Browsing by Author "Cirkovic, Valentina (7102074128)"

The aim of this prospective cohort study was to determine the prevalence of gut colonization with multidrug-resistant (MDR) bacteria, risk factors for colonization, infection risk, and outcomes among preterm neonates hospitalized at a tertiary-care center in Serbia. During the period from December 2017 to April 2018, 103 neonates were screened for rectal carriage at admission and on the seventh day of life. Characterization of MDR strains was done by conventional microbiology and molecular methods. Out of 61 (59.2%) colonized neonates, 12 (11.6%) were found colonized at admission, while 49 (47.6%) became colonized at the study site. Among a total of 72 MDR isolates, extended-spectrum beta-lactamase (ESBL)-producing enterobacteria prevailed (56/72, 77%), followed by Acinetobacter baumannii (14/72, 19%). The majority of ESBL-producing strains carried multiple genes (blaTEM/blaCTX-M-15 or blaTEM/blaSHV). Longer previous hospitalization and delivery by cesarean section were associated with MDR colonization, while mechanical ventilation was a risk factor for colonization at the study site. Infections due to MDR bacteria were more frequent among colonized than non-colonized neonates, but not significantly, and mortality was low (1%) in the studied neonates. These results indicate that hospitalized preterm neonates in Serbia are rapidly colonized with a diversity of MDR species and resistance phenotypes/genotypes. © 2021 by the authors. Licensee MDPI, Basel, Switzerland.

The aim of this prospective cohort study was to determine the prevalence of gut colonization with multidrug-resistant (MDR) bacteria, risk factors for colonization, infection risk, and outcomes among preterm neonates hospitalized at a tertiary-care center in Serbia. During the period from December 2017 to April 2018, 103 neonates were screened for rectal carriage at admission and on the seventh day of life. Characterization of MDR strains was done by conventional microbiology and molecular methods. Out of 61 (59.2%) colonized neonates, 12 (11.6%) were found colonized at admission, while 49 (47.6%) became colonized at the study site. Among a total of 72 MDR isolates, extended-spectrum beta-lactamase (ESBL)-producing enterobacteria prevailed (56/72, 77%), followed by Acinetobacter baumannii (14/72, 19%). The majority of ESBL-producing strains carried multiple genes (blaTEM/blaCTX-M-15 or blaTEM/blaSHV). Longer previous hospitalization and delivery by cesarean section were associated with MDR colonization, while mechanical ventilation was a risk factor for colonization at the study site. Infections due to MDR bacteria were more frequent among colonized than non-colonized neonates, but not significantly, and mortality was low (1%) in the studied neonates. These results indicate that hospitalized preterm neonates in Serbia are rapidly colonized with a diversity of MDR species and resistance phenotypes/genotypes. © 2021 by the authors. Licensee MDPI, Basel, Switzerland.

Here, we report on a serendipitous finding of a chryso-like virus associated with Culex pipiens mosquitos in the course of study aimed to detect and characterize West Nile virus (WNV) circulating in mosquitos in Serbia, Southern Europe. Upon initial detection of unexpected product in a PCR protocol for partial WNV NS5 gene amplification, further confirmation and identification was obtained through additional PCR and Sanger sequencing experiments. Bioinformatic and phylogenetic analysis identified the obtained sequences as Xanthi chryso-like virus (XCLV). The finding is particular for the fact that it associates XCLV with a new potential vector species and documents a novel geographical area of its distribution. © 2023 by the authors.

Here, we report on a serendipitous finding of a chryso-like virus associated with Culex pipiens mosquitos in the course of study aimed to detect and characterize West Nile virus (WNV) circulating in mosquitos in Serbia, Southern Europe. Upon initial detection of unexpected product in a PCR protocol for partial WNV NS5 gene amplification, further confirmation and identification was obtained through additional PCR and Sanger sequencing experiments. Bioinformatic and phylogenetic analysis identified the obtained sequences as Xanthi chryso-like virus (XCLV). The finding is particular for the fact that it associates XCLV with a new potential vector species and documents a novel geographical area of its distribution. © 2023 by the authors.

