White Nodular Structure Research Articles

Immunohistochemical Investigation of Lipid Peroxidation in Renal Coccidiosis of Geese

Objective: In this study, we aimed to evaluate the oxidative damage caused by lipid peroxidation due to renal coccidiosis by histopathological and immunohistochemical methods.Material-Method: The material of this study was made up of tissue samples taken from 139 geese whose average age was 10 weeks, who were brought to our department dead between 2013-2020. Tissue samples taken were fixed in 10% buffered formaldehyde solution. 5 µm-thick sections were taken from the paraffin blocks prepared after routine tissue follow-up procedures. Hematoxylin & Eosin staining was applied to the sections in order to detect histopathological changes. Sections were examined and photographed under a light microscope. Results: Various clinical signs such as fever, respiratory distress, weakness, anorexia, tremors, inability to get up from the ground, balance disorders, rotational movement, diarrhea, wheezing were detected in geese. In systemic necropsies of geese, large and small white nodular structures were detected in the kidney. In histopathological examinations, coccidiosis agents (E. truncata) were found in the tubular epithelium of the kidney. Necrosis and mononuclear cell infiltration were observed in the tubules due to the presence of E. truncata. In addition, edema and hyperemia in the lungs, multifocal necrosis in the liver, cell infiltration in the portal spaces, and enteritis were other important histopathological findings. In one case, aspergillosis was detected together with renal coccidiosis. We observed that MDA expression was more severe in the oocyst stages, which is the mature form of the parasite, compared to other parasitic life stages. Conclusion: Based on the results obtained from this study, it was revealed that renal coccidiosis in geese caused lipid peroxidation / oxidative damage through the increase in MDA expression.

Mass propagation of microtubers from suspension cultures of Pinellia ternata cells and quantitative analysis of succinic acid in Pinellia tubers

The conditions for efficient tuber production from suspension cultures of Pinellia ternata cells which is one of medicinal herbs were established and succinic acid in tubers propagated in vitro was determined. Leaf explants formed white nodular structures and off-white calluses at a frequency of 90.6 % when cultured on Murashige and Skoog medium supplemented with 0.54 μM α-naphthaleneacetic acid (NAA); however, this frequency declined substantially with increasing NAA concentration, up to 16.2 μM, at which the frequency reached zero percent. In combination treatments with 4.44 μM 6-benzyladenine (BA) and NAA, however, the frequency of white nodular structure and off-white callus formation from leaf explants did not decrease, even at 16.2 μM NAA. Suspension cultures of P. ternata cells were established from leaf-derived off-white calluses in MS liquid medium containing 5.4 μM NAA and 4.44 μM BA. Upon plating onto MS basal medium, over 90 % of cell aggregates gave rise to microtubers and developed into plantlets. Regenerated plantlets were transplanted in potting soil and grown to maturity in a growth chamber, with a survival rate of >90 %. The highest succinic acid content in suspension culture-derived microtubers was 45 g/kg of extract. Compared with wild P. ternata medicinal tubers, the succinic acid content was very similar. The in vitro P. ternata microtuber proliferation system established in this study is thus an efficient alternative for the mass production of medicinal tubers.

In Vitro Response of Cotyledon Explants of Cashew (Anacardium occidentale L.) Cultured in Different Concentrations of BAP

This experiment was carried out to study the embryogenic response of cashew ( Anacardium occidentale L.) cotyledon explants cultured in different concentrations of BAP under in vitro conditions. Sterilized cotyledon segments (10 mm long, 5 mm width) were cultured on full strength MS basal medium containing various concentrations (0-10 mg/L) of BAP. The result revealed the medium without BAP exhibited profuse root organogenesis which decreased with the increase in the concentration of BAP. Greenish or yellowish white nodular structures appeared on the edge of the surface concentrated towards the convex side of the explants. Consequently, these structures gave rise to somatic embryoids and the degree of embryoid formation increased with the increasing concentration of BAP. MS medium with 8 mg/L BAP was the most optimal concentration for the induction of somatic embryos among the tested. Cytological study confirmed the development of somatic embryoids directly from cultured cotyledon explants. Journal of Food and Agriculture 2010 Vol. 03, No. 1 & 2, Page 7-13 DOI: http://dx.doi.org/10.4038/jfa.v3i1-2.5165

Establishment of high frequency plant regeneration system from leaf explants of Pinellia koreana via bulblets formation

