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Open Access
ACTIVITY-BASED PROBES TO UTILIZE THE PROTEOLYTIC ACTIVITY OF CATHEPSIN G IN BIOLOGICAL SAMPLES
(Frontiers in Chemistry, 2021-02-25) Burster, Timo; Gärtner, Fabian; Knippschild, Uwe; Zhanapiya, Anuar
Neutrophils, migrating to the site of infection, are able to release serine proteases after being activated. These serine proteases comprise cathepsin G (CatG), neutrophil elastase protease 3 (PR3), and neutrophil serine protease 4 (NSP4). A disadvantage of the uncontrolled proteolytic activity of proteases is the outcome of various human diseases, including cardiovascular diseases, thrombosis, and autoimmune diseases. Activity-based probes (ABPs) are used to determine the proteolytic activity of proteases, containing a set of three essential elements: Warhead, recognition sequence, and the reporter tag for detection of the covalent enzyme activity–based probe complex. Here, we summarize the latest findings of ABP-mediated detection of proteases in both locations intracellularly and on the cell surface of cells, thereby focusing on CatG. Particularly, application of ABPs in regular flow cytometry, imaging flow cytometry, and mass cytometry by time-of-flight (CyTOF) approaches is advantageous when distinguishing between immune cell subsets. ABPs can be included in a vast panel of markers to detect proteolytic activity and determine whether proteases are properly regulated during medication. The use of ABPs as a detection tool opens the possibility to interfere with uncontrolled proteolytic activity of proteases by employing protease inhibitors.
Open Access
ACTIVITY-BASED PROBES TO UTILIZE THE PROTEOLYTIC ACTIVITY OF CATHEPSIN G IN BIOLOGICAL SAMPLES
(Frontiers in Chemistry, 2021) Burster, Timo; Gärtner, Fabian; Knippschild, Uwe; Zhanapiya, Anuar
Neutrophils, migrating to the site of infection, are able to release serine proteases after being activated. These serine proteases comprise cathepsin G (CatG), neutrophil elastase protease 3 (PR3), and neutrophil serine protease 4 (NSP4). A disadvantage of the uncontrolled proteolytic activity of proteases is the outcome of various human diseases, including cardiovascular diseases, thrombosis, and autoimmune diseases. Activity-based probes (ABPs) are used to determine the proteolytic activity of proteases, containing a set of three essential elements: Warhead, recognition sequence, and the reporter tag for detection of the covalent enzyme activity-based probe complex. Here, we summarize the latest findings of ABP-mediated detection of proteases in both locations intracellularly and on the cell surface of cells, thereby focusing on CatG. Particularly, application of ABPs in regular flow cytometry, imaging flow cytometry, and mass cytometry by time-of-flight (CyTOF) approaches is advantageous when distinguishing between immune cell subsets. ABPs can be included in a vast panel of markers to detect proteolytic activity and determine whether proteases are properly regulated during medication. The use of ABPs as a detection tool opens the possibility to interfere with uncontrolled proteolytic activity of proteases by employing protease inhibitors.
Open Access
ACTIVITY-BASED PROBES TO UTILIZE THE PROTEOLYTIC ACTIVITY OF CATHEPSIN G IN BIOLOGICAL SAMPLES
(Front. Chem., 2021-02-25) Burster, Timo; Gärtner, Fabian; Knippschild, Uwe; Zhanapiya, Anuar
Neutrophils, migrating to the site of infection, are able to release serine proteases after being activated. These serine proteases comprise cathepsin G (CatG), neutrophil elastase protease 3 (PR3), and neutrophil serine protease 4 (NSP4). A disadvantage of the uncontrolled proteolytic activity of proteases is the outcome of various human diseases, including cardiovascular diseases, thrombosis, and autoimmune diseases. Activity-based probes (ABPs) are used to determine the proteolytic activity of proteases, containing a set of three essential elements: Warhead, recognition sequence, and the reporter tag for detection of the covalent enzyme activity–based probe complex. ...