West Nile Virus (WNV) is considered one of the most widely distributed arboviruses worldwide. In 2018, Serbia was among the European countries reporting the highest number of WNV cases. This study aimed to characterize WNV strains circulating in Serbia, and to estimate the pathways and dynamics of WNV-2 spread in the Balkans and globally through the phylogenetic approach. Fifty newly generated NS5 Serbian sequences were found to belong to WNV lineage 2. Phylodynamic analyses of the Balkan clade indicated the potential for an increase in genetic diversity and structure of virus populations. Presented phylogeographic analyses implied four instances of long-distance WNV-2 migration from Africa to Europe, during the 1940s to 1950s, while further dissemination of WNV-2 originated in Hungary in mid-1970s and subsequently spread to Austria and Serbia. Extensive dispersion took place during the 1990s–2000s, as the virus spread from Austria to central and western Europe, and from Hungary to the Balkans. Continuous phylogeography analysis of the Balkans WNV-2 clade implied the central role of Serbia for WNV dissemination across the Balkan peninsula. Furthermore, previously undocumented instance of intercontinental migration of WNV-2 from Europe to Asia was implied. In-depth phylogenetic investigation into the global distribution of WNV-2 may provide valuable insights into the prediction and prevention of potential epidemics. © 2024 Wiley Periodicals LLC.

West Nile Virus (WNV) is considered one of the most widely distributed arboviruses worldwide. In 2018, Serbia was among the European countries reporting the highest number of WNV cases. This study aimed to characterize WNV strains circulating in Serbia, and to estimate the pathways and dynamics of WNV-2 spread in the Balkans and globally through the phylogenetic approach. Fifty newly generated NS5 Serbian sequences were found to belong to WNV lineage 2. Phylodynamic analyses of the Balkan clade indicated the potential for an increase in genetic diversity and structure of virus populations. Presented phylogeographic analyses implied four instances of long-distance WNV-2 migration from Africa to Europe, during the 1940s to 1950s, while further dissemination of WNV-2 originated in Hungary in mid-1970s and subsequently spread to Austria and Serbia. Extensive dispersion took place during the 1990s–2000s, as the virus spread from Austria to central and western Europe, and from Hungary to the Balkans. Continuous phylogeography analysis of the Balkans WNV-2 clade implied the central role of Serbia for WNV dissemination across the Balkan peninsula. Furthermore, previously undocumented instance of intercontinental migration of WNV-2 from Europe to Asia was implied. In-depth phylogenetic investigation into the global distribution of WNV-2 may provide valuable insights into the prediction and prevention of potential epidemics. © 2024 Wiley Periodicals LLC.

Background: Usutu virus (USUV) is an emerging mosquito-borne Flavivirus, with birds as the main zoonotic reservoir. Humans are accidental hosts and mostly develop mild or even asymptomatic infections, although severe complications such as encephalitis can also arise. Detailed characterization of the pathogen's phylogenetics may offer valuable insights into the prediction and prevention of potential epidemics; however, lack of uniformity and the number of available USUV sequences worldwide hamper comprehensive investigation. Aim: The study aimed to investigate USUV spatio-temporal dispersal inter- and intracontinentally and to estimate the dynamics of viral spread within Europe. Methods: Phylogeographic and phylodynamic analyses were done using advanced phylogenetic methods implemented in Beast 1.10.4 and Beast 2.6.4 software packages. Results: Herein, we report on a new USUV isolate from Culex pipiens collected in 2019 from Serbia. The results of this research revealed two newly described intercontinental migration events of USUV from Africa to Germany in the 1970s and from Africa to the Middle East (Israel) in the late 90s. Finally, phylodynamic analysis substantiated the ongoing active expansion of USUV in Europe. Conclusion: The data would imply a high potential for further USUV expansion in Europe. Detailed phylogenetic characterization of the pathogen may offer valuable insights into prediction and prevention of potential epidemics; however, lack of uniformity and number of available USUV sequences worldwide hampers comprehensive investigation. This study draws attention to the need for upscaling USUV surveillance. Copyright © 2023 Siljic, Sehovic, Jankovic, Stamenkovic, Loncar, Todorovic, Stanojevic and Cirkovic.