Pinellia koreana K-H Tae & J-H Kim is a recently discovered Korea endemic medicinal plant species whose natural habitat is rapidly destroyed by industrial development. Described in this paper are culture conditions for high frequency plant regeneration via bulblet formation from leaf explant cultures of P. koreana. Leaf explants formed white nodular structures and off-white calluses at a frequency of 91.2% when cultured on MS medium supplemented with 2 mg/L BA and 0.5 mg/L NAA. However, the frequency of white nodular structures and off-white calluses formation was slightly decreased with an increasing concentration of NAA up to 4 mg/L, where the frequency reached 31.7%. Most petiole explants did not form white nodular structures and off-white calluses except the combination treatment of 2 mg/L BA and 2 mg/L NAA. Upon transfer onto MS basal medium, over 90% of nodular structures gave rise to numerous bulblets and developed into plantlets. Plantlets regenerated from bulblets were transplanted to potting soil and grown to maturity at a survival rate of over 95% in a growth chamber. Therefore, the in vitro plant regeneration system of P. koreana obtained in this study will be useful for mass propagation and long-term preservation of genetic resources of P. koreana. *Corresponding author Tel 042-860-4646 Fax 042-860-4677 E-mail: kimsw@kribb.re.kr Introduction Pinellia koreana K-H Tae & J-H Kim is a new plant species discovered at Chrisan in Korea (Tae and Kim, 2005). This plant differs from other plants belonging to the same genus Pinellia by having 3-foliolate leaves with bilobed lateral leaflets at maturity. The genus Pinellia is herbaceous medicinal plants belonging to the Araceae. The chemical composition of Pinellia is not well established as there are only a few reports on the occurrence of small amounts of alkaloids ephedrine in the tuber. Also a glycoprotein fraction was reported to have notable antiemetic effects (Kurata et al. 1998). And root contains toxins (presence of calcium oxylate), that are destroyed by drying or cooking. Pinellia is mostly used for indigestion, nausea, vomiting, gastritis, and ulcer (Lee and Cho 1987; Bown 1995). The genus of Pinellia consists of a few species in eastern Asia (Kitamura et al. 1980). Pinellia is propagated by tubers and bulbils. However, the yield and quality of tubers are gradually reduced over cultivation because of viral diseases. Therefore, introduction of a mass propagation system using in vitro culture technology such as development of virus-free strains and genetic manipulation of tubers is necessary to enhance the productivity of tubers. Despite its medicinal importance, few scientific studies of P. koreana have been reported. Only several in vitro culture studies have reported on the related species P. ternata including protoplast culture and plant regeneration (He et al. 1996), formation of micro-tubers and plantlet regeneration by thidiazuron (Yoo and Lim 1997), and clonal propagation by bulbils (Chen et al. 1989; Tsay et al. 1989). Also natural habitats of P. koreana decreased gradually because of industrialization. Tissue culture techniques may provide an alternative mean for its mass multiplication and ex-situ conservation of genetic resources. This study describes culture conditions for high frequency plant regeneration via bulblet formation from leaf explants cultures of P. koreana. 식물생명공학회지 제36권 제2호 J Plant Biotechnol Vol. 36, No. 2, 193-196 (2009) 194 ‧ Journal of Plant Biotechnology Figure 1. Plant regeneration from leaf explants of P. koreana via bulblets formation. A: Leaf explants; B: Off-white callus formation; C: White nodular structure formation; D: Bulblets formation from leaf-derived callus; E: Leaf and root development from bulblets; F: Regenerated plantlets transferred to soil; G: Morphological variation in leaf shape of regenerated plantlets; H: Morphological variation of plants after 2 year cultivation; I: Flowering of mature plants. Scale bars represents 1 mm (A, B, C, D and E), 2 cm (F, H) and 5 cm (G, I), respectively Materials and methods

Plant regeneration from zygotic embryo-derived embryogenic cell suspension cultures of Ranunculus kazusensis

Culture conditions for high frequency plant regeneration via somatic embryogenesis from cell suspension cultures of Ranunculus kazusensis are described. Zygotic embryos formed white nodular structures and pale-yellow calluses at a frequency of 84.9% when cultured on half-strength Schenk and Hildebrandt (SH) medium supplemented with 0.1 mg l−1 2,4-dichlorophenoxyacetic acid (2,4-D). However, the frequency of white nodular structure and off-white callus formation decreased with an increasing concentration of 2,4-D up to 10 mg l−1, when the frequency reached 25%. Cell suspension cultures were established from zygotic embryo-derived pale-yellow calluses using half-strength SH medium supplemented with 0.1 mg l−1 of 2,4-D. Upon plating onto half-strength SH basal medium, over 90% of cell aggregates gave rise to numerous somatic embryos and developed into plantlets. Regenerated plantlets were successfully transplanted to potting soil and grown to maturity at a survival rate of over 90% in a growth chamber. The plant regeneration system established in this study can be applied to mass propagation and conservation of this species.

