Signaling and Gene Expression for Water-Tolerant Legume Nodulation

In the symbiotic interaction with rhizobia, legumes develop nodules in which nitrogen fixation takes place. Upon submersion, most temperate legumes are incapable of nodulation, but tropical legumes that grow in waterlogged soils have acquired water stress tolerance for growth and nodulation. One well-studied model plant, the tropical, semi-aquatic Sesbania rostrata, develops stem-located adventitious root primordia that grow out into adventitious roots upon submergence and develop into stem nodules after inoculation with the microsymbiont, Azorhizobium caulinodans. Sesbania rostrata also has a nodulated underground root system. On well-aerated roots, nodules form via root hair curling infection in the zone, just above the root tip, where root hairs develop; on hydroponic roots, an alternative process is used, recruiting a cortical intercellular invasion program at the lateral root bases that skips the epidermal responses. This intercellular cortical invasion entails infection pocket formation, a process that involves cell death features and reactive oxygen species. The plant hormones ethylene and gibberellin are the major signals that act downstream from the bacterial nodulation factors in the nodulation and invasion program. Both hormones block root hair curling infection, but cooperate to stimulate lateral root base invasion and play a role in infection thread formation, meristem establishment, and differentiation of meristem descendants.     

Early development ofRhizobium-induced root nodules ofParasponia rigida. I. Infection and early nodule initiation

Summary The first of two major steps in the infection process in roots ofParasponia rigida (Ulmaceae) following inoculation byRhizobium strain RP501 involves the invasion ofRhizobium into the intercellular space system of the root cortex. The earliest sign of root nodule initiation is the presence of clumps of multicellular root hairs (MCRH), a response apparently unique amongRhizobium-root associations. At the same time or shortly after MCRH are first visible, cell divisions are initiated in the outer root cortex of the host plant, always subjacent to the MCRH. No infection threads were observed in root hairs or cortical cells in early stages. Rhizobial entry through the epidermis and into the root cortex was shown to occur via intercellular invasion at the bases of MCRH. The second major step in the infection process is the actual infectionper se of host cells by the rhizobia and formation of typical intracellular infection threads with host cell accommodation. This infection step is probably the beginning of the truly symbiotic relationship in these nodules. Rhizobial invasion and infection are accompanied by host cortical cell divisions which result in a callus-like mass of cortical cells. In addition to infection thread formation in some of these host cortical cells, another type of rhizobial proliferation was observed in which large accumulations of rhizobia in intercellular spaces are associated with host cell wall distortion, deposition of electron-dense material in the walls, and occasional deleterious effects on host cell cytoplasm.     

Dissection of Bacterial Wilt on Medicago truncatula Revealed Two Type III Secretion System Effectors Acting on Root Infection Process and Disease Development

Ralstonia solanacearum is the causal agent of the devastating bacterial wilt disease, which colonizes susceptible Medicago truncatula via the intact root tip. Infection involves four steps: appearance of root tip symptoms, root tip cortical cell invasion, vessel colonization, and foliar wilting. We examined this pathosystem by in vitro inoculation of intact roots of susceptible or resistant M. truncatula with the pathogenic strain GMI1000. The infection process was type III secretion system dependent and required two type III effectors, Gala7 and AvrA, which were shown to be involved at different stages of infection. Both effectors were involved in development of root tip symptoms, and Gala7 was the main determinant for bacterial invasion of cortical cells. Vessel invasion depended on the host genetic background and was never observed in the resistant line. The invasion of the root tip vasculature in the susceptible line caused foliar wilting. The avrA mutant showed reduced aggressiveness in all steps of the infection process, suggesting a global role in R. solanacearum pathogenicity. The roles of these two effectors in subsequent stages were studied using an assay that bypassed the penetration step; with this assay, the avrA mutant showed no effect compared with the GMI1000 strain, indicating that AvrA is important in early stages of infection. However, later disease symptoms were reduced in the gala7 mutant, indicating a key role in later stages of infection.     