Background: Usutu virus (USUV) is an emerging mosquito-borne Flavivirus, with birds as the main zoonotic reservoir. Humans are accidental hosts and mostly develop mild or even asymptomatic infections, although severe complications such as encephalitis can also arise. Detailed characterization of the pathogen's phylogenetics may offer valuable insights into the prediction and prevention of potential epidemics; however, lack of uniformity and the number of available USUV sequences worldwide hamper comprehensive investigation. Aim: The study aimed to investigate USUV spatio-temporal dispersal inter- and intracontinentally and to estimate the dynamics of viral spread within Europe. Methods: Phylogeographic and phylodynamic analyses were done using advanced phylogenetic methods implemented in Beast 1.10.4 and Beast 2.6.4 software packages. Results: Herein, we report on a new USUV isolate from Culex pipiens collected in 2019 from Serbia. The results of this research revealed two newly described intercontinental migration events of USUV from Africa to Germany in the 1970s and from Africa to the Middle East (Israel) in the late 90s. Finally, phylodynamic analysis substantiated the ongoing active expansion of USUV in Europe. Conclusion: The data would imply a high potential for further USUV expansion in Europe. Detailed phylogenetic characterization of the pathogen may offer valuable insights into prediction and prevention of potential epidemics; however, lack of uniformity and number of available USUV sequences worldwide hampers comprehensive investigation. This study draws attention to the need for upscaling USUV surveillance. Copyright © 2023 Siljic, Sehovic, Jankovic, Stamenkovic, Loncar, Todorovic, Stanojevic and Cirkovic.

Transmission of human immunodeficiency virus (HIV) between individuals may have important legal implications and therefore may come to require forensic investigation based upon phylogenetic analysis. In criminal trials results of phylogenetic analyses have been used as evidence of responsibility for HIV transmission. In Serbia, as in many countries worldwide, exposure and deliberate transmission of HIV are criminalized. We present the results of applying state of the art phylogenetic analyses, based on pol and env genetic sequences, in exploration of suspected HIV transmission among three subjects: a man and two women, with presumed assumption of transmission direction from one woman to a man. Phylogenetic methods included relevant neighbor-joining (NJ), maximum likelihood (ML) and Bayesian methods of phylogenetic trees reconstruction and hypothesis testing, that has been shown to be the most sensitive for the reconstruction of epidemiological links mostly from sexually infected individuals. End-point limiting-dilution PCR (EPLD-PCR) assay, generating the minimum of 10 sequences per genetic region per subject, was performed to assess HIV quasispecies distribution and to explore the direction of HIV transmission between three subjects. Phylogenetic analysis revealed that the viral sequences from the three subjects were more genetically related to each other than to other strains circulating in the same area with the similar epidemiological profile, forming strongly supported transmission chain, which could be in favour of a priori hypothesis of one of the women infecting the man. However, in the EPLD based phylogenetic trees for both pol and env genetic region, viral sequences of one subject (man) were paraphyletic to those of two other subjects (women), implying the direction of transmission opposite to the a priori assumption. The dated tree in our analysis confirmed the clustering pattern of query sequences. Still, in the context of unsampled sequences and inherent limitations of the applied methods, we cannot unambiguously prove that HIV-1 transmission occurred directly between two individuals. Further exploration of the known and suspected transmission cases is needed in order to define methodologies and establish their reliability. © 2016 Elsevier Ireland Ltd

