Adaptation to space conditions of novel bacterial species isolated from the International Space Station revealed by functional gene annotations and comparative genome analysis.

BACKGROUND: The extreme environment of the International Space Station (ISS) puts selective pressure on microorganisms unintentionally introduced during its 20+ years of service as a low-orbit science platform and human habitat. Such pressure leads to the development of new features not found in the Earth-bound relatives, which enable them to adapt to unfavorable conditions. RESULTS: In this study, we generated the functional annotation of the genomes of five newly identified species of Gram-positive bacteria, four of which are non-spore-forming and one spore-forming, all isolated from the ISS. Using a deep-learning based tool-deepFRI-we were able to functionally annotate close to 100% of protein-coding genes in all studied species, overcoming other annotation tools. Our comparative genomic analysis highlights common characteristics across all five species and specific genetic traits that appear unique to these ISS microorganisms. Proteome analysis mirrored these genomic patterns, revealing similar traits. The collective annotations suggest adaptations to life in space, including the management of hypoosmotic stress related to microgravity via mechanosensitive channel proteins, increased DNA repair activity to counteract heightened radiation exposure, and the presence of mobile genetic elements enhancing metabolism. In addition, our findings suggest the evolution of certain genetic traits indicative of potential pathogenic capabilities, such as small molecule and peptide synthesis and ATP-dependent transporters. These traits, exclusive to the ISS microorganisms, further substantiate previous reports explaining why microbes exposed to space conditions demonstrate enhanced antibiotic resistance and pathogenicity. CONCLUSION: Our findings indicate that the microorganisms isolated from ISS we studied have adapted to life in space. Evidence such as mechanosensitive channel proteins, increased DNA repair activity, as well as metallopeptidases and novel S-layer oxidoreductases suggest a convergent adaptation among these diverse microorganisms, potentially complementing one another within the context of the microbiome. The common genes that facilitate adaptation to the ISS environment may enable bioproduction of essential biomolecules need during future space missions, or serve as potential drug targets, if these microorganisms pose health risks. Video Abstract.

Targeted isolation of Methanobrevibacter strains from fecal samples expands the cultivated human archaeome.

Archaea are vital components of the human microbiome, yet their study within the gastrointestinal tract (GIT) is limited by the scarcity of cultured representatives. Our study presents a method for the targeted enrichment and isolation of methanogenic archaea from human fecal samples. The procedure combines methane breath testing, in silico metabolic modeling, media optimization, FACS, dilution series, and genomic sequencing through Nanopore technology. Additional analyzes include the co-cultured bacteriome, comparative genomics of archaeal genomes, functional comparisons, and structure-based protein function prediction of unknown differential traits. Successful establishment of stable archaeal cultures from 14 out of 16 fecal samples yielded nine previously uncultivated strains, eight of which are absent from a recent archaeome genome catalog. Comparative genomic and functional assessments of Methanobrevibacter smithii and Candidatus Methanobrevibacter intestini strains from individual donors revealed features potentially associated with gastrointestinal diseases. Our work broadens available archaeal representatives for GIT studies, and offers insights into Candidatus Methanobrevibacter intestini genomes' adaptability in critical microbiome contexts.

Novel species identification and deep functional annotation of electrogenic biofilms, selectively enriched in a microbial fuel cell array.

In this study, electrogenic microbial communities originating from a single source were multiplied using our custom-made, 96-well-plate-based microbial fuel cell (MFC) array. Developed communities operated under different pH conditions and produced currents up to 19.4 A/m3 (0.6 A/m2) within 2 days of inoculation. Microscopic observations [combined scanning electron microscopy (SEM) and energy dispersive spectroscopy (EDS)] revealed that some species present in the anodic biofilm adsorbed copper on their surface because of the bioleaching of the printed circuit board (PCB), yielding Cu2 + ions up to 600 mg/L. Beta- diversity indicates taxonomic divergence among all communities, but functional clustering is based on reactor pH. Annotated metagenomes showed the high presence of multicopper oxidases and Cu-resistance genes, as well as genes encoding aliphatic and aromatic hydrocarbon-degrading enzymes, corresponding to PCB bioleaching. Metagenome analysis revealed a high abundance of Dietzia spp., previously characterized in MFCs, which did not grow at pH 4. Binning metagenomes allowed us to identify novel species, one belonging to Actinotalea, not yet associated with electrogenicity and enriched only in the pH 7 anode. Furthermore, we identified 854 unique protein-coding genes in Actinotalea that lacked sequence homology with other metagenomes. The function of some genes was predicted with high accuracy through deep functional residue identification (DeepFRI), with several of these genes potentially related to electrogenic capacity. Our results demonstrate the feasibility of using MFC arrays for the enrichment of functional electrogenic microbial consortia and data mining for the comparative analysis of either consortia or their members.

