imagine (Institute of molecular genetics and genetic engineering)
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Mouse Tissue of Origin Single Cell Classification System
No full textSingle cell transcriptomics (scRNA-seq) technology can concurrently measure gene
expression from hundreds of thousands of individual cells. The aim of this project is
to build a system for classification of tissue and organ of origin of single cell types and
subtypes for the mouse samples. The classification system maps the cells using scRNAseq
data and supervised machine learning methods.
The first step was to develop a hierarchical mouse cell type map to comprehend the biology
and heterogeneity of different cell types. The second step was to build mouse scRNAseq
reference datasets containing high-quality and well-annotated scRNA-seq datasets
representing different mouse strains, animal ages, sexes, and biological conditions. Third,
his work established computational workflows that integrated standardization, quality
control, clustering, annotation, and classification model building of scRNA-seq data. The
standardization work involved a protocol for data processing that mapped different gene
versions, names, quantities, and data formats to standardized formats. Quality control
involved data visualization, filtering errors and uninformative measurements based on
cell distribution, detection of outliers, and using standard gene markers to filter unrelated
cells. The final step was to build a classification system based on supervised machine
learning. The step deployed the feed-forward multilayer artificial neural network with
logistic regression learning algorithms. The resulting model built using approximately
117 thousand cells from 15 different tissues achieved classification accuracy of 93.6%.
Most misclassified cells were immune cells that are known to migrate across tissues and
organs. The classification accuracy of tissue of origin of immune cells differs according to
the tissue with high classification accuracy (>95%) for aorta, heart, islets, liver, pancreas,
and thymus. Classification accuracy of immune cells from blood, colon, lung, pancreatic
lymph nodes, small intestine, and spleen as tissue of origin was lower.
This research demonstrated that supervised machine learning methods could achieve
high accuracy in classifying mouse single cell types from most tissues and organs.Book of abstracts: 5th Belgrade Bioinformatics Conference, Serbia, Belgrade,17-20 june 2024
5th Belgrade Bioinformatics Conference
No full textWe are pleased to announce the successful conclusion of the 5th Belgrade Bioinformatics Conference - BelBi2024, where numerous high-quality scientific contributions were presented. We sincerely thank all participants and proudly present a book of abstracts that not only reflects the scientific richness and diversity of the conference, but also serves as a lasting memento of this remarkable event. This international conference was jointly organized by several research institutions, faculties, and scientific societies from Serbia. It covered a wide range of topics from the fields of computational biology, bioinformatics, biomedical informatics, and health informatics. The main goal of BelBi 2024 was to promote contacts between scientists of all levels, provide a platform for the exchange of experiences and present the latest advances in their fields. We hope that BelBi2024 was a valuable platform for researchers from all over the world to meet, build new collaborations and expand professional networks. We are grateful and proud that we were able to welcome over 250 researchers from 21 countries from three continents. The conference included 24 scientific sessions with more than 68 oral presentations (including eight keynote lectures), 54 poster presentations, three hands-on workshops and three satellite events – the MICOS-EU competition, the TranSYS final conference and Shere the IDEA session. We also organized two industry presentations and two panel discussions - “Building Skills for the Future: Masters 4.0 in Bioinformatics” and “BIO4 Campus: Transforming Science into Business”. We also presented the first BelBi art exhibition inspired by scientific discoveries, entitled “IMGGE Magnificent Cell Dance”. And finally, we are particularly proud of the “Future Keynote Speakers” program, which enabled students from faculties across Serbia to attend this year’s keynote lectures and panel discussions for free. We would like to thank all the members of the International Advisory Board and the International Program Committee for their efforts and help that contributed to the success of this event. We are very grateful to the Ministry of Science, Technological Development and Innovation of the Republic of Serbia, the SAIGE project and the Chamber of Commerce and Industry of Serbia for their support. Finally, the local organizing committee is very grateful to all sponsors of the conference - BGI & MGI, Elta’90MS, PacBio & East Diagnostics, Alfa Genetics, Vivogen, LKB, Altium, Telekom Srbija, Labena, AlphaMed, Galen Fokus, Superlab, Kefo, RNIDS, Danau Lab Beograd, RTC and Biomedica, and we hope that they will stay with us for many years to come. Thank you once again to all who contributed to the success of BelBi2024. We look forward to seeing you at future conferences. Warm regards, Belgrade, July 2024Book of abstracts: 5th Belgrade Bioinformatics Conference, Serbia, Belgrade,17-20 june 2024
