Molecular evolution and structure-function relationships of myoglobin in diving mammals

Remarkable feats of breath-hold endurance are observed in diving mammals, with some species routinely diving for an hour. During most mammalian dives metabolism remains aerobic in nature, which is accomplished by restricting the blood flow to parts of the body through peripheral vaso-constriction and bradycardia. This mechanism preserves essential oxygen (02) for heart and brain function, but also means some parts of the body, including locomotory muscles, become isolated and have to rely on O2 stored within the tissues. Due to the isolation of skeletal muscles, mammalian divers must be able to buffer large quantities of H+ ions due to the production of CO2 during aerobic metabolism and acidic end products of anaerobic metabolism once muscle O2 stores have been consumed. The protein responsible for storing molecular 02 is myoglobin (Mb), a small 17 kDa monomeric globular haemoprotein with the primary function of reversible O2 binding and facilitated diffusion of O2 to the mitochondria. A hallmark of mammalian divers is increased Mb concentrations ([Mb]. with divers exhibiting concentrations up to thirty times those seen in non-diving species. Previous research has found that proteins at high concentrations are prone to form aggregations leading to non-functioning protein. This raises the question of Mb solubility at such high concentrations as observed in mammalian divers. The central hypothesis of this thesis is that mammalian myoglobin has undergone previously unrecognised, parallel and adaptive evolution in several lineages of mammalian divers that has profoundly increased their maximal physiological diving capacity. To test the hypothesis, Mb amino acid sequences of 124 mammals, including 24 newly determined sequences, are analysed for the content and individual buffering properties of their ionisable amino acids. This is used to calculate the specific Mb buffer value (~Mb) for each species, which is experimentally verified by acid-base titration of purified Mb. Together with known [Mb], the contribution of Mb to whole muscle buffer capacity WmuscleMb) is then quantified. Amino acid sequences are assessed for substitutions that increase modelled net Mb charge in mammalian divers compared to terrestrial species and predictions are confirmed by measuring electrophoretic mobility of purified Mb. Observed changes in Mb buffer properties and net surface charge are mapped on a composite mammalian phylogeny to test whether they are significantly linked to the evolution of diving behaviour. Observed molecular changes in Mb amino acid sequence are integrated with diving capacity, producing a model that allows prediction of maximal dive duration from Mb amino acid sequence and body mass. Using ancestral Mb sequence reconstructions and body mass estimates, the model is applied to infer the evolution of maximal diving capacity in the cetacean lineage. Results suggest a general trend towards increasing ~Mb due to increased Mb histidine content in mammalian divers.βmuscleMb is significantly higher in divers compared to terrestrial species, and can account for up to 45%, of the increase in whole muscle buffering observed in diving mammals. This study shows a remarkable trend in all diving species to significantly increase the net charge of the Mb protein, which would convey increased Mb solubility. This is supported by a significant correlation between Mb net charge and maximal [Mb]. Evolutionary analysis shows that high Mb net charge is significantly linked to the occurrence of diving. Contrary to previous findings, the model developed here finds that increases in [Mb] convey greater increases in dive duration than similar increases in body mass. This study provides novel in sights into how cumulative substitutions on the molecular surface of Mb can have profound adaptive effects on the physiological properties conveyed to the whole animal. It suggests that adaptation to diving in multiple lineages of mammals involved not only evolution of increased expression levels of Mb, but also substantial qualitative changes to the protein to avoid aggregation and increase solubility and buffering

