Inactivation of Spores and Vegetative Forms of Clostridioides Difficile by Chemical Biocides: Mechanisms of Biocidal Activity, Methods of Evaluation, and Environmental Aspects

Clostridioides difficile infections (CDIs) are the most common cause of acquired diseases in hospitalized patients. Effective surface disinfection, focused on the inactivation of the spores of this pathogen, is a decisive factor in reducing the number of nosocomial cases of CDI infections. An efficient disinfection procedure is the result of both the properties of the biocidal agent used and the technology of its implementation as well as a reliable, experimental methodology for assessing the activity of the biocidal active substance based on laboratory models that adequately represent real clinical conditions. This study reviews the state of knowledge regarding the properties and biochemical basis of the action mechanisms of sporicidal substances, with emphasis on chlorine dioxide (ClO2). Among the analyzed biocides, in addition to ClO2, active chlorine, hydrogen peroxide, peracetic acid, and glutaraldehyde were characterized. Due to the relatively high sporicidal effectiveness and effective control of bacterial biofilm, as well as safety in a health and environmental context, the use of ClO2 is an attractive alternative in the control of nosocomial infections of CD etiology. In terms of the methods of assessing the biocidal effectiveness, suspension and carrier standards are discussed.

The SARS-CoV-2 nucleocapsid protein preferentially binds long and structured RNAs

The SARS-CoV-2 nucleocapsid protein (NCAP) functions in viral RNA genome packaging, virion assembly, RNA synthesis and translation, and regulation of host immune response. RNA-binding is central to these processes. Little is known how NCAP selects its binding partners in the myriad of host and viral RNAs. To address this fundamental question, we employed electrophoresis mobility shift and competition assays to compare NCAP binding to RNAs that are of SARS-CoV-2 vs. non-SARS-CoV-2, long vs. short, and structured vs. unstructured. We found that although NCAP can bind all RNAs tested, it primarily binds structured RNAs, and their association suppresses strong interaction with single-stranded RNAs. NCAP prefers long RNAs, especially those containing multiple structures separated by single-stranded linkers that presumably offer conformational flexibility. Additionally, all three major regions of NCAP bind RNA, including the low complexity domain and dimerization domain that promote formation of NCAP oligomers, amyloid fibrils and liquid-liquid phase separation. Combining these observations, we propose that NCAP-NCAP interactions that mediate higher-order structures during packaging also drive recognition of the genomic RNA and call this mechanism recognition-by-packaging. This study provides a biochemical basis for understanding the complex NCAP-RNA interactions in the viral life cycle and a broad range of similar biological processes.

Novel mechanistic insights into the role of Mer2 as the keystone of meiotic DNA break formation

In meiosis, DNA double strand break (DSB) formation by Spo11 initiates recombination and enables chromosome segregation. Numerous factors are required for Spo11 activity, and couple the DSB machinery to the development of a meiosis-specific “axis-tethered loop” chromosome organization. Through in vitro reconstitution and budding yeast genetics we here provide architectural insight into the DSB machinery by focussing on a foundational DSB factor, Mer2. We characterise the interaction of Mer2 with the histone reader Spp1, and show that Mer2 directly associates to nucleosomes, likely highlighting a contribution of Mer2 to tethering DSB factors to chromatin. We reveal the biochemical basis of Mer2 association with Hop1, a HORMA domain-containing chromosomal axis factor. Finally, we identify a conserved region within Mer2 crucial for DSB activity, and show that this region of Mer2 interacts with the DSB factor Mre11. In combination with previous work, we establish Mer2 as a keystone of the DSB machinery by bridging key protein complexes involved in the initiation of meiotic recombination.