Assessment of seasonal airborne resistome dynamics in response to air pollution exposure in the Belgrade metropolitan area (AirPollRes)
Get PDFAntimicrobial resistance (AMR) and air pollution have been identified as one of the most
serious threats to human health worldwide. The scarce data demonstrating their
interdependence indicates a need to obtain evidence from a broader global area, especially
from regions exposed to high air pollution. Considering that Serbia is a country struggling with
excessive antibiotics use and misuse, a high percentage of multidrug-resistant bacterial isolates,
and poor air quality, the Serbian capital Belgrade has been recognized as an interesting
research model for the effects of air pollution on the airborne transmission of AMR. After
optimization of air sampling and DNA extraction protocol, the air samples will be collected at
nine locations in the Belgrade metropolitan area selected according to air pollution level during
four seasons. The state-of-the-art shotgun metagenomic sequencing and bioinformatic analysis
of obtained sequences will provide information about microbial community composition of
airborne metagenomes. In addition, sequenced airborne metagenomes will be analyzed for the
abundance and diversity of resistomes (antibiotic and biocide/metal resistance genes) and
mobilomes using several databases and tools. Correlation analyses will offer us insight into the
effect of air pollution and seasonal variations on abundance and diversity of airborne
pathogens, resistome and mobilome in the Belgrade metropolitan area. In-depth approach of
the AirPollRes project will provide the first insights into intersection of AMR and air pollution in
the Belgrade metropolitan area, which is highly vulnerable to these health threats. As the
AirPollRes is a pioneering project in this field, the expected short-term impact is the
introduction of routine monitoring of pathogenic microbes and resistance determinants in the
air in Belgrade, while the longer-term impact will be reflected in the improvement of human
and animal health, allowing for a longer life with higher quality.Principal Investigator: Dr Katarina Novovic, IMGGEDuration period: 2024-202
Effect of thermal management on vitellogenesis and maturation in indoor-reared pikeperch (Sander lucioperca)
Get PDFThe present study aimed to assess the effects of thermal manipulation on the vitellogenesis and final oocyte maturation (FOM) in pikeperch. Two groups of fish were stocked in two separate tanks of the climate chamber. One group was stocked at 10-month age (THERMAL), while the other was continually kept under stable photothermal conditions until 19-month-age (CONTROL) and then transferred to a climate chamber. The progress of vitellogenesis was assessed via evaluation of the oocyte diameter at the mid- and late-autumn phases, and the gonadosomatic index (GSI) at the late-autumn phase. Finally, females from the CONTROL group were hormonally stimulated before (WARMING) and after (STABLE) increase of the temperature from wintering to spawning, and the FOM progress, ovulation and plasma levels of 17 α, 20β dihydroxy progesterone (DHP) were assessed. Significantly larger oocytes at the mid-autumn phase (878.8 ± 40.1 μm vs 836.5 ± 46.5 μm) as well as the GSI at the end of the autumn phase (10.5 ± 1.7% vs 7.6 ± 1.1%) were noticed in THERMAL fish. Significantly faster FOM was seen in fish under the WARMING post-stimulation regime, and these fish had higher DHP levels at the moment of hormonal stimulation (5.4 ± 1.4 ng/ml vs 3.8 ± 1.2 ng/ml). According to the obtained results, it appears that photothermal induction of fish at a younger stage might have a positive impact on the first spawning, while the WARMING thermal regime seems to be more efficient in stimulating the FOM in fish upon first wintering
Azole rings linked to COX inhibitors via hydrazone bridge: Synthesis, stereochemical analysis, and investigation of antimicrobial activity
