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Introduction: Carbapenem resistance in Acinetobacter baumannii can be mediated by carbapenemases of class A, class B metallo-β-lactamases (MBLs), and class D carbapenem-hydrolyzing oxacillinases (CHDL). The aim of the study was to investigate the antimicrobial susceptibility and β-lactamase production of carbapenem-resistant A. baumannii isolates (CRAB) from the Children’s Hospital Zagreb, Croatia. Methods: A total of 12 A. baumannii isolates collected between August 2016 and March 2018 were analyzed. Antibiotic susceptibility was determined by the broth microdilution method. The presence of MBLs was explored by combined disk test with EDTA. The presence of carbapenemases of class A, B, and D was explored by PCR. The occurrence of the ISAba1 upstream of the blaOXA-51-like or blaOXA-23-like was determined by PCR mapping. Epidemiological typing was performed by determination of sequence groups (SG). Genotyping was performed by SG determination, rep-PCR, and MLST. Results: All CRAB were resistant to piperacillin/tazobactam, ceftazidime, cefotaxime, ceftriaxone, cefepime, imipenem, meropenem, gentamicin, and ciprofloxacin. Moderate resistance rates were observed for ampicillin/sulbactam (67%) and tigecycline (42%). The isolates were uniformly susceptible to colistin. PCR revealed the presence of genes encoding OXA-24-like CHDL in nine and OXA-23-like CHDL in three isolates. blaOXA-51 genes were preceded by ISAba1. PCR for the common MBLs in Acinetobacter was negative. All isolates belonged to SG 1 corresponding to ICL (International Clonal Lineage) II. Rep-PCR identified four major clones. Conclusions: The study found OXA-24-like β-lactamase to be the dominant CHDL among children’sCRAB. The predominant spread of OXA-24-like is in contrast with the recent global dissemination of OXA-23 reported all over the world. In contrast to the previous studies in which emergency of OXA-24-like positive isolates was monoclonal, we found considerable genetic diversity of the isolates.

Diatoms and bacteria are known for being the first colonizers of submerged surfaces including the skin of marine reptiles. Sea turtle carapace and skin harbour diverse prokaryotic and eukaryotic microbes, including several epizoic diatoms. However, the importance of diatom-bacteria associations is hardly investigated in biofilms associated with animal hosts. This study provides an inventory of diatoms, bacteria, and diatom-associated bacteria originating from loggerhead sea turtles using both metabarcoding and culturing approaches. Amplicon sequencing of the carapace and skin samples chloroplast gene rbcL and 16S rRNA gene detected in total 634 diatom amplicon sequence variants (ASVs) and 3,661 bacterial ASVs indicating a high diversity. Cultures of putative epizoic and non-epizoic diatoms contained 458 bacterial ASVs and their bacterial assemblages reflected those of their host. Diatom strains allowed for enrichment and isolation of bacterial families rarely observed on turtles, such as Marinobacteraceae, Alteromonadaceae, and Alcanivoracaceae. When accounting for phylogenetic relationships between bacterial ASVs, we observed related diatom genera might retain similar microbial taxa in culture, regardless of the turtle's skin or carapace source. These data provide deeper insights into the sea turtle-associated microbial communities, and reveal the potential of epizoic biofilms as a source of novel microbes and possibly important diatom-bacteria associations.

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Diatoms and bacteria are known for being the first colonizers of submerged surfaces including the skin of marine reptiles. Sea turtle carapace and skin harbour diverse prokaryotic and eukaryotic microbial taxa, including several epizoic diatoms. However, the importance of diatom-bacteria associations is hardly investigated in biofilms associated with animal hosts. This study provides a detailed inventory of diatoms, bacteria, and diatom-associated bacteria originating from several loggerhead sea turtles using a combination of metabarcoding and culturing approaches. Carapace and skin samples rbcL and 16S rRNA amplicon sequencing showed a high diversity of diatoms and bacteria, respectively. Cultures of putative epizoic and non-epizoic diatom strains contained from 18 to 101 bacterial amplicon sequence variants (ASVs) and their bacterial assemblages strongly reflected those of their source host. Diatom strains allowed for enrichment and isolation of rare-in-source bacterial families such as Marinobacteraceae, Alteromonadaceae, and Alcanivoracaceae. When accounting for phylogenetic relationships between bacterial ASVs, we observed related diatom genera might retain related microbial taxa in culture, regardless of the source environment. These data provide deeper insights into several levels of sea turtle epizoic diatom and bacterial communities, and reveal the potential of epizoic biofilms as a source of novel microbes and possibly important diatom-bacteria associations.