Peak selection matters in principal component analysis: A case study of syntrophic microbes

Abstract

In situ liquid time-of-flight secondary ion mass spectrometry (ToF-SIMS) is a powerful technique to study surface characterization of living biofilms in hydrated conditions. However, ToF-SIMS data analysis is still a great challenge in complicated bacterial biofilms, because many interference peaks from the medium may result in inaccurate interpretation. In this study, two syntrophic Geobacter populations are investigated using in situ liquid ToF-SIMS to reveal the biofilm surface changes between them due to direct interspecies electron transfer. By comparing spectral principal component analysis (PCA) results of all peaks and selected peaks, the authors find that spectral peak overlay is an effective strategy to reduce the matrix effect in handling complex ToF-SIMS data. Additionally, the spectral PCA results of high intensity and high resolution data obtained from liquid ToF-SIMS are compared. Selected peaks, amino acid peaks, and water cluster peaks spectral PCA produce nice separation among samples in both high intensity and high resolution data sets. However, the high resolution data show better separation between coculture planktonic and coculture aggregates, confirming that the higher mass accuracy is useful in the analysis of microbial samples. In conclusion, the results show that peak selection is critical for acquiring effective microbial information and interpretation of syntrophic Geobacter using spectral data from in situ liquid ToF-SIMS.

In Vivo Molecular Insights into Syntrophic Geobacter Aggregates

“…Among the detected biological signals, we are most interested in those that differ between coculture aggregates and other samples. Therefore, several rounds of PCA were conducted using the peak spectral overlay strategy for peak selection, with the goal of removing the maximal numbers of interference peaks derived from two kinds of medium salts (Figure S5a–d). When compared with all peak PCA, the medium effects were reduced in selected peak PCA .…”

“…Therefore, several rounds of PCA were conducted using the peak spectral overlay strategy for peak selection, with the goal of removing the maximal numbers of interference peaks derived from two kinds of medium salts (Figure S5a–d). When compared with all peak PCA, the medium effects were reduced in selected peak PCA . Selected peak PCA in the positive and negation modes (Figures , , and S6a,b) show that the coculture aggregates differ from various control samples, including coculture planktonic cells, pure cultured biofilms, pure cultured planktonic cells, and medium solutions, indicating that a unique molecular composition was formed in these syntrophic communities.…”

“…When compared with all peak PCA, the medium effects were reduced in selected peak PCA. 23 Selected peak PCA in the positive and negation modes (Figures 2, 3, and S6a,b) show that the coculture aggregates differ from various control samples, including coculture planktonic cells, pure cultured biofilms, pure cultured planktonic cells, and medium solutions, indicating that a unique molecular composition was formed in these syntrophic communities. Combined with the respective loadings plots, peaks that account for those discriminations were revealed.…”

“…Spectral Principal Component Analysis (PCA) was conducted with selected peaks that derived from spectral overlay. 23 Aromatic amino acids and polymers relevant to electrical conductance were revealed to be more abundant in coculture aggregates, indicating that DIET was more prevalent in the interface of the dense syntrophic communities. In addition, phosphatidylethanolamine (PE) relevant fragments were observed with higher abundance in coculture aggregates.…”

“…Unlike the bulk information obtained from LC–MS/MS or denatured structural information on dried samples acquired by traditional static SIMS, surface characteristic biological molecules from living cells were observed in this study, including amino acids, lipids, polymers, and water clusters. Spectral Principal Component Analysis (PCA) was conducted with selected peaks that derived from spectral overlay . Aromatic amino acids and polymers relevant to electrical conductance were revealed to be more abundant in coculture aggregates, indicating that DIET was more prevalent in the interface of the dense syntrophic communities.…”