Measles outbreak, Montenegro January-July 2018: Lessons learned.

In 2017, the Regional Verification Commission for Measles and Rubella Elimination (RVC) of the World Health Organization confirmed that measles elimination was sustained in Montenegro, and the previous endemic transmission remained interrupted. However, the RVC was extremely concerned over the continuing low vaccination coverage reported for this country. In this study, we describe the most recent measles epidemic in Montenegro using the epidemiological data collected from January 1 to July 31, 2018. The outbreak is largely attributable to a dangerous accumulation of susceptible subjects across the country and represents a high-risk factor for re-establishing endemic transmission in the Balkan area. This study showed how a vaccine-preventable communicable disease outbreak can have a dramatic impact and severe consequences on regional public health system performance in terms of the sanitary spending point of view. A detailed update is provided on the epidemiological situation in this Central European area, not available until now.

Hepatitis C virus genotype 3A in a population of injecting drug users in Montenegro: Bayesian and evolutionary analysis.

Few reports are available on HCV molecular epidemiology among IDUs in Eastern Europe, and none in Montenegro. The aim of this study was to investigate the HCV genotype distribution in Montenegro among IDUs and to perform Bayesian and evolutionary analysis of the most prevalent HCV genotype circulating in this population. Sixty-four HCV-positive IDUs in Montenegro were enrolled between 2013 and 2014, and the NS5B gene was sequenced. The Bayesian analysis showed that the most prevalent subtype was HCV-3a. Phylogenetic data showed that HCV-3a reached Montenegro in the late 1990s, causing an epidemic that exponentially grew between the 1995 and 2005. In the dated tree, four different entries, from 1990 (clade D), 1994 (clade A) to 1999 (clade B) and 2001 (clade C), were identified. In the NS5B protein model, the amino acids variations were located mainly in the palm domain, which contains most of the conserved structural elements of the active site. This study provides an analysis of the virus transmission pathway and the evolution of HCV genotype 3a among IDUs in Montenegro. These data could represent the basis for further strategies aimed to improve disease management and surveillance program development in high-risk populations.

Genetic diversity of the haemagglutinin (HA) of human influenza a (H1N1) virus in montenegro: Focus on its origin and evolution.

In 2009 an influenza A epidemic caused by a swine origin H1N1strain, unusual in human hosts, has been described. The present research is aimed to perform the first phylogenetic investigation on the influenza virus A (H1N1) strains circulating in Montenegro, from December 1, 2009, when the first case of death due to H1N1 was confirmed, and the epidemic began causing a total of four fatalities. The phylogenetic analysis of the strains circulating showed the absence of a pure Montenegrin cluster, suggesting the occurrence of multiple re-introductions in that population from different areas till as far as the early 2010. The time to most recent common ancestor (TMRCA) for the complete dataset has been dated in early 2008, pre-dating the first Montenegrin identification of H1N1 infection. These data suggest that virus was spreading undetected, may be as a consequence of unidentified infections in returning travelers. Anyhow, the estimated TMRCA of Montenegrin strains is fully consistent to that found in different areas. Compatibly with the time coverage of the study period here analyzed, molecular dynamic of Montenegrin strains follows similar trend as in other countries. J. Med. Virol. 88:1905-1913, 2016. © 2016 Wiley Periodicals, Inc.

Hepatitis B virus genotype and subgenotype prevalence and distribution in Montenegro.

The Mediterranean area and the Balkans in particular show the highest level of genetic heterogeneity of HBV in Europe. Data about the circulation of HBV genotypes in Montenegro are lacking. It was studied the prevalence and distribution of HBV genot/subgenotypes in a total of 150 HBV infected patients living in Montenegro. Phylogenetic analysis of 136 successfully amplified P sequences showed a high degree of genetic heterogeneity of HBV in Montenegro. Subgenotype D2 (36.8%) and D3 (32.3%) were the most prevalent, followed by genotype A (subgenotype A2 in all of the cases-19.8%). Eight patients were infected with recombinant strains. HBV-D1 which is the most spread HBV subgenotype in the south-eastern Mediterranean countries, seems to be relatively rare in Montenegro, suggesting a penetration of HBV more probably from North-East or West than from Eastern Mediterranean countries. The relatively different prevalence of D3 and A2 among subjects infected through sexual route, seems to confirm the association of these subgenotypes with different route of transmissions (mainly parenteral for D3 and mainly sexual for A2) even in Montenegro. The low prevalence of D2 among children and its absence in perinatal transmission, suggests that this subgenotype circulated prevalently in the past. If this is due to changes in the most prevalent way of transmission and in the recent different contacts of Montenegro with other European countries, it remains to be established by other larger studies.

Human papillomavirus genotype spectrum in studied group of Montenegrin women.

