MISCELLANEOUS Genotypic Differences in Root Hair Deformation and Subsequent Nodulation for Red Clover under Different Additions of Starter N Fertilization R. M. M. S. Thilakarathna 1 , Y. A. Papadopoulos 2 , S. A. E. Fillmore 3 & B. Prithiviraj 4 1 Department of Biology, Dalhousie University, Halifax, NS, Canada 2 Agriculture & Agri-Food Canada, Nova Scotia Agricultural College, Truro, NS, Canada 3 Agriculture & Agri-Food Canada, Kentville, NS, Canada 4 Department of Environmental Sciences, Nova Scotia Agricultural College, Truro, NS, Canada Introduction An essential element of sustainable agriculture is the effective management of N fertilizer. Owing to new trends in low-input agriculture and concern over the negative effects of high N inputs, many farmers consider using pasture legumes as a source of biological N fixation (BNF). Perennial forage legumes have great potential to enhance the sustainability of grassland farming systems (Carlsson and Huss-Danell 2003, Paynel et al. 2008). Red clover (Trifolium pratense L.) is an important legume in northern latitudes where much of agriculture is based on livestock production. It is an excellent forage legume for hay-making, silage and grazing, and it can be used in rotation to maintain soil fertility. Red clover is one of the few agriculturally important species in the genus Trifolium (Taylor and Quesenberry 1996) wherein symbi- otic BNF plays a crucial role. Symbiotic N fixation involving legumes and rhizobia depends upon a complex series of processes: successful colonization of effective rhizobium strains in the soil, infection of the host plant and nodule initiation and development (Fisher and Long 1992, Ledgard and Steele 1992). Nodulation in legumes is activated in response to rhizobial signalling molecules, called nodulation (Nod) factors, which induce root hair deformation (curling) (Esseling et al. 2003). Bacterial infection generally occurs through root hair cells curling around the rhizobia, entrapping attached bacteria. These bacteria grow and form infection foci from which infection threads are initi- ated. Infection threads have a plant origin structure and can permeate cell boundaries, which enables bacteria to Keywords nodulation; red clover cultivars; root hair deformation; starter nitrogen Correspondence Y. A. Papadopoulos Agriculture and Agri-Food Canada, Nova Scotia Agricultural College, Truro, NS, Canada Tel.: +1 902 896 2452 Fax: +1 902 895 6734 Email: yousef.papadopoulos@agr.gc.ca Accepted January 25, 2012 doi:10.1111/j.1439-037X.2012.00505.x Abstract Red clover cultivars, including diploid and tetraploid, are commonly used in legume-based pasture mixtures. However, information on nodulation under different starter N regimens is limited. We hypothesized that there is genetic variability among different red clover cultivars for nodulation. A root hair deformation assay was conducted using three diploid (AC Christie, Tapani and CRS15) and three tetraploid (Tempus, CRS18 and CRS39) red clover cultivars by inoculating them with Rhizobium leguminosarum biovar trifolii. Nodulation and morphological characteristics of two selected red clover cultivars, AC Christie and Tempus, were determined under five starter N concentrations (0, 0.2, 0.4, 0.8 and 1.6 mg per plant). Inoculation with rhizobia increased root hair deformation with significant interaction across cultivars. Nodulation was delayed under high starter N concentrations, and genotypic differences were evident for days-to-nodule initiation. There was a positive quadratic response to starter N for AC Christie and a negative quadratic response for Tempus for nodulation. Tempus had more active nodules (92 %) than AC Christie (73 %). The genetic variability of red clover cultivars should be considered in N fixa- tion studies and their response to availability of initial N. J. Agronomy & Crop Science (2012) ISSN 0931-2250 ª 2012 Blackwell Verlag GmbH 1