Loss of pectin is an early event during infection of cocoyam roots by Pythium myriotylum

Cocoyam (Xanthosoma sagittifolium) is an important tuber crop in most tropical zones of Africa and America. In Cameroon, its cultivation is hampered by a soil-borne fungus Pythium myriotylum which is responsible for root rot disease. The mechanism of root colonisation by the fungus has yet to be elucidated. In this study, using microscopical and immunocytochemical methods, we provide a new evidence regarding the mode of action of the fungus and we describe the reaction of the plant to the early stages of fungal invasion. We show that the fungal attack begins with the colonisation of the peripheral and epidermal cells of the root apex. These cells are rapidly lost upon infection, while cortical and stele cells are not. Labelling with the cationic gold, which binds to negatively charged wall polymers such as pectins, is absent in cortical cells and in the interfacial zone of the infected roots while it is abundant in the cell walls of stele cells. A similar pattern of labelling is also found when using the anti-pectin monoclonal antibody JIM5, but not with anti-xyloglucan antibodies. This suggests that early during infection, the fungus causes a significant loss of pectin probably via degradation by hydrolytic enzymes that diffuse and act away from the site of attack. Additional support for pectin loss is the demonstration, via sugar analysis, that a significant decrease in galacturonic acid content occurred in infected root cell walls. In addition, we demonstrate that one of the early reactions of X. sagittifolium to the fungal invasion is the formation of wall appositions that are rich in callose and cellulose.     

The HCL gene of Medicago truncatula controls Rhizobium-induced root hair curling

The symbiotic infection of the model legume Medicago truncatula by Sinorhizobium meliloti involves marked root hair curling, a stage where entrapment of the microsymbiont occurs in a chamber from which infection thread formation is initiated within the root hair. We have genetically dissected these early symbiotic interactions using both plant and rhizobial mutants and have identified a M. truncatula gene, HCL, which controls root hair curling. S. meliloti Nod factors, which are required for the infection process, induced wild-type epidermal nodulin gene expression and root hair deformation in hcl mutants, while Nod factor induction of cortical cell division foci was reduced compared to wild-type plants. Studies of the position of nuclei and of the microtubule cytoskeleton network of hcl mutants revealed that root hair, as well as cortical cells, were activated in response to S. meliloti. However, the asymmetric microtubule network that is typical of curled root hairs, did not form in the mutants, and activated cortical cells did not become polarised and did not exhibit the microtubular cytoplasmic bridges characteristic of the pre-infection threads induced by rhizobia in M. truncatula. These data suggest that hcl mutations alter the formation of signalling centres that normally provide positional information for the reorganisation of the microtubular cytoskeleton in epidermal and cortical cells.     

Live-cell imaging elaborating epidermal invasion and vascular proliferation/colonization strategy of Verticillium dahliae in host plants

The soilborne ascomycete fungus Verticillium dahliae causes destructive vascular wilt disease in hundreds of dicotyledonous plant species. However, our understanding of the early invasion from the epidermis to the vasculature and the prompt proliferation and colonization in the xylem tissues remains poor. To elaborate the detailed infection strategy of V. dahliae in host plants, we traced the whole infection process of V. dahliae by live-cell imaging combined with high-resolution scanning electron microscopy. The 4D image series demonstrated that the apex of invading hyphae becomes tapered and directly invades the intercellular space of root epidermal cells at the initial infection. Following successful epidermal invasion, the invading hyphae extend in the intercellular space of the root cortex toward the vascular tissues. Importantly, the high-resolution microscopic and live-cell images demonstrated (a) that conidia are formed via budding at the apex of the hyphae in the xylem vessels to promote systemic propagation vertically, and (b) that the hyphae freely cross adjacent xylem vessels through the intertracheary pits to achieve horizontal colonization. Our findings provide a solid cellular basis for future studies on both intracellular invasion and vascular colonization/proliferation during V. dahliae infection and pathogenesis in host plants.     

Epidermal and cortical roles of NFP and DMI3 in coordinating early steps of nodulation in Medicago truncatula

Legumes have evolved the capacity to form a root nodule symbiosis with soil bacteria called rhizobia. The establishment of this symbiosis involves specific developmental events occurring both in the root epidermis (notably bacterial entry) and at a distance in the underlying root cortical cells (notably cell divisions leading to nodule organogenesis). The processes of bacterial entry and nodule organogenesis are tightly linked and both depend on rhizobial production of lipo-chitooligosaccharide molecules called Nod factors. However, how these events are coordinated remains poorly understood. Here, we have addressed the roles of two key symbiotic genes of Medicago truncatula, the lysin motif (LysM) domain-receptor like kinase gene NFP and the calcium- and calmodulin-dependent protein kinase gene DMI3, in the control of both nodule organogenesis and bacterial entry. By complementing mutant plants with corresponding genes expressed either in the epidermis or in the cortex, we have shown that epidermal DMI3, but not NFP, is sufficient for infection thread formation in root hairs. Epidermal NFP is sufficient to induce cortical cell divisions leading to nodule primordia formation, whereas DMI3 is required in both cell layers for these processes. Our results therefore suggest that a signal, produced in the epidermis under the control of NFP and DMI3, is responsible for activating DMI3 in the cortex to trigger nodule organogenesis. We integrate these data to propose a new model for epidermal/cortical crosstalk during early steps of nodulation.     

