imagine (Institute of molecular genetics and genetic engineering)
Not a member yet
3088 research outputs found
Sort by
Searching for the baseline miCRObiota
No full textMicrobiota, a complement of microorganisms living on and in us, has recently been
correlated with plethora of health outcomes and health in general. To fully understand
microorganisms with detrimental effects on human health it is necessary to establish a
healthy “baseline” of the population. In our previous studies investigating correlation of
obesity and gut microbiota composition and maintenance of microbiota equilibrium in vitro
it became evident that publicly available microbiota data is lacking, especially its metadata,
and it would be beneficial to create baseline microbiota composition of local population
with metadata of interest. For that purpose, a pilot study of healthy working-age Croatian
population was carried out. In the pilot study 60 volunteers aged between 18 and 65 years
had participated providing faecal samples, for determining gut microbiota composition, and
filling an online questionnaire about relevant health parameters, lifestyle, dietary habits
and adherence to the Mediterranean diet. The composition of participant’s microbiota did
not provide novel insights, as composition on the higher taxonomic levels was similar to
existing data, but it provided on finer resolution and its linkage with collected metadata.
Implementation of the project also gave important feedback about weaknesses of the
study and the challenges which will be addressed in the follow-up research. Results of the
study are planned to be freely available, together with all anonymized metadata.Book of abstracts: 5th Belgrade Bioinformatics Conference, Serbia, Belgrade,17-20 june 2024
Application of Kolmogorov-Arnold Networks in Cervical Cancer Diagnostics
No full textIn this study, we explore the application of Kolmogorov-Arnold Networks (KAN) in the
field of cervical cancer diagnostics. Cervical cancer, a major health concern worldwide,
requires efficient and accurate diagnostic methods for early detection and treatment.
Traditional machine learning approaches, such as multilayer perceptron (MLP) and
K-nearest neighbors (KNN), have shown high classification performance but often at the
cost of complex and resource-intensive architectures. In contrast, KAN offers a simpler
yet effective alternative.
Utilizing a publicly available dataset of cervical cancer data, which includes 859 samples
with 36 input attributes and diagnostic outputs defined as Hinselmann, Schiller, cytology,
and biopsy, we implemented KAN for classification. Given the significant class imbalance
in the dataset, we also applied various class balancing techniques.
Our results indicate that KAN can achieve high classification performance with mean area
under the receiver operating characteristic curve (AUC) and mean Matthew’s correlation
coefficient (MCC) scores comparable to those obtained with more complex architectures.
Specifically, the KAN models demonstrated robust diagnostic capabilities, achieving AUC
and MCC scores above 0.9.
The simplicity of KAN architectures, combined with their strong performance metrics,
underscores their potential as a practical tool in medical diagnostics. These findings
suggest that Kolmogorov-Arnold Networks could be effectively utilized for cervical cancer
screening and diagnosis, providing a balance of high accuracy, robustness, and reduced
computational complexity. This approach could facilitate more accessible and efficient
diagnostic processes, particularly in resource-limited settings.Book of abstracts: 5th Belgrade Bioinformatics Conference, Serbia, Belgrade,17-20 june 2024
PCR-based nanopore sequencing method for characterizing short tandem repeat expansions
No full textShort tandem repeat (STR) expansions cause >60 rare neurological diseases and pose a
methodological challenge due to their length (0.15->3 kb), complex sequence structure,
and a continuous increase in length in somatic cells during a patient’s life. These genetic
features are the main source of extreme variability in the presentation and progression
of the diseases. To make genetic diagnosis more accessible and improve prognosis in
individual patients, we are developing a nanopore sequencing method to characterize STR
expansions deeply.
The method is based on PCR enrichment and a bioinformatics pipeline developed in-house.
The library containing up to six patient samples was prepared using Native Barcoding Kit
24 V14. Sequencing was performed on R10.4.1 Flongle cells on a portable Mk1C device
(Oxford Nanopore Technologies). Raw data were basecalled with Guppy. Reads were
analyzed as complex strings with an STR between predefined locus-specific flanking
sequences. Regular expressions were used to determine the length and structure of the
STRs. Reads were categorized based on the STR length and aligned without reference.
Polishing steps for overcoming per-read errors were based on results from our wet lab
experiments.
The length and structure of STR expansion in patients with spinocerebellar ataxia type 8
(e.g., (CTA)6(CTG)56-60CCG(CTG)53-56) and myotonic dystrophy type 1 (DM1)
(e.g., (CTG)350-700(CCGCTG)3(CTG)4(CCGCTG)2CTGCCG(CTG)18) were accurately determined
compared to orthogonal methods. For somatically highly unstable DM1 mutation, a
reliable distribution of allele length was achieved with at least 200X coverage, allowing
accurate estimation of modal allele size and degree of somatic instability.