Open Access
Antimicrobial susceptibility of Brucella melitensis in Kazakhstan
(Nazarbayev University School of Sciences and Humanities, 2017-12-28) Ramankulov, Yerlan; Shevtsov, Alexandr; Syzdykov, Marat; Kuznetsov, Andrey; Shevtsova, Elena; Berdimuratova, Kalysh; Kasim, Mukanov
Background: Kazakhstan belongs to countries with a high level of brucellosis among humans and farm animals. Although antibiotic therapy is the main way to treat acute brucellosis in humans there is still little information on a circulation of the antibiotic-resistant Brucella strains in the Central Eurasia. In this article we describe an occurrence of the drug resistance of Brucella melitensis isolates in Kazakhstan which is among the largest countries of the region. Methods: Susceptibilities to tetracyclin, gentamycin, doxycyclin, streptomycin and rifampicin were investigated in 329 clinical isolates of Brucella melitensis using E-test method. Results: All isolates were susceptible to streptomycin, tetracycline and doxycycline. 97.3% of the Brucella isolates were susceptible to gentamycin, although only 37.4% of isolates were susceptible to rifampicin. 21.9% of isolates had intermediate resistance, and 26.4% of isolates were resistant to this antibacterial drug. Conclusion: Isolates of Brucella melitensis circulating in Kazakhstan are susceptible to streptomycin, doxicyclin, tetracyclin and gentamycin. At the same time the resistance to rifampicin is widespread, almost half of the isolates were rifampicin-resistant (including the intermediate resistance).
Open Access
Antiviral Resistance of Splenocytes in Aged Mice
(Nazarbayev University School of Sciences and Humanities, 2017-03-30) Burster, Timo
We compared the susceptibility to viral infection of splenocytes, isolated from young versus old CBA mice, and evaluated the antiviral actions of lactoferrin in splenocytes infected with Encephalomyocarditis virus (EMCV). Recombinant mouse lactoferrin (rmLF) and bovine lactoferrin (bLF) were used. There were no differences in the susceptibility to EMCV infection in the studied age categories. Both types of lactoferrins were protective in young and old mice. The study confirmed the undisturbed viral resistance in old mice and the protective actions of lactoferrin in viral infection. The antiviral action of the homologous mouse lactoferrin was demonstrated for the first time.
Open Access
APBioNet's annual International Conference on Bioinformatics (InCoB) returns to India in 2018
(Nazarbayev University School of Sciences and Humanities, 2019-04-18) Schönbach, Christian; Ahmad, Shandar; Gromiha, Michael M.; Raghava, Gajendra P.S.; Ranganathan, Shoba
InCoB, one of the largest annual bioinformatics conferences in the Asia-Pacific region since its launch in 2002, returned to New Delhi, India after 12 years, with a conference attendance of 314 delegates. The 2018 conference had sessions on Big Data and Algorithms, Next Generation Sequencing and Omics Science, Structure, Function and Interactions, Disease and Drug Discovery and Plant and Agricultural Bioinformatics. The conference also featured an industry track as well as panel discussions on Women in Bioinformatics and Democratization vs. Quality control in academic publishing. Asia Pacific Bioinformatics Interaction & Networking Society (APbians) was launched as an APBionet Special Interest Group. Of the 52 oral presentations made, 22 were accepted in supplemental issues of BMC Bioinformatics, BMC Genomics or BMC Medical Genomics and are briefly reviewed here. Next year's InCoB will be held in Jakarta, Indonesia from September 10-12, 2019.
Open Access
APPLICATION OF AN ACTIVITY-BASED PROBE TO DETERMINE PROTEOLYTIC ACTIVITY OF CELL SURFACE CATHEPSIN G BY MASS CYTOMETRY DATA ACQUISITION
(ACS Omega, 2020) Gärtner, Fabian; Knippschild, Uwe; Burster, Timo
During an immune response, cathepsin G (CatG) takes on the role of adaptive and innate immunity and the outcome depends on the localization of CatG. Soluble, cell surface-bound, or intracellular CatG is also responsible for pathophysiology conditions. We applied the activity-based probe MARS116-Bt to mass cytometry by time-of-flight to analyze CatG activity on the cell surface of immune cells. The phosphonate warhead of MARS116-Bt binds covalently to the serine amino acid residue S195 of the catalytic center and thereby CatG activity can be detected. This method contributes to observing the activation or inhibition status of cells during pathogenesis of diseases and enables accurate data acquisition from complex biological samples with a vast panel of cell subset markers in a single-cell resolution.
