Robert T. Lartey

Dryland residue and soil organic matter as influenced by tillage, crop rotation, and cultural practice

Novel management practices are needed to increase dryland soil organic matter and crop yields that have been declining due to long-term conventional tillage with spring wheat (Triticum aestivum L.)-fallow system in the northern Great Plains, USA. The effects of tillage, crop rotation, and cultural practice were evaluated on dryland crop biomass (stems + leaves) yield, surface residue, and soil organic C (SOC) and total N (STN) at the 0–20xa0cm depth in a Williams loam (fine-loamy, mixed, superactive, frigid, Typic Argiustolls) from 2004 to 2007 in eastern Montana, USA. Treatments were two tillage practices [no-tillage (NT) and conventional tillage (CT)], four crop rotations [continuous spring wheat (CW), spring wheat-pea (Pisum sativum L.) (W-P), spring wheat-barley (Hordeum vulgaris L.) hay-pea (W-B-P), and spring wheat-barley hay-corn (Zea mays L.)-pea (W-B-C-P)], and two cultural practices [regular (conventional seed rates and plant spacing, conventional planting date, broadcast N fertilization, and reduced stubble height) and ecological (variable seed rates and plant spacing, delayed planting, banded N fertilization, and increased stubble height)]. Crop biomass and N content were 4 to 44% greater in W-B-C-P than in CW in 2004 and 2005 and greater in ecological than in regular cultural practice in CT. Soil surface residue amount and C and N contents were greater in NT than in CT, greater in CW, W-P, and W-B-C-P than in W-B-P, and greater in 2006 and 2007 than in 2004 and 2005. The SOC and STN concentrations at 0–5xa0cm were 4 to 6% greater in CW than in W-P or W-B-P in NT and CT from 2005 and 2007. In 2007, SOC content at 10–20xa0cm was greater in W-P and W-B-P than in W-B-C-P in CT but STN was greater in W-B-P and W-B-C-P than in CW in NT. From 2004 to 2007, SOC and STN concentrations varied at 0–5xa0cm but increased at 5–20xa0cm. Diversified crop rotation and delayed planting with higher seed rates and banded N fertilization increased the amount of crop biomass returned to the soil and surface residue C and N. Although no-tillage increased surface residue C and N, continuous nonlegume cropping increased soil C and N levels at the surface layer compared with other crop rotations. Continued return of crop residue from 2004 to 2007 may increase soil C and N levels but long-term studies are needed to better evaluate the effect of management practices on soil C and N levels under dryland cropping systems in the northern Great Plains.

Effects of Agaricus lilaceps Fairy Rings on Soil Aggregation and Microbial Community Structure in Relation to Growth Stimulation of Western Wheatgrass (Pascopyrum smithii) in Eastern Montana Rangeland

Stimulation of plant productivity caused by Agaricus fairy rings has been reported, but little is known about the effects of these fungi on soil aggregation and the microbial community structure, particularly the communities that can bind soil particles. We studied three concentric zones of Agaricus lilaceps fairy rings in Eastern Montana that stimulate western wheatgrass (Pascopyrum smithii): outside the ring (OUT), inside the ring (IN), and stimulated zone adjacent to the fungal fruiting bodies (SZ) to determine (1) soil aggregate proportion and stability, (2) the microbial community composition and the N-acetyl-β-d-glucosaminidase activity associated with bulk soil at 0–15xa0cm depth, (3) the predominant culturable bacterial communities that can bind to soil adhering to wheatgrass roots, and (4) the stimulation of wheatgrass production. In bulk soil, macroaggregates (4.75–2.00 and 2.00–0.25xa0mm) and aggregate stability increased in SZ compared to IN and OUT. The high ratio of fungal to bacteria (fatty acid methyl ester) and N-acetyl-β-d-glucosaminidase activity in SZ compared to IN and OUT suggest high fungal biomass. A soil sedimentation assay performed on the predominant isolates from root-adhering soil indicated more soil-binding bacteria in SZ than IN and OUT; Pseudomonas fluorescens and Stenotrophomonas maltophilia isolates predominated in SZ, whereas Bacillus spp. isolates predominated in IN and OUT. This study suggests that growth stimulation of wheatgrass in A. lilaceps fairy rings may be attributed to the activity of the fungus by enhancing soil aggregation of bulk soil at 0–15xa0cm depth and influencing the amount and functionality of specific predominant microbial communities in the wheatgrass root-adhering soil.