Effects of cytokinin on adventitious root formation in callus cultures ofVigna unguiculata (L.) walp

Yellowish compact callus, induced from cowpea hypocotyls on Murashige and Skoog(MS) medium (1962) containing 0.2 mg/l(0.93 μM) kinetin and 0.4 mg/l (1.81 μM) 2,4-dichlorophenoxyacetic acid (2,4-D), was subcultured on MS medium containing cytokinin alone, auxin alone, or auxins plus cytokinins in order to determine the effect of cytokinins on root organogenesis in callus cultures. The callus actively proliferated on the same medium but did not show any organogenic activity macroscopically as well as microscopically. On medium with N6-benzyladenine (BA) and 1-naphthaleneacetic acid (NAA), the yellowish compact callus first changed to pale green compact callus and then many green spots appeared on its surface under light culture. But the yellowsih compact callus remained yellowish and white spots appeared on its surface in dark culture. These spots gradually became white nodular structures. Adventitious root formation from the nodular structures occurred not only on the same medium, but also on medium with either auxin or cytokinin but not both. Yellowish compact callus on medium with auxin alone was transformed to yellowish friable callus, which did not develop adventitious roots. The yellowish friable callus could gain rhizogenic activity only after morphological modification to pale green compact callus on medium with auxin plus cytokinin. The modified callus did not form adventitious roots on medium with auxins but only with cytokinins. Therefore, it is suggested that cytokinins have stimulating effects on root formation from callus that previously did not show rhizogenic activity on medium with auxins alone. In addition, the rhizogenic potential of cowpea callus was discriminated from that of leaf explants, which formed adventitious roots directly on medium with auxin alone.

Embryogenic Callus Formation from Protoplasts Derived from Suspension Cells of Apomictic Guineagrass (Panicum maximum Jacq.).

Protoplasts were isolated from suspension cells of apouuictic guineagrass (Pauicum maximum Jacq.). The suspension culture used as donor Inaterial was originally initiated from immature embryoderived embryogenic callus. Prior to protoplast isolation, suspension cells were conditioned with Murashige and Skoog (MS) Iiquid medium without sucrose and growth regulators. This pretreatment lead to a dramatic increase in protoplast yield and colony formation. Cell division and colony formation from such pretreated protoplasts were found to be best in agarose-solidified modified KM8p medium. Protoplast derived colonies developed into callus on solidified MS medium supplemented with 1 mg/l 2, 4-D. After 2 months in culture, calli formed compact white nodular structures some of which developed into somatic embryos. Although some of the somatic embryos developed small leafy structures, whole plants could not be regenerated.

Recovery of γ-Linolenic Acid from Somatic Embryos of Borage

Abstract Somatic embryos of borage (Borago officinalis L., Boraginaceae) were induced directly from immature zygotic embryos and indirectly from callus. Embryogenic callus maintained on liquid basal medium supplemented with 4.5 μm 2,4-D and 10% (v/v) coconut water (CW) produced globular structures that became rhizogenic upon transfer to 2,4-D-free basal medium. Embryogenic callus maintained on semisolid basal medium supplemented with 4.5 μm 2,4-D and 10% CW continued to produce somatic embryos, but development was abnormal. Globular structures often failed to develop cotyledons, and those that developed were small and fused; hypocotyls tended to be large and elongated. Root meristems appeared normal, but shoot meristems were not formed. Carbon source (sucrose, glucose, or maltose) and ABA did not normalize somatic embryogenesis. A highly embryogenic, non-browning clone produced various tissue types when 2,4-D was withdrawn that varied in total fatty acids: white nodular structures (12.6%), cotyledonary structures (22.5%), white callus (5.0%), green leafy growth (3.1%), and translucent globular growth (5.1%). γ-Linolenic acid, as a percentage of total fatty acids, was highest in cotyledonary structures (19.9%) and lowest in white callus (10.2%). Chemical names used: abscisic acid (ABA); 2,4-dichlorophenoxyacetic acid (2,4-D); coconut water (CW).