Our nanopore sequencing method can reliably characterize clinically relevant features of
STR expansions: length, sequence structure and degree of somatic instability. The method
outperforms gold standard methods (repeat-primed PCR, Southern blot), captures somatic
variability more reliably compared to Cas9-based enrichment and is more accessible for
clinical settings compared to PacBio and Illumina (where applicable) sequencing methods.Book of abstracts: 5th Belgrade Bioinformatics Conference, Serbia, Belgrade,17-20 june 2024
Antitumor effect of glycolysis inhibition in combination with lysosome permeabilization and oxidative phosphorylation suppression
No full textTumorske ćelije svoje povećane energetske potrebe prevashodno zadovoljavaju aerobnom glikolizom, ali
kada je glikoliza inhibirana njihova metabolička plastičnost im omogućava da lako pređu na oksidativnu
fosforilaciju. Veliki, nestabilni lizozomi tumorskih ćelija su bogati hidrolitičkim enzimima i učestvuju u
metastaziranju i razvoju rezistencije na citostatike. Međutim, izlivanje lizozomalnih enzima u citoplazmu
ubija tumorske ćelije. Kombinovanje lekova koji deluju na različite specifičnosti tumorskih ćelija povećava
njihovu terapeutsku efikasnost, a smanjuje neželjene efekte i razvoj rezistencije. Pokazano je da inhibitor
glikolize 2-dezoksi-D-glukoza (2DG) u kombinaciji sa supresorom oksidativne fosforilacije rotenonom
(ROT), ili sa lizozomalnim deterdžentom N-dodecilimidazolom (NDI) sinergistički ubija B16 melanomske i
U251 gliomske ćelije nekrozom, smanjuje nivo ATP, aktivira AMPK i povećava produkciju mitohondrijskog
superoksida. 2DG+ROT stimuliše otpuštanje heksokinaze II sa membrane mitohondrija, otvaranje VDAC
kanala, izlazak superoksida i citohroma c iz mitohondrija u citoplazmu, aktivaciju kaspaza i apoptozu, koja
zbog nedostatka ATP prelazi u nekrozu. Deplecija ATP indukovana 2DG+ROT tretmanom aktivira AMPK i
suprimira mTORC1, ali paradoksalno inhibira autofagiju. S druge strane, 2DG+NDI indukuje
permeabilizaciju membrane lizozoma, izlazak lizozomalnih enzima u citoplazmu, depolarizaciju
mitohondrija i supresiju oksidativne fosforilacije, što dovodi do potpune energetske deplecije i nekroze.
Inhibicija glikolize u kombinaciji sa supresijom oksidativne fosforilacije ili destabilizacijom lizozoma mogla
bi se koristiti u antitumorskoj terapiji.Tumor cells meet their increased energy needs primarily through aerobic glycolysis, but when glycolysis
is inhibited their metabolic plasticity allows them to easily switch to oxidative phosphorylation. Large,
unstable lysosomes of tumor cells are rich in hydrolytic enzymes and participate in metastasis and the
development of resistance to cytostatics. However, the leakage of lysosomal enzymes into the cytoplasm
kills tumor cells. Combining drugs that act on different tumor cell specificities increases their therapeutic
efficacy while reducing side effects and resistance development. It has been shown that the glycolysis
inhibitor 2-deoxy-D-glucose (2DG), in combination with the oxidative phosphorylation suppressor
rotenone (ROT), or with the lysosomal detergent N-dodecylimidazole (NDI), synergistically kills B16
melanoma and U251 glioma cells through necrosis, reduces ATP levels, activates AMPK, and increases
mitochondrial superoxide production. 2DG+ROT stimulates the release of hexokinase II from the
mitochondrial membrane, opening of VDAC channels, release of superoxide and cytochrome c from
mitochondria into the cytoplasm, activation of caspases, and apoptosis, which, due to ATP depletion,
transitions into necrosis. ATP depletion induced by 2DG+ROT treatment activates AMPK and suppresses
mTORC1, but paradoxically inhibits autophagy. On the other hand, 2DG+NDI induces permeabilization of
the lysosomal membrane, release of lysosomal enzymes into the cytoplasm, mitochondrial depolarization,
and suppression of oxidative phosphorylation, which leads to complete energy depletion and necrosis.