Observations on the Responses of Lentil Root Cells to Hyphae of Fusarium oxysporum

The infection of lentil roots by Fusarium oxysporum Schlecht and the responses of the host cells to invading hyphae were examined by light microscopy. Hyphae from inoculum placed on the zone of cell elongation entered the roots at the juncture of epidermal cells within 8 h after inoculation. Although swollen hyphal apices were observed on the epidermal cells, root penetration occurred without formation of these structures or appressoria. The sheath of material found on the surface of uninoculated roots was absent from inoculated roots penetrated by hyphae. Prior to penetration, the epidermal cells became irregular in shape and their cytoplasm appeared to be plasmolysed or granular. Hyphae were observed in the cortex 10—12 h after inoculation and non–penetrated cortical cells were distinctly lobate. Often these lobed cells had a broad, peripheral band of diffuse cytoplasm. When hyphae were first observed in the cortical cells, the walls were ruptured and only slightly stained or unstained by toluidine blue. The inability of such walls to bind the stain may have been the result of the removal of wall components by fungal enzymes. Although extensive proliferation of hyphae was evident throughout the cortex after 24 h of incubation, the endodermis and vascular cylinder were free of hyphae for at least 72 h. Hyphae from inoculum placed on the root hairs or the root apex failed to penetrate the roots during the first 24 h of incubation. The cytological results herein are discussed in relation to the infection of field plantings by this pathogen.     

Infection and invasion of roots by symbiotic, nitrogen-fixing rhizobia during nodulation of temperate legumes.

Bacteria belonging to the genera Rhizobium, Mesorhizobium, Sinorhizobium, Bradyrhizobium, and Azorhizobium (collectively referred to as rhizobia) grow in the soil as free-living organisms but can also live as nitrogen-fixing symbionts inside root nodule cells of legume plants. The interactions between several rhizobial species and their host plants have become models for this type of nitrogen-fixing symbiosis. Temperate legumes such as alfalfa, pea, and vetch form indeterminate nodules that arise from root inner and middle cortical cells and grow out from the root via a persistent meristem. During the formation of functional indeterminate nodules, symbiotic bacteria must gain access to the interior of the host root. To get from the outside to the inside, rhizobia grow and divide in tubules called infection threads, which are composite structures derived from the two symbiotic partners. This review focuses on symbiotic infection and invasion during the formation of indeterminate nodules. It summarizes root hair growth, how root hair growth is influenced by rhizobial signaling molecules, infection of root hairs, infection thread extension down root hairs, infection thread growth into root tissue, and the plant and bacterial contributions necessary for infection thread formation and growth. The review also summarizes recent advances concerning the growth dynamics of rhizobial populations in infection threads.     

GFP sheds light on the infection process of avocado roots by Rosellinia necatrix

In order to monitor Rosellinia necatrix infection of avocado roots, we generated a plasmid vector (pCPXHY1eGFP) constitutively expressing EGFP and developed a protoplast transformation protocol. Using this protocol, four R. necatrix isolates were efficiently transformed and were shown to stably express EGFP homogeneously while not having any observable effect on pathogenicity. Confocal laser scanning microscopy (CLSM) images of avocado roots infected with the highly virulent isolate CH53-GFP demonstrated that fungal penetration of avocado roots occurs simultaneously at several random sites, but it occurs preferentially in the crown region as well as throughout the lenticels and in the junctions between epidermal cells. Not only were R. necatrix hyphae observed invading the epidermal and cortical root cells, but they were also able to penetrate the primary and secondary xylem. Scanning electron microscopy (SEM) images allowed detailed visualisation of the hyphal network generated by invasion of R. necatrix through the epidermal, cortical and vascular cells, including hyphal anastomosis and branching points. To our knowledge, this is the first report describing the construction of GFP-tagged strains belonging to the genus Rosellinia for monitoring white root rot using CLSM and SEM.     