Metabolic engineering of a novel strain of electrogenic bacterium Arcobacter butzleri to create a platform for single analyte detection using a microbial fuel cell.

Electrogenic bacteria metabolize organic substrates by transferring electrons to the external electrode, with subsequent electricity generation. In this proof-of-concept study, we present a novel strain of a known, electrogenic Arcobacter butzleri that can grow primarily on acetate and lactate and its electric current density is positively correlated (R2 = 0.95) to the COD concentrations up to 200 ppm. Using CRISPR-Cas9 and Cpf1, we engineered knockout Arcobacter butzleri mutants in either the acetate or lactate metabolic pathway, limiting their energy metabolism to a single carbon source. After genome editing, the expression of either acetate kinase, ackA, or lactate permease, lctP, was inhibited, as indicated by qPCR results. All mutants retain electrogenic activity when inoculated into a microbial fuel cell, yielding average current densities of 81-82 mA/m2, with wild type controls reaching 85-87 mA2. In the case of mutants, however, current is only generated in the presence of the substrate for the remaining pathway. Thus, we demonstrate that it is possible to obtain electric signal corresponding to the specific organic compound via genome editing. The outcome of this study also indicates that the application of electrogenic bacteria can be expanded by genome engineering.

The parameters of carotid plaques' calcifications and clinical nature of the lesions in the context of revascularization treatment. The enlargement of calcifications is still the most important.

INTRODUCTION: It remains a challenge to determine criteria according to which patients with asymptomatic carotid stenosis could be properly qualified for revascularization treatment. Carotid calcification assessment seems to be here quite attractive. The aim of the study was the histological analysis of various parameters of calcifications in human carotid plaques in relation to the symptomatic/asymptomatic nature of the lesions. MATERIAL AND METHODS: The study involved carotid plaques taken from patients who have undergone endarterectomy of the internal carotid artery. RESULTS: Calcified plaques (with enlarged calcifications) were significantly more frequently asymptomatic than non-calcified plaques. The remaining calcification characteristics played no role. Calcified lesions disclosed the dominance of the fibrous component and the small lipid core significantly more frequently than non-calcified plaques. The percentage of females in the patients group with calcified lesions was significantly higher than in the group with non-calcified plaques. The percentage of males was lower. The former patients group used statins and angiotensin inhibitors significantly more frequently than patients with non-calcified plaques. Enlarged calcifications were independently associated with the asymptomatic nature of the carotid plaques. CONCLUSIONS: The enlargement of calcifications in carotid plaques is the only calcification parameter important for the clinical outcome of carotid atherosclerosis. Patients with calcified carotid plaques have a significantly lower risk of ischemic stroke than patients with non-calcified lesions.

High expression of CX3C chemokine receptor 1 (CX3CR1) in human carotid plaques is associated with vulnerability of the lesions.

Introduction: In data based on ex vivo studies and animal models, fractalkine is considered an important mediator in the development and destabilization of atherosclerotic plaques. We do not know how it is associated with human carotid plaques morphology. Material and methods: The study included 126 carotid plaques taken from 126 patients who underwent endarterectomy of internal carotid arteries. We assessed the following characteristics: inflammatory infiltration, connective tissue elements, foam cells, lipid core, plaque vascularisation, calcifications, intraplaque haemorrhage, thrombi built into the plaque structure, and mural thrombi. CX3CR1 expression in plaques as a response to fractalkine was assessed. Results: Compared to those with a low fractalkine receptor expression, plaques with its high expression exhibited more intensive inflammatory infiltrations. They were more likely to contain inflammatory than fibrous components. They were characterized by a large foam cell component and were less calcified. Intraplaque haemorrhages, the large lipid core and mural as well as intraplaque thrombi were more frequent within them. Conclusions: High expression of the fractalkine receptor within human carotid plaques is associated with morphological parameters of plaque instability. Thus we conclude that fractalkine may be involved in vulnerability of human carotid plaque. .

Relationship between toll-like receptor 2 R753Q and T16934A polymorphisms and Staphylococcus aureus nasal carriage.

