Carbon Sequestration and Soil Organic Carbon Storage

Grogan and Mathews (2001) reviewed available soil C sequestration models and concluded then that the CENTURY model seemed to have the best potential for adaptation to bioenergy crop systems because of its integrated plant-soil approach and the availability of specific forestry subroutines. Not surprisingly, the model has been incorporated into many crop growth models, such as EPIC and DAYCENT to simulate SOC dynamics along with crop growth (Del Grosso et al. 2005; Izaurralde et al. 2006).

Available lands for biofuel production are very limited; therefore, meeting the feedstock needs of the bioenergy industry may require the conversion of agricultural lands to bioenergy crops. Land-use change consequently affects the ecosystem C balance. Corn, soybeans, and wheat are the three major crops with the highest production among food crops in U. S., and switchgrass and Miscanthus are two second-generation biofuel crops with the potential to produce a large amount of biofuel feedstocks and mitigate carbon emissions (Parrish and Fike 2005; Tilman et al. 2009; Pimentel et al. 2010). Davis et al. (2011) used the DAYCENT model to estimate the amount of N leaching and storage of soil C when replacing lands currently growing corn with perennial biofuel grasses. Overall, the model predicted a significant decrease in N leaching up to 24% and an increase in SOC up to 2.8% when land use changed from corn to switchgrass. These results are consistent with field studies showing that perennial grasses like switchgrass can store 1.1 Mg C ha1 yr-1 in the upper 1 m of the soil on conservation reserve program lands (Gebhart et al. 1994). In another field study, McLaughlin and Walsh (1998) reported that soils under switchgrass production had SOC sequestration rates reaching 20 to 30 times greater than soils under annual row crops. Furthermore, McLaughlin et al. (2002) revealed that switchgrass grown on bioenergy research plots could add 1.7 Mg C ha-1 yr-1.

Few insects have been identified as potential pests of switchgrass, and early studies indicate the species is not a preferred host for many insect species (Davis 1914; Walkden 1943). Switchgrass typically is an inferior host relative to other warm-season crops (Nabity et al. 2011; Prasifka et al. 2011a), and fall armyworm (Spodoptera frugiperda (J. E. Smith) (Lepidoptera: Noctuidae)) has reduced survivorship on switchgrass compared with other grasses (Nabity et al. 2011). Variation in armyworm resistance among switchgrass strains cultivars also has been observed (Dowd and Johnson 2009).

While insect damage has been considered minimal, there is potential for this to increase if or when switchgrass for bioenergy systems scale up (Parrish and Fike 2005). Little published data is available on insect pressures during establishment. However, corn flea beetle Chaetocnema pulicaria (Chrysomelidae) damage has been common in Virginia plantings (Dale Wolf, personal communication) and insect pressures during the seedling stage likely represent greatest insect threat to the switchgrass stand productivity.

Grasshoppers (Saltatoria) are known to feed on switchgrass, but the extent of the damage has not been quantified (Parrish and Fike 2005). Schaeffer et al. (2011), in a baseline study of insects in Nebraska switchgrass stands, found that about 60% of arthropods collected were of the orders Thysanoptera and Hymenoptera; leafhoppers, grasshoppers, grass flies and wire worms were noted as the most abundant of potential pest species.

Life stages, geographic distribution, and the symptoms of a stem­boring caterpillar, Blastobasis repartella (Dietz.), recently have been described (Adamski et al. 2010; Prasifka et al. 2010). Switchgrass is the only known host for this caterpillar (Prasifka et al. 2011b), and in a distributional survey B. repartella was found both in cultivated and natural switchgrass stands in eight northern USA states (Prasifka et al. 2010). The species was not observed at southern locations (Arkansas, Louisiana, Oklahoma, and Texas) but lack of observation could not rule out presence at these latitudes. In four northern states (Illinois, Nebraska, South Dakota, and Wisconsin) 1 to 7% of tillers were damaged by B. repartella.

A new species of gall midge (Chilophaga virgate Gagne (Diptera: Cecidomyiidae)) was recently discovered in South Dakota, USA (Boe and Gagne 2011). Proportion of tillers infested with the gall midge in 10 switchgrass genotypes ranged from 7 to 22%. The mass of infested tillers was 35% lower than that of normal tillers, and infested tillers produced no appreciable seed. Such insect pests and associated yield reductions may well become more evident or more common as switchgrass is grown more extensively.