Potential Human Health Implications of Swine Health

Thrs study measured the relationship between subclinical pig health at slaughter and carcass contamination. 280 randomly selected carcasses were swabbed at three points during slaughter: skin pre-scald; pelvic cavity following removal of the distal colon and rectum, and pleural cavity, immediately before the final carcass rinse. Swabs were cultured quantitatively for Campylobacter spp. and Enterococcus spp. Campylobacter spp. were recovered from the pleural cavity in 58.9% (33/56) and 44 .6% (25/56) of pools from the bung cavity Enterococcus spp. were recovered from 66.1% (37/56) and 38.7% (22/56) of pleural and bung samples, respectively. The most common lesion cdentified was pleuritis/adhesions, with a total of 7.1% (186/2,625 total head). Linear regress1on showed that for every percentage point increase in lesions, there was a significant 4.4% increase in Enterococcus spp. and 5.1% increase 1n Campylobacter spp. contamination. Additionally, significant relationships were identified between pleuritis and the quantity (log CFU) of Enterococcus spp. present in the bung cavity or Campylobacter spp. in the pleural cavity

Analysis of the FGF gene family provides insights into aquatic adaptation in cetaceans

Cetacean body structure and physiology exhibit dramatic adaptations to their aquatic environment. Fibroblast growth factors (FGFs) are a family of essential factors that regulate animal development and physiology; however, their role in cetacean evolution is not clearly understood. Here, we sequenced the fin whale genome and analysed FGFs from 8 cetaceans. FGF22, a hair follicle-enriched gene, exhibited pseudogenization, indicating that the function of this gene is no longer necessary in cetaceans that have lost most of their body hair. An evolutionary analysis revealed signatures of positive selection for FGF3 and FGF11, genes related to ear and tooth development and hypoxia, respectively. We found a D203G substitution in cetacean FGF9, which was predicted to affect FGF9 homodimerization, suggesting that this gene plays a role in the acquisition of rigid flippers for efficient manoeuvring. Cetaceans utilize low bone density as a buoyancy control mechanism, but the underlying genes are not known. We found that the expression of FGF23, a gene associated with reduced bone density, is greatly increased in the cetacean liver under hypoxic conditions, thus implicating FGF23 in low bone density in cetaceans. Altogether, our results provide novel insights into the roles of FGFs in cetacean adaptation to the aquatic environment.ope

Gastrointestinal Carriage Is a Major Reservoir of Klebsiella pneumoniae Infection in Intensive Care Patients.

BACKGROUND: Klebsiella pneumoniae is an opportunistic pathogen and leading cause of hospital-associated infections. Intensive care unit (ICU) patients are particularly at risk. Klebsiella pneumoniae is part of the healthy human microbiome, providing a potential reservoir for infection. However, the frequency of gut colonization and its contribution to infections are not well characterized. METHODS: We conducted a 1-year prospective cohort study in which 498 ICU patients were screened for rectal and throat carriage of K. pneumoniae shortly after admission. Klebsiella pneumoniae isolated from screening swabs and clinical diagnostic samples were characterized using whole genome sequencing and combined with epidemiological data to identify likely transmission events. RESULTS: Klebsiella pneumoniae carriage frequencies were estimated at 6% (95% confidence interval [CI], 3%-8%) among ICU patients admitted direct from the community, and 19% (95% CI, 14%-51%) among those with recent healthcare contact. Gut colonization on admission was significantly associated with subsequent infection (infection risk 16% vs 3%, odds ratio [OR] = 6.9, P < .001), and genome data indicated matching carriage and infection isolates in 80% of isolate pairs. Five likely transmission chains were identified, responsible for 12% of K. pneumoniae infections in ICU. In sum, 49% of K. pneumoniae infections were caused by the patients' own unique strain, and 48% of screened patients with infections were positive for prior colonization. CONCLUSIONS: These data confirm K. pneumoniae colonization is a significant risk factor for infection in ICU, and indicate ~50% of K. pneumoniae infections result from patients' own microbiota. Screening for colonization on admission could limit risk of infection in the colonized patient and others

Antimicrobial resistant <i>Klebsiella pneumoniae</i> carriage and infection in specialized geriatric care wards linked to acquisition in the referring hospital