BACKGROUND: The association among specific single-nucleotide polymorphisms (SNPs) in TLR2 R753Q (rs5743708) and T16934A (rs4696480) and the nasal carriage of Staphylococcus aureus was studied in adults before CABG. METHODS: The TLR2 polymorphisms were genotyped in 299 consecutive patients prepared for a CABG operation. Genotyping was performed using restriction fragment length polymorphism (RFLP) analysis of PCR-amplified fragments. Two nasal swab cultures were taken within 2 weeks before the operation. Subjects were classified as Staphylococcus aureus carriers if at least one culture was positive while those patients with both cultures found to be negative were classified as non-carriers. RESULTS: The prevalence of nasal S. aureus carriage in the final cohort was 22.1% (66/299), while no MRSA was detected in our study group. No significant differences in the TLR2 polymorphisms were observed between the study and the control groups. No associations were found between TLR2 haplotypes and the covariates of age, sex, NYHA, weight, height, BMI, CAD, smoking status and ESlog score. No differences were found between carriers and noncarriers regarding the allelic distribution of the TLR2 T-16934A SNP. Almost 93% of the patients who were screened for the presence of the TLR2 Arg753Gln (rs5743708) were GG wild type homozygous. Twenty one subjects from the study group (7.1%) were GA heterozygous, while no patient in either group was homozygous for the TLR2 Arg753Gln (rs5743708) mutation. TLR2 Arg753Gln genotyping showed that GA heterozygous patients were detected more frequently in the group of Staphylococcus aureus nasal carriers than in non-carrier adults. CONCLUSION: Our results suggest that the carrier status for the GA variant of the TLR2 Arg753Gln (rs5743708) polymorphism may be a risk factor for Staphylococcus aureus carriage.

Methods of pain assessment in adult intensive care unit patients - Polish version of the CPOT (Critical Care Pain Observation Tool) and BPS (Behavioral Pain Scale).

Many patients treated in the intensive care unit (ICU) experience pain that is a source of suffering and leaves a longterm imprint (chronic pain, post-traumatic stress disorder). Nearly 30% of patients experience pain at rest, while the percentage increases to 50% during nursing procedures. Pain in ICU patients can be divided into four categories: continuous ICU treatment-related pain/discomfort, acute illness-related pain, intermittent procedural pain and pre-existing chronic pain present before ICU admission. As daily nursing procedures and interventions performed in the ICU may be a potential source of pain, it is crucial to use simple pain monitoring tools. The assessment of pain intensity in ICU patients remains an everyday challenge for clinicians, especially in sedated, intubated and mechanically ventilated patients. Regular assessment of pain intensity leads to improved outcome and better quality of life of patients in the ICU and after discharge from ICU. The gold standard in pain evaluation is patient self-reporting, which is not always possible. Current research shows that the two tools best validated for patients unable to self-report pain are the Behavioral Pain Scale (BPS) and the Critical Care Pain Observation Tool (CPOT). Although international guidelines recommend the use of validated tools for pain evaluation, they underline the need for translation into a given language. The authors of this publication obtained an official agreement from the authors of the two behavioral scales - CPOT and BPS - for translation into Polish. Validation of these tools in the Polish population will aid their wider use in pain assessment in ICUs in Poland.

[The issue of expert opinions in underage pornography materials involving underage persons below 15 years of age]. / Zagadnienie opiniowania w sprawach dotyczacych tresci pornograficznych z udzialem osób maloletnich ponizej 15. roku zycia.

The number of cases in which the estimation of age of the persons pictured in pornography materials is mandatory has been increasing for the last few years. The aim of the publication was to indicate the possibilities which are currently available for the forensic experts in this matter. The ones published heretofore mainly had focused on the difficulties which are inseparable elements of such expertise. Current publication has a demonstrative character and has been based upon the experience of two Forensic Medicine Departments. Such expertise have been constantly prepared in both of them for the last few years and none of them had to be changed till now. The currently available methods which allow for age estimation of the persons upon their photographical and movie images are far from perfection. Still authors remain skeptic about the possibility that significant progress in that matter can be achieved in predictable future. In such situation the most effective application of existing techniques becomes essential to minimize the risk of false-positive and false-negative results from appearing. Some of the difficulties pointed out by other authors are not important in practice.

[Application of thermovision in forensic medicine on the basis of the research carried out in Forensic Medicine Department, Pomeranian Medical University in Szczecin]. / Zastosowanie termowizji w medycynie sadowej na przykladzie badan prowadzonych w Zakladzie Medycyny Sadowej Pomorskiej Akademii Medycznej w Szczecinie.

