Responses of the Physiological and Biochemical Characteristics of Alfalfa Roots with Different Root Collar Diameters to Low Temperatures Under Simulated Winter Irrigation

Background: Due to the frequent occurrence of extremely low temperatures in northern China, the problem of alfalfa mortality during overwintering requires urgent solutions to support the development of large-scale alfalfa production in the region. Understanding the effects of winter irrigation and the alfalfa root collar diameter on physiological and biochemical cold resistance mechanisms in alfalfa will be of great significance to finding such solutions.Methods: In this study, alfalfa roots with different root collar diameters (3.50±0.10 mm, 5.25±0.10 mm, 7.00±0.10 mm) were subjected to simulated winter irrigation (0 ml, 20 ml, 40 ml) and different low-temperature stress treatments (4°C, -10°C, -15°C, -20°C) in the laboratory. The relationships between alfalfa root collar activity, relative conductivity and the nonstructural carbon and nitrogen contents and antioxidant enzyme activity in the collar were explored.Results: Low-temperature stress was the most important factor causing significant changes in the physiological and biochemical indexes of the alfalfa root collar, followed by the root collar diameter. Low-temperature stress and the root collar diameter had significant effects on nonstructural carbon and nitrogen levels and enzyme activity (except POD activity) in alfalfa root collars, and winter irrigation had extremely significant effects on the nonstructural nitrogen contents of alfalfa root collars. The larger the diameter of the root collar was, the higher the soluble protein content and the stronger the SOD activity. Simulated winter irrigation increased the C/N ratio in the alfalfa root collars. The increases in nonstructural carbon content, C/N ratio and SOD activity in alfalfa root collars helped improve alfalfa cold resistance.Conclusions: Alfalfa resists low-temperature stress by adjusting the proportions of nonstructural carbon and nitrogen as well as the SOD activity in the root collar. Therefore, fertilization, early sowing and other management measures should be adopted to ensure healthy alfalfa growth and thereby increase the root collar diameter of alfalfa grown in cold regions in northern China. Moreover, timely supplementary irrigation to reach soil saturation in winter can play an important role in improving cold resistance in alfalfa.

Identification and assessment of a biocontrol agent, Ochrobactrum intermedium I-5, for management of alfalfa root rot caused by Fusarium tricinctum

Alfalfa root rot caused by Fusarium tricinctum is one of the most important soil-borne diseases resulting in significant losses to alfalfa agriculture worldwide. Fungicides used in management of disease affect the environment and human health. In this study, a strain of Ochrobactrum intermedium (I-5), isolated from alfalfa rhizosphere soil, exhibited strong antifungal activity against a number of causative pathogens of alfalfa root rot, and showed the strongest antagonistic activity against F. tricinctum (the longest radius/shortest radius ratio of 3.09). When applied at 10%, a filtrate of I-5 liquid culture significantly reduced the spore production and germination and mycelial growth of F. tricinctum, and the inhibition rate was 76.67%, 78.93% and 55.77%, respectively. Furthermore, a filtrate and suspension of the strain, when applied at 10%, reduced alfalfa root rot by more than 73%. The strain clearly promoted the activities of invertase, urease, cellulose, and neutral phosphatase in alfalfa rhizosphere soil and significantly reduced the damage to rhizosphere soil quality attributable to alfalfa root rot. Moreover, the strain clearly promoted the growth of alfalfa, without causing any evident damage to plants. The active substance produced by the strain was relatively insensitive to heat and ultraviolet irradiation and displayed optimal efficacy at pH 8. To the best of our knowledge, this is the first study describing the use of O. intermedium for the biological control of alfalfa root rot. O. intermedium (I-5) has considerable potential for application in the control of alfalfa root rot and improvement of the quality of cultivated alfalfa.

Exploration of Endophytes from Alfalfa (Medicago sativa L.) as Biocontrol Agents

Endophytes are increasingly investigated as biocontrol agents for agricultural production. The identification of new endophytes with high effectiveness against plant disease is very important. A total of 362 strains of endophytes, including fungi, bacteria, and actinomycete, were isolated from alfalfa (Medicago sativa L.) collected in Hebei, Inner Mongolia and Ningxia provinces of China. The three strains of endophytic bacteria (NA NX51R-5, NA NX90R-8, and NA NM1S-1) with strong biocontrol capability with >50% effectiveness were screened against the common alfalfa root rot pathogen Fusarium oxysporum F. sp. medicaginis in alfalfa seedling germination experiments on MS medium and pot experiments. Using phylogenetic analysis, the isolates of NA NM1S-1 and NA NX51R-5 were identified as Bacillus spp. by 16S rDNA, while NA NX90R-8 was found to be Pseudomonas sp.

