Structural investigation of the capsular polysaccharide of Klebsiella serotype k23

Klebsiella K23 capsular polysaccharide has been investigated by the techniques of hydrolysis, methylation, Smith degradation-periodate oxidation, and base-catalysed degradation, either on the original or the carboxyl-reduced polysaccharide. The structure was found to consist of a tetrasaccharide repeating-unit, as shown below. The anomeric configurations of the sugar residues were determined by ₁H-and ¹³C-n.m.r. spectroscopy on the original and degraded polysaccharides.

     

Endocytosis,digestive vacuolar movement and exocytosis on refeeding starvedTetrahymena pyriformis GL-9

Summary Evidence is presented in support of the hypothesis that the contents of digestive vacuoles in refed starvedTetrahymena pyriformis GL-9 are egested from the cell in approximately the sequence of their order of formation. The investigations involved measurements of the rates of disappearance of digestive vacuoles from the cells and the subsequent appearance of egested globules in the surrounding medium using both cultures and individual cells. The cells were first fed peptone and latex particles for a period and then this type of vacuole formation was suppressed by the addition of excess carmine particles (or the process was repeated with the particles in reverse order). Thus two types of morphologically distinct digestive vacuoles could be produced and observed microscopically. These observations suggest that the temporal nature of the movement of the digestive vacuoles through the cell result in the temporal nature of egestion and that no selective mechanism occurs at egestion. Thus digestive vacuoles are thought to pass through the cell from cytopharynx to cytoproct in approximately the order formed and at approximately constant rate. Under conditions of excess nutrients, where the cells become filled with digestive vacuoles, they seem to be able to maintain an approximately uniform number of digestive vacuoles within themselves by maintaining approximately constant and equal rates of vacuole formation and egestion. The maximum rates of latex or carmine vacuole formation or egestion found in single cells were approximately 0.3–0.4 vacuoles per cell per minute. The results are discussed.     

Citrate lyase from Streptococcus diacetilactis

Citrate lyase (EC 4.1.3.6) was purified 38-fold from cell-free extracts of Streptococcus diacetilactis. The enzyme was homogeneous in analytical ultracentrifugation and polyacrylamide gel electrophoresis The final enzyme preparation contained acetate: HS-citrate lyase ligase—an acetylating enzyme which converts inactive HS-citrate lyase into enzymatically active acetyl-S-citrate lyase. This enzyme activity was purified 25-fold over the crude extract and seemed to be associated with citrate lyase. Partially purified citrate lyase from Leuconostoc citrovorum contained also its acetylating enzyme. Purified citrate lyases from Klebsiella aerogenes and Rhodopseudomonas gelatinosa were devoid of acetylating enzyme activity. The HS-form of citrate lyase from S. diacetilactis was completely acetylated and hence activated by incubation with ATP and acetate for 25 min at 25° C. The enzyme did not acetylate the HS-lyases from R. gelatinosa and K. aerogenes. In contrast to the citrate lyases from R. gelatinosa and K. aerogenes the enzymes from S. diacetilactis and L. citrovorum showed onlya very weak reaction inactivation. It is assumed that this is due to the association of the acetylating enzymes with these lyases.     

Human brucellosis: Widespread information deficiency hinders an understanding of global disease frequency

BackgroundFor decades, human brucellosis has been recognized worldwide as a significant cause of morbidity, yet the annual incidence of this disease remains unknown. We analyzed this frequency, using international reports (2005–2019), identifying information gaps, and distinguishing a possible path forward.Methodology/Principal findingsA novel approach to estimating the incidence of this disease was explored. We utilized annual health data extracted from the World Organization for Animal Health (OIE)–World Animal Health Information System (WAHIS) database, assessing the dataset completeness and representativeness of the data for the world population. Additionally, we assessed the reported country level human brucellosis case counts and the factors that influenced the observed changes over time. Our analysis revealed incomplete and unrepresentative information, preventing the estimation of annual human brucellosis case incidence at the global level. In the OIE-WAHIS database, only 48.4% of the required reports have been submitted as of 2019, with approximately 47.3% of the world population represented. Additionally, geographic regions were disproportionate in completeness, representativeness, and actual reported case counts. Africa and Asia constituted the majority of reported cases, while simultaneously submitting the lowest percentage of reports as well as covering the lowest percentage of their populations within those reports, when compared to the rest of the world.Conclusions/SignificanceThe global annual frequency of human brucellosis cases remains elusive. Furthermore, there exists great heterogeneity in diagnostic, surveillance, and reporting systems worldwide, calling into question the validity of available information. This study reveals that the Neglected Zoonotic Disease priority status for brucellosis should be restored.     