Open Access
An aptasensor for the detection of Mycobacterium tuberculosis secreted immunogenic protein MPT64 in clinical samples towards tuberculosis detection
(Nazarbayev University School of Sciences and Humanities, 2019-11-07) Sypabekova, Marzhan; Dukenbayev, Kanat; Tsepke, Anna; Akisheva, Akmaral; Oralbayev, Nurlan; Kanayeva, Damira
This work presents experimental results on detection of Mycobacterium tuberculosis secreted protein MPT64 using an interdigitated electrode (IDE) which acts as a platform for capturing an immunogenic protein and an electrochemical impedance spectroscopy (EIS) as a detection technique. The assay involves a special receptor, single stranded DNA (ssDNA) aptamer, which specifically recognizes MPT64 protein. The ssDNA immobilization on IDE was based on a co-adsorbent immobilization at an optimized ratio of a 1/100 HS-(CH6)(6)-OP(O)(2)O-(CH2CH2O)(6)-5'-TTTTT-aptamer-3'/6-mercaptohexanol. The optimal sample incubation time required for a signal generation on an aptamer modified IDE was found to be at a range of 15-20 min. Atomic Force Microscopy (AFM) results confirmed a possible formation of an aptamer - MPT64 complex with a 20 nm roughness on the IDE surface vs. 4.5 nm roughness for the IDE modified with the aptamer only. A limit of detection for the EIS aptasensor based on an IDE for the detection of MPT64 in measurement buffer was 4.1 fM. The developed EIS aptasensor was evaluated on both serum and sputum clinical samples from the same TB (-) and TB (+) patients having a specificity and sensitivity for the sputum sample analysis 100% and 76.47%, respectively, and for the serum sample analysis 100% and 88.24%, respectively. The developed aptasensor presents a sensitive method for the TB diagnosis with the fast detection time.
Open Access
BCL-XL ACTIVITY INFLUENCES OUTCOME OF THE MITOTIC ARREST
(Frontiers in Pharmacology, 2022-09-15) Suleimenov, M.; Bekbayev, S.; Ten, M.; Suleimenova, N.; Tlegenova, M.; Nurmagambetova, A.; Kauanova, S.; Vorobjev, I.
Microtubule-targeting (MT) drugs taxanes and vinca alkaloids are widely used as chemotherapeutic agents against different tumors for more than 30 years because of their ability to block mitotic progression by disrupting the mitotic spindle and activating the spindle assembly checkpoint (SAC) for a prolonged period of time. However, responses to mitotic arrest are different—some cells die during mitotic arrest, whereas others undergo mitotic slippage and survive becoming able for proliferation. Using normal fibroblasts and several cancer cell types we determined two critical doses, T1 and T2, of mitotic inhibitors (nocodazole, Taxol, and vinorelbine). T1 is the maximal dose cells can tolerate undergoing normal division, and T2 is the minimal mitostatic dose, wherein > 90% of mitotic cells are arrested in mitosis. In all studied cell lines after treatment with mitotic inhibitors in a dose above T2 cells had entered mitosis either die or undergo mitotic slippage. We show that for all three drugs used cell death during mitotic arrest and after slippage proceeded via mitochondriadependent apoptosis. We determined two types of cancer cells: sensitive to mitotic arrest, that is, undergoing death in mitosis (DiM) frequently, and resistant to mitotic arrest, that is, undergoing mitotic slippage followed by prolonged survival. We then determined that inhibition of Bcl-xL, but not other antiapoptotic proteins of the Bcl-2 group that regulate MOMP, make resistant cells susceptible to DiM induced by mitotic inhibitors. Combined treatment with MT drugs and highly specific Bcl-xL inhibitors A-1155643 or A-1331852 allows achieving 100% DiM in a time significantly shorter than maximal duration of mitotic arrest in all types of cultured cells tested. We further examined efficacy of sequential treatment of cultured cells using mitotic inhibitors followed by inhibitors of Bcl-xL anti-apoptotic protein and for the first time show that sensitivity to Bcl-xL inhibitors rapidly declines after mitotic slippage. Thus sequential use of mitotic inhibitors and inhibitors of Bcl-xL anti-apoptotic protein will be efficient only if the Bcl-xL inhibitor will be added before mitotic slippage occurs or soon afterward. The combined treatment proposed might be an efficient approach to anti-cancer therapy.
