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Genotyping and Evaluation of Pleurotus Ostreatus (Oyster Mushroom) Strains

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Genotyping and Evaluation of Pleurotus Ostreatus (Oyster Mushroom) Strains Genotyping and Evaluation of Pleurotus ostreatus (oyster mushroom) strains Anton S.M. Sonnenberg, Patrick M. Hendricks and Etty Sumiati Applied Plant Research Mushroom Research Unit PPO no. 2005-18 December 2005 © 2005 Wageningen, Applied Plant Research (Praktijkonderzoek Plant & Omgeving B.V.) All rigts reserved. No part of this publication may be reproduced, stored in a retrieval system or transmitted, i n any form of by any means, electronic, mechanical, photocopying, recording or otherwise, without the prior written permission of Applied Plant Research. Praktijkonderzoek Plant & Omgeving takes no responsibility for any injury or damage sustained by using data from this publication. PPO Publication no. 2005-18 This study was financed by the Ministry of Agriculture, Nature and Food Quality Project no.: 620154 Applied Plant Research ( Praktijkonderzoek Plant & Omgeving BV) Mushroom Research Unit Adress : Peelheideweg 1, America, The Netherlands : PO Box 6042, 5960 AA Horst, The Netherlands Tel. : +31 77 4647575 Fax : +31 77 464 1567 E-mail : [email protected] Internet : www.ppo.wur.nl © Praktijkonderzoek Plant & Omgeving B.V. 2 Inhoudsopgave pagina 1 SUMMARY.................................................................................................................................... 4 2 INTRODUCTION ............................................................................................................................ 4 3 GENOTYPING COLLECTIONS ......................................................................................................... 5 4 FRUITING TESTS........................................................................................................................... 6 4.1 Yield ..................................................................................................................................... 6 4.2 Colour................................................................................................................................... 6 4.3 Quality .................................................................................................................................. 7 5 CONCLUSIONS............................................................................................................................. 7 6 TABLES AND FIGURES …………………… ………………………….…………………………………8 © Praktijkonderzoek Plant & Omgeving B.V. 3 1 Summary Oyster mushrooms (Pleurotus ostreatus) represent one of the most popular mushroom species grown in Indonesia. There is a need for strains that are better adapted to the climate conditions at Java, where most mushrooms in Indonesia are produced. Strains that can produce mushrooms at 22 to 28 oC and have a good yield and quality would improve the profit of mushroom growers substantially. In order to find strains that can be used for Indonesian growing conditions a large number of available strains were genetically examined. Two sources were used in this study: - A collection of strains commercially used in Indonesia. These strains are collected by Dr Etty Sumiati from farmers and institutes at Java. - The collection of fungal strains present at PPO, Horst, The Netherlands. This collection contains a substantial number of strains representing the genus Pleurotus. Most of these strains have not been genotyped or evaluated for the production of fruiting bodies. Unfortunately, no strains were obtained from natural environment in Indonesia. Most strains sent by Etty Sumiati appeared to be infected at arrival at PPO. We have identified the species type of the strains representing the genus Pleurotus by ITS-RFLP in order to identify unambiguously the species P. ostreatus (common oyster mushroom). Next, all P. ostreatus strains were genotyped by ISSR to examine the genetic variation between strains. It appeared that the PPO collection contains 167 P. ostreatus strains representing 68 different genotypes. The 24 strains collected from Indonesia represent 7 different genotypes but more than 50% is represented by one variety. This variety, i.e. type florida, originate from wild (Florida, USA) and can be grown at higher temperatures. It lacks, however, the yield and quality present in most strains used in Europe and the USA. Based on the genetic data and data on the origin of strains a selection of 40 strains was made that has been tested in 2 parallel cells (16 and 22 oC production temperature). ……….. 