No full textNonsteroidal anti-inflammatory drugs (NSAIDs) alleviate inflammation and pain through the inhibition of cyclooxygenase (COX) enzymes. Besides these widely recognized therapeutic utilizations, NSAIDs have been reported to display moderate antimicrobial activity and enhance antimicrobial efficacy when administered in combination with commercial antimicrobial drugs. In the present study, we designed novel potential antimicrobial agents by linking some NSAIDs (ibuprofen, flurbiprofen, and naproxen) to various azole rings (pyrazole, imidazole, triazole, and benzimidazole) via hydrazone functionality. The hydrazone linker was introduced into the chemical scaffold of the title molecules by the reaction between hydrazides obtained from NSAIDs and in-house synthesized azole-carrying benzaldehydes. The structures of the target compounds were elucidated by a combination of spectral methods. The NOESY spectra and stereochemical analyses performed using DFT method confirmed the presence of the target molecules as a mixture of E(C=N)-E(N-N)-synperiplanar and E(C=N)-E(N-N)-antiperiplanar conformers in DMSO-d6 solution. 1H and 13C NMR chemical shift values in DMSO were calculated using the GIAO method and compared with the experimental NMR data. Finally, some derivatives were demonstrated to inhibit Candida albicans filamentation and/or bacterial communication system known as quorum sensing. For COX inhibitor-azole hybrids with antimicrobial potency, naproxen appeared to be the most appropriate NSAID, while bulky benzimidazole was not found as a preferable azole ring
Genome sequence diversity of SARS-CoV-2 in Serbia: insights gained from a 3-year pandemic study
Get PDFThe severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), responsible for the COVID-19 pandemic, has been evolving rapidly causing emergence of new variants and health uncertainties. Monitoring the evolution of the virus was of the utmost importance for public health interventions and the development of national and global mitigation strategies. Here, we report national data on the emergence of new variants, their distribution, and dynamics in a 3-year study conducted from March 2020 to the end of January 2023 in the Republic of Serbia. Nasopharyngeal and oropharyngeal swabs from 2,398 COVID-19-positive patients were collected and sequenced using three different next generation technologies: Oxford Nanopore, Ion Torrent, and DNBSeq. In the subset of 2,107 SARS-CoV-2 sequences which met the quality requirements, detection of mutations, assignment to SARS-CoV-2 lineages, and phylogenetic analysis were performed. During the 3-year period, we detected three variants of concern, namely, Alpha (5.6%), Delta (7.4%), and Omicron (70.3%) and one variant of interest—Omicron recombinant “Kraken” (XBB1.5) (<1%), whereas 16.8% of the samples belonged to other SARS-CoV-2 (sub)lineages. The detected SARS-CoV-2 (sub)lineages resulted in eight COVID-19 pandemic waves in Serbia, which correspond to the pandemic waves reported in Europe and the United States. Wave dynamics in Serbia showed the most resemblance with the profile of pandemic waves in southern Europe, consistent with the southeastern European location of Serbia. The samples were assigned to sixteen SARS-CoV-2 Nextstrain clades: 20A, 20B, 20C, 20D, 20E, 20G, 20I, 21J, 21K, 21L, 22A, 22B, 22C, 22D, 22E, and 22F and six different Omicron recombinants (XZ, XAZ, XAS, XBB, XBF, and XBK). The 10 most common mutations detected in the coding and untranslated regions of the SARS-CoV-2 genomes included four mutations affecting the spike protein (S:D614G, S:T478K, S:P681H, and S:S477N) and one mutation at each of the following positions: 5′-untranslated region (5’UTR:241); N protein (N:RG203KR); NSP3 protein (NSP3:F106F); NSP4 protein (NSP4:T492I); NSP6 protein (NSP6: S106/G107/F108 - triple deletion), and NSP12b protein (NSP12b:P314L). This national-level study is the most comprehensive in terms of sequencing and genomic surveillance of SARS-CoV-2 during the pandemic in Serbia, highlighting the importance of establishing and maintaining good national practice for monitoring SARS-CoV-2 and other viruses circulating worldwide.The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), responsible for the COVID-19 pandemic, has been evolving rapidly causing emergence of new variants and health uncertainties. Monitoring the evolution of the virus was of the utmost importance for public health interventions and the development of national and global mitigation strategies. Here, we report national data on the emergence of new variants, their distribution, and dynamics in a 3-year study conducted from March 2020 to the end of January 2023 in the Republic of Serbia. Nasopharyngeal and oropharyngeal swabs from 2,398 COVID-19- positive patients were collected and sequenced using three different next generation technologies: Oxford Nanopore, Ion Torrent, and DNBSeq. In the subset of 2,107 SARS-CoV-2 sequences which met the quality requirements, detection of mutations, assignment to SARS-CoV-2 lineages, and phylogenetic analysis were performed. During the 3-year period, we detected three variants of concern, namely, Alpha (5.6%), Delta (7.4%), and Omicron (70.3%) and one variant of interest—Omicron