Dryland soil nitrogen cycling influenced by tillage, crop rotation, and cultural practice

Management practices may influence dryland soil N cycling. We evaluated the effects of tillage, crop rotation, and cultural practice on dryland crop biomass (stems and leaves) N, surface residue N, and soil N fractions at the 0–20xa0cm depth in a Williams loam from 2004 to 2008 in eastern Montana, USA. Treatments were two tillage practices (no-tillage [NT] and conventional tillage [CT]), two crop rotations (continuous spring wheat [Triticum aestivum L.] [CW] and spring wheat-barley [Hordeum vulgaris L.] hay-corn [Zea mays L.]-pea [Pisum sativum L.] [W-B-C-P]), and two cultural practices (regular [conventional seed rates and plant spacing, conventional planting date, broadcast N fertilization, and reduced stubble height] and ecological [variable seed rates and plant spacing, delayed planting, banded N fertilization, and increased stubble height]). Nitrogen fractions were soil total N (STN), particulate organic N (PON), microbial biomass N (MBN), potential N mineralization (PNM), NH4–N, and NO3–N. Crop biomass N was 30xa0% greater in W-B-C-P than in CW in 2005. Surface residue N was 30–34xa0% greater in NT with the regular and ecological practices than in CT with the regular practice. The STN, PON, and MBN at 10–20 and 0–20xa0cm were 5–41xa0% greater in NT or CW with the regular practice than in CT or CW with the ecological practice. The PNM at 5–10xa0cm was 22xa0% greater in the regular than in the ecological practice. The NH4–N and NO3–N contents at 10–20 and 0–20xa0cm were greater in CT with W-B-C-P and the regular practice than with most other treatments in 2007. Surface residue and soil N fractions, except PNM and NO3–N, declined from autumn 2007 to spring 2008. In 2008, NT with W-B-C-P and the regular practice gained 400xa0kgxa0Nxa0ha−1 compared with a loss of 221xa0kgxa0Nxa0ha−1 to a gain of 219xa0kgxa0Nxa0ha−1 in other treatments. No-tillage with the regular cultural practice increased surface residue and soil N storage but conventional tillage with diversified crop rotation and the regular practice increased soil N availability. Because of continuous N mineralization, surface residue and soil N storage decreased without influencing N availability from autumn to the following spring.

Direct Polymerase Chain Reaction-Based Detection of Cercospora beticola in Field Soils

Cercospora beticola, the causal agent of Cercospora leaf spot of sugar beet, survives as pseudostromata in infected sugar beet residues in the soil. Under optimal conditions, overwintering propagules germinate and produce conidia that are dispersed as primary inoculum to initiate infection in sugar beet. We developed a polymerase chain reaction (PCR) technique for rapid detection of C. beticola in field soils. Total DNA was first isolated from soil amended with C. beticola culture using the PowerSoil DNA Kit. The purified DNA was subjected to PCR in Extract-N-Amp PCR mix with CBACTIN primers over 35 cycles. The amplified products were resolved and compared by electrophoresis in 1% agarose gels. The PCR fragment size of C. beticola from the amended field soil correlated in size with the amplicon from control C. beticola culture DNA extract. Additionally, sample soils were collected from sugar beet fields near Sidney, MT and Foxholm, ND. Total DNA was extracted from the samples and subjected to PCR and resolved as previously described. The amplicons were purified from the gels and subjected to BigDye Terminator Cycle sequencing. All sequences from field soils samples, C. beticola-amended field soil, and pure culture were compared by alignment with a C. beticola actin gene sequence from GenBank. The result of the alignment confirmed the amplicons as products from C. beticola. Rapid screening for the presence of C. beticola in the soil by PCR will improve research capabilities in biological control, disease forecasting, and management of this very important sugar beet pathogen.

First Report of Rhizoctonia spp. Causing a Root Rot of the Invasive Rangeland Weed Lepidium draba in North America

