Aluminum (Al) toxicity in acid soils is a major constraint to global agricultural production, affecting ≈30% of the world’s arable land area. To study Al tolerance in barrel medic (Medicago truncatula), we assessed responses to excess Al in 91 accessions collected from different geographic regions. Root elongations were used to characterize the sensitivity of each accession. Seedlings were grown in an agarose medium that contained three levels of Al (50, 100, and 200 µm), and root elongation was measured at 72 hours after exposure to Al. The ratio of root elongation in the presence and absence of Al [relative root growth (RRG)] differed among accessions. At 50 µm Al, we observed the greatest range of intraspecific variation. Aluminum sensitivity of 30 accessions was tested further by hematoxylin staining. Relative root growth was regressed linearly against the visual staining score, and a significant, negative, linear relationship was found between RRG at 50 or 100 µm Al and the intensity of staining scores. Twelve selected accessions differing in their resistance were grown in Al-toxic soil to confirm their Al response. Such information could be useful in breeding or selecting for improved Al tolerance in barrel medic, as well as other crop species.
Yawadee Srimake and Susan C. Miyasaka
Susan C. Miyasaka and Carol M. Webster
Aluminum toxicity is one of the major factors limiting plant growth in acid soils. Taro [Colocasia esculenta (L.) Schott] cultivars `Lehua maoli' and `Bun long ' were grown in hydroponic solution at six levels of aluminum (0, 110, 220, 440, 890, and 1330 μM Al), to determine the differential response of taro to Al. Increasing Al levels resulted in significantly depressed fresh and dry weights of leaves, petioles, and roots, as well as leaf areas and root lengths. Significant cultivar differences were found, with `Lehua maoli' exhibiting greater leaf fresh weights and root lengths in the presence of Al, compared to `Bun long'. These cultivar differences were not associated with differences in Al concentrations of the leaves, petioles, or roots.
Susan C. Miyasaka and Randall T. Hamasaki
To determine promising olive (Olea europaea) cultivars for oil production in Hawaii, seven trees each of 10 cultivars (Arbequina, Arbosana, Coratina, Frantoio, Koroneiki, Leccino, Mission, Moraiolo, Pendolino, and Taggiasca) were planted in Feb. and July 2011 at the Lalamilo Experiment station on Hawaii Island (lat. 20.0176°N, long. 155.6827°W, elevation 2700 ft). In addition, two trees each of these 10 cultivars were planted in June 2011, with the exception of Arbequina, which was planted in July 2012, at the Maui Agricultural Research Center in Kula, Maui (lat. 20.7564°N, long. 156.3289°W, elevation 3100 ft). At Lalamilo, after ≈2 years of growth in the field (2013), three cultivars of olives (Arbequina, Arbosana, and Koroneiki) flowered, fruited, and produced oil yields of greater than 20%. These same cultivars flowered and fruited in 2014 and 2015. There was no significant difference among cultivars in fresh weight fruit yield averaged over 2 years (2013 and 2014), ranging from 2.14 to 2.45 kg/tree. During December to March, calculation of chilling hours below 12.5 °C was 141 hours during 2012–13 and 161 hours during 2013–14. The other seven cultivars did not flower and fruit during these 2 years of growth at Lalamilo, perhaps due to a greater requirement for chilling hours. At Kula, after 3 years of growth (2015), nine cultivars of olives with the exception of Moraiolo flowered and fruited. Mean fresh weight fruit yield in 2015 ranged from 0.25 to 22.06 kg/tree for various cultivars grown in Kula, Maui. In 2013, the oil from three cultivars grown at Lalamilo was analyzed for free fatty acids (FFA), peroxide value (PV), ultraviolet absorption for conjugated double bonds, 1,2-diacylglycerol (DAG), and pyropheophytins (PPP). Oil quality was within the range of extra-virgin olive oil. There is a need to investigate further the effects of temperature and management on flowering and fruiting of olive cultivars grown in Hawaii at various elevations. In particular, ‘Arbequina’, ‘Arbosana’, and ‘Koroneiki’ appear to have a lower requirement for chilling hours than other cultivars tested.
