Susana R. Milla-Lewis

Inheritance of Evolved Glyphosate Resistance in a North Carolina Palmer Amaranth (Amaranthus palmeri) Biotype

Inheritance of glyphosate resistance in a Palmer amaranth biotype from North Carolina was studied. Glyphosate rates for 50% survival of glyphosate-resistant (GR) and glyphosate-susceptible (GS) biotypes were 1288 and 58u2009gu2009ha−1, respectively. These values for F1 progenies obtained from reciprocal crosses (GR×GS and GS×GR were 794 and 501u2009gu2009ha−1, respectively. Dose response of F1 progenies indicated that resistance was not fully dominant over susceptibility. Lack of significant differences between dose responses for reciprocal F1 families suggested that genetic control of glyphosate resistance was governed by nuclear genome. Analysis of F1 backcross (BC1F1) families showed that 10 and 8 BC1F1 families out of 15 fitted monogenic inheritance at 2000 and 3000u2009gu2009ha−1 glyphosate, respectively. These results indicate that inheritance of glyphosate resistance in this biotype is incompletely dominant, nuclear inherited, and might not be consistent with a single gene mechanism of inheritance. Relative 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) copy number varied from 22 to 63 across 10 individuals from resistant biotype. This suggested that variable EPSPS copy number in the parents might be influential in determining if inheritance of glyphosate resistance is monogenic or polygenic in this biotype.

Use of AFLP Markers to Assess Genetic Diversity in Palmer Amaranth (Amaranthus palmeri) Populations from North Carolina and Georgia

Abstract Glyphosate-resistant Palmer amaranth is a serious problem in southern cropping systems. Much phenotypic variation is observed in Palmer amaranth populations with respect to plant growth and development and susceptibility to herbicides. This may be related to levels of genetic diversity existing in populations. Knowledge of genetic diversity in populations of Palmer amaranth may be useful in understanding distribution and development of herbicide resistance. Research was conducted to assess genetic diversity among and within eight Palmer amaranth populations collected from North Carolina and Georgia using amplified fragment length polymorphism (AFLP) markers. Pair-wise genetic similarity (GS) values were found to be relatively low, averaging 0.34. The highest and the lowest GS between populations were 0.49 and 0.24, respectively, while the highest and the lowest GS within populations were 0.56 and 0.36, respectively. Cluster and principal coordinate (PCO) analyses grouped individuals mostly by population (localized geographic region) irrespective of response to glyphosate or gender of individuals. Analysis of molecular variance (AMOVA) results when populations were nested within states revealed significant variation among and within populations within states while variation among states was not significant. Variation among and within populations within state accounted for 19 and 77% of the total variation, respectively, while variation among states accounted for only 3% of the total variation. The within population contribution towards total variation was always higher than among states and among populations within states irrespective of response to glyphosate or gender of individuals. These results are significant in terms of efficacy of similar management approaches both in terms of chemical and biological control in different areas infested with Palmer amaranth. Nomenclature: Palmer amaranth, Amaranthus palmeri S. Wats

Transferability of SSR and RGA Markers Developed in Cynodon spp. to Zoysia spp.

Bermudagrass (Cynodon spp.) and zoysiagrass (Zoysia spp.), which are both used as warm-season turfgrasses in the USA, are members of subfamily Chloridoideae and reported to be at least 55% genetically similar. To assess if molecular tools between the two species can be interchanged, 93 primer pairs corresponding to bermudagrass simple sequence repeats (SSRs) and resistance gene analogs (RGAs) were used to amplify DNA from eight zoysiagrass cultivars representing three species, Zoysia japonica, Z. matrella, and Z. japonica × Z. pacifica. From these 93 bermudagrass primer pairs, 11 produced clear amplicons in Zoysia cultivars. Alleles from the Zoysia accessions were scored, genetic similarities were calculated, and a cluster analysis was performed. Using these markers, genetic similarity estimates between zoysiagrass cultivars and bermudagrass accessions ranged from 0.16 to 0.22. The transferability of bermudagrass SSRs and RGAs to zoysiagrass cultivars was low (7–12%), but certain markers permitted detection of genetic variability among zoysiagrass cultivars.

Overexpression of ubiquitin-like LpHUB1 gene confers drought tolerance in perennial ryegrass

HUB1, also known as Ubl5, is a member of the subfamily of ubiquitin-like post-translational modifiers. HUB1 exerts its role by conjugating with protein targets. The function of this protein has not been studied in plants. A HUB1 gene, LpHUB1, was identified from serial analysis of gene expression data and cloned from perennial ryegrass. The expression of this gene was reported previously to be elevated in pastures during the summer and by drought stress in climate-controlled growth chambers. Here, pasture-type and turf-type transgenic perennial ryegrass plants overexpressing LpHUB1 showed improved drought tolerance, as evidenced by improved turf quality, maintenance of turgor and increased growth. Additional analyses revealed that the transgenic plants generally displayed higher relative water content, leaf water potential, and chlorophyll content and increased photosynthetic rate when subjected to drought stress. These results suggest HUB1 may play an important role in the tolerance of perennial ryegrass to abiotic stresses.