Inhibition of glycolysis combined with suppression of oxidative phosphorylation or destabilization of
lysosomes could be used in antitumor therapy
Role of the glucose-6-phosphate translocase in the activation of autophagy in glycogen storage disease type Ib
No full textGlukozo-6-fosfat translokaza (G6PT), kodirana genom SLC37A4, neophodna je za transport glukozo-6-
fosfata iz citoplazme u lumen endoplazmatičnog retikuluma, gde se dalje hidrolizuje uz pomoć
glukozo-6-fosfataze. Ovaj proces igra ključnu ulogu u održavanju homeostaze glukoze, posebno u jetri,
bubrezima i drugim tkivima. Međutim, osim svojih metaboličkih funkcija, novi dokazi sugerišu da protein
G6PT može takođe biti uključen u regulaciju procesa energetske homeostaze ćelija, kao što je autofagija,
mehanizam koji ćelije koriste za razgradnju i recikliranje oštećenih organela, pogrešno savijenih proteina
i drugih ćelijskih komponenti. U ovom radu predstavljena su dosadašnja saznanja koja govore u prilog
postojanju dodatne funkcije G6PT u aktivaciji autofagije nezavisno od njene transportne aktivnosti.The glucose-6-phosphate translocase (G6PT), encoded by the SLC37A4 gene, is essential for transporting
glucose-6-phosphate from the cytoplasm into the lumen of the endoplasmic reticulum, where it is
hydrolyzed by glucose-6-phosphatase. This process plays a key role in maintaining glucose homeostasis,
particularly in the liver, kidney, and other tissues. However, beyond its metabolic functions, emerging
evidence suggests that G6PT may also be involved in the regulation of cellular homeostasis processes,
such as autophagy, a mechanism that cells use to degrade and recycle damaged organelles, misfolded
proteins, and other cellular components
Interaction between healthy and diseased bronchial epithelial cells with Lactiplantibacillus plantarum BGPKM22 reveals distinct expression profiles by dual RNA sequencing
No full textAlthough low density features lung microbiota, its composition is crucial for the
development and maintenance of healthy human lungs. In chronic obstructive pulmonary
disease (COPD) lung microbiota is changed, correlating with the disease severity and
exacerbations. But whether changed lung microbiota causes COPD, or microbiota changes
due to disease is unclear. Moreover, the effect of COPD on microbes is unknown.
We aimed to scrutinize the effects of interspecies interaction between healthy and
diseased human bronchial epithelial cells with bacteria featured by preferred properties,
Lactiplantibacillus plantarum BGPKM22, by dual RNA sequencing.
Primary healthy and diseased bronchial epithelial cells, from a healthy subject and COPD
patient, respectively, were exposed to Lactip. plantarum BGPKM22. Dual RNA sequencing
and data processing were performed by Novogene (Beijing, China). Clean reads were
aligned to the human genome and Lactip. plantarum SK151 using Hisat2 tool. Differential
gene expression analysis was performed using the edgeR R package. The adherence of
BGPKM22 to the cells and its resistance to oxidative stress were determined.
In healthy and diseased cells interaction with BGPKM22 caused a change in expression of
52 and 45 genes, respectively. The genes IQCN, LINC01554, KCNB1, and CDK7 indicated a
specific response of human bronchial epithelial cells exposed to BGPKM22, regardless of
the health status. Markedly more genes showed a change in expression in BGPKM22 in
interaction with healthy than with diseased cells, 486 and 101, respectively. The adhesion
of BGPKM22 was better to healthy than to diseased cells. The fitness of BGPKM22
increased only after interaction with healthy cells.
Utilization of BGPKM22 can alleviate symptoms and replenish diminished lung microbiota
in COPD. Preferential affinity of BGPKM22 towards healthy cells can explain diminished
lung microbiota in COPD. Beneficial effects of BGPKM22 can remain unexploited by host
due to decreased affinity and fitness of BGPKM22 in interaction with diseased cells.
