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Microbiology Independent Research Journal (MIR Journal)

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Vol 6, No 1 (2019)
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1-9 266
Abstract

The number of infections caused by microorganisms that are resistant to antibiotics and synthetic antibacterial drugs is growing fast worldwide. This is one of the most important and urgent problems in health care. The main efforts of researchers around the world are focused on solving this issue. Nitrofurans represent one of the most effective classes of antibacterial drugs. We have synthesized 4 analogues of nifuroxazide – a well known nitrofuran antibiotic – and confirmed their structures by NMR, IR spectroscopy, and mass-spectrometry. All of the obtained compounds were studied for antimicrobial and antifungal activity.
Activity against Escherichia coli, Staphylococcus aureus, Staphylococcus haemolyticus, and Pseudomonas aeruginosa was evaluated by the agar diffusion method. The synthesized compounds suppressed the growth of all the studied bacterial strains except Escherichia coli; the diameter of the inhibition zones ranged from 13.5 to 28 mm depending on the concentration of the tested compound and bacterial strain. One of the compounds studied in this project – the pyridine analogue of nifuroxazide – exceeded the activity of the standard (nifuroxazide) against the Staphylococcus aureus. The inhibitory activity of the synthesized compounds against the Candida albicans and Cryptococcus neoformans yeasts was determined using the microdilution method. The results were assessed according to the indicator color change. None of the studied compounds showed activity against these cultures.
The obtained results confirm that substituted nifuroxazides have significant antimicrobial activity and, therefore, can be considered as promising candidates for developing new antibacterial drugs.

10-17 361
Abstract

The number of infections caused by microorganisms that are resistant to antibiotics and synthetic antibacterial drugs is growing fast worldwide. This is one of the most important and urgent problems in health care. The main efforts of researchers around the world are focused on solving this issue. Nitrofurans represent one of the most effective classes of antibacterial drugs. We have synthesized 4 analogues of nifuroxazide – a well known nitrofuran antibiotic – and confirmed their structures by NMR, IR spectroscopy, and mass-spectrometry. All of the obtained compounds were studied for antimicrobial and antifungal activity.
Activity against Escherichia coli, Staphylococcus aureus, Staphylococcus haemolyticus, and Pseudomonas aeruginosa was evaluated by the agar diffusion method. The synthesized compounds suppressed the growth of all the studied bacterial strains except Escherichia coli; the diameter of the inhibition zones ranged from 13.5 to 28 mm depending on the concentration of the tested compound and bacterial strain. One of the compounds studied in this project – the pyridine analogue of nifuroxazide – exceeded the activity of the standard (nifuroxazide) against the Staphylococcus aureus. The inhibitory activity of the synthesized compounds against the Candida albicans and Cryptococcus neoformans yeasts was determined using the microdilution method. The results were assessed according to the indicator color change. None of the studied compounds showed activity against these cultures.
The obtained results confirm that substituted nifuroxazides have significant antimicrobial activity and, therefore, can be considered as promising candidates for developing new antibacterial drugs.

18-27 194
Abstract

Influenza A virus belongs to a family of enveloped viruses with an RNA genome of negative polarity consisting of 8 RNA segments. The transcription of this RNA genome results in the synthesis of positive-sense mRNAs that translate up to 16 unique viral proteins with the help of splicing and translational shift mechanisms. The 8th NS segment encodes the NS1 protein (27 kDa), which is an active interferon antagonist, and the nuclear export protein NEP (14 kDa) through the standard negative polarity pathway. In addition, an alternative open reading frame for the synthesis of a third viral protein (NSP, negative-strand protein) by means of a direct translation of genome polarity RNA (the so-called positive polarity genome strategy) was identified in the NS segment. Since it is unknown as to whether the NSP protein can be synthesized in the infected organism post viral infection, the generation of spleen leucocytes specific to this protein was studied in mice after two sequential infections with influenza A viruses of H1N1 and H3N2 subtypes. It was found that leucocyte clones specifically recognizing a peptide domain in the central region of the NSP protein (amino acid positions 82-119) were generated in mice infected with influenza A viruses. In silico prediction has shown strong major histocompatibility complex-1 (MHC-I) and MHC-II specific epitopes in this central domain of the NSP. Comparative analysis of the influenza H3N2 viruses circulating in humans during 1968-2018 has shown high NSP variability, which was similar to that shown for the hemagglutinin (HA) and neuraminidase (NA) proteins. The highest variability was found to be in the N- and C-terminal parts of the NSP. These observations suggest that synthesis of the NSP protein occurs in infected animals and further support a bipolar (ambisense) strategy of the RNA genome of human influenza A virus.