Susan C. Miyasaka, Charles E. McCulloch and Scot C. Nelson
Taro leaf blight (TLB), caused by the oomycete pathogen Phytophthora colocasiae, is a worldwide disease that threatens the sustainable cultivation of the tropical root crop taro (Colocasia esculenta). To evaluate taro germplasm from Asia, Hawai‘i, and several South Pacific Islands for resistance to TLB, 119 cultivars were planted along the Hamakua Coast of Hawai‘i (mean annual rainfall of 130 inches) in plots containing five or 10 plants that were replicated over time from 1993 through 2005. Fresh and dry weights of corms were measured after about nine months, with rotten portions removed and weighed. When epidemics of TLB occurred (in nine out of 12 years), visual estimates of disease severity on leaves were assessed using a modified Horsfall–Barratt scale. The correlations between mean dry weight yields for each cultivar and mean severity of TLB, and, respectively, between mean yields and mean severity of corm rots were calculated. As severity of TLB or severity of corm rots increased (suggesting increased susceptibility of particular cultivars to TLB or corm rots), mean dry weight yields decreased significantly (r 2 = 0.37 and 0.22, respectively). “Multiple comparisons with the best” (MCB) were conducted on fresh and dry weight yields, severity of TLB, severity of corm rots, percentage dry matter of corm, and consumer acceptance. Five cultivars were found to be “among the best” with: 1) fresh or dry weight yields that did not differ from the highest level; 2) severity ratings for TLB that were significantly lower than the highest level, suggesting TLB resistance; and 3) percentage of corm rots that were lower than the highest level, suggesting disease resistance. These cultivars, four of which originated from Palau, were Dirratengadik, Merii, Ngesuas, Ochelochel, and Sawa Bastora. Two commercial cultivars from Hawai‘i, Bun Long and Maui Lehua, had fresh and dry weight yields that were significantly lower than the maximum and severity of TLB injury that did not differ from the highest level, indicating that conventional breeding of taro to improve TLB resistance could improve yields of commercial taro cultivars, particularly in areas where epidemics of TLB occur.
Steven A. Hill, Susan C. Miyasaka and Russell S. Yost
Cupric sulfate pentahydrate (CuSO4·5H2O) has been proposed for use in Hawaii as a molluscicide to control golden apple snail (Pomacea canaliculata Lamarck) infestations of taro [Colocasia esculenta (L.) Schott]. Two hydroponic, greenhouse studies were conducted to determine the effects of solution Cu2+ levels on taro growth, the solution Cu2+ toxicity threshold, and useful diagnostic indicators of Cu toxicity. In the first experiment, taro cultivars Lehua maoli and Pololu were grown at nine levels of Cu2+ ranging from 0.5 to 25.0 μm. In the second experiment, `Lehua maoli' was grown at six levels of Cu2+ ranging from 0.25 to 2.5 μm. Significant (P ≤ 0.05) toxic effects included reduced dry matter production, leaf area, and root length: root dry weight ratio, and both impaired photosynthesis and a generalized reduction of cation accumulation in leaf blade tissue. The solution Cu2+ toxicity threshold (based on 90% of relative total dry weight) for young taro plants was 1.2 μm. Because Cu does not accumulate in leaf blade tissues with increasing solution Cu2+ levels, leaf Cu concentration cannot be used as a diagnostic indicator of Cu toxicity in taro.
Xiaoling He, Susan C. Miyasaka, Maureen M.M. Fitch, Sawsan Khuri and Yun J. Zhu
Production of taro [Colocasia esculenta (L.) Schott], a tropical root crop, is declining in many areas of the world as a result of the spread of diseases such as Taro leaf blight (TLB). Taro cv. Bun Long was transformed through Agrobacterium tumefaciens with the oxalate oxidase (OxO) gene gf2.8 from wheat (Triticum aestivum). Insertion of this gene was confirmed by polymerase chain reaction (PCR) and Southern blot analysis. One independent transformed line contained one gene insertion (g5), whereas a second independent line contained four copies of the gene. Reverse transcriptase PCR (RT-PCR) confirmed the expression of this gene in line g5. Histochemical analysis of the enzyme oxalate oxidase confirmed its activity increased in the leaves of line g5. A bioassay for resistance to TLB used zoospores of Phytophthora colocasiae to inoculate tissue-cultured plantlets. Transgenic line g5 showed the complete arrest of this disease; in contrast, the pathogen killed non-transformed plants by 12 days after inoculation. A second bioassay, in which spores of P. colocasiae were inoculated onto disks of leaves of one-year-old potted plants, confirmed that transgenic line g5 had greatly increased resistance to this pathogen. This is the first report to demonstrate that genetic transformation of a crop species with an OxO gene could confer increased resistance to a pathogen (P. colocasiae) that does not secrete oxalic acid (OA).