For endophytic fungal growth, surface-sterilized tissues are cut into small pieces or homogenized using a homogenizer and plated on Potato dextrose agar (PDA) or other fungal culture media, such as MEA medium (2% malt extract and 1.8% agar) (Vallino et al. 2009) supplemented with several antibiotics, such as ampicillin (100 mg/L), chloramphenicol (50 mg/L) and streptomycin (50 mg/L) to prevent bacterial growth (Ghimire et al. 2011; Craven K, personal communication). Plates containing tissues are incubated at 25-28°C for up to one month. Observations should be taken every day in order to isolate individual strains for further identification. The growing fungal colonies are then re-plated on fresh medium to obtain individual colonies.

Sexual and asexual reproduction systems and associated mechanisms are essential for the breeders to determine both general features of a breeding program and specific procedures of hybridization, selection and cultivar development. The inflorescence of switchgrass is a typical open and diffuse panicle of 20-60 cm in length and 20-40 cm in width (Fig. 1). Each panicle consists of many to hundreds of spikelets, with two dissimilar florets in each spikelet (Tyrl et al. 2002). The lower floret is staminate while the upper one is perfect (Fig. 2). Stigma of each perfect floret exerts out of lemmas about 1-3 days earlier than anthers.

But the stigma is still receptive when anthers shed pollen grains on the same floret. The morphological structure and blooming behavior of florets provide specific conditions which may favor outcrossing over selfing. Switchgrass has long been recognized as a naturally cross-pollinating species (Jones and Brown 1951). The peak period of pollen shedding in a day occurs from 10 am to 4 pm (Jones and Brown 1951). Dispersal of pollen grains is facilitated by wind. Occasionally, pollen grains can form a visible tunnel (like a swirl) in the air over a large switchgrass planting. Anthesis duration of a panicle is about one to two weeks long, which may be affected by genotypes and environmental factors.

Talbert et al. (1983) reported that bagged inflorescences on average produced less than 1% seed yield as compared to open-pollinated inflorescences on the same plants in an experiment of 33 plants. Their results indicated that switchgrass produces seed primarily through cross pollination while self-pollination is minimal. Using verified tetraploid and octoploid plants in a greenhouse, Taliaferro and Hopkins (1996) concluded that there is a strong genetic barrier between tetraploid and octoploid plants. Martinez-Reyna and Vogel (2002) reported the cross­fertility between octoploid and tetraploid plants is inhibited by a post­fertilization incompatibility system. They observed tetraploid by octoploid or reciprocally fertilized zygotes have a much slower growth than tetraploid by tetraploid or octoploid by octoploid zygotes. However, the cross-fertility between tetraploid upland and lowland plants is substantial or quite normal, and switchgrass outcrossing behavior is enforced by self-incompatibility (Taliaferro and Hopkins 1996; Martinez-Reyna and Vogel 2002). However,

seed set of selfing octoploid plants was as high as 6% (a range of 0-35%) while less than 1% (0-1.5%) selfed seed is produced in bagged inflorescences of tetraploid plants (Taliaferro and Hopkins 1996). Martinez-Reyna and Vogel (2002) noted the self-incompatibility in switchgrass is controlled by a gametophytic prefertilization incompatibility system, which is similar to the S-Z incompatibility system commonly seen in many members of the Poaceae plants. It appears tetraploid plants tend to be less self-fertile than octoploid plants. In later experiments, Taliaferro (2002) reported some lowland switchgrass plants of ‘Alamo’ and ‘Kanlow’ produced more than 20 selfed (S1) seeds from bagged plants while selfing plants of ‘Caddo’, ‘Blackwell’ and ‘Cave-in-Rock’ produced 100 or more seeds.