Transmission of human immunodeficiency virus (HIV) between individuals may have important legal implications and therefore may come to require forensic investigation based upon phylogenetic analysis. In criminal trials results of phylogenetic analyses have been used as evidence of responsibility for HIV transmission. In Serbia, as in many countries worldwide, exposure and deliberate transmission of HIV are criminalized. We present the results of applying state of the art phylogenetic analyses, based on pol and env genetic sequences, in exploration of suspected HIV transmission among three subjects: a man and two women, with presumed assumption of transmission direction from one woman to a man. Phylogenetic methods included relevant neighbor-joining (NJ), maximum likelihood (ML) and Bayesian methods of phylogenetic trees reconstruction and hypothesis testing, that has been shown to be the most sensitive for the reconstruction of epidemiological links mostly from sexually infected individuals. End-point limiting-dilution PCR (EPLD-PCR) assay, generating the minimum of 10 sequences per genetic region per subject, was performed to assess HIV quasispecies distribution and to explore the direction of HIV transmission between three subjects. Phylogenetic analysis revealed that the viral sequences from the three subjects were more genetically related to each other than to other strains circulating in the same area with the similar epidemiological profile, forming strongly supported transmission chain, which could be in favour of a priori hypothesis of one of the women infecting the man. However, in the EPLD based phylogenetic trees for both pol and env genetic region, viral sequences of one subject (man) were paraphyletic to those of two other subjects (women), implying the direction of transmission opposite to the a priori assumption. The dated tree in our analysis confirmed the clustering pattern of query sequences. Still, in the context of unsampled sequences and inherent limitations of the applied methods, we cannot unambiguously prove that HIV-1 transmission occurred directly between two individuals. Further exploration of the known and suspected transmission cases is needed in order to define methodologies and establish their reliability. © 2016 Elsevier Ireland Ltd

Abacavir is an effective antiretroviral drug and one of the most commonly used nucleoside reverse transcriptase inhibitors in Serbia. A percentage of the treated patients experience a potentially life-threatening hypersensitivity reaction, which was shown to be associated with the presence of the class I MHC allele, HLA-B∗57:01; hence genotyping for HLA-B∗57:01 prior to starting abacavir is nowadays recommended in international HIV treatment guidelines. In Serbia, this testing became available in 2013. This study was designed to estimate the prevalence of the HLA-B∗57:01 allele in Serbian HIV-1-infected patients. The presence of the HLA-B∗57:01 allele was analyzed in 273 HIV-1-infected patients aged 18 years or more, who were abacavir naïve. Buccal swab samples were obtained from all participants and assayed for the presence of HLA-B∗57:01 using a commercially available HLA-B∗57:01 real-time PCR kit. The presence of the HLA-B∗57:01 allele was found in 22 of 273 tested individuals (8%; 95% CI 5.4-11.9%). This is the first study that estimated the HLA-B∗57:01 prevalence among HIV-infected patients in Serbia. The very high prevalence of HLA-B∗57:01 found in our study strongly supports HLA-B∗57:01 genotyping, which should be implemented prior to the initiation of an abacavir-containing therapy to reduce the risk of potentially life-threatening hypersensitivity reactions. © 2017 S. Karger AG, Basel.

Abacavir is an effective antiretroviral drug and one of the most commonly used nucleoside reverse transcriptase inhibitors in Serbia. A percentage of the treated patients experience a potentially life-threatening hypersensitivity reaction, which was shown to be associated with the presence of the class I MHC allele, HLA-B∗57:01; hence genotyping for HLA-B∗57:01 prior to starting abacavir is nowadays recommended in international HIV treatment guidelines. In Serbia, this testing became available in 2013. This study was designed to estimate the prevalence of the HLA-B∗57:01 allele in Serbian HIV-1-infected patients. The presence of the HLA-B∗57:01 allele was analyzed in 273 HIV-1-infected patients aged 18 years or more, who were abacavir naïve. Buccal swab samples were obtained from all participants and assayed for the presence of HLA-B∗57:01 using a commercially available HLA-B∗57:01 real-time PCR kit. The presence of the HLA-B∗57:01 allele was found in 22 of 273 tested individuals (8%; 95% CI 5.4-11.9%). This is the first study that estimated the HLA-B∗57:01 prevalence among HIV-infected patients in Serbia. The very high prevalence of HLA-B∗57:01 found in our study strongly supports HLA-B∗57:01 genotyping, which should be implemented prior to the initiation of an abacavir-containing therapy to reduce the risk of potentially life-threatening hypersensitivity reactions. © 2017 S. Karger AG, Basel.

