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The genes of the major histocompatibility complex ( MHC) are a key component of the mammalian immune system and have become important molecular markers for measuring fitness-related genetic variation in wildlife populations. Because of human persecution and habitat fragmentation, the grey wolf has become extinct from a large part of Western and Central Europe, and remaining populations have become isolated. In Croatia, the grey wolf population, part of the Dinaric-Balkan population, shrank nearly to extinction during the 20th century, and is now legally protected. Using the cloning-sequencing method, we investigated the genetic diversity and evolutionary history of exon 2 of MHC class II DLA-DRB1, DQA1 and DQB1 genes in 77 individuals. We identified 13 DRB1, 7 DQA1 and 11 DQB1 highly divergent alleles, and 13 DLA-DRB1/ DQA1/ DQB1 haplotypes. Selection analysis comparing the relative rates of non-synonymous to synonymous mutations ( dN/ dS) showed evidence of positive selection pressure acting on all three loci. Trans-species polymorphism was found, suggesting the existence of balancing selection. Evolutionary codon models detected considerable difference between alpha and beta chain gene selection patterns: DRB1 and DQB1 appeared to be under stronger selection pressure, while DQA1 showed signs of moderate selection. Our results suggest that, despite the recent contraction of the Croatian wolf population, genetic variability in selectively maintained immune genes has been preserved. [ABSTRACT FROM AUTHOR]

In the previous studies OXA-23-like and OXA-24-like β-lactamase were reported among Acinetobacter baumannii in both hospitals and long-term care facilities (LTCF) in Croatia. The aim of this study was to analyze clinical and sewage A. baumannii isolates from two nursing homes in Zagreb, with regard to antibiotic susceptibility and resistance mechanisms, to determine the route of spread of carbapenem-resistant isolates. Nine clinical isolates were collected from February to May 2017 whereas in April 2017, ten A. baumannii isolates were collected from sewage of two nursing homes in Zagreb. Antibiotics susceptibility was determined by broth microdilution method. The presence of carbapenemase and extended spectrum β-lactamases (ESBL) encoding genes was explored by PCR. Conjugation and transformation experiments were performed as previously described. Genotyping was performed by SG determination, PFGE and MLST. Seven clinical isolates were positive for bla OXA24-like whereas two clinical and environmental carbapenem-resistant isolates, respectively, were found to possess bla OXA-23-like genes. Attempts to transfer imipenem resistance were unsuccessful indicating chromosomal location of bla OXA-23 gene. All carbapenem-resistant isolates belonged to SG- 1 (IC-2) whereas the rest of the isolates susceptible to carbapenems were allocated to SG- 2 (IC-1). PFGE analysis revealed low degree of genetic variability within both IC- I and IC- II. MLST corroborated that two environmental OXA-23 isolates belong to the ST-195. This study showed dissemination of OXA-23 producing A. baumannii from the nursing home into the urban sewage. Disinfection of nursing home sewage should be recommended in order to prevent the spread of resistance genes into the community sewage.

This paper reports the first case of Klebsiella pneumoniae carbapenemase (KPC)-2-producing K. pneumoniae in river water in Croatia. In total, four KPC-2-producing K. pneumoniae isolates were analysed. All isolates shared a similar genetic background, belonging to ST258. Isolates displayed uniform, multi-drug-resistant profiles susceptible to colistin. bla SHV-1 , aac(3')-II, aac(6')-Ib and aph(3')-Ia genes were detected in all isolates. In all isolates, the bla KPC-2 gene was localized on a single non-conjugative IncFII plasmid that varied in size (∼140, ∼230, ∼225 and ∼220 kb). K. pneumoniae was viable in river water for up to 50 days, confirming its ability to survive and disseminate in the environment., (Copyright © 2019 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.)

Background: Acinetobacter baumannii is one of the most prevalent causes of severe hospital-acquired infections and is responsible for the dramatic increase in carbapenem resistance in Croatia in the last 5 years. Such data have encouraged multicenter research focused on the organism's ability to form biofilm, susceptibility to antibiotics, and particular genotype lineage., Methods: Biofilm formation in 109 unrelated clinical isolates of A. baumannii recovered in six cities of Southern Croatia was investigated. Genotyping was performed by pulsed-field gel electrophoresis and antibiotic profile was tested by applying the disc diffusion method and confirmed by determining the minimum inhibitory concentrations. The ability to form biofilm in vitro was determined from overnight cultures of the collected isolates on microtiter plates, after staining with crystal violet, and quantified at 570 nm after solubilization with ethanol. The statistical relevance was calculated in an appropriate program with level of statistical confidence., Results: There was no significant difference in biofilm formation due to the genotype lineage. Isolates collected from intensive care units (ICUs) and isolated from respiratory samples were more likely to create a biofilm compared with isolates from other departments and other samples. There was a significant difference in the ability to produce biofilm in relation to antibiotic resistance pattern. A large proportion of A. baumannii isolates that were resistant to ampicillin/sulbactam, carbapenems, and amikacin were found to be biofilm-negative. In contrast, isolates susceptible and intermediately susceptible to ampicillin/sulbactam, carbapenems, and amikacin were biofilm producers., Conclusion: Clinical isolates of A. baumannii from respiratory samples in ICUs with a particular susceptibility pattern are more prone to form biofilm.

The relationship between the physiological group of gelatinolytic bacteria and the abundance of heterotrophic bacteria in freshwater ecosystems was described, based on analysis of 1082 different freshwater samples collected in Croatia. Percentages of gelatinolytic bacteria among the population of heterotrophic bacteria showed a significant negative correlation with the abundance of heterotrophic bacteria. The relation between the physiological group (gelatinolytic bacteria) and heterotrophic bacteria can be considered to be an indicator of the pollution degree of freshwaters. A high relative content of gelatinolytic bacteria (> 76%) always indicates the colony-forming units of heterotrophic bacteria < 1000/mL, which corresponds to the high water quality; gelatinolytic bacteria < 11% indicate polluted waters. Isolated strains of aerobically grown gelatinolytic bacteria were Gram-negative rod-shaped or Gram-positive endospore-forming rod-shaped cells.

In a cross-sectional study of growth, 5,155 children (2,591 females, 2,564 males) from the town of Zagreb (Croatia) were measured. Four traits of linear dimensionality (stature, sitting height, arm and leg lengths) were studied in the age span of 3 to 18 years. A significant average annual increase of all four anthropometric parameters were observed up to 14 and 15 years of age in girls and 16 years of age in boys, showing that girls had a shorter growing period. In the prepubertal period until 9 years of age, gender differences were negligible. At the age of 10, boys were overgrown by girls in all parameters due to the earlier onset of puberty in girls. The growth gains for girls, when compared with those for boys, show a different pattern across variables. The female growth advantage remained in a two years period for the limbs length, but in a three year period for stature and the longest, for 4 years, for sitting height. The male predominance in size had an onset at the age of 13 for the limbs and in the age of 14 for stature and sitting height. The patterns of sexual dimorphism in stature and sitting height during growing years are similar to those observed in other populations of Europe. Growth of Croatian children and youth is very similar to that of the tallest European populations.