Interference and Control of Glyphosate-Resistant and -Susceptible Palmer Amaranth (Amaranthus palmeri) Populations under Greenhouse Conditions

Abstract Interference for 40 d after emergence (DAE) of corn, cotton, peanut, and snap bean by four glyphosate-resistant (GR) and four glyphosate-susceptible (GS) Palmer amaranth populations from Georgia and North Carolina was compared in the greenhouse. Greater interference from Palmer amaranth, measured as crop height and fresh weight reduction, was noted in cotton and peanut compared with corn or snap bean. Crop height 15 to 40 DAE was reduced similarly by GR and GS populations. Crop fresh weight, however, was reduced 25 and 19% in the presence of GS and GR populations, respectively. Measured as percent reduction in fresh weight, GR and GS populations of Palmer amaranth were controlled similarly by glufosinate, lactofen, paraquat, and trifloxysulfuron applied POST. Atrazine and dicamba controlled GR populations more effectively than GS populations. Nomenclature: Atrazine; dicamba; glufosinate; lactofen; paraquat; trifloxysulfuron; Palmer amaranth, Amaranthus palmeri S. Wats.; corn, Zea mays L.; cotton, Gossypium hirsutum L.; peanut, Arachis hypogaea L..; snap bean, Phaseolus vulgaris L.

Use of sequence-related amplified polymorphism (SRAP) markers for comparing levels of genetic diversity in centipedegrass (Eremochloa ophiuroides (Munro) Hack.) germplasm

Centipedegrass (Eremochloa ophiuroides (Munro) Hack.) has great potential as a low-input turf within the U.S. because of its lower management requirements and its tolerance to an array of environmental stresses. Only a handful of centipedegrass cultivars have been released to date, however. This is mainly due to limited morphological variation present in U.S. centipedegrass germplasm. With the objective of broadening the genetic base of this species, a germplasm collection trip was conducted in seven Chinese provinces in 1999. Although the resulting accessions exhibited morphological variation for a number of traits, little is known about levels of molecular variability in these accessions or how those levels compare to diversity in materials previously present in the U.S. Sequence-related amplified polymorphism (SRAP) markers were used in the current study to investigate these issues. Eleven primer combinations yielded 279 scored fragments. Genetic diversity, in terms of number of alleles and Dice similarity values, was highest in the Henan and U.S. groups. AMOVA indicated that while both the among and within components of variance were significant (Pxa0<xa00.0001), most of the variation (94%) could be explained by differences within groups. The PCO plot showed large differences in levels of diversity in all groups evaluated. Overall, our results indicate that while the U.S. collection had high levels of diversity compared to other groups, there are alleles in the Chinese groups not represented in the U.S. collection. These materials could represent additional sources of variation to be used in centipedegrass cultivar development programs.

St. Augustinegrass Germplasm Resistant to Blissus insularis (Hemiptera: Blissidae)

ABSTRACT n St. Augustinegrass (Stenotaphrum secundatum (Walter) Kuntze) is an economically important turfgrass in the southeastern United States. However, this turf species is prone to southern chinch bug, Blissus insularis Barber (Heteroptera: Blissidae) outbreaks. This insect is the most destructive pest of St. Augustinegrass wherever this turfgrass is grown. Host plant resistance has historically been an effective management tool for southern chinch bug. Since 1973, the ‘Floratam’ St. Augustinegrass cultivar effectively controlled southern chinch bug in the southeast. However, southern chinch bug populations from Florida and Texas have now circumvented this resistance, through mechanisms still unknown. Therefore, identifying and deployingnewcultivars with resistance to the southern chinch bug is imperative to combat this pest in an economically and environmentally sustainable manner. Currently, the number of cultivars with resistance against southern chinch bug is limited, and their efficacy, climatic adaptability, and aesthetic characters are variable. Hence, the main focus of this study is the identification of alternative sources of resistance to southern chinch bugs in previously uncharacterized St. Augustinegrass plant introductions (PIs) and its closely related, crossbreeding species, Pembagrass (Stenotaphrum dimidiatum (L.) Brongniart). The PIs exhibited a wide range of responses to southern chinch bug feeding, as indicated by damage ratings. Damage ratings for seven PIs grouped with our resistant reference cultivars. Moreover, nine PIs exhibited antibiosis, based on poor development of southern chinch bug neonates, when compared with our susceptible reference cultivars. Altogether our study has produced strong support to indicate these materials are good candidates for future southern chinch bug resistance breeding in St. Augustinegrass.

