Lyophilization as a method for pathogens long term preservation

Abstract

Lyophilization (freeze-drying) is one of the most suitable methods used for a long term preservation of pathogens. The aim of this paper was the application of lyophilization for storage of three significant plant pathogens: Fusarium graminearum, Helminthosporium gramineum, and Pseudomonas syringae pv. gylicinea, respectively. The plant material was collected continuously (during a four year period 2002-2006), depending on a plant development stage, from different localities in Vojvodina. Pathogens were isolated from diseased parts with characteristic symptoms, and placed on nutritive media specific for a certain pathogen, using standard phytopathological methods. Lyophilization was carried out in marked and coded ampoules by freezing and drying of pathogen suspension and nutritive medium. Revitalization of lyophilized isolates was done after four days. High percentage of revitalization was characteristic for all studied isolates, and it ranged from 85-92%, confirming that lyophilized pathogens would be capable of keeping viability for a long time in the collection. Besides above mentioned pathogens, there were 200 isolates in the collection, originating mostly from field and vegetable crops. Each isolate that was put into the Collection, was followed by all the necessary data such as: name of the pathogen, number of isolates, locality, host plant year of isolation, name of the researcher and other relevant data.Liofilizacija (freeze-drying) je jedna od najpogodnijih metoda koja se koristi za dugorocno cuvanje patogena. Cilj rada je primena metoda liofilizacije u cuvanju tri znacajna biljna patogena: Fusarium graminearum Helminthosporium gramineum i Pseudomonas syringae pv. gylicinea. Prikupljanje uzoraka biljnog materijala vrseno je u kontinuitetu (tokom cetiri godine, 2002-2006) sa razlicitih lokaliteta na podrucju Vojvodine, u zavisnosti od razvojne faze biljaka. Izolacija patogena vrsena je iz obolelih biljnih delova sa karakteristicnim simptomima na hranljive podloge specificne za odredjenog patogena koriscenjem standardnih fitopatoloskih metoda. Postupak liofilizacije vrsen je u obelezenim i sifriranim ampulama smrzavanjem i susenjem suspenzije patogena i hranljivog medijuma. Revitalizacija liofilizovanih izolata izvrsena je cetiri dana nakon liofilizacije. Za sve proucene izolate karakteristican je visok procenat revitalizacije i iznosi 85-92%, sto potvrdjuje da ce liofilizovani patogeni dugorocno zadrzati vitalnost u kolekciji. U okviru kolekcije pored navedenih patogena postoji 200 izolata koji poticu uglavnom sa ratarskih i povrtarskih biljnih vrsta. Svaki izolat koji se nalazi u kolekciji unet je u bazu podataka sa svim potrebnim podacima, kao sto su: naziv patogena, broj izolata, lokalitet, biljka domacin, godina izolacije, ime istrazivaca i drugi relevantni podaci.