Open Access
BIOCHEMICAL AND ELECTROCHEMICAL CHARACTERIZATION OF BIOFILMS FORMED ON EVEROLIMUS-ELUTING CORONARY STENTS
(Enzyme and Microbial Technology, 2022) Akhmetzhan, Gauhar; Olaifa, Kayode; Kitching, Michael; Cahill, Paul A.; Pham, Tri T.; Ajunwa, Obinna M.; Marsili, Enrico
Drug-eluting stents (DES) are mostly used in percutaneous coronary intervention, which is the main treatment for coronary artery occlusion. This procedure aims to restore the natural lumen, while minimizing the risk of restenosis. However, stent insertion increases the risk for infections, due to contamination of the device or insertion hub with normal skin flora. While coronary stent infection is a rare complication, it can be fatal. Currently, there is little information on biofilm formation on everolimus-eluting stents. Although everolimus is not designed as an antimicrobial agent, its antimicrobial activity should be investigated. In this study, biofilm formation on everolimus-eluting and bare metal stents (BMS) is characterized through biochemical and electrochemical methods. DES and BMS are inoculated with Pseudomonas aeruginosa and Staphylococcus epidermidis, both independently and in co-culture. Biofilms formed on DES were 49.6 %, 12.9 % and 47.5 % higher than on BMS for P. aeruginosa, S. epidermidis and their co-culture, respectively. Further, the charge output for DES was 18.9 % and 59.7 % higher than BMS for P. aeruginosa and its co-culture with S. epidermidis, respectively. This observation is most likely due to higher surface roughness of DES, which favors biofilm formation. This work shows that bioelectrochemical methods can be used for rapid detection of biofilms on drug-eluting and bare metal stents, which may find application in quality assessment of stents and in characterization of stents removed after polymicrobial infections.
Open Access
A bioinformatics potpourri
(Nazarbayev University School of Sciences and Humanities, 2018-01-19) Schönbach, Christian; Li, Jinyan; Ma, Lan; Horton, Paul; Sjaugi, Muhammad Farhan
The 16th International Conference on Bioinformatics (InCoB) was held at Tsinghua University, Shenzhen from September 20 to 22, 2017. The annual conference of the Asia-Pacific Bioinformatics Network featured six keynotes, two invited talks, a panel discussion on big data driven bioinformatics and precision medicine, and 66 oral presentations of accepted research articles or posters. Fifty-seven articles comprising a topic assortment of algorithms, biomolecular networks, cancer and disease informatics, drug-target interactions and drug efficacy, gene regulation and expression, imaging, immunoinformatics, metagenomics, next generation sequencing for genomics and transcriptomics, ontologies, post-translational modification, and structural bioinformatics are the subject of this editorial for the InCoB2017 supplement issues in BMC Genomics, BMC Bioinformatics, BMC Systems Biology and BMC Medical Genomics. New Delhi will be the location of InCoB2018, scheduled for September 26–28, 2018.
Open Access
CAMOSTAT DOES NOT INHIBIT THE PROTEOLYTIC ACTIVITY OF NEUTROPHIL SERINE PROTEASES
(Pharmaceuticals, 2022) Assylbekova, Akmaral; Zhanapiya, Anuar; Grzywa, Renata; Sienczyk, Marcin; Schönbach, Christian
Coronavirus disease 2019 (COVID-19) can lead to multi-organ failure influenced by co morbidities and age. Binding of the severe acute respiratory syndrome coronavirus 2 spike protein (SARS-CoV-2 S protein) to angiotensin-converting enzyme 2 (ACE2), along with proteolytic digestion of the S protein by furin and transmembrane protease serine subtype 2 (TMPRSS2), provokes inter nalization of SARS-CoV-2 into the host cell. Productive infection occurs through viral replication in the cytosol and cell-to-cell transmission. The catalytic activity of TMPRSS2 can be blocked by the trypsin-like serine protease inhibitor camostat, which impairs infection by SARS-CoV-2. At the site of infection, immune cells, such as neutrophils, infiltrate and become activated, releasing neutrophil ser ine proteases (NSPs), including cathepsin G (CatG), neutrophil elastase (NE), and proteinase 3 (PR3), which promote the mounting of a robust immune response. However, NSPs might be involved in in fection and the severe outcome of COVID-19 since the uncontrolled proteolytic activity is responsible for many complications, including autoimmunity, chronic inflammatory disorders, cardiovascular diseases, and thrombosis. Here, we demonstrate that camostat does not inhibit the catalytic activity of CatG, NE, and PR3, indicating the need for additional selective serine protease inhibitors to reduce the risk of developing severe COVID-19.