Recently, molecular markers have been available and used to accurately identify breeding origin of progeny of two controlled crosses (Okada et al. 2010b; Liu and Wu 2012). Okada et al. (2010b) found about 4% progeny of a full-sib cross between a ‘Kanlow’ (female) and an ‘Alamo’ (male) parents are selfed. In an attempt to make a full-sib cross-fertilized mapping population, two lowland plants: ‘NL94 LYE16x13’ (NL94) and ‘SL93 7×15’ (SL93) were grown in a large growth chamber before they were blooming. From the seed harvested from the female parent NL94 plant, 456 progeny were grown in a greenhouse (Liu and Wu 2012). Using 12 simple sequence repeat markers, they identified 279 of the progeny population were selfed progeny of NL94 whereas only 39% (177) were crossed between the two parents. The selfed progeny percentage is much higher than those reported before. However, whether the NL94 plant would produce a similar amount of selfed and crossed progeny when grown under field conditions and when other crossing compatible switchgrass plants are available, is not known. The authors speculated that limited wind flow in the growth chamber enhanced NL94 self-fertilization and reduced cross fertilization. Similarly, it has been observed that some other switchgrass genotypes have a self­pollination rate as high as 50% as cited by Casler et al. (2011).

Switchgrass can be reproduced in asexual ways. In breeding programs, switchgrass clones are normally produced by digging selected field-grown stands and separating into individual ramets, which are used to establish new crossing blocks. But it is difficult to use the labor intensive method to produce a large amount of clones. A micropropagation method to produce switchgrass by nodal culture has been reported (Alexandrova et al. 1996). With nodal segments cultured on an optimized MS medium, 500 plantlet clones can be produced from one parent plant within a period of three months. In our own experience, mature nodes of switchgrass genotypes can be cut into 1- or 2-nodal segments, when grown into a good soil medium and maintained moisture for two or three weeks, new clonal plants will grow out from the buds on the nodes (Fig. 3a and 3b). The greenhouse-type nodal propagating methods may be useful in producing clonal plants of the same size, which are important in quantitative trait loci (QTL) mapping experiments.

It appears there is large variation for breeding behavior in switchgrass. This is not surprising because switchgrass is genetically very diverse. Switchgrass is a predominantly wind-facilitated outcrossing species. Conventional breeding and selection methods have been developed and used based on the major mating system. It is also possible to inbreed switchgrass, at least in some genotypes. Breeding methods exploring the mating systems will be discussed in detail in latter sections.

MiRNAs are important regulators playing important roles in plant development and plant responses to biotic and abiotic stresses as well as in regulating miRNAs themselves and other small RNAs (Bartel 2009; Chen 2009; Chuck et al. 2009; Poethig 2009; Voinnet 2009; Zhu et al. 2009). Many miRNA families have multiple members with different temporal and spatial expression patterns (Palatnik et al. 2007; Nag et al. 2009; Voinnet

2009) . Mature miRNAs usually have multiple targets, which constitute a complicated regulatory network impacting different aspects of plant life (Axtell and Bowman 2008; Bartel 2009; Rubio-Somoza and Weigel 2011).

Based on reactor configuration, gasifiers can be classified into fixed-bed (downdraft and updraft), fluidized-bed, and entrained-bed. Fluidized-bed gasifier can be bubbling or circulating-bed. In fixed-bed gasifiers, biomass is fed from top so the biomass moves downward through the reactor. Ash and residual char pass through and are collected at the bottom grates. The gas flow in the fixed-bed gasifier can be in upward or downward direction. When the gas flow is in upward direction, it is called an "updraft" gasifier; whereas when the flow is in downward direction, it is called "downdraft" gasification. Fixed-bed gasifiers have a relatively simpler design and are best for small-scale applications of up to 1.5 MWe (Bridgwater 2006). Fluidized — bed gasifiers require a bed of fluidizing medium such as silica or olivine, where the biomass and fluidizing agent, such as air are introduced. Due to the bed fluidization, fluidized-bed gasifiers have high heat and mass transfer rates which result in higher efficiency as compared to the fixed-bed gasifier especially as the scale is increased.

Based on the heat source for the gasification reaction, gasifiers can be classified into directly or indirectly heated. In the case of directly heated gasifiers, no external heat is supplied. Oxygen (or any gas containing oxidizing agent) is supplied into the reactor which results in partial oxidation of biomass. Partial oxidation of biomass provides the heat required for the gasification endothermic reactions. In the case of indirectly heated gasifier, heat is provided by external heat source by combustion of fuels, such as char, natural gas or using electricity.

