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Extraradical hyphae are generally thin, but profuse around roots. The overall spore development is quite similar in the two genera, but Acaulospora develop its spores laterally from the neck of a predifferentiated ‘sporiferous saccule’ that is formed terminally on a fertile hypha, while in Enthrospora, spores are borne from within the neck of the saccule (Figure 2). Spores are produced singly, and on rare occasions in loose aggregates. There is an outer layer to the spore wall, which is continuous with the wall of the saccule neck. This layer usually sloughs with age or manipulation, leaving the spores without hyphal attachment (INVAM 2013; Ruissen notes, 2012a).
Archaeospora, Paraglomus Arbuscules and intraradical hyphae consistently stain lightly, and definite vesicles are not formed in any of the species examined to date. Archaeospora have a patchy distribution of arbuscules, which consist of narrow trunk hyphae with fine branching near tips in both genera. Intracellular hyphae are often tightly coiled, but can also coil more infrequent and looser, with irregular branching. The hyphae have variable widths, depending on their growing pattern, and spread both intra - and inter-cellularly in Archaespora. Extraradical hyphae stain darkly, and are often in profuse abundance around the roots.
The spores of Paraglomus develop terminally on a cylindrical to slightly flared subtending hypha.
They are produced singly, and on rare occasions in loose aggregates of a few spores. The sub-cellular spore structure and development is identical to that of Glomus species, having layers of the spore wall that are continuous with layers of the subtending hypha after blastic expansion of the hyphal tip. In some species the subtending hypha of mature spores is so thin that it is hard to see or separate from the spore. Archaeospora spores are also produced singly and more rarely in loose aggregates, and originate laterally from the neck of a sporiferous saccule that is formed terminally on a fertile hypha.
Spore development can be similar to that of Acaulospora species, with spores eventually detaching from their hyphae and remaining sessile in the soil. The spores can also develop similar to that of Glomus species, on a hyphal ‘pedicel’ branching from the subtending hypha of the soporiferous saccule (Figure 2). In the latter case, Glomus – like spores may also be formed from external hyphae, some of which also can form soporiferous saccules (INVAM 2013; Ruissen notes, 2012b).
Gigaspora, Scutellospora These genera do not form intraradical vesicles, and are thus not ‘VAM fungi’. Instead, auxiliary cells (thin–walled cells which compartmentalize lipids) are formed singly or in clusters by branching from external hyphae, often differentiating on germ tubes from spores prior to establishment of mycorrhizal colonization. Colors are hyaline to dark brown. The cells are very abundant around roots during early colonization, but become less frequent and sometimes absent as sporulation increases, suggesting that one of their tasks is to provide carbon macromolecules independent of the host during spore formation.
Gigaspora have auxiliary cells with a spiny surface, while Scutellospora form broad concavities to varying degrees that makes the cells appear almost smooth to having wide knobs. Arbuscules stain darkly. They generally have swollen trunks with branches tapering abruptly at tips, and the network can be abundant for a long time after the roots have ceased growth. With arbuscule senescence, the fine tips are degraded but the trunk may remain intact in cells as tightly packed coils. Intraradical hyphae are often coiled throughout the root, but their coiling is most prominent at entry points. The hyphae vary in width, but often appear knobby or have projections. Infection units merge and form a uniform colonization throughout the root cortex. Extraradical hyphae are either coarse and wide, or fine hyphae. Both are abundant during auxiliary formation, whereas fine hyphae are less evident in older cultures. Most of the fungal biomass is found in the external hyphae, which are bridging over long distances with few deriving branches. Gigaspora and Scutellospora form large spores that are usually 200 µm after maturation, and range from white to dark red in color. The spores develop singly and blastically, from the tip of a bulbous, sporogenous cell that is formed terminally on a fertile hypha growing relatively distant from the root (Figure 2). Subcellular organization consists only of a bilayered spore wall. In Gigaspora, germ tubes arise from a thin papillate, or warty, layer developing from the inner surface. Spores are without ornamentations. Some Scutellospora species, on the other hand, have ornamentations of the outer layer of their spore wall, and the inner layer may vary in color.
