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1.
Highly specific qPCR and amplicon sequencing method for detection of quarantine citrus pathogen Phyllosticta citricarpaapplicable for air samples
Janja Zajc, Zala Kogej Zwitter, Sara Fišer, Cene Gostinčar, Antonio Vicent, Anaïs Galvañ Domenech, Luca Riccioni, Neil Boonham, Maja Ravnikar, Polona Kogovšek, 2023, izvirni znanstveni članek

Povzetek: The fungus Phyllosticta citricarpa is a quarantine pathogen in the EU and is of high economic importance in many parts of the world where favourable climate conditions drive the development of citrus black spot (CBS) disease. Disease symptoms include necrotic lesions on leaves and fruits. Low disease pressure can reduce crop market-ability, while higher disease pressure can cause premature fruit drop, significantly increasing crop losses. The wind-dispersed spores of P. citricarpa are especially prob-lematic for rapid pathogen dispersal, but also provide an opportunity for early detec-tion of the disease spreading into a new area. In this study we have developed and validated a quantitative PCR (qPCR) assay based on the TEF1-α sequence. Specificity testing demonstrated that it is currently the only qPCR assay that does not cross- react with closely related Phyllosticta species. The assay is sensitive and can detect a single copy of the TEF1 gene in a reaction, it is highly repeatable and reproducible and can be used for testing of the sticky tapes from spore traps as well as citrus fruit sam-ples. High-throughput sequencing (HTS) of the DNA barcodes ITS1 and TEF1 was also explored for the detection and discrimination of P. citricarpa. The limit of detection of the HTS was 1000 spores on a daily spore trap tape. This study makes an important improvement to the diagnostics of the CBS and the methods developed can also be applied to improve the surveillance and early detection of the pathogen when linked to spore samplers in the field.
Ključne besede: detection, fungal spore sampling, internal transcribed region (ITS), translation elongation factor 1-α (TEF1)
Objavljeno v DiRROS: 12.07.2024; Ogledov: 21; Prenosov: 20
.pdf Celotno besedilo (1,49 MB)
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2.
Effects of desiccation and freezing on microbial ionizing radiation survivability : considerations for Mars sample return
William H. Horne, Tine Grebenc, Rok Tkavc, Cene Gostinčar, Nina Gunde-Cimerman, 2022, izvirni znanstveni članek

Povzetek: Increasingly, national space agencies are expanding their goals to include Mars exploration with sample return. To better protect Earth and its biosphere from potential extraterrestrial sources of contamination, as set forth in the Outer Space Treaty of 1967, international efforts to develop planetary protection measures strive to understand the danger of cross-contamination processes in Mars sample return missions. We aim to better understand the impact of the martian surface on microbial dormancy and survivability. Radiation resistance of microbes is a key parameter in considering survivability of microbes over geologic times on the frigid, arid surface of Mars that is bombarded by solar and galactic cosmic radiation. We tested the influence of desiccation and freezing on the ionizing radiation survival of six model microorganisms: vegetative cells of two bacteria (Deinococcus radiodurans, Escherichia coli) and a strain of budding yeast (Saccharomyces cerevisiae); and vegetative cells and endospores of three Bacillus bacteria (B. subtilis, B. megaterium, B. thuringiensis). Desiccation and freezing greatly increased radiation survival of vegetative polyploid microorganisms when applied separately, and when combined, desiccation and freezing increased radiation survival even more so. Thus, the radiation survival threshold of polyploid D. radiodurans cells can be extended from the already high value of 25 kGy in liquid culture to an astonishing 140 kGy when the cells are both desiccated and frozen. However, such synergistic radioprotective effects of desiccation and freezing were not observed in monogenomic or digenomic Bacillus cells and endospores, which are generally sterilized by 12 kGy. This difference is associated with a critical requirement for survivability under radiation, that is, repair of genome damage caused by radiation. Deinococcus radiodurans and S. cerevisiae accumulate similarly high levels of the Mn antioxidants that are required for extreme radiation resistance, as do endospores, though they greatly exceed spores in radioresistance because they contain multiple identical genome copies, which in D. radiodurans are joined by persistent Holliday junctions. We estimate ionizing radiation survival limits of polyploid DNA-based life-forms to be hundreds of millions of years of background radiation while buried in the martian subsurface. Our findings imply that forward contamination of Mars will essentially be permanent, and backward contamination is a possibility if life ever existed on Mars.
Ključne besede: ionizing radiation, life on Mars, astrobiology, radiation, fungi, bacteria, radiotolerance
Objavljeno v DiRROS: 03.11.2022; Ogledov: 716; Prenosov: 337
.pdf Celotno besedilo (922,44 KB)
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