A barley ROP GTPase ACTIVATING PROTEIN associates with microtubules and regulates entry of the barley powdery mildew fungus into leaf epidermal cells

Little is known about the function of host factors involved in disease susceptibility. The barley (Hordeum vulgare) ROP (RHO of plants) G-protein RACB is required for full susceptibility of the leaf epidermis to invasion by the biotrophic fungus Blumeria graminis f. sp hordei. Stable transgenic knockdown of RACB reduced the ability of barley to accommodate haustoria of B. graminis in intact epidermal leaf cells and to form hairs on the root epidermis, suggesting that RACB is a common element of root hair outgrowth and ingrowth of haustoria in leaf epidermal cells. We further identified a barley MICROTUBULE-ASSOCIATED ROP-GTPASE ACTIVATING PROTEIN (MAGAP1) interacting with RACB in yeast and in planta. Fluorescent MAGAP1 decorated cortical microtubules and was recruited by activated RACB to the cell periphery. Under fungal attack, MAGAP1-labeled microtubules built a polarized network at sites of successful defense. By contrast, microtubules loosened where the fungus succeeded in penetration. Genetic evidence suggests a function of MAGAP1 in limiting susceptibility to penetration by B. graminis. Additionally, MAGAP1 influenced the polar organization of cortical microtubules. These results add to our understanding of how intact plant cells accommodate fungal infection structures and suggest that RACB and MAGAP1 might be antagonistic players in cytoskeleton organization for fungal entry.     

Development of N2-fixing nodules on the wetland legume Lotus uliginosus exposed to conditions of flooding

Seeds of the wetland legume, Lotus uliginosus , were germinated and grown in vermiculite which was either continuously flooded or well-drained. Plants from both treatments were infected by Mesorhizobium loti strain DUS341 via a 'classical' root hair pathway, although some flooded plants appeared to be infected via enlarged epidermal cells. Subsequent to infection by M. loti , nodule meristems, which had developed within the root outer cortex, were penetrated by infection threads that released bacteria into the meristematic cells. The infection threads and infection droplets were immunogold labelled with monoclonal antibodies (MAC265 and MAC236) that recognize epitopes (at approx. 155/170 and 170/210 kDa, respectively) on a glycoprotein component of the matrix that surrounded the bacteria within the threads or droplets. Although labelling of infection threads or infection droplets with MAC236 was stronger than that with MAC265, both antibodies strongly labelled material occluding intercellular spaces in the cortices of developing nodules that had not yet expressed nitrogenase (as determined by a lack of signal after immunogold labelling with an antibody raised against nitrogenase component II). After 60 d, nitrogenase activity, shoot and root dry weights, and nodule fresh weight per plant did not differ between the treatments. After a further 30 d submergence, the flooded stems developed extensive aerenchyma and there was profuse development of (nodulated) adventitious roots. Nodules also formed at the junction of adventitious roots and the subtending stem and these were connected vascularly to a small stalk of tissue which gave rise to both a nodule and an adventitious root. The flooded nodules had prominent lenticels, and possible air pathways from the atmosphere to the nitrogen-fixing bacteroids are discussed.     

Barriers to the radial diffusion of ions in maize roots

Summary The water-extractable and ion-exchangeable fractions of the free space of maize roots for sodium ions has been determined. The free space of whole roots, excised roots and isolated stelar and cortical tissues, has been compared and the results examined for any evidence of a barrier between the cortex and the stele. Similarly the free space of whole roots and excised roots, from which the epidermal and outer cortical cells have been removed by shaving, has been compared and the results examined for any evidence of an epidermal barrier.Whole roots gave a free space value some 20% lower than excised roots. It was calculated that this difference could be accounted for if the cortical tissues only were considered in estimating the whole root value, that is if the stele was considered as participating in the excised root but not in the whole root. Samples in which isolated cortical and stelar material were measured together, or separately and the value calculated, gave similar values to those obtained for excised roots. These results are interpreted as evidence that a barrier to free diffusion exists between the cortical and the stelar tissues at or near the endodermis. Shaving both whole and excised roots increased the free space by about 35%. However, as this value was similar for both, it was concluded that the increase was due to the contribution of damaged cortical cells and does not indicate that the epidermis is an effective barrier to the diffusive entry of sodium ions into the root.     

