Marine Microbes in the Production of Recalcitrant Dissolved Organic Matter

Introduction

Dissolved organic matter (DOM) is defined as organic particles smaller than 0.45 μm in size. DOM provides nutrients to bacteria, thus acting as a significant contributor to bacterial growth in marine environment. DOM pools can be divided into three groups based on their turnover times: labile, semi-labile and recalcitrant or refractory (these two terms will be used interchangeably).

DOM	Abundance (%)	Turnover	Composition	Location
Labile	<1%	minutes to weeks	Amino acids, D-glucose, etc.	Surface water
Semi-labile	50%	months to years	Polysaccharides, proteins, etc.	>1000m
Recalcitrant	50%	years to millenia	Mostly unknown	<1000m to ocean sediments

There are several mechanisms proposed for the production of dissolved organic matter. However, there is only a small amount of information known about refractory DOM (RDOM). In 2010, Jiao "et al". published a paper that suggests a mechanism involving marine microbes in producing RDOM, which they call "Microbial Carbon Pump" (MCP). Microbial carbon pump is depicted as a type of biological carbon pump, a process in which CO₂ is fixed by primary producers and transported down to the depth of the oceans as DOM. According to this study, there are three major pathways of microbial carbon pump: (1) exudation of microbial cells, (2) viral lysis of microbial cells, and (3) particulate organic matter (POM) degradation carried out by microbes. The MCP model differs from the conventional biological pump not only in a way that it solely focuses on microbial interactions, but also in a way that MCP takes place regardless of the depth in the water column whereas the biological pump stresses on vertical transport of DOM.

Relationship to prior literature

The study done by Jiao et al. persuasively introduces the concept of microbial carbon pump as a bacteria-driven mechanism which converts labile form of DOM into recalcitrant DOM. The idea had a great effect in the field of marine microbiology as well as marine geology and even meteorology, inspiring many other studies. However, the paper is largely conceptual and lacks experimental data to support the hypothesis, thus, further study to supplement the concept of microbial carbon pump with experimental results will enable it to become more than just a "conceptual framework". Another gap found in the research is the fact that the composition of RDOM is largely unknown. Although there has been several studies done on the composition of RDOM recently, most of its components remain as a mystery as well as what makes RDOM "refractory". With this incomplete understanding of RDOM, Jiao's study will not be completed since it is not possible to excavate the detailed mechanism behind MCP without a full understanding what is actually being generated (ie. RDOM). Moreover, in correspondence to Jiao et al.'s study, Chen (2011) argues that there are some points that need to be taken into further consideration, mostly regarding to Jiao's view towards the distribution and age of RDOM. In Jiao et al.'s paper, the figures included (fig. 2 and 4) imply that only the labile DOM occupies the ocean surface. However, Chen points out that this can be misleading as a substantial fraction of DOM at the surface seawater is indeed RDOM with an age of 2,500 - 3,100 years (Sargasso sea and north-central pacific) (Bauer et al., 1992). Another point Chen makes is that the 3 MCP pathways ((1) exudation of microbial cells, (2) viral lysis, and (3) POM degradation) depicted in Jiao et al. (2010) do not make the carbon necessarily older. Rather, the influx of extremely old DOM via deep water flow between the Pacific and the Atlantic oceans largely accounts for the aging of RDOM, , which ultimately questions the effectiveness of the microbial carbon pump in generating carbon reservoir.

In 2001, Ogawa et al. published an experimental report on recalcitrant DOM production by prokaryotes, at the time when microbial production of DOM was not much being discussed. As one of the first papers to generate data on microbial RDOM, there is no doubt that Ogawa's study had a strong impact on the process of building up the microbial carbon pump concept. To carry out the experiment, Ogawa made experimental samples with artificial seawater containing C, N, and P sources with inoculations from Gulf of Mexico as well as Sagami Bay, Japan. At the end of the year-long incubation, up to 50% of bacterial dissolved organic carbon (DOC) remained, suggesting an efficient production of refractory form of DOM from labile DOM. Although such data obtained in laboratory settings can differ from experiments in situ, the significance of this study lies in the context that it showed microbes are indeed capable of producing RDOM from labile DOM, opening a whole new view toward possible sources of RDOM. For this reason, Ogawa's study most certainly triggered further research on the role of microorganisms in RDOM production and the mechanism behind it (MCP for instance).