Digital Imaging to Evaluate Root System Architectural Changes Associated with Soil Biotic Factors

Root system architecture is critical for plant growth, which is influenced by several edaphic, environmental, genetic, and biotic factors including beneficial and pathogenic microbes. Studying root system architecture and the dynamic changes that occur during a plant’s lifespan, especially for perennial crops growing over multiple growing seasons, is still a challenge because of the nature of their growing environment. We describe the utility of an imaging platform called RhizoVision Crown to study root system architecture of alfalfa, a perennial forage crop threatened by Phymatotrichopsis root rot (PRR) disease. Phymatotrichopsis omnivora is the causal agent of PRR disease that reduces alfalfa stand longevity. During the lifetime of the stand, PRR disease rings enlarge and the field can be categorized into three zones based upon plant status: asymptomatic, disease front and survivor. To study root system architectural changes associated with PRR, a 4-year-old 25.6-ha alfalfa stand infested with PRR was selected at the Red River Farm, Burneyville, OK during October 2017. Line transect sampling was conducted from four actively growing PRR disease rings. At each disease ring, six line transects were positioned spanning 15 m on either side of the disease front with one alfalfa root crown sampled at every 3 m interval. Each alfalfa root crown was imaged with the RhizoVision Crown platform using a backlight and a high-resolution monochrome CMOS camera enabling preservation of the natural root system integrity. The platform’s image analysis software, RhizoVision Analyzer, automatically segmented images, skeletonized, and extracted a suite of features. Data indicated that the survivor plants compensated for damage or loss to the taproot through the development of more lateral and crown roots, and that a suite of multivariate features could be used to automatically classify roots as from survivor or asymptomatic zones. Root growth is a dynamic process adapting to ever changing interactions among various phytobiome components. By utilizing the low-cost, efficient, and high-throughput RhizoVision Crown platform, we quantified these changes in a mature perennial forage crop.

Digital imaging to evaluate root system architectural changes associated with soil biotic factors

Root system architecture (RSA) is critical for plant growth, which is influenced by several edaphic, environmental, genetic and biotic factors including beneficial and pathogenic microbes. Studying root architecture and the dynamic changes that occur during a plant's lifespan, especially for perennial crops growing over multiple growing seasons, is still a challenge because of the nature of their growing environment in soil. We describe the utility of an imaging platform called RhizoVision Crown to study RSA of alfalfa, a perennial forage crop affected by Phymatotrichopsis Root Rot (PRR) disease. Phymatotrichopsis omnivora is the causal agent of PRR disease that reduces alfalfa stand longevity. During the lifetime of the stand, PRR disease rings enlarge and the field can be categorized into three zones based upon plant status: asymptomatic, disease front and survivor. To study root architectural changes associated with PRR, a four-year old 25.6-hectare alfalfa stand infested with PRR was selected at the Red River Farm, Burneyville, OK during October 2017. Line transect sampling was conducted from four actively growing PRR disease rings. At each disease ring, six line transects were positioned spanning 15 m on either side of the disease front with one alfalfa root sampled at every 3 m interval. Each alfalfa root was imaged with the RhizoVision Crown platform using a backlight and a high-resolution monochrome CMOS camera enabling preservation of the natural root architectural integrity. The platform's image analysis software, RhizoVision Analyzer, automatically segmented images, skeletonized, and extracted a suite of features. Data indicated that the survivor plants compensated for damage or loss to the taproot through the development of more lateral and crown roots, and that a suite of multivariate features could be used to automatically classify roots as from survivor or asymptomatic zones. Root growth is a dynamic process adapting to ever changing interactions among various phytobiome components, by utilizing a low-cost, efficient and high-throughput Rhizo-Vision Crown platform we showed quantification of these changes occurring in a mature perennial forage crop.