Biological effects of high-diluted substances and periodic table of elements

Background and Aims. There are several experimental evidences for the effects of high-diluted substances (see e.g. C. Taddei-Ferretti, A. Cotugno 1997, on effects of high-diluted drugs on the prevention and control of mice teratogenicity induced by purine derivatives; N.C. Sukul, C. Taddei-Ferretti, S.P. Sinha Babu, A. De, B. Nandi, A. Sukul, R. Dutta-Nag 2000, on high-diluted Nux vomica countering alcohol-induced loss of righting reflex in toads). Also the physical characterization and mechanism of action of high-diluted drugs have been studied (see e.g. N.C. Sukul, A. Sukul, High dilution effects: Physical and biochemical basis 2004). However, further experimental researches are needed to clarify how physical characteristics of a drug are linked to its global biological effects. Considerations on some high-diluted mineral remedies will be developer here. Methods. In Organon, sect. 119, S. Hahnemann writes: «As certainly each species of plants is different from every other one with regard to external appearance, way of life and growth, taste and smell, and as certainly each mineral, each salt is different from the others with regard to external, internal, physical and chemical qualities [...], so certainly all these vegetal and mineral substances have pathogenetic – and thus also curative – effects different among themselves [...]». This statement may be taken as basis for considering the characteristics of some elements, as ordered in the periodic table, in relation to those of some high-diluted mineral remedies. Conclusions. The elements were previously ordered in the periodic table according to the atomic weight chemically determined, and later more precisely according to the atomic number (number of protons). Then also the electronic configuration was taken into account: properties depending on atomic mass and deep electrons are not periodical, while chemical and several physical properties are linked to external electrons which have periodical configuration. In particular, let us consider the group of elements C, P, S, Cl and the group of elements Ca, Mg, K, Na. One may conclude that the four elements of the first group (respectively receiver-or-donor of 4 electrons, receiver of 3, of 2, of 1 electron), which, according to H. Bernard, are linked to the fixed human constitutions, are close among themselves in the periodic table, while they are very distant from the four elements of the second group (respectively donor of 2, of 2, of 1, of 1 electron), which are close among themselves and are linked to the changing constitutional stages.

Second-Hand Risk Health Tragedies and Human Right Violations due to Ethanol Consumption. Biochemical basis for metabolic and behavioral Tragedies due to Ethanol Consumption

Everywhere a nonsmoker who is an alcohol consumer, complains of secondhand smoke, without being aware of second-hand risk health tragedies and human rights violations provoked by alcohol consumption. Here we analyze the concept, mainly unexplored, of dramatic adverse health effects and of human rights violations against third parties generated for alcohol consumption by others; and also, the harm due to the chemical transient prefrontal lobotomy generated by alcohol consumption. Alcohol consumption has been a part of the everyday human diet for centuries, especially because of the fact that alcoholic beverages are a safe means of hydration wherever clear water has not been available [1]. Old patients could, simultaneously be part of the alcohol consumers and/or secondhand victims. Before deciding to analyze the geriatric problems, we propose the allegorical model, based on Scott, Ellison, and Sinclair, as it was published in Nature Aging, in July 2021. We divide the theoretical analysis with four fundamental alternatives [2]: The extension of life (Struldbrug case). In Jonathan Swift's 1726 novel “Gulliver's Travels”, the struldbrugs were humans born apparently normal. The Struldbruggs, however immortal however they age normally, live in continuously deteriorating health. This takes us to the philosophical alternative of: “living or lasting” [2]. To lower morbidity (Dorian Gray case). Narratively, in “The Picture of Dorian Gray” a philosophical novel by Oscar Wilde, Dorian Gray owns a portrait of himself and while the picture ages, Dorian Gray does not change, maintaining his health and appearance until death [2]. Slowing aging (Peter Pan case), In this extreme case, where aging is not just slowed but canceled, the mortality and the health become independent of the age, and thus the individual is ‘forever young’. This constitutes the ‘Peter Pan’ case, after the play and novel about a boy who never grows old. This closely corresponds to the Hypocaloric diet claiming that it slows aging [2]. To reverse aging physical damage is repaired instead of slowed. This is a close analogy to the “Theseus Boat” as well as the regeneration of salamanders and lizards and transplants from donors. Desiderative, this is the future of organoids and the engineering of the pluripotent cell [2].

Lysosomal storage diseases. Mucopolysaccharidosis type III, sanfilippo syndrome

The review describes the clinical, biochemical and molecular genetic characteristics of autosomal recessive mucopolysaccharidosis type III, or Sanfilippo syndrome. This is a genetically heterogeneous group of rare, but similar in nature, diseases caused by a deficiency of one of the four lysosomal enzymes involved in the degradation of heparan sulfate. All types of mucopolysaccharidosis III are characterized by severe degeneration of the central nervous system in combination with mild somatic manifestations, which is explained by the accumulation of high concentrations of heparan sulfate in the lysosomes of various cells, including the central nervous system. The primary biochemical defect in the most common type of mucopolysaccharidosis IIIA, occurring with a frequency of 1 : 105 and presented in 60% of all cases of the disease, is heparan-N-sulfatase, or sulfamidase deficiency. Mucopolysaccharidosis IIIB type occurs twice less often and accounts for about 30% of all cases of Sanfilippo syndrome. It is caused by the presence of inactivating mutations in the lysosomal -N-acetylglucosaminidase gene. Mucopolysaccharidosis IIIC and IIID are 4% and 6%, and occur at frequencies of 0.7 and 1.0 : 106. Mucopolysaccharidosis IIIC is caused by inactivating mutations in the gene of membrane-bound lysosomal acetyl-CoA:-glucosaminid-N-acetyltransferase, or N-acetyltransferase. Mucopolysaccharidosis IIID is based on the deficiency of lysosomal N-acetylglucosamine-6-sulfatase. The role of experimental models in the study of the biochemical basis of the pathogenesis of Sanfilippo syndrome and the development of various therapeutic approaches are discussed. The possibility of neonatal screening, early diagnosis, prevention and pathogenetic therapy of these severe lysosomal diseases are considered. As an example, a clinical case of diagnosis and treatment of a child with type IIIB mucopolysaccharidosis is presented.