Higher substrate concentrations affect the agitation, lower oxygen transfer rates and reduce the availability of enzymes, hence limiting the rate of cellulolytic enzyme synthesis (Oashima et al. 1990). The enzyme synthesis rate is directly proportional to substrate intake. Cellulose and xylans are insoluble polymers that could lead to a longer lag phase during enzyme fermentation. A supplement of soluble sugars alongside lignocellulosic biomass plays an important role in inducing enzyme production. Both the carbon and nitrogen sources are equally important requirements in the enzyme synthesis. Urea and aqueous ammonium hydroxide are potential nitrogen sources.

An economically efficient biorefinery could be expected to require a steady flow of feedstock throughout the year. The logistics of providing a daily flow of several thousand Mg of bulky biomass could be challenging. Hwang (2007) enhanced a model (Tembo et al. 2003; Mapemba et al. 2007; Epplin et al. 2007; Mapemba et al. 2008) designed to approximate a just-in-time system for delivery of feedstock.

Providing a continuous flow of feedstock could be a major challenge for a biofuel production system that relies exclusively on switchgrass for feedstock. Most published estimates of switchgrass production costs assume that switchgrass would be harvested once per year when yield per hectare is maximized resulting in a narrow harvest window. A wide harvest window could reduce the fixed costs of harvest machinery per Mg of feedstock relative to a narrow harvest window and reduce storage cost. However, switchgrass harvestable dry matter yield and fertilizer requirements differ across harvest month (Haque et al. 2009; Haque 2010). The maximum expected dry matter yield of switchgrass grown in the U. S. Southern Plains is obtained by harvesting in either September or October (Table 10). In most years, harvest during April, May, and June would damage the plants and result in lower expected yields in subsequent years. The downside of an extended harvest window is that the expected yield from harvest in July is approximately 80 percent of maximum, and if switchgrass is left to stand in the field, dry matter losses of 5 percent per month are expected from November through March. Established stands of switchgrass that are harvested in July are expected to require about 90 kg/ha/yr of nitrogen to achieve the plateau yield, whereas fields harvested from October through March are expected to require approximately 67 kg/ha/yr (Haque 2010).

Conventional budgeting is necessary but insufficient to determine if the cost savings from extending harvest over nine months are sufficient to offset the losses in harvestable yield. A more comprehensive modeling approach is required to fully evaluate these tradeoffs. To address these tradeoffs, models may be constructed that encompass the entire chain of economic activities from acquisition of land use to delivery of baled switchgrass. Modeling may be conducted to determine if the additional fertilizer cost and the lower yield from an extended harvest window can be offset by the reduction in cost resulting from fewer harvest machines. The model optimally selects the number of harvest machines and the quantity of biomass to harvest by month and county during the potential July through March harvest window.

The 1992 Annual Progress Report of the DOE’s Biofuels Feedstock Development Program (Wright et al. 1993) supported the selection of switchgrass as a model bioenergy crop by stating, "the examination of data on yield potential, production economics, and regional site potential, led in 1991 to the selection of a perennial forage grass switchgrass as a model species for further research", recognizing that "more than one species will certainly be required ultimately, switchgrass was seen as an excellent beginning with the available programmatic resources". Several important characteristics such as being a widely adapted native species, a demonstrated capacity for high yields on relative poor quality sites, a significant capacity to improve soil quality by sequestering carbon, improved erosion control, reduced fertilizer and pesticide requirements, a capacity for providing wildlife cover, and a strong potential appeal to landowners supported this decision (Wright et al. 1993). Extensive research has continued to support the feasibility of switchgrass for bioenergy (Mitchell et al. 2012b).

Bioenergy efficiency and sustainability is held to a different standard than energy produced from petroleum since renewable fuels must have lower greenhouse gas (GHG) emissions and higher net energy values (NEV) than petroleum based transportation fuels (Mitchell et al. 2010a). The NEV, net energy yield (NEY), and the ratio of the biofuel output to petroleum input [petroleum energy ratio (PER)] have been used to quantify the energy efficiency and sustainability of ethanol produced from switchgrass (Schmer et al. 2008). Farrell et al. (2006) developed an energy model using estimated agricultural inputs and simulated yields and predicted switchgrass produced 700% more output than input energy. Schmer et al. (2008) validated the modeled results with actual inputs from switchgrass grown at the field scale on 10 farms in Nebraska, South Dakota, and North Dakota. They concluded that switchgrass produced 540% more renewable energy than non-renewable energy consumed over a 5-year period, had a PER of 13.1, and that average GHG emissions from switchgrass-based ethanol was 94% lower than estimated GHG emissions for gasoline (Schmer et al. 2008).