2 Introduction The oyster mushroom (Pleurotus) is one of the most cultivated mushroom types worldwide. The genus consists of a number of species. However, more than 90% of the Pleurotus crops grown world wide concerns the common oyster mushroom, P. ostreatus. Its ease of cultivation on different substrates makes it a popular species for small farmers. No expensive climate controlled growing houses are needed because of the availability of varieties that can complete their life cycle under different climatic conditions. The common oyster mushroom is also one of the most cultivated mushroom species in Indonesia, especially on Java. It is grown inland on elevated regions (1000 or more meters above sea level) where temperatures do not exceed 22-28 0C. In Europe and the USA, oyster mushroom varieties are grown at low temperatures (fruiting bodies are produced at temperatures of 15 to 18 oC). Mushroom farmers obtain high quality strains from companies specialized in the production of spawn (pure fungal inoculum for substrate). A large number of strains are available but previous research has shown that they represent only a limited number of genotypes. There are no spawn companies in Indonesia and import of spawn is too expensive for most growers. Strains are usually obtained by taking along strains from contacts in Western countries (or recently China and Taiwan) and they are stored on slant tubes in a refrigerator or even at room temperature. This is not an optimal way of maintaining strains often resulting in decrease of quality after some time. Next to strain qualities, also a lack of varieties © Praktijkonderzoek Plant & Omgeving B.V. 4 specially adapted to the Indonesian growing/climate conditions is a problem. There is only one commercial variety available (“florida” variety) that is adapted to cultivation at higher temperatures. The goal of this research is to see if strains can be found that are more suitable for Indonesian cultivation conditions, i.e. can produce fruiting bodies above 22 oC and have a good yield and quality. Two types of sources for strains are used in this study: the collection of PPO and the collection of commercial strains used in Indonesia (collected by IVEGRI at Lembang). Since space and money is limited in this project, we first genotypes the available Pleurotus species and identified all P. ostreatus strain. Subsequently, the genetically variation within the species P. ostreatus was determined. A selection of 40 genetically different strains were tested on fruiting in two parallel cell test at 16 and 22 oC respectively. 3 Genotyping Collections The collection of PPO contains ca. 5000 strains representing more than 100 species. Approximately 270 strains are Pleurotus species (including 24 strains sent by Etty Sumiati/IVEGRI). Most of these are the widely cultivated common oyster mushroom P. ostreatus. The collection P.ostreatus of PPO consists of old and present-day commercial lines, gifts from other collections and private persons and strains collected from natural environment in different parts in the world. Especially in the last years, a number of strains have been obtained from China and Korea. Most strains of this collection have not been genotyped and not evaluated in a crop. Dr Etty Sumiati has collected 31 fungal strains from different sources in Indonesia of species that are grown commercially. Most of these are P. ostreatus species. She also has collected a number of strains from the wild, mainly in Java. Unfortunately, most of these strains appeared to be infected after arrival at PPO (those that were purified were not Pleurotus species). Since this subproject focus now on P. ostreatus we have first checked on the correct species name. All Pleurotus spp. were genotyped using ITS-RFLP as described earlier (Report: “Evaluation of shiitake strains of the PPO collection”; Hortin Mushroom 2005-1). Four different restriction enzymes were used to digest the amplified ribosomal DNA region. In this way all species could be discriminated into the four groups in accordance with intersterility groups (Appendix: Table 1; Figure 1). The latter name is derived from the compatibility test between strains. Strains from identical species are compatible (fertile) whereas strains from different species can not be crossed (intersterile). In table 2 (Appendix) a list is presented of commercial strains collected by Dr Etty Sumiati from different farms and institutes in Java. All these strains were identified as P. ostreatus by ITS-RFLP. In order to evaluate what strains have potentials for the Indonesian cultivation conditions we tested strains at 2 different fruiting temperatures, 16 and 22 oC. In this way we could also see what strains have potentials for the European and USA market or what strains could be a base for a breeding program for both climatic conditions. Since we have a limited space in our experimental farm and the budget is also limited we had to make a selection of the 168 identified as P. ostreatus species. For this, all strains were first analyzed on genetic relatedness using the ISSR technique as described earlier (Report: “Evaluation
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