The risk of high-grade lesions and carcinoma is known to correlate with specific high-risk HPV genotypes. The distribution of HPV types varies between countries and little is known about HPV distribution in our country. Therefore, the purpose of this study was to determine the range and frequency of HPV genotypes in studied group of women in Montenegro. HPV genotypes were determined using the method of enzyme restriction of PCR products amplified with group-specific primers MY09/MY11 and restricted with seven different restriction endonucleases. Out of the total number of women HPV infection was found in 1/5 of participants (20%). Genotyping performed in HPV DNA positive women shows that the HPV genotype 16 is dominant and present in more than 1/3 of the participants (36.8%). The second most frequent HPV infection is with HPV genotype 58 and it is found in 10.5% of participants. HPV 31 and HPV 6 infections are present in 7.9% of women, while infections with other genotypes were demonstrated individually by 2.6%. Multiple HPV infection was demonstrated in 18.4% and they are dominant in younger women (aged 25 to 30 years).

SARS-CoV-2 Omicron Variant in Croatia-Rapid Detection of the First Case and Cross-Border Spread.

BACKGROUND: Due to rapid spread, the Omicron variant has become the dominant SARS-CoV-2 variant responsible for infections worldwide. We present the first detection of the Omicron variant in Croatia which resulted in rapid cross-border spreading. METHODS: Whole-genome sequencing was performed using the Illumina MiniSeq sequencing system. SARS-CoV-2 lineages were identified using the PANGOLIN and GISAID databases. RESULTS: The first case of the Omicron variant (BA.1.17) emerged in Croatia after a workshop held in Zagreb in November 2021. The patient reported a history of previous COVID-19 and received two doses of an mRNA vaccine. Three additional cases were detected among Croatian participants of the workshop. At the beginning of December, SARS-CoV-2 infection was confirmed in one participant from Montenegro and her husband. Phylogenetic analysis showed that the detected Omicron variants were closely related to the first Croatian case, confirming the connection with the workshop outbreak and rapid cross-border spreading. Subsequent analyses of SARS-CoV-2 positive samples in Croatia showed the rapid introduction of the Omicron variant and depletion of the Delta variant resulting in the fifth pandemic wave. CONCLUSIONS: Genomic monitoring and early detection of novel SARS-CoV-2 variants are essential to implement timely epidemiological interventions and reduce further transmission in the population.

Severe acute respiratory infection surveillance in Montenegro, 2014-2017.

AIM: The study aim was to analyze the epidemiology and clinical characteristics of severe acute respiratory infection (SARI) cases and to compare demographic and clinical characteristics as well as outcomes of influenza-positive SARI cases to those of influenza-negative SARI cases in Montenegro. METHODS: SARI surveillance was established in 2014 in nine healthcare institutions. Retrospective analysis of case-based surveillance data pertaining to all reported SARI cases during three seasons was conducted. RESULTS: Among the 90 identified SARI cases, 64 (71%) were influenza positive. Death outcome was reported in 25 (28%) of all registered SARI cases. Cardiovascular disease was more prevalent among the patients in the influenza-positive SARI group (36% vs. 12%, p = .021), as was concurrence of two or more chronic medical conditions (57% vs. 30%, p = .042). These patients were also more likely to be immunocompromised (16% vs. 0%, p = .057) and have viral pneumonia (14.4% vs. 20.3%, p = .017), compared to those in the influenza-negative SARI group. Younger age, presence of cardiovascular disease and being immunocompromised were patient characteristics independently associated with SARI related to influenza. CONCLUSION: Continued and extended monitoring of SARI is necessary in order to fully assess the burden of flu disease, define risk groups and establish better control measures.

Antibiotic susceptibility of Acinetobacter species in intensive care unit in Montenegro.

The global increase in multidrug resistance of Acinetobacter has created widespread problems in the treatment of patients in intensive care units (ICUs). The aim of this study was to assess the current level of antimicrobial susceptibility of Acinetobacter species in ICU of Clinical Centre of Montenegro and determine their epidemiology. Antibiotic susceptibility was tested in 70 isolates of Acinetobacter collected from non-repeating samples taken from 40 patients. The first nine isolates were genotyped by repetitive sequence-based PCR (rep-PCR). Tigecycline was found to be the most active antimicrobial agent with 80.6% of susceptibility. All the isolates were multidrug resistant with fully resistance to cefalosporinas, piperacillin and piperacillin/tazobactam. More than half of them (58.5%) were probably extensively resistant. Seven out of nine examined strains were clonally related by rep-PCR. Our results showed extremely high rate of multidrug resistance (MDR) of Acinetobacter isolates and high percentage of its clonally spreading.

Tachycardia in a newborn with enterovirus infection.

