Abstract
BackgroundTissue culture has been viewed as a relatively safe method of transporting conifer germplasm as the risk of pathogens associated with tissue cultured material has been assumed to be low. Despite these assumptions, it is unknown whether tissue cultured propagation material contains microorganisms or whether such microorganisms could grow out onto the media used.MethodsTo determine whether asymptomatic tissue culture material contains fungal microorganisms, 30 different Pinus radiata D.Don genotypes from four different types of tissue culture material were analysed for the presence of fungal DNA or mycelium. In addition, thirteen fungal and oomycete isolates were cultured on standard tissue propagation media.ResultsFungal DNA was detected in all samples tested. No fungal or bacterial microorganisms were able to be grown from any of the tissue culture material tested. However, confocal microscopy showed the presence of filaments that could have been fungal mycelium. Growth of thirteen fungal and oomycete isolates on standard tissue propagation media indicated these media can severely limit the growth of some of these microorganisms.ConclusionThe results from this study show that tissue culture material does contain asymptomatic fungal microorganisms but the exact risk from these microorganisms is unclear.
Highlights
Tissue culture has been viewed as a relatively safe method of transporting conifer germplasm as the risk of pathogens associated with tissue cultured material has been assumed to be low
Tissue culture preparation Four types of tissue culture material were prepared from Pinus radiata: embryogenic tissue derived from immature zygotic embryos; adventitious-axillary organogenesis shoots derived from cotyledons dissected from mature zygotic embryos; epicotyl-axillary organogenesis shoots derived from the epicotyls of mature zygotic embryos; and mature organogenesis shoots derived from field grown material and established as in-vitro shoot cultures
Growth on general fungal media No fungal or bacterial growth was observed from any of the tissue culture material plated onto the four general fungal media (PDA, malt extract agar (MEA), carrot agar (CA) and prune agar (PA)) tested
Summary
Tissue culture has been viewed as a relatively safe method of transporting conifer germplasm as the risk of pathogens associated with tissue cultured material has been assumed to be low. Despite these assumptions, it is unknown whether tissue cultured propagation material contains microorganisms or whether such microorganisms could grow out onto the media used. In an effort to find alternative methods to import new Pinus spp. or Pseudotsuga menziesii material into New Zealand, the importation of live material in tissue culture has been suggested. Tissue culture has been viewed as a relatively safe method of transporting conifer germplasm for a variety of reasons. The likelihood that fungi could survive the intensive nature of many currently used tissue culture procedures is thought to be low
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