Open Access
CAPIVASERTIB RESTRICTS SARS-COV-2 CELLULAR ENTRY: A POTENTIAL CLINICAL APPLICATION FOR COVID-19
(International Journal of Biological Sciences, 2021) Sun, Fang; Mu, Chenglin; Kwok, Hang Fai; Xu, Jiyuan; Wu, Yingliang; Liu, Wanhong; Sabatier, Jean-Marc; Annweiler, Cédric; Li, Xugang; Cao, Zhijian; Xie, Yingqiu
Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has led to more than 150 million infections and about 3.1 million deaths up to date. Currently, drugs screened are urgently aiming to block the infection of SARS-CoV-2. Here, we explored the interaction networks of kinase and COVID-19 crosstalk, and identified phosphoinositide 3-kinase (PI3K)/AKT pathway as the most important kinase signal pathway involving COVID-19. Further, we found a PI3K/AKT signal pathway inhibitor capivasertib restricted the entry of SARS-CoV-2 into cells under non-cytotoxic concentrations. Lastly, the signal axis PI3K/AKT/FYVE finger-containing phosphoinositide kinase (PIKfyve)/PtdIns(3,5)P2 was revealed to play a key role during the cellular entry of viruses including SARS-CoV-2, possibly providing potential antiviral targets. Altogether, our study suggests that the PI3K/AKT kinase inhibitor drugs may be a promising anti-SARS-CoV-2 strategy for clinical application, especially for managing cancer patients with COVID-19 in the pandemic era
Open Access
Cell Encapsulation Within Alginate Microcapsules: Immunological Challenges and Outlook
(Frontiers Media, 2019-12-03) Ashimova, Assem; Yegorov, Sergey; Negmetzhanov, B.; Hortelano, Gonzalo
Cell encapsulation is a bioengineering technology that provides live allogeneic or xenogeneic cells packaged in a semipermeable immune-isolating membrane for therapeutic applications. The concept of cell encapsulation was first proposed almost nine decades ago, however, and despite its potential, the technology has yet to deliver its promise. The few clinical trials based on cell encapsulation have not led to any licensed therapies. Progress in the field has been slow, in part due to the complexity of the technology, but also because of the difficulties encountered when trying to prevent the immune responses generated by the various microcapsule components, namely the polymer, the encapsulated cells, the therapeutic transgenes and the DNA vectors used to genetically engineer encapsulated cells. While the immune responses induced by polymers such as alginate can be minimized using highly purified materials, the need to cope with the immunogenicity of encapsulated cells is increasingly seen as key in preventing the immune rejection of microcapsules. The encapsulated cells are recognized by the host immune cells through a bidirectional exchange of immune mediators, which induce both the adaptive and innate immune responses against the engrafted capsules. The potential strategies to cope with the immunogenicity of encapsulated cells include the selective diffusion restriction of immune mediators through capsule pores and more recently inclusion in microcapsules of immune modulators such as CXCL12. Combining these strategies with the use of well-characterized cell lines harboring the immunomodulatory properties of stem cells should encourage the incorporation of cell encapsulation technology in state-of-the-art drug development.
Open Access
Cell surface cathepsin G can be used as an additional marker to distinguish T cell subsets
(SPANDIDOS, 2019-04) Penczek, Adriane; Burster, Timo
The serine protease cathepsin G (CatG) is involved in numerous processes associated with the innate and adaptive immune system. During an immune response, neutrophils secrete CatG, which can bind to the cell surface of immune cells to provoke the proteolytic processing of cytokines and chemokines in order to stimulate lymphocytes. The present study analyzed peripheral blood mononuclear cells to characterize T cell populations in terms of their CatG content by flow cytometry. It was identified that CatG was exclusively present on the cell surface of a subset of T regulatory cells (Tregs), cluster of differentiation (CD) 39(+) Tregs, which expressed CatG in contrast to CD39(-) Tregs. Additionally, CatG was expressed on double positive CD4(+)CD8(+) T cells, T helper (Th) 9 cells and Th22 cells, implicating CatG as a novel marker to distinguish certain T cell subsets.