recombinant “Kraken” (XBB1.5) (<1%), whereas 16.8% of the samples belonged to other SARS-CoV-2 (sub)lineages. The detected SARS-CoV-2 (sub)lineages resulted in eight COVID-19 pandemic waves in Serbia, which correspond to the pandemic waves reported in Europe and the United States. Wave dynamics in Serbia showed the most resemblance with the profile of pandemic waves in southern Europe, consistent with the southeastern European location of Serbia. The samples were assigned to sixteen SARS-CoV-2 Nextstrain clades: 20A, 20B, 20C, 20D, 20E, 20G, 20I, 21J, 21K, 21L, 22A, 22B, 22C, 22D, 22E, and 22F and six different Omicron recombinants (XZ, XAZ, XAS, XBB, XBF, and XBK). The 10 most common mutations detected in the coding and untranslated regions of the SARS-CoV-2 genomes included four mutations affecting the spike protein (S:D614G, S:T478K, S:P681H, and S:S477N) and one mutation at each of the following positions: 5′-untranslated region (5’UTR:241); N protein (N:RG203KR); NSP3 protein (NSP3:F106F); NSP4 protein (NSP4:T492I); NSP6 protein (NSP6: S106/G107/F108 - triple deletion), and NSP12b protein (NSP12b:P314L). This national-level study is the most comprehensive in terms of sequencing and genomic surveillance of SARS-CoV-2 during the pandemic in Serbia, highlighting the importance of establishing and maintaining good national practice for monitoring SARS-CoV-2 and other viruses circulating worldwide
Characterization of Nanohybridosomes from Lipids and Spruce Homogenate Containing Extracellular Vesicles
Get PDFIntroduction: Lipid nanovesicles associated with bioactive phytochemicals from spruce needle homogenate (here called nano-sized
hybridosomes or nanohybridosomes, NSHs) were considered.
Methods: We formed NSHs by mixing appropriate amounts of lecithin, glycerol and supernatant of isolation of extracellular vesicles
from spruce needle homogenate. We visualized NSHs by light microscopy and cryogenic transmission electron microscopy and
assessed them by flow cytometry, dynamic light scattering, ultraviolet–visual spectroscopy, interferometric light microscopy and liquid
chromatography–mass spectrometry.
Results: We found that the particles consisted of a bilayer membrane and a fluid-like interior. Flow cytometry and
interferometric light microscopy measurements showed that the majority of the particles were nano-sized. Dynamic light
scattering and interferometric light microscopy measurements agreed well on the average hydrodynamic radius of the
particles Rh (between 140 and 180 nm), while the concentrations of the particles were in the range between 1013 and
1014/mL indicating that NSHs present a considerable (more than 25%) of the sample which is much more than the yield of
natural extracellular vesicles (EVs) from spruce needle homogenate (estimated less than 1%). Spruce specific lipids and
proteins were found in hybridosomes.
Discussion: Simple and low-cost preparation method, non-demanding saving process and efficient formation procedure suggest that
large-scale production of NSHs from lipids and spruce needle homogenate is feasible.
Plain Language Summary: Cells shed into their exterior nanoparticles (here referred to as extracellular vesicles – EVs) that
are free to move, reach distant cells and are taken up by them. As they carry bioactive constituents, EVs may have important
impact on the recipient cells. The mechanisms of EV formation and mediation can be employed in designing therapeutic,
prophylactic and diagnostic methods for various medical issues. EVs can be harvested from biological samples; however,
their yield is small,12 and there are potential side effects. Artificial vesicles – liposomes – have high yield; however, in vivo,
they can be degraded before reaching the target and their reproducibility is yet insufficient. In order to combine advantages of
both types of nanoparticles, we have composed nanohybridosomes (NSHs) from soya lecithin, water and supernatant of
isolation of EVs from spruce needle homogenate, visualized them by cryogenic electron microscopy and characterized them
with respect to their size, concentration and protein/nucleic acid content. We have applied a recently developed interferometric light microscopy to determine the hydrodynamic radius and the concentration of EVs. We found that the majority of
composed particles are nano-sized and that they enclose more than 25% of the incoming volume of liquid, which is considerably more than about 1% that can be harvested by isolation of EVs from spruce needle homogenate by (ultra)
centrifugatio
Revolutionizing Spinal Muscular Atrophy Prevention in Serbia: Implementing a Mandatory Statewide Newborn Screening
Get PDFSpinal muscular atrophy (SMA) is the prevalent genetic cause of childhood mortality. Pioneering treatments yield utmost advantages only within the presymptomatic phase, underlining the medical and ethical significance of newborn screening.
