The evolution of determinate and indeterminate nodules within the Papilionoideae subfamily

Guiling Ren

Research output: Thesisinternal PhD, WU


We selected the legume Indigofera argentea for our research. It was collected in the desert of Jizan in Saudi Arabia. We selected this species for two reasons. It is well adapted to heat and drought and therefore it has the potential to study, in the future, mechanisms that confer tolerance to these abiotic stresses. Further, Indigofera represents an early branching lineage within the indigoferoid/milletioid clade. Therefore, Indigofera is a key genus in studying the evolution of nodulation within the Papilionoideae subfamily.

In Chapter 1, a general introduction is given on nitrogen fixing symbiosis of legumes and rhizobia. In this introduction, we focus on the process of nodule initiation and organogenesis. Two main nodule types, determinate and indeterminate nodule, are introduced based on the knowledge of the few well-studied legumes species. Further, terminal differentiation of rhizobia that is induced by NCR peptides of the host is introduced.

In Chapter 2, we characterized the desert legume I. argentea and developed a platform by which future studies on mechanisms controlling abiotic stress become available. We developed an Agrobacterium rhizogenes-mediated root transformation procedure and did a de novo transcriptome assembly using RNA of various organs.

In Chapter 3, about 60 rhizobium strains have been isolated from nodulated I. argentea plants that were collected in the desert. The strains were characterized by 16S sequencing and their nodulation abilities were studied. One of the efficient nodulating Bradyrhizobium strains was selected for further studies.

In Chapter 4, nodule development of Indigofera and Tephrosia species was analysed and described in detail. Species from both genera have an indeterminate growth. However, it was shown that this is not due to a meristem that is formed at the primordium stage, which is the basis of indeterminate growth of IRLC species like Medicago. The indeterminate growth was shown to be due to secondary clusters of dividing infected cells that were formed from nodule parenchyma cells. Therefore, it evolved independently from the indeterminate growth from IRLC species which is controlled by a persistent meristem composed of non-infected cells.

In Chapter 5, it is shown that bacteroids from I. argentea nodules are markedly enlarged as described for IRLC (e.g. Medicago) and Aeschynomene species. This increase in size was correlated with endoreduplication and terminal differentiation. Further, it was shown that in nodules of I. argentea 4 NCR genes are expressed. Therefore it is probable that these terminal differentiation is controlled by the NCR peptides and this evolved independently in the IRLC clade, Aeschynomene and Indigofera.

In Chapter 6, I discuss my results and put them in a broader perspective. I summarize and discuss the determinate and indeterminate nodule evolution in the Papilionoideae subfamily. 

We have shown in Chapter 4 that Indigofera and Tephrosia species have a nodule development that is very similar to that of determinate nodules and their indeterminate growth is due to the formation of secondary clusters of dividing cells. Species from other clades of the Papilionoideae subfamily were analyzed. Based on these analyses, I conclude that the ancestor of the Papilionoideae subfamily formed determinate nodules that had the ability to form secondary clusters of dividing cells. Further, NCR-triggered bacteroid evolution in the Papilionoideae subfamily is discussed.

Original languageEnglish
QualificationDoctor of Philosophy
Awarding Institution
  • Wageningen University
  • Bisseling, Ton, Promotor
  • Geurts, Rene, Co-promotor
Award date23 Jan 2018
Place of PublicationWageningen
Print ISBNs9789463432306
Publication statusPublished - 2018

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