Microtubule reorganization as a response to implementation of NO signals in plant cells

The effects of an exogeneous NO donor, sodium nitroprusside, on the orientation and organization of cortical microtubules in Arabidopsis thaliana root cells expressing GFP-MAP4 were studied in vivo. It was found that sodium nitroprusside treatment (10–500 μM, 24 h) caused the acceleration of primary root growth and enhanced initiation of root hairs in the differentiation zone. The influence of sodium nitroprusside revealed in changes in the orientation and organization of cortical microtubules in different types of cells of A. thaliana root. The most sensitive to sodium nitroprusside exposure were microtubules in epidermal cells of the elongation zone, where native transverse orientation of cortical microtubules turned into random, oblique, or longitudinal relative to the primary root axis. We suggest that NO, as one of the intracellular secondary messengers, triggers cell differentiation by reorientation of cortical microtubules, possibly via tubulin nitrotyrosination.     

Symptomatic Evidence for Differential Root Invasion by Fusarium Crown and Root Rot Pathogens Between Common Tomato Lycopersicon esculentum and Its Varieties

The pathogenic isolates (Kin2001a, Kin2001b and Kin2003) of Fusarium oxysporum f. sp. radicis‐lycopersici were obtained from hydroponically cultured seedlings of pear tomato (Lycopersicon esculentum var. pyriforme) infected at different times and their pathogenicity examined in an in vitro assay system on cotyledonal seedlings of pear tomato, cherry tomato (L. esculentum var. cerasiforme) and common tomato (L. esculentum). With the in vitro assay, infection and subsequent disease progress could be microscopically observed. Pear and cherry tomatoes suppressed invasion by all isolates at the junctions of epidermal cells along the root, comparable with the resistant cultivars of common tomato. The pathogen entered pear and cherry tomatoes at the tips of lateral roots and tap roots, in contrast to infection of susceptible cultivars of common tomato. In Kin2003‐inoculated roots, the top of the lateral rootlets first became discoloured, followed by the cortical parenchyma, central xylem vessel and finally the crown. This dark‐brown discolouration expanded rapidly and severe rot developed in the discoloured regions. In contrast, the dark‐brown discolouration in Kin2001b‐infected roots expanded into the cortical parenchyma cells abutting the originally infected lateral rootlets and at a much slower rate. Kin2001a was in a new group that entered via the cortical cleavage formed by the emergence of lateral rootlets, in addition to the tips of taproots and lateral roots. In this in vitro assay system, the Japanese pathogenic isolates collected from different districts of Japan were characterized and classified by the mode of host invasion. Of 13 isolates, four were placed with Kin2003, six with Kin2001a and three with Kin2001b.     

Root Infection and Systemic Colonization of Maize by Colletotrichum graminicola

Colletotrichum graminicola is a filamentous ascomycete that causes anthracnose disease of maize. While the fungus can cause devastating foliar leaf blight and stalk rot diseases, little is known about its ability to infect roots. Previously published reports suggest that C. graminicola may infect maize roots and that root infections may contribute to the colonization of aboveground plant tissues, leading to disease. To determine whether C. graminicola can infect maize roots and whether root infections can result in the colonization of aboveground plant tissues, we developed a green fluorescent protein-tagged strain and used it to study the plant root colonization and infection process in vivo. We observed structures produced by other root pathogenic fungi, including runner hyphae, hyphopodia, and microsclerotia. A mosaic pattern of infection resulted from specific epidermal and cortical cells becoming infected by intercellular hyphae while surrounding cells were uninfected, a pattern that is distinctly different from that described for leaves. Interestingly, falcate conidia, normally restricted to acervuli, were also found filling epidermal cells and root hairs. Twenty-eight percent of plants challenged with soilborne inoculum became infected in aboveground plant parts (stem and/or leaves), indicating that root infection can lead to asymptomatic systemic colonization of the plants. Many of the traits observed for C. graminicola have been previously reported for other root-pathogenic fungi, suggesting that these traits are evolutionally conserved in multiple fungal lineages. These observations suggest that root infection may be an important component of the maize anthracnose disease cycle.     

