Micología Aplicada International ISSN: 1534-2581 [email protected] Colegio de Postgraduados México Dede, A. P. O.; Okungbowa, F. I. In vitro growth of Ceratocystis paradoxa in oil palm (Elaeis guineensis) fruit extract media Micología Aplicada International, vol. 19, núm. 2, july, 2007, pp. 51-55 Colegio de Postgraduados Puebla, México Available in: http://www.redalyc.org/articulo.oa?id=68519203 How to cite Complete issue Scientific Information System More information about this article Network of Scientific Journals from Latin America, the Caribbean, Spain and Portugal Journal's homepage in redalyc.org Non-profit academic project, developed under the open access initiative MICOLOGIA APLICADA INTERNATIONALCERATOCYSTIS, 19(2), PARADOXA 2007, pp. 51-55 © 2007, BERKELEY, CA, U.S.A. www.micaplint.com IN VITRO GROWTHGROWTH OFOF CERATOCYSTIS PARADOXA IN OIL PALM (ELAEISLAEIS GUINEENSISGUINEENSIS) FRUITFRUIT EXTRACTEXTRACT MEDIAMEDIA A. P. O. DEDE AND F. I. OKUNGBOWA Department of Botany, Faculty of Life Sciences, University of Benin, P.M.B. 1154, Ugbowo, Benin City, Nigeria. E-mail: fi [email protected]. Phone: 234-8055376204. Accepted for publication June 25, 2007 ABSTRACT The fungus Ceratocystis paradoxa, a pathogen of the oil palm and other economic crops, grows well in vitro on the oil palm fruit extract media (solid and liquid forms). A stock culture of the fungus (IMI specimen no. 79166) was used in this study, and its growth in oil palm fruit basal medium, potato dextrose and corn meal media were compared at three different temperatures (15 C, 28±2 C, 35 C). After 72 h of incubation at room temperature (28±2 C), the fungus had equal mean radial growth on potato dextrose agar (PDA) and oil palm fruit extract agar (PFEA) with a growth increase of 38 mm, while an increase of 11 mm was recorded for corn meal agar (CMA). At 15 C, growth was generally slow but the highest mean radial growth (increase of 10.8 mm) was recorded for PFEA, while 3.2 mm and 6.0 mm for CMA and PDA, respectively. No growth was observed on the three solid media at 35 C. However, the fungus grew at 35 C with mean increases in dry weight of 197 mg, 56 mg, and 271 mg, for potato dextrose broth (PDB), corn meal broth (CMB) and palm fruit extract broth (PFEB), respectively. PFEB yielded the highest mycelial dry weight of 80 mg at 15 C. There were signifi cant differences between mycelial dry weight in PFEB and the control at all temperatures, as well as between PFEB and the other media at the same temperature. Palm fruit extract can be used as a substitute for commercial media for in vitro growth of this fungus. Key words: Ceratocystis paradoxa, fruit extract, in vitro growth, Nigeria, oil palm. 52 A. P. O. DEDE AND F. I. OKUNGBOWA INTRODUCTION Oxoid, England) plates. Slants in sterile McCartney bottles were made from a 2-day Ceratocystis paradoxa (Dade)C. Moreau old culture and stored in the refrigerator (5-8 belongs to the Class Ascomycota, C), after incubation for three days at room Order Microascales, and is distributed temperature (28±2 C). From these slants, worldwide. C. paradoxa is parasitic on a subcultures were made on PDA when variety of economic and food crops among required and used for this study. which are Ananas sativa, Cocos nucifera, Preparation of culture media. PDA and Elaeis guineensis, Ipomoea batatas, Musa corn meal agar (CMA, Oxoid, England) sapientum, Phoenix africanus, Sandoricum were prepared following the manufacturer’s indicum, Sorghum vulgare, Theobroma instructions. cacao and Zea mays. In the oil palm, C. Palm fruit extract agar (PFEA) and broth paradoxa is associated with dry basal rot in (PFEB). Two hundred grams of oil palm which it attacks the stem, leaves and fruits, fruits (ElaeisElaeis guineensisguineensis Jacq., Dura variety) causing premature fruit drop4. were boiled until very soft. The fruits were Growth of an organism in culture is then squeezed in water so as to extract the infl uenced by the nutrient composition of oil-rich mesocarp. A volume of 450 ml was the medium and the availability of these obtained. Then 10 g of agar powder was nutrients, as well as other environmental dissolved in 50 ml distilled water, and added factors, such as temperature and inhibitory to the 450 ml palm fruit extract to get a total factors1. Natural materials used to volume of 500 ml in a round-bottomed supplement growth media of in vitro studies fl ask. The 500 ml mixture was sterilized supported good growth of microfungi in in an autoclave at 121 C for 20 min. The earlier work2. With declining resources, it is liquid versions of the solid media were made