Ellobius species formed a monophyletic group with the high bootstrap value (100%) in all examinations.

Asymptotic solution for the problem of sound propagation in a sea with an underwater canyon.

The problem of the sound propagation in shallow-water waveguide with a seabottom featuring canyon-type inhomogeneity of a specific form is considered. The sound pressure in such waveguide is represented in a form of modal expansion and the equations for modal coefficients are derived. In case of a single-mode adiabatic propagation, it is possible to neglect the mode interaction and omit the coupling terms in these equations. The uncoupled equation for the mode amplitude admits an explicit analytical solution via the separation of variables.