Complete Genome Sequencing of Acinetobacter sp. Strain LoGeW2-3, Isolated from the Pellet of a White Stork, Reveals a Novel Class D Beta-Lactamase Gene.

Whole-genome sequencing of Acinetobacter sp. strain LoGeW2-3, isolated from the pellet of a white stork (Ciconia ciconia), reveals the presence of a plasmid of 179,399 bp encoding a CRISPR-Cas (clustered regularly interspaced short palindromic repeats and associated genes) system of the I-F type, and the chromosomally encoded novel class D beta-lactamase OXA-568.

GENOME ANNOUNCEMENT

While studying the ecology of the nosocomial pathogen Acinetobacter baumannii (1), we isolated Acinetobacter sp. strain LoGeW2-3 from the pellet of a white stork (Ciconia ciconia) collected in Loburg, Germany, in the year 2015, following recently described protocols (2). Since partial 16S rRNA and rpoB gene sequencing (GenBank accession no. KT809317 and KT809318, respectively) did not indicate the strain’s belonging to any described species, PacBio RS single-molecule real-time (SMRT) sequencing was commissioned at GATC (Konstanz, Germany). Genomic DNA was isolated as recently described (3). SMRT sequencing resulted in 84,981 reads with a total of 1,145,091,354 sequenced bases and 276-fold coverage. Genome assembly using the Hierarchical Genome Assembly Process (HGAP) version 3 yielded a circular chromosome with a size of 3,178,335 bp and a circular plasmid of 179,399 bp. NCBI Prokaryotic Genome Annotation Pipeline analysis revealed a total of 3,240 genes, including 3,122 coding sequences and 118 RNA genes, of which 93 define tRNAs, as well as 7 complete rRNA gene sets and 4 noncoding RNAs. PacBio modification and motif analysis identified N-6-methylated adenines in motifs TGAANNNNNCTG and CAGNNNNNTTCA and an unknown modification within the motif VNCGGTGTANND (modified bases underlined).

The plasmid encodes a CRISPR-Cas (clustered regularly interspaced short palindromic repeats and associated genes) system of the I-F type (4). CRISPRDetect version 2.1 (5) identified a CRISPR array ranging from nucleotides 50488 to 45838 in reverse orientation on the plasmid and harboring 77 spacer sequences with a predominant length of 32 nucleotides. Most of the spacer sequences (61%) show highest similarity to database entries of eukaryotic origin, casting into doubt a role in targeting plasmid and phage sequences.

Twenty putative genomic islands were predicted on the chromosome by at least one method applying IslandViewer version 4 (6).

Acinetobacter species found in diverse environmental habitats are considered to contribute to the mobilization of antibiotic resistance genes into clinically relevant Acinetobacter species (7–9). A search for putative resistance genes in the genome of Acinetobacter sp. strain LoGeW2-3 applying ResFinder version 3.0 (10) revealed the presence of a beta-lactamase gene with an overall identity of 79% to blaOXA-363 of Acinetobacter lwoffii. The novel allele received the designation blaOXA-568 and the product was named class D beta-lactamase OXA-568. In its native background, no resistance phenotype could be attributed to blaOXA-568 following standard procedures, as recently described (11).

Average nucleotide identity calculations based on BLAST+ (ANIb) analysis (12) indicate that A. schindleri CIP 107287 is the closest relative for which whole-genome data are available (87.11% identity with 80.65% of the chromosome of Acinetobacter sp. strain LoGeW2-3 aligned) and support our assumption that strain LoGeW2-3 is the first representative of a novel Acinetobacter species.

Accession number(s).

The complete genome sequence of Acinetobacter sp. strain LoGeW2-3 has been deposited at GenBank under the accession no. CP024011 (chromosome) and CP024012 (plasmid).