The exotic, invasive perennial rangeland weed Lepidium draba spreads rapidly and reduces native species diversity. The extensive root system of L. draba constitutes 76% of its biomass (4). Thus, searches have been done for biocontrol agents that target root tissue or that may interact with a weevil, Ceutorhynchus assimilis, that causes galls in the crown area of L. draba. An association of Rhizoctonia spp. with root tissue of plants galled by the weevil has been documented in Europe (3). The possible presence of soilborne pathogens similar to those found in the native range has been the subject of L. draba surveys in the United States. One such survey in 2008 detected a few plants with reddened and chlorotic foliage in a stand near Shepherd, MT. Such symptoms typically indicate the occurrence of soilborne diseases on L. draba in the native range of the weed (2). The site had shown a gradual increase in the range of detectable pathogens beginning with foliar pathogens in 1997. In 2010, at the Shepherd site, L. draba plants with similar (but more severe) symptoms to those seen in 2008 were noted in a different area of the stand. Excavation of the roots in both years revealed brown, sunken crown and root cankers. Pieces of root tissue were excised from the lesions and plated on acidified PDA and Ko and Hora medium. A non-sporulating fungus was isolated from three plants. Colonies of the isolates on PDA were typical of known Rhizoctonia spp. The 2010 isolates were determined to be multinucleate using DAPI and were paired with 14 tester (including subgroups) isolates of AG-1 to AG-4 on water agar. Anastomosis was observed between the multinucleate isolates and the AG-2-1 tester isolate. Sequence analysis of ITS of the rDNA of a multinucleate isolate (GenBank KJ545577) indicated 99% similarity with an accession of R. solani AG 2-1 (AB547381). The 2008 isolates were binucleate. A binucleate isolate, KJ545578, had 100% similarity with an isolate of Rhizoctonia spp. AG-A (AY927356). Pathogenicity tests consisted of planting 6-week-old seedlings of L. draba, one per pot, in ten 85-cm-diameter pots of pasteurized soil mix infested with Rhizoctonia-colonized barley grain that had been dried and milled. An inoculum level of ~8 CFU/g (1) of air-dried soil was established by most probable number calculations from fourfold dilutions of infested soil. Controls were the same number of plants in pasteurized potting mix. Results were recorded after 3 months in a greenhouse at 20-25°C. The test was repeated. Typically, R. solani caused mortality of six to eight plants, from which it was re-isolated, whereas binuclate isolates caused stunting and lower dry weight of L. draba. Control plants remained asymptomatic. This is the first report of R. solani and binucleate Rhizoctonia spp. on L. draba in North America. References: (1) A. J. Caesar et al. Plant Dis. 93:1350, 2009. (2) A. J. Caesar et al. Biol. Control 52:140, 2010. (3) A. J. Caesar et al. Plant Dis. 96:145, 2011. (4) R. F. Miller et al. Agronomy J. 86:487, 1994.

First Report of Spot Form Net Blotch Caused by Pyrenophora teres f. maculata on Barley in the Mon-Dak Area of the United States

Pyrenophora teres Drechs. causes net blotch of barley, a common foliar disease in cultivation zones around the world. The disease occurs in two forms, namely a net form net blotch (NFNB) caused by P. teres f. teres and a spot form net blotch (SFNB) caused by P. teres f. maculata. As in other parts of the northern Great Plains, in the Mon-Dak area (western North Dakota and eastern Montana), NFNB is prevalent. SFNB was first reported in western Montana in 1983 (1) and more recently in eastern North Dakota in 2010 (3) but not in the Mon-Dak area. In the summer of 2011, unusual spot lesions that were surrounded by necrosis or chlorosis were observed on different barley cultivars in fields at Williston, ND, Nesson Valley, ND, and Sidney, MT areas. Diseased leaves from various barley cvs. from the three locations were transferred to water agar and incubated at room temperature for 24 h to induce sporulation. Morphological examination of conidia (45 to 169 × 15 to 21 μm) did not show significant differences from a known isolate of P. teres f. teres 0-1 (provided by Tim Friesen, ARS, Fargo, ND). For pathogenicity testing, six 14-day-old plants of barley cv. Tradition were sprayed until runoff with a 2,000 spore/ml suspension of two isolates from each location and the control P. teres f. maculata isolate DEN2.6 (provided by Tim Friesen). Plants were incubated first in a lighted humidity chamber for 24 h and then in a greenhouse for 7 days at 21°C. Regardless of inoculum source, spot lesions surrounded by necrosis or chlorosis, typical of SFNB, appeared on the inoculated leaves within 7 days. Fungi isolated from symptomatic leaves were identified as P. teres and the morphology of the conidia was undistinguishable from those of P. teres f. teres. All control plants which were sprayed with sterile distilled water were symptomless. The pathogenicity test was repeated. Rapid PCR detection and amplicon sequencing (2) of the internal transcribed spacer (ITS) region of ribosomal genes was performed on field and pathogenicity test leaf lesion samples to confirm the presence of P. teres f. maculata. DNA templates were prepared using the Extract-N-Amp Plant PCR Kits (Sigma Chemical Co., St. Louis, MO) and subjected to PCR using ITS1 and ITS4 primers. Amplicons were then purified and sequenced. The 585-bp nucleotide sequences of P. teres f. maculata from Mon-Dak area were submitted to GenBank under accession nos. PtmNES1 (JX187587), PtmSDY1 (JX187588), PtmSDY2 (JX187589), and PtmWIL1 (JX187590). The sequences from the four locations shared 100% similarity and also with P. teres f. maculata (EF452471) from GenBank while showing 10 nucleotide differences (99% similarity) with P. teres f. teres (EF452472).The results represent first report of SFNB in the Mon-Dak. Barley is one of the most important crops in the area. Resistance of the NFNB and SFNB of barley are controlled by different genes (4). Based on this report, SFNB therefore have to be considered in selection of barley cultivars for cultivation in the area. References: (1) H. E Bockelman et al. Plant Dis. 67:696, 1983. (2) R. T. Lartey et al. J. Sugar Beet Res. 40:1, 2003. (3) Z. H. Liu and T. L. Friesen. Plant Dis. 94:480, 2010. (4) O. M. Manninen et al. Genome. 46:1564, 2006.

