Микробные эукариот являются источником фотосинтеза производные углерода и топ хищных видов в постоянно покрытых льдом озер Антарктиды. В настоящем докладе описывается подход обогащение культуры, чтобы изолировать метаболически универсальный эукариот микробных из Антарктики озера, озера Бонни, и оценивает потенциал неорганических фиксации углерода помощью радиоизотопного анализа для рибулозо-1 ,5-bisphophate карбоксилазы оксигеназы (RUBISCO) деятельности.
Lake Bonney is one of numerous permanently ice-covered lakes located in the McMurdo Dry Valleys, Antarctica. The perennial ice cover maintains a chemically stratified water column and unlike other inland bodies of water, largely prevents external input of carbon and nutrients from streams. Biota are exposed to numerous environmental stresses, including year-round severe nutrient deficiency, low temperatures, extreme shade, hypersalinity, and 24-hour darkness during the winter 1. These extreme environmental conditions limit the biota in Lake Bonney almost exclusively to microorganisms 2.
Single-celled microbial eukaryotes (called “protists”) are important players in global biogeochemical cycling 3 and play important ecological roles in the cycling of carbon in the dry valley lakes, occupying both primary and tertiary roles in the aquatic food web. In the dry valley aquatic food web, protists that fix inorganic carbon (autotrophy) are the major producers of organic carbon for organotrophic organisms 4, 2. Phagotrophic or heterotrophic protists capable of ingesting bacteria and smaller protists act as the top predators in the food web 5. Last, an unknown proportion of the protist population is capable of combined mixotrophic metabolism 6, 7. Mixotrophy in protists involves the ability to combine photosynthetic capability with phagotrophic ingestion of prey microorganisms. This form of mixotrophy differs from mixotrophic metabolism in bacterial species, which generally involves uptake dissolved carbon molecules. There are currently very few protist isolates from permanently ice-capped polar lakes, and studies of protist diversity and ecology in this extreme environment have been limited 8, 4, 9, 10, 5. A better understanding of protist metabolic versatility in the simple dry valley lake food web will aid in the development of models for the role of protists in the global carbon cycle.
We employed an enrichment culture approach to isolate potentially phototrophic and mixotrophic protists from Lake Bonney. Sampling depths in the water column were chosen based on the location of primary production maxima and protist phylogenetic diversity 4, 11, as well as variability in major abiotic factors affecting protist trophic modes: shallow sampling depths are limited for major nutrients, while deeper sampling depths are limited by light availability. In addition, lake water samples were supplemented with multiple types of growth media to promote the growth of a variety of phototrophic organisms.
RubisCO catalyzes the rate limiting step in the Calvin Benson Bassham (CBB) cycle, the major pathway by which autotrophic organisms fix inorganic carbon and provide organic carbon for higher trophic levels in aquatic and terrestrial food webs 12. In this study, we applied a radioisotope assay modified for filtered samples 13 to monitor maximum carboxylase activity as a proxy for carbon fixation potential and metabolic versatility in the Lake Bonney enrichment cultures.
Последние молекулярные исследования показали высокое разнообразие одноклеточных эукариот в целом ряде сред 3, 19, 20, однако из-за отсутствия изолятов по всему кругу protist обитания функциональной роли этих отдельных видов в пищевых цепях в основном неизвестно. В этом исследовании, м…
The authors have nothing to disclose.
Авторы выражают благодарность И. Priscu, А. Chiuchiolo и Мак-Мердо LTER лимнологии команде за помощь в сборе и сохранении образцов в Антарктиде. Мы благодарим Ratheon Полярный Услуги и PHI вертолетов для материально-технического обеспечения. Свет снимках были получены в центре Майами перспективных микроскопии и обработки изображений центра. Эта работа была поддержана NSF Управления Полярный грантов Программы 0631659 и 1056396.
Name of the reagent | Company | Catalogue number | Comments |
BBM | Sigma | B5282 | |
BG11 | Sigma | C3061 | |
F/2 | Sigma | G9903 | |
GF/F filter, 25 mm | Fisher Scientific | 09-874-64 | |
GF/F filter, 47 mm | Fisher Scientific | 09-874-71 | |
Polyethersulfone filter, 0.45 μm pore, 47 mm | Pall Life Sciences | 61854 | |
Sterile cell culture flask, 25 cm2 | Corning | 430639 | |
Diurnal growth chamber | VWR | 35960-076 | |
Zirconia/silica beads, 0.1 mm diamter | BioSpec Products | 11079101z | |
Mini-Bead beater | BioSpec Products | 3110BX | |
Screw-cap microcentrifuge tube (1.5 μL) | USA Scientific | 1415-8700 | |
NaH14CO3 | ViTrax | VC 194 | Keep in aliquots of 400 μL at -20°C |
RuBP | Sigma | R0878-100mg | Dissolve in 10 mM Tris-propionic acid (pH 6.5) |