A comparative study of ultrasound-assisted extraction (USE) with the mixture pentane:ether (1:2) and hydrodistillation (HD) with the same trapping mixture is presented for the isolation of volatile compounds from two unifloral honeys of Robinia pseudoacacia L. and Castanea sativa L. All HD isolates contained many thermally derived artefacts (especially phenylacetaldehyde with lower percentages of furfural, cis- and trans-linalool oxides and others). USE method gave the most representative profile of all honey volatiles (without artefacts). In addition, USE enabled extraction of low molecular weight semivolatile markers (especially benzoic, vanillic and phenylacetic acids) that were not extracted by HD. In this regard, low percentage of benzoic acid (0.7-7.4%), vanillic acid (0.0-1.6%) and phenylacetic acid (0.5-4.1%) was determined in Rp USE extracts, while Cs USE extracts contained phenylacetic acid (20.2-23.5%) as the major constituent with low percentage of benzoic acid (2.5-5.5%).

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11%]). Overexpression of the natural AmpC cephalosporinase was common (42%), but only a few ST235 or ST111 isolates produced VIM-1 or VIM-2 metallo-␤-lactamases or PER-1 or GES-7 extendedspectrum ␤-lactamases.

This report describes that the early stage of KPC-producing K. pneumoniae dissemination in Croatia is associated with a prolific PFGE type belonging to ST258. So far, the spread of an outbreak strain is limited to the northwest region of the country.

Here, we report a retrospective study conducted to elucidate emergence, epidemiology, and molecular mechanisms of resistance underlying the early spread of OXA-48 carbapenemase-producing Enterobacteriaceae in Croatia. Retrospective screening for OXA-48 producers was performed on a collection of 296 nonrepetitive, carbapenem-nonsusceptible enterobacterial isolates collected from January 2011 to December 2012 from 40 participating centers in Croatia. Antimicrobial susceptibility profiles and production of carbapenemases were assessed phenotypically. Pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing were used for epidemiological analysis. Resistance genes were characterized by polymerase chain reaction (PCR) and sequencing. Plasmid localization of bla in isolates and transconjugants was investigated by S1-PFGE and Southern hybridization. PCR mapping was used for identification of genetic platform surrounding bla. Out of 296 carbapenem-nonsusceptible isolates, bla gene was detected in 12 Klebsiella pneumoniae isolates. All OXA-48-producing isolates showed varying resistance to carbapenems and 11 were multidrug resistant. All coproduced additional beta-lactamases, including CTX-M-15, which was detected in eight isolates. Isolates were delineated in five clonal types by PFGE corresponding to five sequence types (STs) assigned ST15, ST16, ST37, ST528, and ST1418. All OXA-48 isolates conjugated successfully and other resistance determinants were not cotransferred. bla was carried on a ∼60 kb IncL/M plasmid and was detected within Tn1999.2 composite transposon. OXA-48, a class D carbapenemase, is emerging as a potentially significant contributor among carbapenem-resistant Enterobacteriaceae in Croatia, alongside class A and B carbapenemases. Polyclonal genetic background of K. pneumoniae isolates carrying ∼60 kb incL/M plasmid indicates that dissemination of the bla gene is not driven exclusively by the spread of a single clone.