Measurement error and resolution in quantitative stable isotope probing: implications for experimental design

Published by Ecoss on

Quantitative stable isotope probing (qSIP) estimates the degree of incorporation of an isotope tracer into nucleic acids of metabolically active organisms and can be applied to microorganisms growing in complex communities, such as the microbiomes of soil or water. As such, qSIP has the potential to link microbial biodiversity and biogeochemistry. As with any technique involving quantitative estimation, qSIP involves measurement error; a more complete understanding of error, precision and statistical power will aid in the design of qSIP experiments and interpretation of qSIP data. We used several existing qSIP datasets of microbial communities found in soil and water to evaluate how variance in the estimate of isotope incorporation depends on organism abundance and on the resolution of the density fractionation scheme. We also assessed statistical power for replicated qSIP studies, and sensitivity and specificity for unreplicated designs. We found that variance declines as taxon abundance increases. Increasing the number of density fractions reduces variance, although the benefit of added fractions declines as the number of fractions increases. Specifically, nine fractions appear to be a reasonable tradeoff between cost and precision for most qSIP applications. Increasing replication improves power and reduces the minimum detectable threshold for inferring isotope uptake to 5 atom%. Finally, we provide evidence for the importance of internal standards to calibrate the %GC to mean weighted density regression per sample. These results should benefit those designing future SIP experiments, and provide a reference for metagenomic SIP applications where financial and computational limitations constrain experimental scope.Importance One of the biggest challenges in microbial ecology is correlating the identity of microorganisms with the roles they fulfill in natural environmental systems. Studies of microbes in pure culture reveal much about genomic content and potential functions, but may not reflect an organism’s activity within its natural community. Culture-independent studies supply a community-wide view of composition and function in the context of community interactions, but fail to link the two. Quantitative stable isotope probing (qSIP) is a method that can link the identity and function of specific microbes within a naturally occurring community. Here we explore how the resolution of density-gradient fractionation affects the error and precision of qSIP results, how they may be improved via additional replication, and cost-benefit balanced scenarios for SIP experimental design.

Categories:

Related Posts

A CIRCUIT SHOWS HOW CLIMATE WARMING MAY ALTER DIFFERENT ECOLOGICAL LEGACIES IN THE ARCTIC BOREAL FOREST, WHICH THEN FEEDBACK INTO CLIMATE WARMING AND CHANGES TO LANDSCAPE PROCESSES.

NAU-led research team receives $9.6M to study how Alaska’s forests change, adapt to warmer future

Ecological memory stored in a landscape can help an ecosystem recover from disturbances like fire and outbreaks of disease. But what happens when climate warming disrupts that process? How long before ecological memories stored in Read more…

A SCATTERPLOT DEPICTS HOW MUCH CARBON OR OXYGEN ISOTOPE DIFFERENT SOIL BACTERIA TOOK INTO THEIR DNA (A PROXY FOR GROWTH) DURING A 7-DAY EXPERIMENT. SMALL POINTS REPRESENT THE GROWTH OF INDIVIDUAL BACTERIAL STRAINS LINKED BY LINES TO THE AVERAGE GROWTH OF THEIR CLOSEST IDENTIFIED GENUS. COLORS WERE ASSIGNED BY A CLUSTERING ALGORITHM TO GROUP GENERA INTO FAST-GROWING OR SLOW-GROWING LIFESTYLES.

Lifestyles of the fast and slow (bacteria): In the wild, most live in the slow lane

A study led by Northern Arizona University offers new evidence that a common framework to sort bacteria into two lifestyles doesn’t easily apply to bacteria living in wild soil. The findings, published in The ISME Journal, show Read more…

A trio of illustrated panels depicts the soil microbial communities in three treatments of a long-term warming experiment north of Flagstaff, Arizona. The panels are labeled in gray text, left to right: control, short-term warming, long-term warming. The control treatment, far left, depicts bacteria, protists, and other microorganisms identified by sequencing the soil’s metagenome in yellow, orange, green, and blue, along with chains of available nutrients. Soil depicted in the center panel, which has undergone short-term warming, shows an increased abundance and diversity of microbes, some of which are growing, incorporating nutrients, and respiring carbon dioxide. The soil that has undergone long-term warming, imagined in the panel on the right, shows fewer microbial species and competitive relationships that are developing for control of the scarcer nutrients.

Does a warmer future favor microbial friend or foe? Ecoss researchers win $3.4M to study interactions in changing soil

In 2002, the Odyssey probe discovered evidence of past ice on Mars. The U.S. Congress authorized the Iraq War resolution. The Anaheim Angels won the World Series. And in a meadow 15 miles north of Read more…