First Report of a Root and Crown Disease of the Invasive Weed Lepidium draba Caused by Phoma macrostoma

The exotic rangeland perennial Lepidium draba occurs as a noxious weed in 22 states, mostly in the western United States. Because chemical control measures against this invasive perennial, a member of the Brassicaceae, have not achieved adequate results, biological control is being pursued. While inventories of arthropods that feed on L. draba have been established, little is known of soilborne pathogens for possible use as biological control agents. To address this deficiency, we have surveyed for diseases of L. draba in the United States and Eurasia to identify and test potential biocontrol agents. In intensive surveys for soilborne diseases in a single infestation that is >20 years old in a cattle pasture in south-central Montana, several chlorotic, stunted plants were noted. Roots of chlorotic plants that exhibited elongated fissures from which other soilborne fungi were isolated also had numerous prominent pycnidia embedded in the crown tissue above the lesions. Examination with a dissecting microscope revealed large ostioles made evident by the wide concave inversions in the short necks of the pycnidia. Culture of root tissue on potato dextrose agar resulted in whitish, becoming pale gray colonies, with a dull peach-to-reddish tinge at the margins, with abundant single pycnidia. Conidia in vitro were mainly unicellular, variable shape, subglobose to ellipsoidal, with several guttules averaging 6 × 2.5 μm. These morphological traits are characteristic of Phoma macrostoma, which is regarded as a weak or wound pathogen. The internal transcribed spacer region of rDNA was amplified using primers ITS1 and ITS4 and sequenced. BLAST analysis of the 575-bp fragment showed a 100% homology with the sequence of an isolate of P. macrostoma that has been investigated extensively for commercialization as a biological control agent of various agricultural weeds (1), including wild mustard (GenBank No. DQ474091). The nucleotide sequence has been assigned GenBank No. HM755951. Pathogenicity tests consisted of making four 1.4-mm-diameter holes in five NaOCl (0.1%)-sterilized root sections of L. draba and pipetting ~50 to 100 μl of a 106 CFU/ml conidial suspension into the incisions, incubating the inoculated roots at 20 to 25°C overnight and planting the root sections, one per pot, in an artificial greenhouse potting mix and placing the pots in the greenhouse at 20 to 25°C. Controls were five root sections that were treated similarly except that sterile water was injected. The experiment was repeated. After 10 days, shoots that grew from inoculated roots were chlorotic and shorter than those produced from control roots. P. macrostoma was isolated from tissue of inoculated roots that became blackened distal to the inoculation points. To examine the host range of P. macrostoma on other brassica species, crowns of 2-week-old seedlings of radish, broccoli, cauliflower, broccoli raab, turnip, kohlrabi, cabbage, Chinese cabbage, mustard greens, and canola were injected with 0.5 ml of a 106 CFU/ml conidial suspension. Plants were grown in the greenhouse at 20 to 25°C for 4 weeks after inoculation and examined for symptoms. The experiment was repeated twice. Blackened root tissue with slight chlorosis occurred only on roots of radish and crowns of broccoli, from which P. macrostoma was reisolated. To our knowledge, this the first report of a disease of L. draba caused by P. macrostoma. Reference: (1) K. L. Bailey et al. U.S. Patent Application Serial No. 60/294,475, Filed May 20, 2001.