Our aim was to investigate gut colonization with carbapenem-resistant Enterobacterales (CRE) in the population of preterm neonates at discharge from a tertiary care center in Serbia. The study included 350 randomly selected neonates/infants discharged in the period April 2018–May 2019. CRE colonization was present in 88/350 (25.1%) of patients. Klebsiella pneumoniae producing KPC and OXA-48 carbapenemase were detected in 45 and 42 subjects, respectively, while NDM producing Escherichia coli was identified in one patient only. All OXA-48 strains harbored blaCTX-M-15, while both blaTEM and blaSHV were present in all but one KPC-producing strain. CRE isolates exhibited a multidrug resistance pattern with uniform fluoroquinolone resistance, universal susceptibility to colistin, and variable susceptibility to aminoglycosides. Administration of carbapenems was common (~50%) and it was strongly associated with colonization, as well as the combinational therapeutic regimens that included meropenem, contrary to ampicillin–sulbactam/colistin therapy and prolonged course of the initial therapy (ampicillin/amikacin ≥ 7 days). Other risk factors for CRE carriage were level of immaturity, admission to neonatal intensive care unit, prolonged hospitalization and invasive procedures. Although the rate of clinically and/or laboratory proven systemic infections was significantly higher among colonized patients, CRE infection was confirmed in one patient only (1.1%) that was colonized with NDM E. coli. Clonal relatedness of CRE isolates was high, with seven and eight clusters detected among KPC (N = 30) and OXA-48 (N = 37) producing strains, respectively. The follow up of the 31 KPC-colonized patients after discharge from hospital revealed common decolonization within one month (~68%). In conclusion, our results demonstrated a high rate of CRE colonization that is most likely related to carbapenem consumption and lack of screening as important infection prevention practice. © 2023 by the authors.

Our aim was to investigate gut colonization with carbapenem-resistant Enterobacterales (CRE) in the population of preterm neonates at discharge from a tertiary care center in Serbia. The study included 350 randomly selected neonates/infants discharged in the period April 2018–May 2019. CRE colonization was present in 88/350 (25.1%) of patients. Klebsiella pneumoniae producing KPC and OXA-48 carbapenemase were detected in 45 and 42 subjects, respectively, while NDM producing Escherichia coli was identified in one patient only. All OXA-48 strains harbored blaCTX-M-15, while both blaTEM and blaSHV were present in all but one KPC-producing strain. CRE isolates exhibited a multidrug resistance pattern with uniform fluoroquinolone resistance, universal susceptibility to colistin, and variable susceptibility to aminoglycosides. Administration of carbapenems was common (~50%) and it was strongly associated with colonization, as well as the combinational therapeutic regimens that included meropenem, contrary to ampicillin–sulbactam/colistin therapy and prolonged course of the initial therapy (ampicillin/amikacin ≥ 7 days). Other risk factors for CRE carriage were level of immaturity, admission to neonatal intensive care unit, prolonged hospitalization and invasive procedures. Although the rate of clinically and/or laboratory proven systemic infections was significantly higher among colonized patients, CRE infection was confirmed in one patient only (1.1%) that was colonized with NDM E. coli. Clonal relatedness of CRE isolates was high, with seven and eight clusters detected among KPC (N = 30) and OXA-48 (N = 37) producing strains, respectively. The follow up of the 31 KPC-colonized patients after discharge from hospital revealed common decolonization within one month (~68%). In conclusion, our results demonstrated a high rate of CRE colonization that is most likely related to carbapenem consumption and lack of screening as important infection prevention practice. © 2023 by the authors.