In 2022, the Centre for Human Molecular Genetics initiated a pilot study of the newborn screening for SMA, working closely alongside the University Children’s Hospital Tirsova and Association SMA Serbia. The aim was to lay the foundation for the implementation of statewide newborn screening for SMA in Serbia by conducting screening for ~8000 infants from the Obstetrics and Gynaecology Clinic Narodni Front over the course of a year. In the subsequent year, we expanded the initiative to include another maternity hospital located outside Belgrade, introducing sample shipment via postal services and extending screening accessibility to a greater number of infants.
In the initial year, 6 950 newborns underwent testing, revealing SMA in two unrelated infants. Subsequently, an older sibling of the first newborn, although asymptomatic at the time, was also tested at the age of 16 months, and SMA diagnosis was confirmed in this child as well. All three children received therapeutic interventions in <1 month from birth. To date, they have exhibited no signs of SMA, and there have been no false
negative outcomes among the newborns who tested negative during the screening. In
the second year, an additional 5 000 newborns underwent testing. As frontrunners in this field in Serbia, we orchestrated harmonized efforts across various tiers of healthcare, established screening and diagnostic algorithms and follow-up protocols. Our extensive efforts were primarily aimed at elevating awareness among all stakeholders about the critical importance of early disease detection. In this transformative journey, we transitioned from being isolated individuals and visionaries who championed a singular idea to an entire community and nation that now acknowledges the paramount significance of newborn screening. As a result, a total of 11 950 infants underwent testing during the 17-month pilot project, culminating in the rapid incorporation of newborn screening for SMA into the national screening program, effective as of September 14th 2023.
Timely detection and treatment can transform SMA into a manageable condition, and there is substantial evidence supporting its inclusion in state-wide screening programs.4th International Congress on Spinal Muscular Atrophy, from 14th to 16th March 2024 in Ghent, Belgiu
Adaptable alginate-based microfibers for 3D in vitro cultures of cancer cells: an anticancer drug testing model
Get PDFThe slow advance in anticancer drug development can be attributed to the limitations of conventional models, predominantly monolayer cell (2D) cultures and animal models, which inadequately recapitulate the complex nature of human malignant tumors. Three-dimensional (3D) in vitro models are invaluable tools in drug screening; however, creating a universal model for all cancer types poses challenges due to the diverse nature of cancers. The aim of this work was to develop a single, versatile model using alginate microfibers to accommodate cultivation of various cancer cells
Biomimetic tumor engineering to enhance drug discovery - BioengineeredTumor
Get PDFDevelopment of novel, effective, and safe anti-tumor drugs is still a slow and cumbersome process, which is often attributed to weaknesses of current preclinical assays and low correlation of the preclinical in vitro and in vivo data with the results obtained in clinical trials. Consequently, there is a clear need for development of more reliable in vitro three dimensional (3D) tumor models, which will capture key features of the in vivo tumor cell microenvironment and provide drug testing results relevant for human patients. The aim of the project “Biomimetic tumor engineering to enhance drug discovery – BioengineeredTumor” funded by the Science Fund of the Republic of Serbia is to develop 2 novel, simple and robust 3D models for cultures of carcinoma and osteosarcoma cells by applying systematic and integrated methodology to comprehensively define the key model components. In specific, the aim is to use different human and animal cancer cell lines in conjunction with alginate-based biomaterials as artificial extracellular matrices imitating tumor environments and to cultivate the obtained constructs in perfusion bioreactors providing enhanced transport of nutrients, gases and biochemical signals to the cells as well as adequate levels of hydrodynamic shear stresses. Thus, the strategic goal is to establish an adaptable platform suited to the use by scientists without technical expertise for long-term in vitro studies of cancer cells for applications in anti-cancer drug discovery and validation, development of personalized medical treatments, and cancer research