Influence on the field of research downstream

Jiao et al.'s research on microbial production of RDOM received a lot of attention as it can be seen from the number of times it has been cited (237 - number retrieved from the Web of Science) since its publication in 2010. The fields of research which referenced the paper include ecology, geology, oceanography, microbiology, even environmental engineering and more. Looking at this result, it is evident that the concept of microbial carbon pump has an enormous potential to be applied in various fields.

Tracing the long-term microbial production of recalcitrant fluorescent dissolved organic matter in seawater (Jorgensen et al. 2014)

Although this study is not a result of a direct influence from Jiao et al. (2010), it certainly adds some supportive information to the paper which lacks experimental data. Production of oceanic RDOM can be traced with fluorescent DOM (FDOM) in relation to increase in oxygen consumption by marine microorganisms (Jiao et al. 2010). In the study by Jorgensen et al. (2014), they examined the humic-like FDOM (represents recalcitrant RDOM because it accumulates in the absence of UV light) generation as a function of microbial respiration in ~1 year incubations of both natural and artificial seawater. As a result of this experiment, a nonlinear relationship between FDOM production and cumulative microbial respiration was shown. In other words, there is a a correlation between the presence of marine microorganisms and generation of RDOM. Additionally, the non-linearity aspect depicts a gradual transition from labile DOM to refractory DOM. While this cannot be a direct evidence of microbial carbon pump, the result does show that microbes are playing a role in RDOM production from labile/ semi-labile DOM.

Inefficient microbial production of refractory dissolved organic matter in the ocean (Osterholz et al. 2015)

On the other hand, it is quite evident that the study by Osterholz et al. was heavily influenced by the conceptual framework formed by Jiao et al. (2010). Interestingly enough, Osterholz et al. deny the efficient production of RDOM by marine microorganisms in direct contrast to what Jiao et al. have proposed in their 2010 paper. The first question that is brought up in this study is whether experimentally formed RDOM can be a true representative of marine RDOM that is known to be kept for millennia. In order to address this problem, Osterholz et al. not only took the timescale into account, but also the similarity of molecular structures with the oceanic RDOM when characterizing the microbial RDOM generated in the lab. As a result, they concluded that most of microbial RDOM do not equal to oceanic RDOM, thus, the efficiency of microbial carbon pump must be much lower than expected. However, By providing with a conceptual framework, Jiao et al. enabled other scientists like Osterholz, to initiate further research in terms of experimental approach toward microbial RDOM production and even RDOM composition which would allow enhancements in designing more accurate experimental models for studying MCP.

It cannot be said that Jiao's study opened up whole new horizons in the field of marine microbiology and oceanography just yet as the idea of microbial carbon pump is still in its early stages of development. However, it has certainly proposed fresh ideas and a new set of guidelines to those who study the biological mechanism behind production of RDOM in the ocean. Also,

Critical reflection

Refractory DOM which contributes to the majority of marine DOM has been left outside the field of knowledge for some time and it still is for a large part. The significance of the study led by Jiao involves that it recognized and conceptualized a mechanism of RDOM generation from bioavailable DOM in the ocean, carried out by microbes. Carbon in the ocean can exist in multiple forms: dissolved CO₂ as carbonic acid H₂CO₃) and dissolved organic carbon originating from carbon fixation by phytoplanktons (Stone 2010, Jiao et al. 2010). RDOM, which is recalcitrant from microbial remineralization, can act as a long-term carbon reservoir and fix atmospheric carbon under water. Upon further investigation, this finding may enable alteration of carbon sequestration in the ocean and control the amount of atmospheric carbon to reduce the accumulation of carbonic acid and global warming. Despite the doubts toward its feasibility, it is believed to incur many follow-up studies to implement the idea of enhancing oceanic carbon storage since ocean acidification and global warming due to increasing atmopheric CO₂ is a sensitive topic that is gaining a worldwide attention.

Annotated Bibliography

1. Jiao, N., Herndl, G. J., Hansell, D. A., Benner, R., Kattner, G., Wilhelm, S. W., ... Azam, F. 2010. Microbial production of recalcitrant dissolved organic matter: long-term carbon storage in the global ocean. Nature Reviews Microbiology, 8(8): 593-599. doi: 10.1038/nrmicro2386

This paper is the conceptual basis of the project, suggesting the concept of microbial carbon pump as the possible model for production of recalcitrant dissolved organic matter.