All solid bodies in the temperature above absolute zero emit electromagnetic radiation whose spectrum depends, among others, on temperature. Thermovision is the method which uses infrared cameras for recording infrared radiation emitted by human skin. The method is widely applied in medicine. In this study we want to present possible applications of the thermovision camera in forensic medicine (to investigate the decrease in the temperature of mortal remains; to study temperature distribution in the material simulating human body tissue exposed to the action of a bullet shot from firearms) as well as emphasize the necessity of establishing thermal norms as a potential source for comparative research.

[Waterhouse-Friderichsen cases in the archives of ZMS PAM--epidemiological conditions and diagnostic difficulties]. / Przypadki zespolu Waterhouse'a-Friderichsena w materialach ZMS PAM--uwarunkowania epidemiologiczne oraz trudnosci diagnostyczne.

15 cases of meningococcal infections from 2003-2004 were discussed in the study. Their therapy ended in failure and they had been typically autopsied in our Forensic Med. Department. During the autopsy material for classical microbiological investigations was collected. That type of death is characterised with pathognomonic and autopsy image. The problem is the microbiological identification of the pathogen group and type. It is connected with the characteristic of the analysed material and the biological features of Neisseria meningitidis. The molecular and genetic methods show their usefulness in this type of diagnostic of the properly selected and collected material which typically are: cerebrospinal fluid, brain tissue with the arachnoid membranes and spleen tissue.

[Artefacts in the anthropologic (craniologic) material at the archives of ZMS PAM--possible diagnostic difficulties]. / Artefakty w materiale antropologicznym (kraniologicznym) ZMS PAM--mozliwe trudnosci diagnostyczne.

In the study we presented some of the more interesting forms of artefacts which had been documented during last 7 years of the anthropological laboratory of the Forensic Med. Department of the Pomeranian Med. Univ. We also present the outgoing conclusions concerning the possible diagnostic difficulties. The descriptions of archaeological as well as autopsy-derived material were included. The anthropological analysis was made using data contained in the classic works of literature, which included cranioscopy. The longitudinal and transversal intersections of skulls were used for the analysis of their deformations.

[Application of anthropometric methods in comparative identification based on monitoring recordings]. / Zastosowanie metod antropometrycznych w identyfikacji porównawczej na podstawie zapisów systemów monitoringu.

In recent years more and more often cases have occurred concerning identification of suspects of various crimes for eg. car theft from parking lots in front of supermarkets or money from ATMS. It often happens that such places are monitored twenty-four hours a day. Still, both the quality of cameras and their location in the wrong places cause the recordings to be illegible. In the present pager, we present two cases of this type, where by applying computer graphics and anthropometric methods contributed to identification of law-breakers.

[Dating the time of death of an unknown individual on the basis of remnants of material culture--a case study]. / Datowanie czasu zgonu NN osobnika na podstawie zabytków kultury materialnej--opis przypadku.

The anthropological laboratory of the Forensic Medicine Department PAM carries out a lot of cases concerning identification on the basis of skeletal remains. Standard procedures are: anthropological analysis, estimation of time of burial of the remains, characteristics of morbid changes and the cause of death as well as skull based face reconstruction and super-projection. Very often some tiny details can be useful while answering questions relating to the origin of the remains, time of death, or identity of an unknown individual. In the described case, the key method was analysis of two pieces of clothing found inside the skull.

[A case of considerable incongruence between the expected and actual height of an individual, who was identified with methods of super-projection and comparative testing of DNA]. / Przypadek znaczacej niezgodnosci miedzy wzrostem naleznym a rzeczywistym u osobnika zidentyfikowanego metodami superprojekcji i badan porównawczych DNA.

Anthropology is used, among others, in forensic medicine for identification of skeletal remains. The case we would like to introduce concerns the remains of an unknown man, which were subjected to identification tests in anthropological laboratory of the Forensic Medicine Department PAM. Anthropological analysis let us define the basic distinctive features, on the basis of which police selected an individual, whose features corresponded with the collected data. To confirm identity we used the super-projection method. We obtained the superposition of the estimation craniometric and cephalometric points which were suitable for, which was absolutely sufficient to regard the super-projection as successful. Comparing anthropometric features of the unknown man and appointed individual we noted considerable differences in height values. Comparative research of DNA, however, confirmed the individual's identity.