Structural and biochemical basis of a marine bacterial glycoside hydrolase family 2 β-glycosidase with broad substrate specificity

Uronic acids are commonly found in marine polysaccharides and increase structural complexity sanand intrinsic recalcitrance to enzymatic attack. The glycoside hydrolase family 2 (GH2) include proteins that target sugar conjugates with hexuronates and are involved in the catabolism and cycling of marine polysaccharides. Here, we reported a novel GH2, Aq GalA from a marine algae-associated Bacteroidetes with broad-substrate specificity. Biochemical analyses revealed that Aq GalA exhibits hydrolyzing activities against β-galacturonide, β-glucuronide, and β-galactopyranoside via retaining mechanisms. We solved the Aq GalA crystal structure in complex with galacturonic acid (GalA) and showed (via mutagenesis) that charge characteristics at uronate-binding subsites controlled substrate selectivity for uronide hydrolysis. Additionally, conformational flexibility of the Aq GalA active site pocket was proposed as a key component for broad substrate enzyme selectivity. Our Aq GalA structural and functional data augments the current understanding of substrate recognition of GH2 enzymes and provided key insights into the bacterial use of uronic acid containing polysaccharides. IMPORTANCE The decomposition of algal glycans driven by marine bacterial communities represents one of the largest heterotrophic transformation of organic matter fueling marine food webs and global carbon cycling. However, our knowledge of the carbohydrate cycling is limited due to structural complexity of marine polysaccharides and the complicated enzymatic machinery of marine microbes. To degrade algal glycan, marine bacteria such as members of Bacteroidetes produce a complex repertoire of carbohydrate-active enzymes (CAZymes) matching the structural specificity of the different carbohydrates. In this study, we investigated an extracellular GH2 β-glycosidase, Aq GalA from a marine Bacteroidetes to identify the key components responsible for glycuronides recognition and hydrolysis. The broad substrate specificity of Aq GalA against glycosides with diverse stereochemical substitutions indicates its potential in processing complex marine polysaccharides. Our findings promote a better understanding of microbially-driven mechanisms of marine carbohydrate cycling.

Biochemical Response to Freezing in the Siberian Salamander Salamandrella keyserlingii

The Siberian salamander Salamandrella keyserlingii Dybowski, 1870 is a unique amphibian that is capable to survive long-term freezing at −55 °C. Nothing is known on the biochemical basis of this remarkable freezing tolerance, except for the fact that it uses glycerol as a low molecular weight cryoprotectant. We used 1H-NMR analysis to study quantitative changes of multiple metabolites in liver and hindlimb muscle of S. keyserlingii in response to freezing. For the majority of molecules we observed significant changes in concentrations. Glycerol content in frozen organs was as high as 2% w/w, which confirms its role as a cryoprotectant. No other putative cryoprotectants were detected. Freezing resulted in ischemia manifested as increased concentrations of glycolysis products: lactate and alanine. Unexpectedly, we detected no increase in concentrations of succinate, which accumulates under ischemia in various tetrapods. Freezing proved to be a dramatic stress with reduced adenosine phosphate pool and high levels of nucleotide degradation products (hypoxanthine, β-alanine, and β-aminoisobutyrate). There was also significant increase in the concentrations of choline and glycerophosphocholine, which may be interpreted as the degradation of biomembranes. Thus, we found that freezing results not only in macroscopical damage due to ice formation, but also to degradation of DNA and biomembranes.

Mechanisms Causing Aging, Current Knowledge and the Way Forward

The rapid advancement in research technologies and bioinformatics over the past few decades has enabled researchers to shed light on the underlying mechanisms behind aging. Whilst the progress in understanding the biochemical processes involved is impressive, a lot more still needs to be uncovered before any potential effective anti-aging treatment can be produced. Unravelling the various root causes of aging is still the most important obstacle to overcome. The data available highlights that the most likely drivers of aging are the proteosome, the ribosome and telomeres. This review focuses largely on these factors and how they contribute to initiating aging and their targeting in potential therapy against the multitude of age-associated disorders. The investigation thus far of these causative factors will be presented. Understanding these root causes and how they cause aging is fundamental to present a way forward, such that the biochemical basis of aging can be discovered, in order to usher in a new wave of therapeutics against complex diseases.