Extreme polyploidy in a large bacterium

Epulopiscium sp. are an important bacterial species due to their link between bacteria and eukaryotic organisms, and unusual size. There are excessively large in comparison to other bacteria it that they are over 600um long, which appears to be a trait that has been negatively selected out in other species of bacteria. Those that do develop a larger overall size tend to maintain a small volume cytoplasm by the inclusion of a vaccuole. Again, Epulopiscium differs here in that they contain a high volume cytoplasm. These factors possibly benefit the fact that the bacteria are found in symbiosis with surgeonfish.

Mendell et al (2008) consider the implications of size in terms of the genome and the cells as a whole. N. tonganus (surgeonfish) were collected by spearfishing on reefs around Liard Island, Australia. Samples of Epulopiscium sp were isolated from the intestine and individual Type B cells were selected using a micropipettor. Samples were 5000 ‘mother’ bacteria with immature offspring and 5000 ‘mother’ bacteria with mature offspring, both groups of offspring being held internally.
The volumes of the cells were recorded by observing them under a microscope. The DNA was extracted using methods based on standard procedures, then quantified using PicoGreen assays. DNA was amplified using TaqMan Quantitative PCR Assays, and real-time quantitative PCR. Statistical analysis was carried out, comparing the genome size with the volume of the cell.

In terms of the number of copies of genes in relation to cytoplasmic volume, there is no relationship amongst small cells, followed by a linear relationship when the cells reach a threshold size. This supports the proposed theory that their genome is composed of several copies of a smaller genome: immature offspring, that have smaller cytoplasmic volumes, develop a complete copy of their genome (represented the lag phase), the small genome replicates throughout the rest of development (the linear phase).

Mature offspring were found to contain almost 3 times the amount of DNA as immature offspring. This size difference suggests the offspring only inherit a small proportion of the parental DNA. Large cells contain on average approximately 50,000 to 120, 000 copies of the specific genes investigated (ftsZ, dnaA, recA), and 370,000 copies of 16S rRNA. The copy numbers of the first 3 were statistically similar, support the theory that extreme polyploidy occurs. The values for small cells varied, most likely since they were only part way through development.

The copies of the genome are not always identical. This variation hints at a link to eukaryotic organisms in that a specific copy of the genome can be responsible for one function of the bacteria and therefore varies slightly, whilst another copy can take responsibility for another function. Polyploidy allows organisms with unstable regions of some of the genome copies to survive with the stability provided by the other copies that are stable.

The 16S rRNA sequence was found to contain coding regions for several operons. Operons are known to respond to inducers and repressors, which control replication. In terms of using the PCR method for this genome, the natural method of replication may not be represented due to the potential for the operons to control replication highly, in a natural state.

The figures stated, such as gene copy numbers, were noted as being estimates due to high variation amongst samples. Bacteria replicate at a high rate, and the fact that the bacteria were not halted at a specific point in their life cycle; they will be at different stages of development, even though they are classified loosely as mature or immature it is difficult to specific when only used two groups. When culturing bacteria, it is possible to stop replication at a specific point, such as in E. coli; Epulopiscium sp. cannot be cultured, therefore this is not possible.

The success of the bacteria as part of a symbiotic relationship and the highly suggested polyploidy present supports the idea that polyploidy is beneficial for symbiotic relationships. Polyploidy allows for rapid cell division and metabolic differentiation. These factors could therefore be vital for successful symbiosis. Polyploidy has also been shown correlate with cell size. Epulopiscium sp. are extremely large, which aids in avoiding predation when in the digestive tract of surgeonfish.

It would be interesting to work with an organism that can be cultured, and that can form a symbiotic relationship, and remove the polyploidy ability of the genome or sequences that are responsible for varying metabolic pathways, observing the effect of the survival of the bacteria. However many bacteria that form thrive in or on a host have be deemed not possible to culture at present, including Vibrio fischeri (in Euprymna scolopes) and  sulphur-oxidising bacteria in Riftia pachyptila. It would be highly beneficial to develop methods to culture these in order to gain further understanding. It would also be interesting to know at what point during development of the bacterial species a symbiotic relationship occurred: does the bacteria become too large to survive without the high nutrient content found in the fish’s intestine, or did the host environment pressurize the bacteria to develop?

Mendell, J. E;  Clements,K. D; Choat, J. C; and  Angert, E. R (2008) Extreme polyploidy in a large bacterium, Proc Natl Acad Sci U S A, 105(18): 6730–6734

http://www.pnas.org/content/105/18/6730.full