Ester Linked Fatty Acid (ELFA) method should be used with caution for interpretating soil microbial communities and their relationships with environmental variables in forest soils

Abstract

As an alternative for phospholipid fatty acid (PLFA) analysis, a simpler ester linked fatty acid (ELFA) analysis has been developed to characterize soil microbial communities. However, few studies have compared the two methods in forest soils where the contribution of nonmicrobial sources may be larger than that of microbial sources. Moreover, it remains unclear whether the two methods yield similar relationships of microbial biomass and composition with environmental variables. Here, we compared PLFA and ELFA methods with respect to microbial biomass and composition and their relationships with environmental variables in six oriental oak (Quercus variabilis) forest sites along a 1500-km latitudinal gradient in East China. We found that both methods had a low sample-to-sample variability and successfully separated overall community composition of sites. However, total, bacterial, and fungal biomass, the fungal-to-bacterial ratio, and the gram-positive to gram-negative bacteria ratio were not significantly or strongly correlated between the two methods. The relationships of these microbial properties with environmental variables (pH, precipitation, and clay) greatly differed between the two methods. Our study indicates that despite its simplicity, the ELFA method may not be as feasible as the PLFA method for investigating microbial biomass and composition and for identifying their dominant environmental drivers, at least in forest soils.

A comparison among EL-FAME, PLFA, and quantitative PCR methods to detect changes in the abundance of soil bacteria and fungi

“…It may be argued that the good correlation we found between EL-FAME and PLFA data is because phospholipids were the dominant lipid fraction in our EL-FAME analysis; however, across all the samples in our experiment, total, bacterial, and fungal PLFAs only represented ∼27, 38, and 15 % of the EL-FAMEs, respectively. These values are a bit lower than those found in other studies (Miura et al, 2017; Li et al, 2020; Yu et al, 2021), although the variability between studies is high, which is explained by interlaboratory differences in protocols and analytical conditions.…”

“…This could favour the dispersal of soil colloids and increase the extraction of fatty acids from plant material and humified soil organic matter, interfering with the quantification of real amounts of bacteria and fungi (Li et al, 2020). The lower resolution that the EL-FAME method has in the quantification of bacteria and fungi in forest soils with respect to the PLFA approach has previously been reported (Yu et al, 2021). Despite this, it is important to highlight that, in our study, the EL-FAME data significantly correlated with the PLFA and qPCR data and that the EL-FAME method was able to successfully capture the impact that both nutrients and antibiotics produced on soil microbial abundances.…”

“…Studies comparing EL-FAME, PLFA, and qPCR methods are useful to disentangle their reliability for quantifying microbial abundance. Previous studies, such as those of Hinojosa et al (2005), Miura et al (2017), Li et al (2020), and Yu et al (2021) have compared EL-FAME- and PLFA-based abundances of bacteria and fungi in soils from different ecosystems and under different degrees of anthropogenetic influence, concluding that although both approaches provide consistent results, PLFA data seem to be more sensitive to reflect environmental changes. The EL-FAME and qPCR methods were purposely compared by Pérez-Guzmán et al (2021), and the PLFA and qPCR approaches by Baldrian et al (2013), Osburn et al (2022), and Zhang et al (2017), among others, demonstrating that, in general, fatty acid-based methods seem to be more reliable than qPCR, although the reliability differences depended on the type of soil and the factor driving the changes in microbial abundance.…”