A SNP-based high-density linkage map of zoysiagrass ( Zoysia japonica Steud.) and its use for the identification of QTL associated with winter hardiness

Zoysia japonica Steud. (2nu2009=u20094×u2009=u200940) is a C4 turfgrass well-adapted for the warm-humid and transitional climatic zones of the USA. Its use is limited to warmer climates because of a relative lack of winter hardiness compared to C3 grasses. Molecular markers associated with this trait would be useful for effective selection of winter hardy germplasm before field testing. A pseudo-F2 mapping population of 175 individuals was developed from crosses between Z. japonica cultivars “Meyer” (freeze-tolerant) and “Victoria” (freeze-susceptible) and used to generate a high-density genetic map of 104 SSR markers and 2359 sequencing-derived SNP markers. The map covers 324xa0Mbp and 2520xa0cM as well as the 20 chromosomes for the zoysiagrass haploid genome. Phenotypic data on winter injury, establishment, and turf quality collected in North Carolina and Indiana in 2014–2016 were used in conjunction with this map to identify quantitative trait loci (QTL) associated with winter hardiness. Fifty-six QTL associated with winter injury, establishment, and turf quality were identified over six environments. Twelve of those were identified in two or more environments. Furthermore, seven regions of interest were identified on chromosomes 8, 11, and 13 where co-location of QTL for three or more traits occurred. Within these regions, analysis with NCBI basic local alignment search tool (BLAST) identified proteins related to cold and other abiotic stresses tolerance. These QTL and associated markers could be valuable in implementing marker-assisted selection for winter hardiness in zoysiagrass breeding programs.

Assessing freeze-tolerance in St. Augustinegrass: temperature response and evaluation methods

Winter hardiness is a major-limiting factor for St. Augustinegrass [Stenotaphrum secundatum (Walt.) Kuntze] grown in the transitional climatic region of the United States. Lab-based freeze tests that mimic the range of field winter survivability in St. Augustinegrass can contribute to the selection of cold hardy genotypes. This study used a whole container method, four freezing temperatures, and two data collection systems to evaluate the freezing response of nine St. Augustinegrass genotypes ranging in their winter hardiness. Results indicated −3 and −4xa0°C with average regrowth ratings of 33.6 and 17.8% respectively, were more suitable temperatures for evaluating freeze survival in St. Augustinegrass than −5 and −6xa0°C with average regrowth ratings of 0.4 and 0%, respectively. Visual ratings of surviving green tissue and regrowth were generally well correlated when evaluated over a six week period post-freeze with Pearson correlation coefficients ranging of 0.17–0.62xa0 for −3xa0°C freeze tests and 0.79–0.93 for −4xa0°C freeze tests. Additionally, measurement of percent green cover using digital imaging techniques commonly utilized in turfgrass field studies were significantly correlated (0.66) with visual ratings averaged across weekly post-freeze evaluation measurements for both −3 and −4xa0°C freezing temperatures. These results provide evidence that digital imaging analyses are useful in estimating surviving green tissue and regrowth in lab-based freeze tests. This study provides additional information regarding freezing temperatures, genotype responses, and data collection methods in St. Augustinegrass, which should aid breeders in the improvement of freeze tolerance in the species.

Response of Herbicide-Resistant Palmer Amaranth (Amaranthus palmeri) Accessions to Drought Stress

Palmer amaranth is a very problematic weed in several crops in the southern USA due to its competitive ability and resistance to herbicides representing different mechanisms of action. Variation in growth and subsequent interference of North Carolina Palmer amaranth accessions has not been examined. A greenhouse experiment determined response of 15 North Carolina Palmer amaranth accessions to drought stress beginning 15 days after seedling emergence (DAE) for a duration of 3, 5, 7, and 9 days. Following exposure to drought, plants were grown under optimal moisture conditions until harvest at 30 DAE. Five accessions each of glyphosate-resistant (GR), acetolactate synthase inhibitor-resistant (ALSR), and acetolactate synthase inhibitor-susceptible and glyphosate-susceptible (ALSS/GS) were compared. Variation in response to drought stress, based on height and dry weight reduction relative to nonstressed controls, was noted among accessions. Stress for 3 or more days affected height and dry weight. Height and dry weight of GR and ALSR accession groups were reduced less by drought than the ALSS/GS accession group. Results suggest a possible relationship between herbicide resistance and ability of Palmer amaranth to withstand drought stress and thus a possible competitive advantage for resistant accessions under limited moisture availability.