To sustain an agricultural production system, carbon inputs must equal or exceed the carbon outputs or soil organic carbon (SOC) will decline and overall system productivity will decline (Mitchell et al. 2010a). Historically, about half of the SOC present in pre-agricultural grasslands was presumed lost in the conversion of perennial grasslands to annual cropland that occurred after European settlement (Mitchell et al. 2010a). Consequently, SOC trend is an excellent indicator of the long-term sustainability of a production system. SOC increases rapidly when annual cropland is converted to switchgrass (Mitchell et al. 2010a). In just 5 years, growing and managing switchgrass for bioenergy on three marginally productive cropland sites in the Central Plains resulted in an average SOC increase of 2.9 Mg C ha1 yr-1 in the top 1.2 m of soil (Liebig et al. 2008). Growing switchgrass increased SOC at rates ranging from 1.7 to 10.1 Mg C ha1 yr-1 throughout North America (Garten and Wullschleger 2000, Zan et al. 2001, Frank et al. 2004; Lee et al. 2007). In irrigated switchgrass in the arid regions of the Pacific Northwest, 5-years of switchgrass cropping resulted in a 1.2 Mg ha-1 increase in SOC in the 0 to 15-cm depth, with no change below 15 cm (Collins et al. 2010).

Modeling efforts and numerous field studies have demonstrated that growing and managing switchgrass for bioenergy on sites formerly in row crop production rapidly and significantly increases SOC, improves soil quality, and promotes long-term sustainability (Liebig et al. 2005, 2008; Schmer et al. 2011; Follett et al. 2012). A limitation with many modeling efforts is that SOC accumulation is usually only predicted for sampling depths of 30 to 40 cm (Follett et al. 2012) and may be underestimating actual SOC accumulation. For example, a 9-year study on rainfed switchgrass and maize had average annual increases in SOC that exceeded 2 Mg C ha1 year-1 for the 0 to 150 soil depth and over 50% of the SOC increase occurred below 30-cm (Follett et al. 2012). The SOC for switchgrass was 2 to 4 times greater in this study than that modeled in life-cycle assessments to date. They concluded that sampling soil to only 30 to 40 cm is inadequate and

future analyses and modeling should include deep soil sampling to fully account for SOC accumulations in both switchgrass and maize (Follett et al. 2012). Chapter 12 addresses specific approaches to modeling switchgrass biomass production.

Switchgrass production for bioenergy is economically feasible (Perrin et al. 2008; Mitchell et al. 2012a, b). A large regional field scale trial was conducted in 50 production environments on 10 farms in Nebraska, South Dakota, and North Dakota (Perrin et al. 2008). Actual on-farm production costs were tracked for each farm, including land costs, which accounted for nearly half of the production costs. The cost of production for switchgrass to the farm gate averaged $66 Mg-1 (Perrin et al. 2012). Five farmers delivered switchgrass to the farm gate at an average cost of $52 Mg-1over the 5-year period. The 5-year average cost for farmers with experience growing switchgrass was $39 Mg1, and one producer grew switchgrass for $34 Mg1. Switchgrass farm-gate costs tend to decline over time with highest costs occurring on a per mass basis during the establishment year; a result of high input costs and low biomass yields (Perrin et al. 2008). When the authors projected field production for 10 years, farm-gate delivery costs were reduced to $46 Mg-1. They concluded that, with experience, farmers could achieve switchgrass production costs of $40 to $55 Mg1. Assuming a conversion rate of 0.329 liters of ethanol per kg of switchgrass, the farm-gate feedstock cost would range from $0.12 to $0.16 L1 (Perrin et al.