Anthony M. Ortiz, Brent S. Sipes, Susan C. Miyasaka and Alton S. Arakaki
To determine the potential to suppress root-knot nematode Meloidogyne javanica, 10 genotypes of seven green manure species were evaluated in a greenhouse study. These species were: black hollyhock (Alcea rosea L.); canola (Brassica napus L.); cabbage (B. oleracea L.); French marigold (Tagetes patula L.), sorghum–sudangrass [Sorghum bicolor (L.) Moench nothosubsp. drummondii (Steud.) de Wet ex Davidse]; sunn hemp (Crotalaria juncea L.); and yellow mustard (Sinapis alba L.). Plants were inoculated with eggs of M. javanica and after 6 weeks, nematode eggs and reproduction factor (Rf = final egg population density/initial egg population density) were determined. Marigolds were non-hosts to M. javanica; other crop species that were poor hosts to M. javanica included canola cv. Dwarf Essex, sorghum–sudangrass cvs. Piper and Sordan 79, black hollyhock cv. Nigra, and sunn hemp. Based on low Rf, four groups of species were selected for further evaluation in the greenhouse to determine the response to both M. javanica and another crop pathogen, Pythium aphanidermatum. These four groups of green manure crops were: 1) seven marigold genotypes; 2) four Brassicaceae genotypes; 3) seven sorghum–sudangrass hybrids; and 4) four other species [black hollyhock, sunn hemp, elecampane (Inula helenium L.), and black-eyed Susan (Rudbeckia hirta L.)]. Plants were inoculated with a factorial combination of M. javanica and P. aphanidermatum (none, each alone, and in combination) and repeated four times in a split-plot experimental design (whole plots were factorial treatments and subplots were green manure crop genotypes). Six weeks after inoculation, plants were harvested and measured for fresh and dry weights of shoots and roots and Rf of M. javanica. Adverse effects of P. aphanidermatum were characterized by dead or dying roots and measured by reduced plant biomass. Negative synergistic effects were observed in several marigold and Brassicaceae genotypes, in which the combined effects of M. javanica and P. aphanidermatum reduced shoot and root growth more severely than either treatment alone. Marigold T. erecta cv. Orangeade, sorghum–sudangrass cvs. Graze-All, Piper, and Sordan 79, and sunn hemp appeared to be resistant to M. javanica and P. aphanidermatum, either alone or in combination. Based on results of greenhouse trials, eight green manure crops (yellow mustard cv. Ida Gold, French marigolds cvs. Nema-gone and Golden Guardian, sorghum–sudangrass cvs. Sordan 79 and Tastemaker, sunn hemp, unplanted plot, and a control plot with weed mat) were selected and grown for 3 months in a field trial in Pepeekeo, HI. Each treatment was replicated four times in a randomized complete block design. Shoot biomass was sampled at 1, 2, and 3 months after planting. Plant–parasitic nematodes were counted before planting and at 4 months after planting. Dry weight biomass averaged across three sampling dates was greatest for the two sorghum–sudangrass hybrids followed by those of two marigold cultivars that did not differ from them. No significant differences in populations of root-knot nematodes were found. Based on this field trial as well as greenhouse trials, marigold cultivars, sorghum–sudangrass hybrids, and sunn hemp appeared to be non-hosts or poor hosts to reniform (Rotylenchulus reniformis) as well as root-knot nematodes and well adapted to the environmental conditions found along the Hamakua Coast of the Hawaii Island.
Xiaoling He, Susan C. Miyasaka, Yi Zou, Maureen M.M. Fitch and Yun J. Zhu
Genetic engineering has the potential to improve disease resistance in taro [Colocasia esculenta (L.) Schott]. To develop a method to produce highly regenerable calluses of taro, more than 40 combinations of Murashige and Skoog (MS) media at full- or half-strength with varying concentrations of auxin [α-naphthaleneacetic acid (NAA) or 2, 4-dichlorophenoxyacetic acid (2, 4-D)], cytokinin [benzyladenine (BA) or kinetin], and taro extract were tested for callus initiation and plant regeneration. The best combination, MS medium with 2 mg·L−1 BA and 1 mg·L−1 NAA (M5 medium), was used to produce regenerable calluses from taro cv. Bun Long initiated from shoot tip explants. After 8 weeks of growth, multiple shoots from these calluses could be induced on MS medium with 4 mg·L−1 BA (M15 medium). The rice chitinase gene (ricchi11) along with the neomycin phosphotransferase (npt II) selectable marker and β-glucuronidase (gus) genes were introduced into these taro calluses through particle bombardment. Transformed calluses were selected on M5 medium containing 50 mg·L−1 geneticin (G418). Histochemical assays for beta-glucuronidase (GUS), polymerase chain reaction (PCR), reverse transcription–PCR, and Southern blot analyses confirmed the presence, integration, and expression of the rice chitinase gene in one transgenic line (efficiency less than 0.1%). Growth and morphology of the transgenic plants appeared normal and similar to non-transformed controls. In pathogenicity tests, the transgenic line exhibited improved resistance to the fungal pathogen, Sclerotium rolfsii, but not to the oomycete pathogen, Phytophthora colocasiae.