A mycorrhizal fungus changes microtubule orientation in tobacco root cells

Summary Cortical cells of mycorrhizal roots undergo drastic morphological changes, such as vacuole fragmentation, nucleus migration, and deposition of cell wall components at the plant-fungus interface. We hypothesized that the cytoskeleton is involved in these mechanisms leading to cell reorganization. We subjected longitudinal, meristem to basal zone, sections of uninfectedNicotiana tabacum roots to immunofluorescence methods to identify the microtubular (MT) structures associated with root cells. Similar sections were obtained from tobacco roots grown in the presence ofGigaspora margarita, an arbuscular mycorrhizal fungus which penetrates the root via the epidermal cells, but mostly develops in the inner cortical cells. While the usual MT structures were found in uninfected roots (e.g., MTs involved in mitosis in the meristem and cortical hoops in differentiated parenchyma cells), an increase in complexity of MT structures was observed in infected tissues. At least three new systems were identified: (i) MTs running along large intracellular hyphae, (ii) MTs linking hyphae, (iii) MTs binding the hyphae to the host nucleus. The experiments show that mycorrhizal infection causes reorganization of root MTs, suggesting their involvement in the drastic morphological changes shown by the cortical cells.     

Nodulation of Aeschynomene afraspera and A. indica by photosynthetic Bradyrhizobium Sp. strain ORS285: the nod-dependent versus the nod-independent symbiotic interaction

Here, we present a comparative analysis of the nodulation processes of Aeschynomene afraspera and A. indica that differ in their requirement for Nod factors (NF) to initiate symbiosis with photosynthetic bradyrhizobia. The infection process and nodule organogenesis was examined using the green fluorescent protein-labeled Bradyrhizobium sp. strain ORS285 able to nodulate both species. In A. indica, when the NF-independent strategy is used, bacteria penetrated the root intercellularly between axillary root hairs and invaded the subepidermal cortical cells by invagination of the host cell wall. Whereas the first infected cortical cells collapsed, the infected ones immediately beneath kept their integrity and divided repeatedly to form the nodule. In A. afraspera, when the NF-dependent strategy is used, bacteria entered the plant through epidermal fissures generated by the emergence of lateral roots and spread deeper intercellularly in the root cortex, infecting some cortical cells during their progression. Whereas the infected cells of the lower cortical layers divided rapidly to form the nodule, the infected cells of the upper layers gave rise to an outgrowth in which the bacteria remained enclosed in large tubular structures. Together, two distinct modes of infection and nodule organogenesis coexist in Aeschynomene legumes, each displaying original features.     

Distribution and pathway for phloem-dependent movement of Melon necrotic spot virus in melon plants

The translocation of Melon necrotic spot virus (MNSV) within tissues of inoculated and systemically infected Cucumis melo L. 'Galia' was studied by tissue-printing and in situ hybridization techniques. The results were compatible with the phloem vascular components being used to spread MNSV systemically by the same assimilate transport route that runs from source to sink organs. Virus RNAs were shown to move from the inoculated cotyledon toward the hypocotyl and root system via the external phloem, whereas the upward spread through the stem to the young tissues took place via the internal phloem. Virus infection was absent from non-inoculated source tissues as well as from both shoot and root apical meristems, but active sink tissues such as the young leaves and root system were highly infected. Finally, our results suggest that the MNSV invasion of roots is due to virus replication although a destination-selective process is probably necessary to explain the high levels of virus accumulation in roots. This efficient invasion of the root system is discussed in terms of natural transmission of MNSV by the soil-borne fungal vector.     

Infection and Invasion of Roots by Symbiotic, Nitrogen-Fixing Rhizobia during Nodulation of Temperate Legumes

Bacteria belonging to the genera Rhizobium, Mesorhizobium, Sinorhizobium, Bradyrhizobium, and Azorhizobium (collectively referred to as rhizobia) grow in the soil as free-living organisms but can also live as nitrogen-fixing symbionts inside root nodule cells of legume plants. The interactions between several rhizobial species and their host plants have become models for this type of nitrogen-fixing symbiosis. Temperate legumes such as alfalfa, pea, and vetch form indeterminate nodules that arise from root inner and middle cortical cells and grow out from the root via a persistent meristem. During the formation of functional indeterminate nodules, symbiotic bacteria must gain access to the interior of the host root. To get from the outside to the inside, rhizobia grow and divide in tubules called infection threads, which are composite structures derived from the two symbiotic partners. This review focuses on symbiotic infection and invasion during the formation of indeterminate nodules. It summarizes root hair growth, how root hair growth is influenced by rhizobial signaling molecules, infection of root hairs, infection thread extension down root hairs, infection thread growth into root tissue, and the plant and bacterial contributions necessary for infection thread formation and growth. The review also summarizes recent advances concerning the growth dynamics of rhizobial populations in infection threads.