Open Access
CETYLTRIMETHYLAMMONIUM BROMIDE (CTAB)-LOADED SIO2–AG MESOPOROUS NANOCOMPOSITE AS AN EFFICIENT ANTIBACTERIAL AGENT
(Nanomaterials, 2021-02-13) Abduraimova, Aiganym; Molkenova, Anara; Duisembekova, Assem; Mulikova, Tomiris; Kanayeva, Damira; Atabaev, Timur Sh.
To date, Ag-based nanomaterials have demonstrated a high potential to overcome antibiotic resistance issues. However, bare Ag nanomaterials are prone to agglomeration in the biological environment, which results in a loss of antibacterial activity over time. Furthermore, it is still challenging to collect small-sized Ag nanomaterials right after the synthesis process. In this study, spherical-shaped Ag nanoparticles (NPs) (~6–10 nm) were attached on the surface of cetyltrimethylammonium bromide (CTAB)-loaded mesoporous silica nanoparticles (MSNs) (~100–110 nm). Antibacterial activity tests suggested that the obtained nanocomposite can be used as a highly efficient antibacterial agent against both Gram-negative and Gram-positive bacterial strains. The minimum inhibitory concentration (MIC) recalculated to pure Ag weight in nanocomposite was found to be ~1.84 µg/mL (for Escherichia coli) and ~0.92 µg/mL (for Staphylococcus aureus)—significantly smaller compared to values reported to date. The improved antibacterial activity of the prepared nanocomposite can be attributed to the even distribution of non-aggregated Ag NPs per volume unit and the presence of CTAB in the nanocomposite pores.
Open Access
A CHIRALITY-DEPENDENT ACTION OF VITAMIN C IN SUPPRESSINGKIRSTEN RAT SARCOMA MUTANT TUMOR GROWTH BY THE OXIDATIVECOMBINATION: RATIONALE FOR CANCER THERAPEUTICS
(International Journal of Cancer, 2019-08-31) Wu, Xinggang; Park, Mikyung; Sarbassova, Dilara A.; Ying, Haoqiang; Lee, Min Gyu; Bhattacharya, Rajat; Ellis, Lee; Peterson, Christine B.; Hung, Mien-Chie; Lin, Hui-Kuan; Bersimbaev, Rakhmetkazhi I.; Song, Min Sup; Sarbassov, Dos D.
Kirsten rat sarcoma (KRAS) mutant cancers, which constitute the vast majority of pancreatic tumors, are characterized by their resistance to established therapies and high mortality rates. Here, we developed a novel and extremely effective combinational therapeutic approach to target KRAS mutant tumors through the generation of a cytotoxic oxidative stress. At high concentrations, vitamin C (VC) is known to provoke oxidative stress and selectively kill KRAS mutant cancer cells, although its effects are limited when it is given as monotherapy. We found that the combination of VC and the oxidizing drug arsenic trioxide (ATO) is an effective therapeutic treatment modality. Remarkably, its efficiency is dependent on chirality of VC as its enantiomer D-optical isomer of VC (D-VC) is significantly more potent than the natural L-optical isomer of VC. Thus, our results demonstrate that the oxidizing combination of ATO and D-VC is a promising approach for the treatment of KRAS mutant human cancers
Open Access
COMBINING IMAGING FLOW CYTOMETRY AND MOLECULAR BIOLOGICAL METHODS TO REVEAL PRESENCE OF POTENTIALLY TOXIC ALGAE AT THE URAL RIVER IN KAZAKHSTAN
(Frontiers Media S.A., 2021-07-21) Mirasbekov, Yersultan; Abdimanova, Aigerim; Sarkytbayev, Kuanysh; Samarkhanov, Kanat; Abilkas, Aidyn; Potashnikova, Daria; Arbuz, Galina; Issayev, Zhanpeis; Vorobjev, Ivan A.; Malashenkov, Dmitry V.; Barteneva, Natasha S.