Susan C. Miyasaka, Charles E. McCulloch, Graham E. Fogg and James R. Hollyer
Taro (Colocasia esculenta L. Schott) is a root crop widely grown in the Tropics. To determine the optimum plot size for taro field trials, fresh and dry weights of individual corms were collected from two field trials conducted under flooded culture and two conducted under upland culture. For a given maximum test plot with a single border row surrounding inner measured plants, all possible combinations of smaller plot sizes were investigated. A plot size was defined as a given number of adjacent plants. A strong linear relationship was found between the natural logarithm of variance of yield and the natural logarithm of plot size. Expressed on the non-log-transformed scale, the point of maximum curvature in this relationship indicates a sudden decrease in advantage to larger plot sizes and is taken as optimum. Calculating maximum curvature mathematically, optimum plot size was 21 inner plants (5.7 m2) for the second flooded trial and 18 inner plants (4.9 m2) for the second upland trial. Another method of estimating optimum plot size minimized the cost per unit of research data by using the index of degree of correlation between neighboring plots. In three of four trials, the optimum plot size ranged from 16 to 24 inner plants (4.3 to 6.5 m2). In this second method, we calculated a non-linear relationship between plot size and outer border plants to estimate the fixed and per-unit cost of a single border row surrounding the inner measured plants. Both methods of calculating optimal plot size sometimes resulted in estimates that exceeded the maximum test plot size for particular field trials, indicating limitations of each method and the importance of managing field trials to ensure uniformity across treatments. No evidence of spatial autocorrelation was found in the corm yield of taro, indicating that the two methods used were adequate in calculating optimum plot size. In addition, we conducted an analysis based on statistical power but found that plot size did not materially affect the power to detect differences between treatments. To our knowledge, this is the first report of optimum plot size for field trials of taro.
Archana P. Pant, Theodore J.K. Radovich, Nguyen V. Hue and Susan C. Miyasaka
Previous work has demonstrated the potential of compost tea to enhance plant growth and nutritional status. One factor thought to contribute to variability in the efficacy of compost tea is the amount of compost per unit volume of water. To address these gaps in our understanding, two greenhouse trials and two field trials were conducted to investigate the effects of various extraction ratios on the growth, mineral nitrogen (N), and phytonutrient content of pak choi (Brassica rapa, Chinensis) and on soil biological properties. In greenhouse experiments, plants were fertilized with a single rate of chicken manure-based thermophilic compost. In field trials, three fertilizer treatments: 1) rendered meat byproduct or Tankage (Island Commodities, Honolulu, HI); 2) soluble fertilizer (16:16:16); and 3) chicken manure-based thermophilic compost were applied. Aerated vermicompost teas were prepared using chicken manure-based vermicompost and water at various ratios. Pak choi plants were treated weekly for 4 weeks with 10%, 5%, 3%, and 1% vermicompost teas in the greenhouse experiments and 10% and 5% teas in the field trials. Applications of vermicompost tea significantly increased plant growth, N content, total carotenoids, and total glucosinolates in plant tissue; this response was greatest in chicken manure-fertilized treatments. Increases in yield and phytonutrient content were associated with increased N uptake. Vermicompost tea also increased soil respiration and dehydrogenase activity over the control (water). Plant growth, phytonutrient content, and microbial activities in soil increased with increasing concentrations of vermicompost tea. Within the range of concentrations evaluated (1%–10%), greatest plant growth response was observed with 5% and 10% vermicompost tea, indicating that the optimal water-to-vermicompost ratio for extraction is lower than 50:1 and is likely in the range of 10:1 to 20:1. The findings suggest that vermicompost tea could be used to improve plant nutrient status and enhance soil biological properties in vegetable production.