Editor's Note: InterFaces is a new section of Interactions that recognizes scientists who have contributed to our field.

Ann M. Hirsch,¹ Euan K. James,² and Janet I. Sprent³

¹Department of Molecular, Cell, and Developmental Biology and Molecular Biology Institute, UCLA, Los Angeles, CA 90095 U.S.A.

²The James Hutton Institute, Invergowrie, Dundee DD2 5DA, U.K.

³Royal Botanical Gardens Edinburgh, 20a Inverleith Row, Edinburgh EH3 5LR, U.K.

Science is based on discovery, but often, the awards and kudos go to the scientists who take the initial findings of others to the point at which they become canon. Yet without the initial discovery and the research that enabled it, our understanding of the intricacies of nature would be incomplete. Science needs pioneers who undertake the study of various phenomena not because they are fashionable or fundable but because the pioneers are curious and want to learn more about the world around them.

Until 2001, only the alpha-proteobacteria (specifically, Rhizobium sensu lato) were known to nodulate legumes. The discovery that year by Moulin et al. (2001) that beta-proteobacteria (specifically, Burkholderia) nodulated legumes generated a great deal of excitement. Numerous investigators looked for more strains, plant assays were performed, and genomes were sequenced. Why had this group of nodulating bacteria, called “beta-rhizobia,” been overlooked? Some thought it was because the two nodulating strains described in Moulin et al. were isolated from Papilionoid legumes growing in the Fynbos of South Africa or in the tropical forests of French Guiana and because so few studies had been made in either country. This is partly true. However, Burkholderia strains were isolated from Mimosoid legume nodules much earlier than the Fynbos and French Guiana representatives but not from Africa or South America. Michael J. Trinick, who graduated in Agricultural Science (BSc Agr; majoring in soil microbiology) from the University of Sydney in 1958 was the first to isolate them from legumes growing in Papua New Guinea, and he gave the strains to Ivan Kennedy of the same university. These strains, some of which were reported as effective “Rhizobium” symbionts of Mimosa in Trinick (1980), were later described in detail as Mimosa-nodulating Burkholderia and Cupriavidus strains (Elliot et al., 2007, 2009).

Trinick conducted studies using serology and various other characteristics of the bacteria, such as vitamin requirements and carbohydrate utilization, and from them, he discovered that nodules isolated from plants growing in poor soils contained bacteria other than Rhizobium sensu stricto. However, Vincent (1970) cautioned readers against isolating nonrhizobial contaminants from nodules. Could this warning have influenced an avenue of discovery that was not publishable until 2001? It seems likely. Furthermore, after Burkholderia species were first isolated from legume nodules, a number of researchers were concerned that they might be closely related to mammalian and plant pathogens, because this genus is well known for its virulence on both plants and humans. However, phylogenetic studies using 16S RNA (Gyaneshwar et al., 2011) and multilocus sequence analysis (MLSA) (Estrada de-los Santos et al., 2013) separated the symbionts from the pathogens, and based on these studies and others, many species were categorized into two new genera: namely, Paraburkholderia (Sawana et al., 2014) and Caballeronia (Dobritsa and Samadpour, 2016). Our most recent effort was to separate a distinct subgroup of nonpathogenic species into a new genus, which we named Trinickia, after Michael Trinick (Estrada-de los Santos et al., 2018).

Trinick also made other important discoveries. After graduating from the University of Sydney, he joined the Department of Agriculture, Stock, and Fisheries in Port Moresby, Papua New Guinea, in 1959 in the plant pathology department. After extensive research into the tropical legumes of this region, he received an MS in agriculture from the University of Sydney in October 1966. At this time, he discovered the promiscuous Rhizobium strain NGR234, which nodulates an exceptionally broad range of legume species. This finding led to many discoveries about variations in genetic factors, such as the type 3 secretion system, which controls host specificity in the nitrogen-fixing symbiosis (Deakin and Broughton, 2009).

Trinick wrote his PhD thesis, “The Ecology of Rhizobium-Interactions between Rhizobium Strains and Other Soil Microorganisms,” after studying the influence of the soil microflora on the survival of strains of R. trifolii, R. meliloti, and R. lupini in the sandy soils of western Australia. He received his doctoral degree in 1970 from the University of Western Australia. While in Papua New Guinea, Trinick also discovered that a nonlegume, Parasponia (Cannabaceae), was nodulated by Rhizobium species. The original description was for a related species, Trema aspera (now cannabina) (Trinick, 1973), but it was later reported that the actual nodulating nonlegume was a sister species of Trema—namely, Parasponia parviflora Miq. (Akkermanns et al., 1978). Research on Parasponia species dominated his later studies at the CSIRO, Division of Plant Industry and Land Management. Investigations with C. A. Appleby, J. B. Whittenburg, B. A. Whittenburg, A. A. Kortt, D. J. Goodchild, and others set up a baseline for studies on this unexpected symbiosis between a no-legume and a Rhizobium strain. Trinick’s discoveries opened new doors to study the evolution of symbiotic nitrogen fixation and the possibility of transferring nodulation ability to other nonlegumes (Van Vetzen et al., 2018).