Algal blooms occur in freshwater bodies throughout the world, often leading to fish kills. Cases of these kills along the Ural River were reported in 2018–2019, involving significant amount of sturgeon in fish farming areas. In this study, the analysis of algal samples from the delta of the Ural River up to 100 km inland was carried out from August to December 2019 using imaging flow cytometry (IFC), molecular biological, and microscopic techniques. We identified the filamentous cyanobacteria Cuspidothrix issatschenkoi, Dolichospermum cf. flos-aquae, Dolichospermum cf. macrosporum, Pseudanabaena limnetica, and Planktothrix spp. as the dominant potentially toxic phytoplankton species, and we also found minor quantities of Cylindrospermopsis raciborskii. For the first time, molecular phylogenetic investigations of field clones of cyanobacteria from Ural River were carried out to establish the taxa of the dominant species and to identify the presence of genes encoding toxins. The complementary analysis with nanopore-based next-generation sequencing overlapped with the results of IFC and was instrumental in revealing minor cyanobacteria taxa. Real-time PCR analysis and sequencing indicated the presence of Microcystis and ADA-clade spp. as well as genes associated with the production of microcystin (mcyE) and the algal neurotoxin saxitoxin (sxtA) originating from cyanobacteria. These findings suggest that toxin-producing cyanobacteria could become a threat in the Ural River near Atyrau, which can significantly affect aquaculture in the region.
Open Access
COMBINING IMAGING FLOW CYTOMETRY AND MOLECULAR BIOLOGICAL METHODS TO REVEAL PRESENCE OF POTENTIALLY TOXIC ALGAE AT THE URAL RIVER IN KAZAKHSTAN
(Frontiers in Marine Science, 2021-07) Mirasbekov, Yersultan; Abdimanova, Aigerim; Sarkytbayev, Kuanysh; Samarkhanov, Kanat; Abilkas, Aidyn; Potashnikova, Daria; Arbuz, Galina; Issayev, Zhanpeis; Vorobjev, Ivan A.; Malashenkov, Dmitry V.; Barteneva, Natasha S.
Algal blooms occur in freshwater bodies throughout the world, often leading to fish kills. Cases of these kills along the Ural River were reported in 2018–2019, involving significant amount of sturgeon in fish farming areas. In this study, the analysis of algal samples from the delta of the Ural River up to 100 km inland was carried out from August to December 2019 using imaging flow cytometry (IFC), molecular biological, and microscopic techniques. We identified the filamentous cyanobacteria Cuspidothrix issatschenkoi, Dolichospermum cf. flos-aquae, Dolichospermum cf. macrosporum, Pseudanabaena limnetica, and Planktothrix spp. as the dominant potentially toxic phytoplankton species, and we also found minor quantities of Cylindrospermopsis raciborskii. For the first time, molecular phylogenetic investigations of field clones of cyanobacteria from Ural River were carried out to establish the taxa of the dominant species and to identify the presence of genes encoding toxins. The complementary analysis with nanopore-based next-generation sequencing overlapped with the results of IFC and was instrumental in revealing minor cyanobacteria taxa. Real-time PCR analysis and sequencing indicated the presence of Microcystis and ADA-clade spp. as well as genes associated with the production of microcystin (mcyE) and the algal neurotoxin saxitoxin (sxtA) originating from cyanobacteria. These findings suggest that toxin-producing cyanobacteria could become a threat in the Ural River near Atyrau, which can significantly affect aquaculture in the region.
Open Access
COMPARATIVE CHARACTERIZATION AND IDENTIFICATION OF POLY-3-HYDROXYBUTYRATE PRODUCING BACTERIA WITH SUBSEQUENT OPTIMIZATION OF POLYMER YIELD
(Polymers, 2022) Rysbek, Aidana; Ramankulov, Yerlan; Kurmanbayev, Askar; Richert, Agnieszka; Abeldenov, Sailau
In this work, the strains Bacillus megaterium RAZ 3, Azotobacter chrocococcum Az 3, Bacillus araybhattay RA 5 were used as an effective producer of poly-3-hydroxybutyrate P(3HB). The purpose of the study was to isolate and obtain an effective producer of P(3HB) isolated from regional chestnut soils of northern Kazakhstan. This study demonstrates the possibility of combining the protective system of cells to physical stress as a way to optimize the synthesis of PHA by strains. Molecular identification of strains and amplification of the phbC gene, transmission electron microscope (TEM), extracted and dried PHB were subjected to Fourier infrared transmission spectroscopy (FTIR). The melting point of the isolated P(3HB) was determined. The optimal concentration of bean broth for the synthesis of P(3HB) for the modified type of Bacillus megaterium RAZ 3 was 20 g/L, at which the dry weight of cells was 25.7 g/L􀀀1 and P(3HB) yield of 13.83 g/L􀀀1, while the percentage yield of P(3HB) was 53.75%. The FTIR spectra of the extracted polymer showed noticeable peaks at long wavelengths. Based on a proof of concept, this study demonstrates encouraging results.