The insulin receptor family in the heart: new light on old insights

Insulin was discovered over 100 years ago. Whilst the first half century defined many of the physiological effects of insulin, the second emphasised the mechanisms by which it elicits these effects, implicating a vast array of G proteins and their regulators, lipid and protein kinases and counteracting phosphatases, and more. Potential growth-promoting and protective effects of insulin on the heart emerged from studies of carbohydrate metabolism in the 1960s, but the insulin receptors (and the related receptor for insulin-like growth factors 1 and 2) were not defined until the 1980s. A related third receptor, the insulin receptor-related receptor remained an orphan receptor for many years until it was identified as an alkali-sensor. The mechanisms by which these receptors and the plethora of downstream signalling molecules confer cardioprotection remain elusive. Here, we review important aspects of the effects of the three insulin receptor family members in the heart. Metabolic studies are set in the context of what is now known of insulin receptor family signalling and the role of protein kinase B (PKB or Akt), and the relationship between this and cardiomyocyte survival versus death is discussed. PKB/Akt phosphorylates numerous substrates with potential for cardioprotection in the contractile cardiomyocytes and cardiac non-myocytes. Our overall conclusion is that the effects of insulin on glucose metabolism that were initially identified remain highly pertinent in managing cardiomyocyte energetics and preservation of function. This alone provides a high level of cardioprotection in the face of pathophysiological stressors such as ischaemia and myocardial infarction.     

The Role of Influenza A Virus Hemagglutinin Residues 226 and 228 in Receptor Specificity and Host Range Restriction

Influenza A viruses can be isolated from a variety of animals, but their range of hosts is restricted. For example, human influenza viruses do not replicate in duck intestine, the major replication site of avian viruses in ducks. Although amino acids at positions 226 and 228 of hemagglutinin (HA) of the H3 subtype are known to be important for this host range restriction, the contributions of specific amino acids at these positions to restriction were not known. Here, we address this issue by generating HAs with site-specific mutations of a human virus that contain different amino acid residues at these positions. We also let ducks select replication-competent viruses from a replication-incompetent virus containing a human virus HA by inoculating animals with 10^10.5 50% egg infectious dose of the latter virus and identified a mutation in the HA. Our results showed that the Ser-to-Gly mutation at position 228, in addition to the Leu-to-Gln mutation at position 226 of the HA of the H3 subtype, is critical for human virus HA to support virus replication in duck intestine.     

Phytochemical composition of Potentilla anserina L. analyzed by an integrative GC-MS and LC-MS metabolomics platform

Potentilla anserina L. (Rosaceae) is known for its beneficial effects of prevention of pre-menstrual syndrome (PMS). For this reason P. anserina is processed into many food supplements and pharmaceutical preparations. Here we analyzed hydroalcoholic reference extracts and compared them with various extracts of different pharmacies using an integrative metabolomics platform comprising GC-MS and LC-MS analysis and software toolboxes for data alignment (MetMAX Beta 1.0) and multivariate statistical analysis (COVAIN 1.0). Multivariate statistics of the integrated GC-MS and LC-MS data showed strong differences between the different plant extract formulations. Different groups of compounds such as chlorogenic acid, kaempferol 3-O-rutinoside, acacetin 7-O-rutinoside, and genistein were reported for the first time in this species. The typical fragmentation pathway of the isoflavone genistein confirmed the identification of this active compound that was present with different abundances in all the extracts analyzed. As a result we have revealed that different extraction procedures from different vendors produce different chemical compositions, e.g. different genistein concentrations. Consequently, the treatment may have different effects. The integrative metabolomics platform provides the highest resolution of the phytochemical composition and a mean to define subtle differences in plant extract formulations.     

Cytoskeletal scaffolding proteins interact with Lynch‐Syndrome associated mismatch repair protein MLH1

The involvement of MLH1 in several mismatch repair‐independent cellular processes has been reported. In an attempt to gain further insight into the protein's cellular functions, we screened for novel interacting partners of MLH1 utilizing a bacterial two‐hybrid system. Numerous unknown interacting proteins were identified, suggesting novel biological roles of MLH1. The network of MLH1 and its partner proteins involves a multitude of cellular processes. Integration of our data with the “General Repository for Interaction Datasets” highlighted that MLH1 exhibits relationships to three interacting pairs of proteins involved in cytoskeletal and filament organization: Thymosin β 4 and Actin γ, Cathepsin B and Annexin A2 as well as Spectrin α and Desmin. Coimmunoprecipitation and colocalization experiments validated the interaction of MLH1 with these proteins. Differential mRNA levels of many of the identified proteins, detected by microarray analysis comparing MLH1‐deficient and ‐proficient cell lines, support the assumed interplay of MLH1 and the identified candidate proteins. By siRNA knock down of MLH1, we demonstrated the functional impact of MLH1–Actin interaction on filament organization and propose that dysregulation of MLH1 plays an essential role in cytoskeleton dynamics. Our data suggest novel roles of MLH1 in cellular organization and colorectal cancerogenesis.