Enterovirus infections are common in the neonatal period. Newborns are at a higher risk of severe disease including meningoencephalitis, sepsis syndrome, cardiovascular collapse, or hepatitis. The mechanism of heart failure in patients with enterovirus infection remains unknown. Early diagnosis may help clinicians predict complications in those infants initially presenting with severe disease. An 11-day-old male newborn was admitted to our neonatal intensive care unit because of tachycardia and crises of cyanosis. His elder brother had febrile illness. The newborn was cyanotic, in respiratory distress, with tachycardia, low blood pressure and prolonged capillary refilling time. Limb pulse oximeter was around 85%. During the first day of hospitalization, the newborn had one febrile episode. Laboratory data: elevated transaminases, markers of inflammation negative, all bacterial cultures negative. Enterovirus RNA was detected in blood sample. Other blood findings were without significant abnormalities. Electrocardiogram showed tachycardia, with narrow QRS complexes (atrial tachycardia) and heart rate up to 280/min. In order to convert the rhythm, the patient was administered adenosine and amiodarone. In the further course of hospitalization, the patient was in good general condition, eucardiac and eupneic. Newborns with tachycardia and a family history of febrile illness should be suspected to have enterovirus infection. Enterovirus infection is a highly contagious and potentially life-threatening infection if not detected early. The use of sensitive molecular-based amplification methods offers potential benefits for early diagnosis and timely treatment.

Reconstruction of the evolutionary dynamics of hepatitis C virus subtypes in Montenegro and the Balkan region.

More than 20 million hepatitis C virus (HCV) carriers live in the countries of the Eastern Mediterranean. We determined HCV genotype distribution among chronically infected patients in Montenegro and investigated the phylodynamics and phylogeography of the most represented HCV subtypes. The HCV-NS5b sequences of the Montenegrin patients were compared with sequences isolated in different known localities of the Mediterranean area, Europe and Asia. A Bayesian approach was used in order to allow the simultaneous estimate of the evolutionary rate, time-scaled phylogeny, demography and ancestral spatial status. The most frequent HCV subtypes among the Montenegrin patients, were 1b (34.7%) and 3a (24.7%), but there was also a significant prevalence of 1a and 4d (19.5%). Subtype 3a was significantly more frequent among younger patients and intravenous drug users (IDUs), whereas subtype 1b was more frequently associated with iatrogenic exposure and older ages. The spatio-temporal analysis of the epidemic suggested that HCV-1b penetrated Europe at the beginning of the XX century, probably through Greece and Cyprus and in the 1920s reached Montenegro, where there was an exponential increase in the effective number of infections between the 1950s and 1970s. The phylogeographic and phylodynamic analysis of HCV 3a showed that its most probable origin was in the Indian sub-continent (Pakistan in our reconstruction) about 300years ago. The evolutionary dynamics analysis showed that HCV-3a reached Montenegro more recently in the late 1970s and underwent multi-phasic growth still persisting. Our data suggest multiple introduction of HCV subtypes in the area, supported by different causes of dispersion: adverse social conditions and unsafe medical practices for HCV-1b and i.v. drug use for HCV-3a.

Phylogeographic reconstruction of HIV type 1B in Montenegro and the Balkan region.

Human immunodeficiency virus (HIV) is one of the most genetically variable human viruses as it is characterized by high rates of mutation, viral replication, and recombination. Phylodynamics is a powerful means of describing the behavior of an infection as a combination of evolutionary and ecological processes. Only a few studies of HIV-1 molecular epidemiology have so far been carried out in the Balkans. In this study, we used Bayesian methods to reconstruct the phylogeography and phylodynamics of HIV-1B in Montenegro and some other Balkan countries on the basis of pol gene sequences retrieved from a public database. The phylogenetic analysis showed that 43% of the isolates grouped in accordance with their geographic area, whereas the majority were interspersed in the tree, thus confirming the multiple introductions of HIV-1B in the Balkans. The Bayesian phylogeographic analysis suggested that HIV-1B entered the Balkans in the early 1970s probably through Greece and other Mediterranean tourist/travel destinations (such as Slovenia). Other Balkan countries, such as Bulgaria and Serbia, may have played an important role in spreading the infection to the entire Eastern Mediterranean area, and possibly to Northeast Europe. This suggests that the Balkans may have played a role as a "gateway" between Western and Eastern Europe.

Genetic diversity of HIV type 1 in Montenegro.

Human immunodeficiency virus type 1 (HIV-1) is characterized by high genetic variability due to its high replication rate and the lack of proofreading activity of the reverse transcriptase enzyme. On the basis of phylogenetic analysis performed on numerous isolates from all over the world, HIV-1 is subdivided into types, subtypes, subsubtypes, circulating recombinant forms, and unique recombinant forms. No data are currently available about the circulation of HIV-1 types in Montenegro. Here, we describe the genetic variability of HIV-1 strains identified in plasma samples of patients from Montenegro. Phylogenetic analysis on 32 HIV-1 sequences was carried out. The prevalent circulating HIV-1 subtype is B. The strains were interspersed within the tree. Two main clades (I and II) may suggest independent introductions of HIV-1 subtype B into Montenegro, although other epidemiological evidence will be needed to assume a small number of introductions. No obvious evidence of clustering by residence, age, or sex was found (data not shown). Nelfinavir resistance was found, though lopinavir is the only PI administered. Continuous monitoring of HIV-1-infected individuals is crucial to a better understand of the epidemiology of the B subtype in Montenegro.