Ma­rine Ar­chaea and the Ni­tro­gen Cy­cle

by Dou­glas Bartlett

What is more stir­ring than to con­tem­plate the cy­cles of mat­ter in na­ture? We sense that we wit­ness vi­tal trans­ac­tions of this planet's me­tab­o­lism, a ful­fill­ing thought. Such mat­ters speak parti­cularly loudly to us mi­cro­bi­ol­o­gists: not only do global fluxes con­cern the el­e­ments of life but are to a great ex­tent dri­ven by mi­cro­bial ac­tiv­i­ties. In this vein, here is some im­por­tant news.

Analy­ses of the ni­tro­gen bud­get in the oceans sug­gest that a whole lot more ni­tro­gen is be­ing fixed than was rec­og­nized. Who does this ex­tra ni­tro­gen fix­a­tion? A re­cent pa­per from the labora­tory of Vic­to­ria Or­phan at Cal Tech pro­vides clues that may help bal­ance the ni­tro­gen bud­get. These re­searchers stud­ied the syn­trophic ag­gre­gate of ar­chaea be­long­ing to the ANME‑2 group and bac­te­ria be­long­ing to the Desulfosarcina/Desulfococcus. These con­sor­tia are a big deal: they con­sume about 80% of the methane re­leased from seeps in ma­rine sed­i­ments. In­trigu­ing here is that their metage­nomic analy­sis re­vealed that they con­tain genes for ni­tro­ge­nase, the key en­zyme in ni­tro­gen fix­a­tion. So, do these part­ner­ships fix ni­tro­gen? And, if so, does this con­tribute sub­stantially to that arm of the ni­tro­gen cy­cle? The an­swer seems to be yes to the first and, "well, to some ex­tent," to the sec­ond.

Mag­ni­tude and distri­bu­tion of ¹⁵N in­cor­po­ra­tion in a ANME‑2/ DSS con­sor­tium from sed­i­ments incu­bated with CH₄ and ei­ther ¹⁵N₂ (A and E) or ¹⁵NH₄⁺ (I and V) as the N source. FISH used probes tar­get­ing ANME‑2 (red) and Desul­fobac­ter­aceae  (green). NanoSIMS im­ages show the ¹²C ¹⁵N / ¹²C ¹⁴N ra­tios of the same mi­cro­bial con­sor­tia to de­monstrate the lo­ca­tion of ¹⁵N incor­po­ration. (The scale range varies be­tween im­ages, with the min. con­sis­tently set to nat­ural abun­dance ¹⁵N/¹⁴N.) Source

How these work­ers went about this re­search is instruc­tive. They in­cu­bated ma­rine sed­i­ment sam­ples with me­thane and one of sev­eral ¹⁵N‑labeled N sources and fol­lowed the en­rich­ment of the con­sor­tia for ¹⁵N over a pe­riod of six months. They de­ter­mined ¹⁵N/¹⁴N by se­condary ion mass spec­trom­e­try (nanoSIMS) imag­ing of sin­gle cells la­beled for bac­te­r­ial and archeal flu­o­res­cent probes. Thus, they could tell the lo­ca­tion of the iso­topic la­bel in each of the mi­cro­bial part­ners. The con­sor­tia of­ten con­sist of spher­i­cal cell ag­gre­gates hav­ing a bac­terial shell and an ar­chaeal core. Their data sug­gest that ni­tro­gen fix­a­tion takes place in the ar­chaeal in­te­rior and that the ar­chaea use that ni­tro­gen to make re­duced ni­tro­gen com­pounds that they ex­port to the bac­te­r­ial layer.

Cells in­cu­bated with­out suf­fi­cient methane or un­der con­di­tions that in­hibit sul­fate re­duc­tion did not incor­porate ¹⁵N dini­tro­gen. PCR and se­quence analy­ses of ni­tro­gen fix­a­tion (nif) genes re­cov­ered from the DNA present in the ¹⁵N‑labeled in­cu­ba­tions dis­played si­milarity to se­quences from methanogenic ar­chaea. The nif operon gene or­der also sug­gested an ar­chaeal source.

An ad­di­tional cu­ri­ous as­pect of ni­tro­gen fix­a­tion in this ag­gre­gate is that ANME‑2 ar­chaea gener­ate less en­ergy per mole of sub­strate ox­i­dized than any other mi­crobe known to un­der­take the en­er­get­i­cally ex­pen­sive process of ni­tro­gen fix­a­tion. The con­sor­tium grew twenty times faster when pro­vided with am­mo­nium rather than ni­tro­gen as a ni­tro­gen source, but the cells main­tained sim­i­lar res­pi­ra­tory rates un­der both con­di­tions, ap­par­ently ad­just­ing to a lower growth rate when stressed with the need to use more ATP to fix their own ni­tro­gen.

There is much more to be found out. The au­thors state: "...our in­ven­tory of ma­rine di­a­zotrophs is in­com­plete and... we are only be­gin­ning to un­der­stand the ex­tent and im­por­tance of ben­thic ma­rine N₂ fix­a­tion." That may be so, but the trip to this des­ti­na­tion will be noth­ing short of fascinat­ing.

Doug Bartlett is Pro­fes­sor of Mi­cro­bi­ol­ogy at the Scripps In­sti­tute of Oceano­graphy, Uni­ver­sity of Cal­i­for­nia at San Diego.