Germ tubes arise from a plate–like germination shield that is associated with the flexible inner wall (INVAM 2013; Parniske 2008; Ruissen notes, 2012c; Ruissen notes, 2012e).
Figure 2: Classification and illustrated spore formation in the two glomeromycetous sub orders;
Glomineae, which form vesicles, and Gigasporineae, forming auxiliary cells. Both morphologial and molecular characters have been taken into account. Source: (INVAM 2013).
Fine endophytes Another group of fungi that form symbiosos within same hosts as AMF are called fine endophytes, formerly named Glomus tenue and classified as glomalean fungi. The function and taxonomical status of these fungi is an unsolved mysterium, but they seem to be more frequent in cold or harsh environments than AMF, which can be abundant in antarctic cold areas but have a very low frequency at the more polar sites. Typical AM forming plants are often abundant in such harsh environments, but their lack of AM shows that the symbiosis is less important in arctic and alpine ecosystems than it is in temperate ecosystems. The relatively high frequency of fine endophytes at high latitudes compared to that of AMF indicates that these fungi are better adapted for establishing successful symbioses under adverse conditions that inculde short growth seasons (Christie & Nicolson 1983; Olsson et al. 2004;
Thippayarugs et al. 1999).
How does AM form, and how does it work?
When a fungus and a plant host are going to form AM symbiosis, both parts have to recognize and accept each other before intimate associations that involve penetration of plant tissue and invasion of individual host cells can be established. First, the plant roots release root exudates that are recognized by the AMF spores and hyphae, which starts to grow, branch and alter their physiological activity, once they perceive this signal. The substance released are called strigolactones, because when they were discovered 50 years ago, this class of compounds was found to induce seed germination of the parasitic plant genus Striga. Faster growth of the AMF hyphae increases the chance of encountering a host, but the hyphae can also be stimulated to grow chemotropically towards a root, because strigolactones hydrolyze quickly in soil and form a steep concentration gradient in the rhizosphere that makes its perception a reliable guide towards the root. Other microbes are also able to recognize inducing signals released from the plant, which therefore must be able to to recognize their rightful symbionts and reject saprotrophs or potential pathogens at the same time. This problem is solved by means of unknown, diffusible symbiosis signals called ‘Myc factors’ that are emitted from AMF hyphae growing towards the roots in response to the plant initial signals. The Myc factors initiate a cascade of signals in the plant cells that leads to altered metabolism and transcription of symbiosis – related genes in the plant, which then actively helps the fungus colonize its roots. Pathogenesis – related proteins may also be released from the plant during early stages of AMF colonization, as a defence reaction to unspecific microbial signals (Parniske 2008; Reinhardt 2007).
Once the fungus comes into contact with the root surface, infection structures named appressoria, or hyphopodia in the case of AMF, is formed outside the epidermal cell layer. The underlying epidermal cell responds to mechanical stimulation together with a local signal emitted from the fungus that contains information about the exact position of the appressoria, and start reorganizing its cell components to form an aggregation of microtubules, actin filaments and ER cisternae; the pre – penetration apparatus (PPA). A fungal hypha that extends from the hyphopodium then penetrate the epidermal cell through the trajectory formed by the PPA, while the cell membrane invaginates, leaving the fungus in an apoplastic pocket that contains remnants of the plant cell wall. The PPA guides the fungus through epidermal cells and towards the cortex, where the hypha leaves the plant cell and enters the apoplast. Lateral, intercellular growth and branching along the root axis eventually lead to the hyphae inducing formation of PPA – like structures in inner cortical cells, through a similar process as with the epidermal cells. The hyphae enter these cells, where they develop into a highly ramified structure with fine terminal tips; the arbuscule. This structure is separated from the host cytoplasm by a plant – derived periarbuscular membrane (PAM) that is continuous with the plasmalemma, the fungal plasma membrane, and the periarbuscular space between them, containing remnants of the fungal cell wall and apoplastic material of the plant. The way that AM develops in a plant root are illustrated in Figure 3.