2. Jørgensen, L., Stedmon, C. A., Granskog, M. A., and Middelboe, M. 2014. Tracing the long-term microbial production of recalcitrant fluorescent dissolved organic matter in seawater. Geophysical Research Letters, 41: 2481-2488. doi: 10.1002/2014GL059428

This paper takes advantage of the fluorescent properties of RDOM to trace the mechanism behind its production and its connection to microbial respiration.

3. Murphy, K. R., Butler, K. D., Spencer, R. G. M., Stedmon, C. A., Boehme, J. R., and Aiken G. R. 2010. Measurement of dissolved organic matter fluorescence in aquatic environments: an interlaboratory comparison. Environmental Science and Technology, 44(24): 9405-9412. doi: 10.1021/es102362t

While fluorescence is a useful tool in measuring the amount and studying the composition of DOM in the ocean, this paper provides a deeper insight into limitations of the method and possible solutions to address the issue.

4. Dittmar, T., and Kattner, G. 2003. Recalcitrant dissolved organic matter in the ocean: major contribution of small amphiphilics. Marine Chemistry, 82(1-2): 115-123. doi: 10.1016/S0304-4203(03)00068-9

Aside from the microbial aspect of DOM, this study reveals more about the biochemical parameters of RDOM in the ocean. Although the primary focus of the project should be the microbial aspect, it is also necessary to know the fundamental basis of RDOM’s biochemistry for the sake of deeper understanding of the subject.

5. Osterholz, H., Niggemann, J., Giebel, H., Simon, M., and Dittmar, T. 2015. Inefficient microbial production of refractory dissolved organic matter in the ocean. Nature Communications, 6: 7422. doi: 10.1038/ncomms8422

This paper characterizes the molecular composition of experimentally produced microbial RDOM in comparison with oceanic RDOM.

6. Xue, Y., Ge, T., and Wang, X. 2015. An effective method of UV-oxidation of dissolved organic carbon in natural waters for radiocarbon analysis by accelerator mass spectrometry. Journal of Ocean University of China, 14(6): 989-993. doi: 10.1007/s11802-015-2935-z

As the sources of DOM are poorly understood and not enough data are present to fully characterize the microbial activities in the production of RDOM, this study suggests the use of radiocarbon (14C) compositions in DOC that are naturally present to trace the sources of DOM and the processes involved in its production.

7. Zhang, Z., Chen, Y., Wang, R., Cai, R., Fu, Y., and Jiao, N. 2015. The fate of marine bacterial exopolysaccharide in natural marine microbial communities. PLoS One, 10(11): e0142690. doi: 10.1371/journal.pone.0142690

To explain the mechanism behind marine microbes’ involvement in RDOM formation, this study tests a proposed hypothesis which the exopolysaccharaides produced by most marine bacteria are resistant to mineralization by microbes.

8. Legendre, L., Rivkin, R. B., Weinbauer, M. G., Guidi, L., Uitz, J. 2015. The microbial carbon pump concept: Potential biogeochemical significance in the globally changing ocean. Progress in Oceanography, 134: 432-450. doi: 10.1016/j.pocean.2015.01.008

This paper discusses about vertical ocean carbon pump as well as the microbial carbon pump as a source of sequestered carbon and their relationship to climate change.

9. Buchan, A., LeCleir, G. R., Gulvik, C. A. and Gonzalez, J. M. 2014. Master recyclers: features and functions of bacteria associated with phytoplankton blooms. Nature Reviews Microbiology, 12: 686-698. doi: 10.1038/nrmicro3326

10. Stone, R. 2010. The invisible hand behind a vast carbon reservoir. Science, 328(5985): 1476-1477. doi: 10.1126/science.328.5985.1476

This article provides a detailed insight into Jiao et al. (2010).

11. Chen, C. A. 2011. Microbial carbon pump: additional considerations. Nature Reviews Microbiology, 9(7): 555. doi: 10.1038/nrmicro2386-c4

Chen corresponds to aspects of Jiao's 2010 study that need possible consideration.

12. Ogawa, H., Amagai, Y., Koike, I., Kaiser, K. and Benner, R. 2001. Production of refractory dissolved organic matter by bacteria. Science. 292(5518): 917-920. doi: 10.1126/science.1057627

This paper acts as an experimental basis of MCP model by recognizing microbial role in RDOM generation