Complete Genome Sequence of Citricoccus sp. Strain SGAir0253, Isolated from Indoor Air in Singapore

Citricoccus sp. strain SGAir0253 was isolated from indoor air collected in Singapore. Its genome sequence was assembled using single-molecule real-time sequencing. It comprises one chromosome of 3.32 Mb and two plasmids of 137 kb and 99 kb. The genome consists of 2,950 protein-coding genes, 49 tRNAs, and 9 rRNAs.

M embers of the bacterial genus Citricoccus are Gram-positive, nonmotile bacteria classified in the phylum Actinobacteria, family Micrococcaceae (1). The first reported draft genome sequence of a Citricoccus sp. was of an isolate from the Cuatro Cienegas Basin (CCB) in Coahuila, Mexico (2). The members of this genus have been isolated and reported from various environments, such as marine macroalgae (3), mold-colonized walls (4), saline wastewater bioreactors (5), wastewater treatment plants (6), medieval wall paintings (1), desert soil (7), and human skin (8). There are no previous reports indicating the presence of Citricoccus spp. in air samples, and therefore, this work provides the first evidence that members of this genus can also be found in air.
The strain SGAir0253 was isolated from an indoor air sample collected in Singapore (global position system coordinates, 1.304842°N, 103.791614°E) using the Andersen single-stage impactor (SKC, USA). The air was impacted onto marine agar (Becton, Dickinson, USA), and further isolation of colonies was carried out by culturing onto Trypticase soy agar (TSA) at 30°C. A single colony was grown in lysogeny broth (Becton, Dickinson) at 30°C overnight and subjected to DNA extraction using the Wizard genomic DNA purification kit (Promega, USA), according to the manufacturer's protocol. Library preparation was performed with the SMRTbell template prep kit 1.0 (Pacific Biosciences), followed by single-molecule real-time (SMRT) sequencing on the Pacific Biosciences RS II platform.
For the following analysis, default parameters were used for all software unless stated otherwise. Quality control of raw reads was done with PreAssembler filter v1 within the Hierarchical Genome Assembly Process (HGAP) version 3 protocol (9) implemented in the PacBio SMRT Analysis 2.3.0 package. HGAP was also used for the de novo assembly of 78,591 subreads, followed by polishing with Quiver (9). The consensus assembly generated three contigs (Table 1). All three contigs were circularized using Circlator 1.1.4 (10).
Taxonomic identification was performed using average nucleotide identity (ANI) analysis (11) against a database of bacterial refseq genomes that was created using a text filter for "type, synonymtype, proxytype," which showed 82% identity with Tepidibacter formicigenes. Phyla-AMPHORA (12) was run using MarkerScanner.pl with an added -DNA flag and MarkerAlignTrim.pl with options -With reference and -output-Format phylip. Phylotyping.pl was run with default parameters, which showed 89.4% identity to Micrococcus luteus. As the identity was below the threshold for species-level identification for ANI results, we also performed 16S identification by extracting the full-length 16S rRNA gene sequence with Barrnap 0.7 (13). The BLASTn (14) alignment of the 16S rRNA gene sequence to the Silva database resulted in 98.7% identity to Citricoccus sp. strain CH26A. Hence, only genus-level assignment was done for this strain.
The NCBI Prokaryotic Genome Annotation Pipeline (PGAP) version 4.2 (15) was used for genome annotation. A total of 3,134 genes were predicted, comprising 2,950 protein-coding genes (PCGs), 9 rRNA operons (3 each of 5S, 16S, and 23S rRNAs), 49 tRNAs, 3 noncoding RNAs, and 123 pseudogenes. Annotation for plasmid 1 predicted proteins carrying antibiotic resistance (class A beta-lactamase, major facilitator superfamily [MFS], and tetracycline [TetR]), as well as protein CopG, which is responsible for copy number regulation of the plasmid. Plasmid 2 contains metal resistance proteins (copper and arsenic). Both plasmids also contain the plasmid mobilization relaxasome protein MobC and a ParA family protein which is essential for plasmid partition.
Data availability. The complete genome sequences of Citricoccus sp. strain SGAir0253 and its plasmids are available in DDBJ/EMBL/GenBank under accession numbers CP039424, CP039425, and CP039426 and SRA accession number SRR9043820.

ACKNOWLEDGMENT
This work was supported by a Singapore Ministry of Education Academic Research Fund tier 3 grant (MOE2013-T3-1-013).