The arbuscule has a high surface – to volume ratio, which makes this symbiotic interface a perfect site for exchanging nutrients and symbiotic signals. Local cell autonomous signals that are produced by the fungus activate expression of genes that, among others, code for transporter proteins mediating this metabolite exchange. Arbuscules generally have a short lifetime, but a single host cell may undergo several rounds of successive fungal invasions. Early degradation of arbuscules is probably a way of discriminating between efficient and inefficient fungal species, because research has suggested that the lifetime of an arbuscule is influenced by its ability to deliver nutrients. A short arbuscule lifetime thus ensures constant renewal of the hyphal network, while connections are made to the most efficient nutrient providers (Javot et al. 2007; Parniske 2008; Reinhardt 2007).
Figure 3: Development of AM symbiosis in a plant root. Fungal spores germinate, and hyphae start branching and growing towards the root when perceiving strigolactones; initial signals that are released from the plant. When the hyphae encounter the root, Myc factors act as a recognition signal to the plant, which then starts to express symbiosis-related genes and alter their metabolic function. A fungal hypha extending from a hyphopodium infection structure penetrates the epidermal cell layer through a pre – penetration apparatus (PPA) formed by cellular reorganization, and guides the fungus into the cortex. Here, the fungus enters the apoplast and grows laterally along the root axis before it invades the inner cortical cells in a similar manner as with the epidermal cells, and branches extensively to form arbuscules. The endodermis is never penetrated. Source: (Parniske 2008).
Due to improved molecular methods, it has now become clearer how AM symbiosis develops, how nutrients are taken up by external soil hyphae, and how these substances are translocated on to the internal hyphae before leaving the fungus through structures that are well adapted to nutrient exchange with the host plant. Yet, the major changes in fungal and plant gene expression that leads to AM formation are not fully resolved. Important details about the physiological mechanisms underlying signalling pathways and nutrient transport are missing, and there is also need for identification of all single components involved (Balestrini & Lanfranco 2006; Parniske 2008; Reinhardt 2007).
Mycorrhizal plants have two ways of taking up nutrients: through the roots and root hairs, also called the direct pathway (DP), or through the mycorrhizal pathway (MP). The pathways have different biochemistry, and the fact that plants often favor the MP for nutrient uptake might be due to a higher efficiency of fungal nutrient transporters. However, when nutrients are absorbed through the MP, they first have to pass a boundary between the soil and fungal hyphae, before transport through the intraradical hyphae eventually leads them to a second boundary between the fungus and plant which must also be passed before nutrient uptake is complete. The DP only requires that nutrients pass the soil-plant cell boundary, but since depletion zones develop quickly in the rhizosphere, and inorganic ion replacement from bulk soil is slow and inefficient, choice of pathways for nutrient uptake is probably related to the ability of root systems to access nutrients from undepleted soil. In this case, the MP is most efficient because of the ability of AM hyphae to comb the soil for nutrients that are placed far away from where the root hairs can reach. (Ruissen 2012a; Smith & Smith 2011) The varying mycorrhizal growth responses (MGR) that may occur to AM colonization are often discussed in relation to a mutualism – parasitism continuum (Johnson et al. 1997). Positive MGRs are usually due to increased nutrient uptake via the MP, but many factors, both molecular and ecological, can influence the final response in the plant. For example; fungal growth and capacity of nutrient uptake/delivery, efficiency of nutrient exchange interfaces, root morphology and ability to produce nutrient mobilizing root exudates, as well as environmental factors, all may influence MGRs.
Thus; the conventional explanation to why plants sometimes show neutral or negative MGRs is that physiological features in both the plant and and the fungus makes the MP less efficient than the DP alone, and maintaining the fungus becomes negative to the plant because the P benefit is lower than the C cost, especially when nutrient levels are high. In this case, the fungus could be regarded as a parasite. However, the plant never eliminates its fungal partner to save photosyntates, although colonization and MP operation may be suppressed with strongly elevated P levels in the soils that make the symbiosis redundant (Amijee et al. 1989; Joner 2012; Nagy et al. 2009; Smith & Smith 2011).