Abundant response of BGPKM22 in interaction with healthy cells, sheds a new light on
potential lung probiotics depending on the host state.Book of abstracts: 5th Belgrade Bioinformatics Conference, Serbia, Belgrade,17-20 june 2024
Our Journey from Individual Efforts to Nationwide Support: Implementing Newborn Screening for Spinal Muscular Atrophy in Serbia
No full textInnovative treatments for spinal muscular atrophy (SMA) yield the utmost advantages only within the presymptomatic phase, underlining the significance of newborn screening (NBS). We aimed to establish statewide NBS for SMA in Serbia. Our stepwise implementation process involved technical validation of a screening assay, collaboration with patient organizations and medical professionals, a feasibility study, and negotiation with public health representatives. Over 12,000 newborns were tested during the 17-month feasibility study, revealing two unrelated SMA infants and one older sibling. All three children received therapeutic interventions during the presymptomatic phase and have shown no signs of SMA. No false-negative results were found among the negative test results. As frontrunners in this field in Serbia, we established screening and diagnostic algorithms and follow-up protocols and raised awareness among stakeholders about the importance of early disease detection, leading to the incorporation of NBS for SMA into the national program on 15 September 2023. Since then, 54,393 newborns have been tested, identifying six SMA cases and enabling timely treatment. Our study demonstrates that effective collaborations between academia, non-profit organizations, and industry are crucial in bringing innovative healthcare initiatives to fruition, and highlights the potential of NBS to revolutionize healthcare outcomes for presymptomatic SMA infants and their families
Double layer bacterial nanocellulose - poly(hydroxyoctanoate) film activated by prodigiosin as sustainable, transparent, UV-blocking material
No full textSynthetic materials alternatives are crucial for reaching sustainable development goals and waste reduction. Biomaterials and biomolecules obtained through bacterial fermentation offer a viable solution. Double-layer active UV-blocking material composed of bacterial nanocellulose as an inner layer and poly(hydroxyoctanoic acid) containing prodigiosin as an active compound was produced by layer-by-layer deposition. This study referred the new material consisted of the three components produced in sustainable manner, by bacterial activity: bacterial bio-pigment prodigiosin, bacterial nanocellulose and poly(hydroytoctanoate) - biopolymer obtained by microbial fermentations. Prior the final double layer film was produced, PHO films containing different PG concentrations as a layer in charge of the bioactivity (0.2, 0.5 and 1 wt%) was casted and systematically characterized (FTIR, DSC, XRD, wettability, SEM, transparency, mechanical tests) to optimize their properties. The formulation with the best UV-blocking properties and less toxicity effect tested using MRC5 cells was chosen as an outer layer in double-layer films production. Water contact angle measurements confirmed that hydrophilic – hydrophobic double layer film was obtained with the improved mechanical properties in comparison to the native BNC. Migration test indicated release of PG in all tested media as a consequence of bilayer formulation, while the PG release from PHO in 10 % ethanol was not detected. All findings from the study suggested this activated, UV-blocking material as a candidate with excellent potential in packaging industry
Insight into the Probiogenomic Potential of Enterococcus faecium BGPAS1-3 and Application of a Potent Thermostable Bacteriocin
No full textThis study aimed to investigate the probiogenomic features of artisanal bacteriocin-producing Enterococcus faecium BGPAS1-3 and the use of the improved pMALc5HisEk expression vector for overexpressing class II bacteriocins and the application of purified bacteriocin 31 in a milk model as a preservative against L. monocytogenes. The BGPAS1-3 strain was isolated from traditional fresh soft cheese manufactured in households on a small scale in rural locations surrounding Pale Mountain City in Bosnia and Herzegovina. The whole-genome sequencing approach and bioinformatics analyses revealed that the strain BGPAS1-3 was non-pathogenic to humans. The presence of bacteriocin operons suggested the ability of the isolate to suppress the growth of pathogens. Coding regions for three maturated bacteriocins (bacteriocin 31, bacteriocin 32, and enterocin P) produced by BGPAS1-3 were amplified and expressed in Escherichia coli ER2523 using the pMALc5HisEk system. All three bacteriocins were successfully overexpressed and purified after enterokinase cleavage but showed different antimicrobial activity. Bacteriocin 31 showed significantly stronger antimicrobial activity compared with bacteriocin 32. It was the only one that proved to be suitable for use as a food preservative against L. monocytogenes in a milk model
Novel telomerase reverse transcriptase gene mutation in a family with aplastic anaemia
No full textTelomerase Reverse Transcriptase (TERT) encodes the telomerase reverse transcriptase enzyme and is the most frequently mutated gene in patients with telomeropathies. Heterozygous variants impair telomerase activity by haploinsufficiency and pathogenic variants are associated with bone marrow failure syndrome and predisposition to acute myeloid leukaemia. Owing to their rarity, telomeropathies are often unrecognised and misdiagnosed. Herein, we report a novel TERT gene variant, c.2605G > A p.(Asp869Asn) in a family with hereditary aplastic anaemia. This report emphasises the importance of routine deep genetic screening for rare TERT variants in patients with a family history of cytopenia or aplastic anaemia, which could identify clinically inapparent telomere disorders