Complete Genome Sequence of Haloarcula hispanica, a Model Haloarchaeon for Studying Genetics, Metabolism, and Virus-Host Interaction

Abstract

Haloarcula hispanica is an extremely halophilic archaeon that has an unusually low restriction barrier and is therefore significant for studying archaeal genetics, metabolism, and virus-host interactions. Here we report the complete genome sequence (3,890,005 bp) of H. hispanica strain CGMCC 1.2049, consisting of two chromosomes and one megaplasmid.

GENOME ANNOUNCEMENT

Haloarcula hispanica was first isolated from a solar saltern in Spain (8). It is exceptional among members of the Halobacteriaceae family in that it exhibits unusually low restriction activity and is therefore significant as one of the most tractable haloarchaea for archaeal studies (2), as well being useful as a vital host for investigations of viruses in hypersaline ecosystems (13). A transformation method for H. hispanica was established 20 years ago (2, 3), whereas the high-efficient knockout system for its genome manipulation was developed recently (11). In particular, H. hispanica has shown the ability to accumulate polyhydroxyalkanoate (PHA) when carbon sources are in excess. Although a number of key genes (phaEC and phaB) involved in PHA biosynthesis in H. hispanica have been identified (6, 7), the entire PHA biosynthetic and regulatory networks in haloarchaea remain to be established. Thus, H. hispanica would be a good haloarchaeon model organism after the genome sequence is made available. Here, we announce the complete genome sequence of H. hispanica CGMCC 1.2049 (= ATCC 33960), which was obtained from the China General Microbiological Culture Collection Center (CGMCC).

The genome sequence was determined mainly by the use of massively parallel pyrosequencing technology and a Roche 454 GS FLX sequencer. A total of 506,320 reads, counting up to 124,501,912 bases (32-fold coverage of the genome), were assembled using GS de novo Assembler software, which generated 29 contigs ranging from 500 to 916,380 bp, with bases having quality scores of 40 and above. Relationships of the contigs were determined by multiplex PCR (14). Gaps were filled by sequencing the PCR products and the pSMART (Lucigen) shotgun library using ABI 3730xl capillary sequencers. Finally, the genome was assembled using Phred, Phrap, and Consed software (http://www.phrap.org/phredphrapconsed.html), and low-quality regions of the genome were resequenced. Putative coding sequences (CDSs) were identified by ZCURVE (5) and Glimmer 3 (4) software, and peptides shorter than 30 amino acids were eliminated. tRNA genes were predicted by tRNAScan-SE software (12) and rRNA genes were identified by RNAmmer 1.2 (10). Functional annotation of CDSs was performed by searching the NCBI nonredundant protein database and the Kyoto Encyclopedia of Genes and Genomes (KEGG) protein database (9) using BLASTP (1).

The H. hispanica genome consists of main chromosome I (2,995,271 bp) (G+C content, 63.7%; coding density, 88.1%), minichromosome II (488,918 bp) (G+C content, 57.0%; coding density, 80.3%), and megaplasmid pHH400 (405,816 bp) (G+C content, 59.9%; coding density, 84.4%). The genome encodes 3,859 predicted proteins, 48 tRNAs, and 3 copies of 16S-23S-5S rRNA on chromosomes (1 copy on chromosome II) plus 1 copy of 5S rRNA on pHH400. Compared to the genome of H. marismortui, another sequenced Haloarcula with two chromosomes and seven plasmids, up to 80% of predicted proteins of H. hispanica have counterparts in H. marismortui, whereas ∼700 predicted proteins are only present in H. hispanica. Interestingly, although their main chromosomes are homologous and their megaplasmids are homologous, their minichromosomes are completely different. Comparative analysis performed with the genome of H. marismortui and other sequenced haloarchaeal genomes may provide further insight into the genetics, metabolism, origin, and evolution of the extremely halophilic archaea.

Nucleotide sequence accession numbers.

The genome sequence of H. hispanica CGMCC 1.2049 was deposited in NCBI GenBank under accession numbers CP002921 (chromosome I), CP002922 (chromosome II), and CP002923 (pHH400).