AbstractBackgroundKlebsiella pneumoniae is a leading cause of extended-spectrum beta-lactamase (ESBL) producing hospital-associated infections, for which elderly patients are at increased risk.MethodsWe conducted a 1-year prospective cohort study, in which a third of patients admitted to two geriatric wards in a specialized hospital were recruited and screened for carriage of K. pneumoniae by microbiological culture. Clinical isolates were monitored via the hospital laboratory. Colonizing and clinical isolates were subjected to whole genome sequencing and antimicrobial susceptibility testing.ResultsK. pneumoniae throat carriage prevalence was 4.1%, rectal carriage 10.8% and ESBL carriage 1.7%. K. pneumoniae infection incidence was 1.2%. The isolates were diverse, and most patients were colonized or infected with a unique phylogenetic lineage, with no evidence of transmission in the wards. ESBL strains carried blaCTX-M-15and belonged to clones associated with hospital-acquired ESBL infections in other countries (ST29, ST323, ST340).One also carried the carbapenemase blaIMP-26. Genomic and epidemiological data provided evidence that ESBL strains were acquired in the referring hospital. Nanopore sequencing also identified strain-to-strain transmission of a blaCTX-M-15 FIBK/FIIK plasmid in the referring hospital.ConclusionsThe data suggest the major source of K. pneumoniae was the patient’s own gut microbiome, but ESBL strains were acquired in the referring hospital. This highlights the importance of the wider hospital network to understanding K. pneumoniae risk and infection control. Rectal screening for ESBL organisms upon admission to geriatric wards could help inform patient management and infection control in such facilities.SummaryPatients’ own gut microbiota were the major source of K. pneumoniae, but extended-spectrum beta-lactamase strains were acquired in the referring hospital. This highlights the potential for rectal screening, and the importance of the wider hospital network, for local risk management.</jats:sec

Deiminated proteins and extracellular vesicles - novel serum biomarkers in whales and Orca

Peptidylarginine deiminases (PADs) are a family of phylogenetically conserved calcium-dependent enzymes which cause post-translational protein deimination. This can result in neoepitope generation, affect gene regulation and allow for protein moonlighting via functional and structural changes in target proteins. Extracellular vesicles (EVs) carry cargo proteins and genetic material and are released from cells as part of cellular communication. EVs are found in most body fluids where they can be useful biomarkers for assessment of health status. Here, serum-derived EVs were profiled, and post-translationally deiminated proteins and EV-related microRNAs are described in 5 ceataceans: minke whale, fin whale, humpback whale, Cuvier's beaked whale and orca. EV-serum profiles were assessed by transmission electron microscopy and nanoparticle tracking analysis. EV profiles varied between the 5 species and were identified to contain deiminated proteins and selected key inflammatory and metabolic microRNAs. A range of proteins, critical for immune responses and metabolism were identified to be deiminated in cetacean sera, with some shared KEGG pathways of deiminated proteins relating to immunity and physiology, while some KEGG pathways were species-specific. This is the first study to characterise and profile EVs and to report deiminated proteins and putative effects of protein-protein interaction networks via such post-translationald deimination in cetaceans, revealing key immune and metabolic factors to undergo this post-translational modification. Deiminated proteins and EVs profiles may possibly be developed as new biomarkers for assessing health status of sea mammals

Possibilities and importance of using computer games and simulations in educational process

The paper discusses if it is possible and appropriate to use simulations (simulation games) and traditional games in the process of education. It is stressed that the terms "game" and "simulation" can and should be taken in a broader sense, although they are chiefly investigated herein as video-computer games and simulations. Any activity combining the properties of game (competition, rules, players) and the properties of simulation (i.e. operational presentation of reality) should be understood as simulation games, where role-play constitutes their essence and basis. In those games the student assumes a new identity, identifies himself with another personality and responds similarly. Game rules are basic and most important conditions for its existence, accomplishment and goal achievement. Games and simulations make possible for a student to acquire experience and practice i.e. to do exercises in nearly similar or identical life situations, to develop cognitive and psycho-motor abilities and skills, to acquire knowledge, to develop, create and change attitudes and value criteria, and to develop perception of other people’s feelings and attitudes. It is obligatory for the teacher to conduct preparations to use and apply simulation games in the process of teaching