A detective story using genomics to find the parent strain of Hbt. salinarum NRC-1 and R1

Many labs have worked with Halobacterium salinarum, and they either use the R1 strain or the NRC-1 strain. When the genomes of these two strains were reported in 2008 (Pfeiffer et al.), it showed they are so similar in sequence that they must have derived from the same isolate, but for various reasons it was not possible to identify that isolate. Here we used publicly available sequence data to hunt down the parent. See https://doi.org/10.1002/mbo3.1365. This is not a simple BLASTn comparison but rather a comprehensive analysis and includes a speculative outline of how the two strains ended up as we see them today.

While nothing to do with this study, here is a colorful picture of purple membrane preparations from Halobacterium salinarum R1, as seen after sucrose gradient centrifugation.

Genome sequence of Hardyhisp2, a Gammapleolipovirus infecting Har. hispanica.

Published in Microbial Resource Announcements in May, 2021 (doi: 10.1128/MRA.00226-21) this describes the genome of Hardyhisp2, a virus isolated in my lab way back in 1998! It came from a water sample of the hypersaline Lake Hardy, Victoria, Australia. As seen in the dot plot (above left) and genome map comparison (above right), the 16,133 bp linear dsDNA genome shows strong sequence similarity to the type species of gammapleolipovirus, His2. The most disparate gene appears to be the one encoding spike protein (VP1 spike), which is used by the virus for cell attachment, but both Hardyhisp2 and His2 infect the same host, Haloarcula hispanica. Perhaps they attach to different cell receptors. Characteristic of this viral genus, the Hardyhisp2 genome carries a gene encoding a protein-primed DNA polymerase, and also has inverted terminal repeats (shaded grey in the figure). From work with His2 (the first pleolipovirus isolated) we know that the genome ends have proteins attached, and the His2 genome can be transfected into host cells to recover infectious virus (Porter and Dyall-Smith, 2008). It is a pity that more genetic investigations have not been pursued on viruses of this group.

A novel halovirus, Hardycor1

In the second paper of the series, we describe the genome of a siphovirus that infects Halorubrum coriense. It was isolated from Lake Hardy way back in 1998, and the genome shows it is quite distinct from other reported caudoviruses of haloarchaea. The graphical abstract above shows a picture of Hardy Lake in the background, and a map of the Hardycor1 genome in the foreground. We also observed a number of active proviruses occurred in our stocks of this virus and other haloviruses, highlighting the dynamic nature of proviruses in haloarchaea. See the paper here, at doi.org/10.3390/genes12020149.

Transcription of Halovirus His2 and its host.

Our microarray study of the transcription of the gammapleolipovirus His2 and its host Har. hispanica has just been published in MicrobiologyOpen. The direct link to it is https://doi.org/10.1002/mbo3.1016. Viral gene expression occurred in three main phases (early, middle, and late), and by 4.5 hr p.i. the majority of genes were actively transcribed. Eighty host genes were differentially regulated ≥twofold post-infection.

Halobacterium salinarum (DSM 3754T) comparison to strains R1 and NRC‐1

Friedhelm Pfeiffer, Gerald Losensky, Anita Marchfelder, Bianca Habermann and Mike Dyall‐Smith

Finally the paper was online today in MicrobiologyOpen at (https://onlinelibrary.wiley.com/doi/full/10.1002/mbo3.974). It is a detailed description of the genome of Hbt. salinarum strain 91‐R6 (DSM 3754), which is the type strain of the genus. We have compared it to the two well-studied laboratory strains R1 and NRC-1, which were sequenced many years ago. The type strain has only rarely been studied, and as Aharon Oren said in 2012, “it is to be regretted that no genomic information is available for the nomenclatural type” – but now it is available for everyone to examine and enjoy!! Genbank accessions are CP038631.1CP038632.1, and CP038633.1.

Genome of the type strain of Hbt. salinarum

In 2012, Aharon Oren wrote a review on the family Halobacteriaceae, where he pointed out with some surprise that the “type species of the type genus of the family and the order” had not yet been sequenced. This was despite the genomes of strains R1 and NRC-1 having been reported years previously. I am happy to say that today this deficit has been remedied, as described in our recent publication in Microbial Resource Announcements, and sequence submissions to Genbank, under accessions  CP038631 (chromosome), CP038632 (pHSAL1), and CP038633 (pHSAL2). Assembly of these replicons was not a straightforward task, as there were very large repeats, and considerable genomic variability. We are currently working on a more detailed analysis of this sequence.

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