2008) . Land and other production costs have increased since the regional field scale study was completed. Perrin et al. (2012) estimated an updated switchgrass farm-gate price of $75 Mg1 and $60 Mg1 for biomass yields of 6.7 Mg ha1 and 13.5 Mg ha1, respectively. Farm-gate costs in growing switchgrass for bioenergy are largely variable with respect to yield, with approximately 25% of total costs being fixed (Perrin et al. 2012). Farm-gate prices are also dependent on land type being converted, regional variations in land costs, yield potential, and rotation time length. An estimated 5.1 x 106 ha to 11.8 x 106 ha could be allocated to switchgrass production in the United States by 2030 assuming a farm gate price of $44 Mg1 to $66 Mg1 (USDOE 2011). Future improvements in large-scale harvest machinery and implementation of farm telematics will likely reduce variable switchgrass harvest and delivery costs. See Chapter 13 for a more detailed discussion of the economics of switchgrass feedstock production.

Conclusions

Switchgrass is the most advanced herbaceous perennial feedstock for bioenergy. Switchgrass research has been conducted for more than 75 years, with a focus on bioenergy for more than 20 years. Mitchell et al.

(2012) reported on the feasibility of growing switchgrass for bioenergy. They reported that all practices for growing switchgrass for biofuels including establishing, managing, and delivering to the biorefinery gate have been developed, with specific management requirements for most US agroecoregions (Mitchell et al. 2012). They concluded that the research to date fully supports that switchgrass for bioenergy is productive, protective of the environment, and profitable for the farmer. Additionally, switchgrass has been seeded on millions of hectares of CRP grasslands since 1986, so it is familiar to many producers. Further research on the processes of converting switchgrass to transportation fuels at the commercial scale is needed. Additionally, field-validating some of the models for deploying switchgrass at the landscape scale are needed to demonstrate the feasibility and environmental benefits, especially for wildlife, of large-scale feedstock production. Switchgrass has high biomass production potential, wide adaptability, low fossil fuel energy requirements, and is compatible with modern agriculture practices making it an ideal herbaceous energy crop for large-scale bioenergy production. Significant research has been conducted on switchgrass genetics, agronomic management, and harvest practices which will be invaluable for an emerging cellulosic bioenergy industry.

The term ‘endophyte’ is derived from the Greek term ‘endo’ (within) and ‘phyte’ (plant), and may apply to both fungi and bacteria that reside in plant tissues during all or part of their life cycle and cause no apparent harm (Wilson 1995). It is estimated that every plant species has at least one associated bacterial endophyte (Strobel et al. 2004), and they belong to diverse classes of bacteria including alpha, beta, and gamma subdivisions of Proteobacteria, Firmicutes, Bacteroidetes, and Actinobacteria (Rosenblueth and Martinez-Romero 2006). These bacteria thrive within plants where they successfully colonize roots, translocate to leaves, stems, and even to reproductive organs where they may be vertically transmitted to the next generation, ensuring a stable interaction with its host plant.

The number of microorganisms present in natural ecosystems is tremendous. In fact, estimates of the number of bacterial endophytes in the Brazilian Atlantic forest indicate the possibility of 2-13 million species in the aboveground plant parts alone (Lambais et al. 2006). Of the bacterial species identified, 97% were previously not described. A single plant species may also have a wide range of different bacterial genera associated. In wheat, culture based studies have shown that 88 bacterial species representing 37 genera inhabit the aboveground plant tissue (Legard et al. 1994), which likely underestimate the number of microorganisms as molecular studies yield much larger population numbers (Rasche et al. 2006). Both culture based and molecular based analyses indicate that alpha and beta Proteobacteria are the most numerous colonizers of the phyllosphere (Thompson et al. 1993). In total, 853 bacterial endophytes were isolated from aboveground parts of four agronomic crops and 27 prairie plants including switchgrass. Cellulomonas, Clavibacter, Curtobacterium, and Microbacterium isolates showed high levels of colonization and had the ability to persist in host plants (Zinniel et al. 2002).