Background: Hantaviruses are etiological agents of emerging zoonotic diseases worldwide, including hemorrhagic fever with renal syndrome (HFRS). A number of hantavirus species is known to be present in Europe. In Serbia, existing data on hantavirus presence and prevalence rely in serological findings. In this study, molecular analysis was performed in order to characterize HFRS causing hantaviruses in Serbia. Methods: Sixty four serum samples of HFRS cases, previously found seropositive to anti-hantaviral antibodies, were included in the study. Partial hantaviral L and S segments were PCR amplified producing 390nt and 598nt amplicons, respectively, in parallel with human beta-actin mRNA as external reverse transcription positive control. Hantavirus specific PCR products were DNA sequenced in both direction and the obtained sequences phylogenetically confirmed and analyzed. Results: PCR detection of hantavirus L and S genome segments was positive in 18/64 and 11/64 tested samples, respectively. Positive PCR results involved samples obtained from different locations, mostly from central and southern parts of Serbia. All the obtained sequences were identified as Dobrava-Belgrade virus (DOBV). In the phylogenetic analysis sequences from Serbia tended to cluster in distinctive, geographically related clusters. Conclusions: Our findings indicate DOBV as the main HFRS causing hantavirus in Serbia, the site of its initial isolation. © 2019

The HIV/AIDS epidemic in Russia is among the fastest growing in the world. HIV epidemic burden is non-uniform in different Russian regions and diverse key populations. An explosive epidemic has been documented among people who inject drugs (PWID) starting from the mid-1990s, whereas presently, the majority of new infections are linked to sexual transmission. Nationwide, HIV sub-subtype A6 (previously called AFSU) predominates, with the increasing presence of other subtypes, namely subtype B and CRF063_02A. This study explores HIV subtype B sequences from St. Petersburg, collected from 2006 to 2020, in order to phylogenetically investigate and characterize transmission clusters, focusing on their evolutionary dynamics and potential for further growth, along with a socio-demographic analysis of the available metadata. In total, 54% (107/198) of analyzed subtype B sequences were found grouped in 17 clusters, with four transmission clusters with the number of sequences above 10. Using Bayesian MCMC inference, tMRCA of HIV-1 subtype B was estimated to be around 1986 (95% HPD 1984–1991), whereas the estimated temporal origin for the four large clusters was found to be more recent, between 2001 and 2005. The results of our study imply a complex pattern of the epidemic spread of HIV subtype B in St. Petersburg, Russia, still in the exponential growth phase, and in connection to the men who have sex with men (MSM) transmission, providing a useful insight needed for the design of public health priorities and interventions. © 2022 by the authors.

The HIV/AIDS epidemic in Russia is among the fastest growing in the world. HIV epidemic burden is non-uniform in different Russian regions and diverse key populations. An explosive epidemic has been documented among people who inject drugs (PWID) starting from the mid-1990s, whereas presently, the majority of new infections are linked to sexual transmission. Nationwide, HIV sub-subtype A6 (previously called AFSU) predominates, with the increasing presence of other subtypes, namely subtype B and CRF063_02A. This study explores HIV subtype B sequences from St. Petersburg, collected from 2006 to 2020, in order to phylogenetically investigate and characterize transmission clusters, focusing on their evolutionary dynamics and potential for further growth, along with a socio-demographic analysis of the available metadata. In total, 54% (107/198) of analyzed subtype B sequences were found grouped in 17 clusters, with four transmission clusters with the number of sequences above 10. Using Bayesian MCMC inference, tMRCA of HIV-1 subtype B was estimated to be around 1986 (95% HPD 1984–1991), whereas the estimated temporal origin for the four large clusters was found to be more recent, between 2001 and 2005. The results of our study imply a complex pattern of the epidemic spread of HIV subtype B in St. Petersburg, Russia, still in the exponential growth phase, and in connection to the men who have sex with men (MSM) transmission, providing a useful insight needed for the design of public health priorities and interventions. © 2022 by the authors.