increasingly diffi cult for mycologists to buy in the same way but without adding agar commercial media for routine laboratory powder. The sterile cooled molten media work in Africa. Comparable alternative were dispensed into sterile (86 mm) plastic sources of media would therefore be useful. petri dishes, while 100 ml of liquid medium The suitability of oil palm fruit extract as was poured into 250 ml conical fl asks, each a media supplement was investigated for treatment in three replicates. growing C. paradoxa in vitro. Inoculation of media. Each plate was point-inoculated at the center with a mycelial disc cut with a sterile 5 mm cork MATERIALS AND METHODS borer. A sterile wire loop was used to transfer mycelial discs to the media. A mycelial disc A fi ve-day old stock culture ofCeratocystis was also dropped in each culture flask. paradoxa (IMI Specimen no. 79166), Water agar plates that did not contain any isolated from an infected oil palm root, nutrients media were inoculated with the was obtained from the culture collection of fungus as control for the solid media, while the Pathology Division, Nigerian Institute distilled water inoculated with the organism for Oil Palm Research (NIFOR), Benin served as control for the liquid media. Each City, Nigeria. Fresh stock cultures were culture fl ask was plugged with sterile cotton prepared from the 5-day old culture by wool wrapped with aluminum foil. Cultures inoculation on potato dextrose agar (PDA, were incubated at room temperature (28±2 MICOL. APL. INT., 19(2), 2007, PP. 51-55 IN VITRO GROWTH OF CERATOCYSTIS PARADOXA 53 C), 15 C, and 35 C, for 72 h, under daylight different solid media. At 28±2 C, the highest alternating with dark. The liquid media growth of 38 mm was recorded for PDA and fl asks were shaken manually by swirling PFEA, while for CMA radial growth was continuously for 2 min, at regular intervals 11 mm and the control 3.6 mm. At 15 C, of 12 h. PFEA supported the highest growth (10.8 Determination of growth. Radial mycelial mm), while PDA and CMA had 6.0 and 3.2 growth of each solid culture was measured mm, respectively, and the control was 2.8 regularly, while mycelial colonies were mm. At 35 C, there was no visible growth. harvested from the liquid media, dried Growth of C. paradoxa at 28±2 C in liquid and weighed, at 24 h intervals. The mean media supplemented with oil palm fruit radial growth and mycelial dry weight for extract (PFEB) yielded 290 mg mycelial dry each medium at different temperatures weight, while in PDB and CMB dry weights were determined. Increase in growth for were 181 and 44 mg, respectively, and 9 each solid medium was determined as mg for the control (Fig. 1). At 15 C, PFEB the difference between one growth radius yielded the highest mycelial dry weight (80 and the previous day’s radius, while for mg), followed by PDB (64 mg), CMB (29 the liquid medium it was calculated as mg), and the control (4 mg), which can be the difference in dry weight of one culture seen in Fig. 2. The fungus grew well at 35C and that of the previous day. Student t-test with dry weights of 197 mg for PDB, 56 mg was used to test the effect of the media for CMB, and 271 mg for PFEB, while dry supplement and temperature on the growth weight in the control was 4 mg (Fig. 3). of the organism. The growth of Ceratocystis paradoxa in the three different liquid media at all temperatures used in the experiment was RESULTS AND DISCUSSION signifi cantly higher when compared with the control. It is not surprising that the fungus Table 1 shows the growth (increase in grew well in PFEA and PFEB, as palm fruit radius) of Ceratocystis paradoxa on three extracts contain suitable amounts of protein, Table 1. Mean radial growth of Ceratocystis paradoxa on different solid media at three different temperatures. Values are means of two independent experiments each replicated three times. Medium Mean radial growth ± SD (mm) 28±2 C 15 C 35 C Potato dextrose agar (PDA) 38.0 ±0.1a 6.0 ±0.4a 0.0 ±0.0 Palm fruit extract agar (PFEA) 38.0 ±0.1a 10.8 ±0.0a 0.0 ±0.0 Corn meal agar (CMA) 11.0 ±0.7b 3.2 ±0.2b 0.0 ±0.0 Control 3.6 ±0.3c 2.8± 0.7b 0.0 ±0.0 SD= Standard deviation of mean of three replicates. Values in the same column with different letters in superscript are signifi cantly different according to Studentt -test (p= 0.05). MICOL. APL. INT., 19(2), 2007, PP. 51-55 54 A. P. O. DEDE AND F. I. OKUNGBOWA difference in growth. In the growth of C. paradoxa in the three liquid media, there was signifi cant difference in growth between 15 C and 28±2 C, but between 28±2 C and 35C there was no signifi cant difference.