Effect of lead (Pb) on the systemic movement of RNA viruses in tobacco (Nicotiana tabacum var. Turkish)

Effect of various lead (Pb) concentrations on the systemic movement of RNA viruses was examined in tobacco plants. Prior to inoculation, plants were grown hydroponically for 6xa0days in Hoagland’s solution supplemented with five concentrations of lead nitrate [Pb(NO3)2]: 0.0 (control), 10, 15, 50, and 100xa0μM. Four different RNA viruses with different cell-to-cell movement mechanisms were used. Two weeks after inoculation lower and upper leaves of each treatment were harvested and examined for the presence of viral coat protein. In plants inoculated with Tobacco mosaic virus, Potato virus X, and Tobacco etch virus, TEM images and western blot assays confirmed the presence of viral coat proteins in the upper leaves of all lead treatments. However, in plants inoculated with Turnip vein-clearing virus (TVCV), no signs of viral particles were detected in the upper leaves of plants treated with 10xa0μM or 15xa0μM lead nitrate. In contrast, plants treated with high concentrations of lead nitrate (50xa0μM or 100xa0μM) showed viral particles in their upper leaves. In plants treated with 10xa0μM or 15xa0μM lead nitrate, callose accumulation was the same as in control plants. This suggests that non-toxic concentrations of lead nitrate may trigger the production of putative cellular factors in addition to callose that interfere with the TVCV systemic movement. In contrast, plants treated with 100xa0μM lead nitrate showed less callose as compared to control plants, facilitating the systemic movement of TVCV.

First Report of Anthracnose Stem Canker of the Invasive Perennial Weed Lepidium draba Caused by Colletotrichum higginsianum in Europe

Exotic perennial Lepidium draba, native to Eurasia, is an invasive weed in dense stands in rangelands and disturbed areas in several states of the western United States and an agricultural weed in the prairie provinces of Canada. To determine strategies, such as a potential multipathogen strategy (1), for biological control of the weed, surveys that included the native range were conducted in spring 2009 to detect diseases that occur on this weed. Several stunted and chlorotic plants were found scattered throughout a stand of L. draba growing in a vacant lot near Riddes, Switzerland (46°0822.99″N, 7°919.02″E): ( http://maps.google.com/maps?source=earth&ll=46.13983490,7.15503250&layer= c&cbll=46.13983490,7.15503250&cbp=1,360,,0,5 ). Affected plants had reddish brown cankers on the lower stems, usually elongated and irregular in shape and slightly sunken. Insect injury was associated with the cankers. Symptoms often occurred on plants that were also infected with Rhizoctonia solani. After surface disinfestation with 0.1% sodium hypochlorite, tissue adjacent to and including lesions were plated on acidified potato dextrose agar and incubated at 20 to 25°C for 1 week. Zonate, dark gray colonies with sparse mycelia resulted that exhibited abundant, faintly pink spore masses with numerous dense clusters of black setae. Spores were single celled, hyaline, cylindrical to oval shaped, and 13.5 to 19.5 × 4 to 5.5 μm. Setae were 1- to 3-septate and 20 to 42 × 3 to 5 μm. These morphological traits correspond to Colletotrichum higginsianum. For pathogenicity tests, three 4-month-old L. draba plants were sprayed until runoff with a 106 conidia/ml suspension of the fungus and incubated for 72 h in plastic bags at 20 to 25°C in a quarantine greenhouse. Within 4 days, water-soaked lesions appeared that coalesced, resulting in chlorosis and collapse of inoculated leaves. Such symptoms are typical of infection by C. higginsianum and similar necrotrophic species (4). Fungi isolated from inoculated leaves were identified as C. higginsianum. To assess the host range of C. higginsianum, three plants each of turnip, radish, mustard greens, kale, broccoli raab, and Chinese cabbage, all in the Brassicaceae to which L. draba belongs, were inoculated with the same conditions used for the pathogenicity tests. Control plants in pathogenicity and host range tests were sprayed with sterile distilled water and all experiments were repeated at least once. All control plants were symptomless. Leaf necrosis occurred on radish and turnip and to a lesser extent on the lower leaves of Chinese cabbage and broccoli; numerous scattered dark necrotic flecks and small grayish leaf spots occurred on kale and mustard greens, respectively. These results are similar to previous studies (2,3) involving a cultivated species as the host in the field. The ITS1, 5.8S, and ITS2 sequences of this fungus (GenBank No. HM044877) were 99% similar to sequences of multiple isolates of C. higginsianum (GenBank Nos. AB042302, AB042303, AB455253, AJ558109, and AJ558110). To our knowledge, this is the first report of C. higginsianum on a wild species of the Brassicaceae, although a Colletotrichum sp. was reported on wild radish in Australia (1). References: (1) A. Maxwell and J. K. Scott. Australas. Plant Pathol. 37:523, 2008. (2) R. OConnell et al. Mol. Plant-Microbe Interact. 17:272, 2004. (3) R. P. Scheffer. N. C. Agric. Exp. Stn. Tech. Bull. 1950. (4) H. Sun and J. Z. Zhang. Eur. J. Plant Pathol. 125:459, 2009.