The discoveries of (1) strain NGR234 and (2) the different behaviors of fast-growing versus slow-growing rhizobial strains and their beta-rhizobia counterparts, as well as (3) detailed studies on Parasponia, established a solid foundation upon which many more recent investigations have been established and future studies will be based. Three cheers and many thanks to Michael Trinick for three incredible breakthroughs in symbiotic nitrogen fixation!

References

Akkermanns, A. D. L., Abdulkadir, S., and Trinick, M. J. 1978. N2-fixing root nodules in Ulmaceae: Parasponia or (and) Trema spp.? Plant Soil 49:711-715.

Deakin, W. J., and Broughton, W. J. 2009. Symbiotic use of pathogenic strategies: Rhizobial protein secretion systems. Nat. Rev. Microbiol. 6:312-320. doi:10.1038/nrmicro2091

Dobritsa, A. P., and Samadpour, M. 2016. Transfer of eleven Burkholderia species to the genus Paraburkholderia and proposal of Caballeronia gen. nov., a new genus to accommodate twelve species of Burkholderia and Paraburkholderia. Int. J. Syst. Evol. Microbiol. 66:2836-2846. doi:10:1094/ijsem.0.001065

Elliott, G. N, Chen, W. M., Chou, J. H., Wang, H. C., Sheu, S. Y., Perin, L., Reis, V. M., Moulin, L., Simon, M. F., and Bontemps, C. 2007. Burkholderia phymatum is a highly effective nitrogen‐fixing symbiont of Mimosa spp. and fixes nitrogen ex planta. New Phytol. 173:168-180. doi:10.1111/j.1469-8137.2006.01894.x

Elliott, G. N., Chou, J.-H., Chen, W.-M., Bloemberg, G. V., Bontemps, C., Martínez- Romero, E., Velázquez, E., Young, J .P. W., Sprent, J. I., and James, E. K. 2009. Burkholderia spp. are the most competitive symbionts of Mimosa, particularly under N-limited conditions. Environ. Microbiol. 11:762-778.

Estrada-de los Santos, P., Vinuesa, P., Martínez-Aguilar, L., Hirsch, A. M., and Caballero-Mellado, J. 2013. Phylogenetic analysis of Burkholderia species by multilocus sequence analysis. Curr. Microbiol. 67:51-60.

Estrada-de los Santos, P., et al. 2018. Whole genome analyses suggest that Burkholderia sensu lato contains two further novel genera in the “rhizoxinica-symbiotica group” (Mycetohabitans gen. nov., and Trinickia gen. nov.): Implications for the evolution of diazotrophy and nodulation in the Burkholderiaceae. Genes 9:389. doi:10.3390/genes9080389

Gyaneshwar, P., Hirsch, A. M., Moulin, L., Chen, W. M., Elliott, G. N., Bontemps, C., Estrada-de los Santos, P., Gross, E., Bueno dos Reis Junior, F., Sprent, J. I., Young, J. P. W., and James, E. K. 2011. Legume nodulating β-proteobacteria: diversity, host range and future prospects. Mol. Plant-Microbe Interactions 24:1276-1288.

Moulin, L., Munive, A., Dreyfus, B., and Boivin-Masson, C. 2001. Nodulation of legumes by members of the β-subclass of Proteobacteria. Nature 411:948-950.

Sawana, A., Adeolu, M., and Gupta, R. S. 2014. Molecular signatures and phylogenomic analysis of the genus Burkholderia: Proposal for division of this genus into the emended genus Burkholderia containing pathogenic organisms and a new genus Paraburkholderia gen. nov. harboring environmental species. Front. Genet. 5:429. doi:10.3389/fgene.2014.00429

Trinick, M. J. 1973. Symbiosis between Rhizobium and the non-legume, Trema aspera. Nature 244:459-460.

Trinick, M. J. 1980. Relationships amongst the fast-growing rhizobia of Lablab purpureus, Leucaena leucocephala, Mimosa spp., Acacia farnesiana and Sesbania grandiflora and their affinities with other rhizobial groups. J. Appl. Bacteriol. 49:39-53.

Van Vetzen, R., Doyle J. J., and Guerts, R. 2018. A resurrected scenario: Single gain and massive loss of nitrogen-fixing nodulation. Trends Plant Sci. doi:10.1016/j.tplants.2018.10.005

Vincent, J. M. 1970. A manual for the practical study of root nodule bacteria. Oxford, UK: Blackwell Scientific.

Archives

Interactions

‭(Hidden)‬ Blog Tools