Diazotrophs, or atmospheric nitrogen-fixing bacteria have been isolated from bioenergy crops, including Miscanthus spp. and Pennisetum purpureum, where Herbaspirillum frisingense sp. nov. (Kirchhof et al. 2001), Azospirillum doebereinerae (Eckert et al. 2001), and Herbaspirillum frisingense (Rothballer et al. 2008) were found. Similarly, different nitrogen-fixing bacteria belonging to genera Stenotrophomona, Pseudomonas and Burkholderia were isolated from sand dune grasses (Ammophila arenaria and Elymus mollis) in Oregon, which may biologically fix nitrogen and promote the growth of these plants under poor soil conditions (Dalton et al. 2004). Nitrogen-fixing bacteria have also been isolated from different plant species, such as Kallar grass (Leptochoa fusa) (Reinhold-Hurek et al. 1993), lodgepole pine (Pinus contorta), western red cedar (Thuja plicata) (Bal et al. 2012), and hybrid poplar (Populous trichocarpa) (Taghavi et al. 2010). While general surveys of endophytic populations in switchgrass have been undertaken (Zinniel et al. 2002), there are no detailed analysis on native bacterial endophytic interactions in switchgrass.

Fungal endophytic populations may also be substantial, particularly in longer lived plants, as 340 genetically distinct taxa were recovered from two tropical understory plant species (Arnold et al. 2000). Endophytic fungi can also have a significant beneficial impact on switchgrass performance (Kleczewski et al. 2012). While much emphasis has been placed on the study of clavicipitaceous fungal endophytes (Neotyphodium/Epichloe) with cool — and warm-season grasses (Rodriguez et al. 2009), two recent surveys of switchgrass endophytes have failed to identify members of the Clavicipitaceae family (Ghimire et al. 2011; Kleczewski et al. 2012), suggesting that the major endophytic fungi inhabiting switchgrass are of the non-clavicipitaceous type, representing primarily ascomycetous fungi (Kleczewski et al. 2012). These endophytes may be found colonizing tissues above — and/or below-ground (Rodriguez et al. 2009). Recently, 18 taxonomic orders of fungal endophytes were isolated from switchgrass plants in northern Oklahoma belonging to the genera Alternaria, Codinaeopsis, Fusarium, Gibberella, Hypoerea and Periconia, and switchgrass shoot tissues showed a significantly higher diversity of fungal endophytic species compared to the root tissues (Ghimire et al. 2011). Similar fungal endophytic genera were isolated from switchgrass plants growing in a range of habitats across Indiana and Illinois, such as Alternaria, Epicoccum, Phoma, Phaeosphaeria and Stagonospora (Kleczewski et al. 2012). Since switchgrass is one of the most promising bioenergy crops, several laboratories in the US have been working on isolation and characterization of bacterial and fungal endophytes from switchgrass. Identifying and harnessing beneficial endophytic microorganisms that have a broad spectrum of plant growth promotion traits and possess various mechanisms for stress tolerance may aid in the development of a low input and sustainable switchgrass feedstock production system, particularly on marginal land.

Mycorrhizae are symbiotic fungi that interact with the roots of vascular plants. These fungi are typically divided into two groups: ectomycorrhizas which have hyphae that do not penetrate individual cells within the root and endomycorrhizas which, as the name implies, have hyphae that penetrate the cell wall and invaginate the cell membrane. Eighty to 92% of land plant species surveyed are associated with mycorrhizal fungi, among them, arbuscular mycorrhizal (AM) fungi are the predominant type (Wang and Qiu 2006), and are placed in the phylum (division) Glomeromycota. AM fungus is characterized by highly branched fungal structures located within the plant root cortical cells. Generally, AM fungi comprise 130 species of fungi classified as Zygomycotina (Simon et al. 1993). AM fungi from the order Glomales (Glomeromycota) are associated with most plant species including angiosperms, gymnosperms, pteridophytes, lycopods, and mosses (reviewed in Hause and Fester 2005). The fungi involved in the AM interaction are obligate biotrophs and reproduce asexually. As obligate biotrophs, AM fungi are not culturable without their host plant, making the study of these organisms difficult. AM fungal associations are important to help switchgrass tolerate unfavorable soil conditions (Parrish and Fike 2005). It has been reported that AM fungi play essential roles in switchgrass growth in acidic soil, which has high levels of exchangeable aluminum and immobile minerals, such as phosphorus (Koslowsky and Boerner 1989; Brejda et al. 1993; Johnson 1998). AM fungal associations may be more critical in warm-season grasses, such as switchgrass because from an evolutionary perspective, both are of tropical origin (Hetrick et al. 1988).