28-36 218
Abstract

Influenza A virus belongs to a family of enveloped viruses with an RNA genome of negative polarity consisting of 8 RNA segments. The transcription of this RNA genome results in the synthesis of positive-sense mRNAs that translate up to 16 unique viral proteins with the help of splicing and translational shift mechanisms. The 8th NS segment encodes the NS1 protein (27 kDa), which is an active interferon antagonist, and the nuclear export protein NEP (14 kDa) through the standard negative polarity pathway. In addition, an alternative open reading frame for the synthesis of a third viral protein (NSP, negative-strand protein) by means of a direct translation of genome polarity RNA (the so-called positive polarity genome strategy) was identified in the NS segment. Since it is unknown as to whether the NSP protein can be synthesized in the infected organism post viral infection, the generation of spleen leucocytes specific to this protein was studied in mice after two sequential infections with influenza A viruses of H1N1 and H3N2 subtypes. It was found that leucocyte clones specifically recognizing a peptide domain in the central region of the NSP protein (amino acid positions 82-119) were generated in mice infected with influenza A viruses. In silico prediction has shown strong major histocompatibility complex-1 (MHC-I) and MHC-II specific epitopes in this central domain of the NSP. Comparative analysis of the influenza H3N2 viruses circulating in humans during 1968-2018 has shown high NSP variability, which was similar to that shown for the hemagglutinin (HA) and neuraminidase (NA) proteins. The highest variability was found to be in the N- and C-terminal parts of the NSP. These observations suggest that synthesis of the NSP protein occurs in infected animals and further support a bipolar (ambisense) strategy of the RNA genome of human influenza A virus.

37-42 235
Abstract

Fighting against pathogenic bacteria that are resistant to antibiotics has become critical for health care worldwide. More than half a million people die every year from infections caused by drug resistant bacteria. Since bacteria acquire resistance to antibiotics very quickly and the development of new antibiotics is a lengthy process, the search for new approaches to stop the spread of bacterial resistance is extremely important. The spread of antibiotic resistance is accomplished mainly by horizontal gene transfer. Scientists are concentrating their efforts on studying the mechanism of this process in order to find a way to stop or reverse it. In this paper, the author gives a brief review of the recent studies on horizontal gene transfer, particularly on incompatibility-based plasmid curing systems. The author examines new possibilities to use the mechanism of horizontal gene transfer for the developing of novel approaches to fight pathogenic bacteria.

43-48 203
Abstract

Viral nucleoprotein (NP) is an abundant essential protein of an influenza virus that has important functional and structural roles. It participates in genomic organization, nuclear trafficking, RNA transcription, and genome replication. From the research point of view, NP is an important protein that is used in the development of new diagnostic methods and vaccination protocols. NP is a promising target for antiviral chemotherapeutic drugs as well. Successful expression of codon-optimized NP genes in E. coli has been reported. In this study, we demonstrated the efficient expression and purification of soluble NPs of influenza A and B viruses in E. coli without the codon-optimization of DNA sequences. This procedure preserves the co-translational protein folding, protein configuration and function. Obtained NPs of influenza A and B viruses were monomers and reacted well with mouse specific antibodies according to Western blot analysis. Our results show that both influenza A and influenza B virus NPs can be efficiently expressed in E. coli without codon-optimization.

49-59 153
Abstract

A broad range of naturally occurring antigenic variants of the influenza virus is caused by its rapid evolutionary variability. The survival of viable influenza virus variants occurs through natural selection. Treatment of influenza infection with modern antiviral drugs – neuraminidase (NA) inhibitors – leads to occurrence of mutations in the NA gene, which result in emergence of virus resistance to these drugs. The goal of this study was to determine the selection pressure on the NA protein of influenza viruses isolated in Ukraine from 2009 to 2015. The main method for assessing the evolutionary pressure on proteins is to quantify the ratio of substitution rates at nonsynonymous (dN) and synonymic (dS) sites. With the help of this method we showed that only a few codons in the NA gene were under the positive selection resulting in mutations at the following sites: for influenza A viruses of A(H1N1)pdm09 subtype – site 40, for viruses of A(H3N2) subtype – sites 93 and 402, for B/Yamagata lineage viruses – sites 74, 99, and 268, and for the B/Victoria lineage viruses – sites 358, 288, and 455. These sites are not associated with NA active site, transmembrane domain, or the antigenic sites of this protein. We concluded that NA inhibitors are not a significant factor in the process of selection of the influenza viruses in Ukraine because sites associated with the resistance of influenza viruses to NA inhibitors were not affected by positive selection. This finding could be explained by the limited use of NA inhibitors for the treatment of influenza infections in Ukraine.

60-69 159
Abstract

A broad range of naturally occurring antigenic variants of the influenza virus is caused by its rapid evolutionary variability. The survival of viable influenza virus variants occurs through natural selection. The treatment of influenza infection with modern antiviral drugs – neuraminidase (NA) inhibitors – leads to the occurrence of mutations in the NA gene, which thereby result in the emergence of virus resistance to these drugs. The goal of this study was to determine the selection pressure on the NA protein of influenza viruses isolated in Ukraine from 2009 to 2015. The main method for assessing the selection pressure on proteins is to quantify the ratio of substitution rates at nonsynonymous (dN) and synonymous (dS) sites. With the help of this method, we showed that only a few codons in the NA gene were under positive selection resulting in mutations at the following sites: for influenza A viruses of the A(H1N1)pdm09 subtype – site 40, for viruses of the A(H3N2) subtype – sites 93 and 402, for Influenza B viruses of the B/Yamagata lineage – sites 74, 99, and 268, and for the viruses of the B/Victoria lineage – sites 358, 288, and 455. These sites are not associated with the NA active site, transmembrane domain, or the antigenic sites of this protein. We concluded that NA inhibitors are not a significant factor in the process of selection of the influenza viruses in Ukraine because the sites associated with the resistance of influenza viruses to NA inhibitors were not affected by positive selection. This finding could be explained by the limited use of NA inhibitors for the treatment of influenza infections in Ukraine. 



ISSN 2500-2236 (Online)