Metabolomics Reveals Strain-Specific Cyanopeptide Profiles and Their Production Dynamics in Microcystis aeruginosa and M. flos-aquae

Abstract

Cyanobacterial blooms that release biologically active metabolites into the environment are increasing in frequency as a result of the degradation of freshwater ecosystems globally. The microcystins are one group of cyanopeptides that are extensively studied and included in water quality risk management frameworks. Common bloom-forming cyanobacteria produce incredibly diverse mixtures of other cyanopeptides; however, data on the abundance, distribution, and biological activities of non-microcystin cyanopeptides are limited. We used non-targeted LC-MS/MS metabolomics to study the cyanopeptide profiles of five Microcystis strains: four M. aeruginosa and one M. flos-aquae. Multivariate analysis and GNPS molecular networking demonstrated that each Microcystis strain produced a unique mixture of cyanopeptides. In total, 82 cyanopeptides from the cyanopeptolin (n = 23), microviridin (n = 18), microginin (n = 12), cyanobactin (n = 14), anabaenopeptin (n = 6), aeruginosin (n = 5), and microcystin (n = 4) classes were detected. Microcystin diversity was low compared with the other detected cyanopeptide classes. Based on surveys of the literature and spectral databases, most cyanopeptides represented new structures. To identify growth conditions yielding high amounts of multiple cyanopeptide groups, we next examined strain-specific cyanopeptide co-production dynamics for four of the studied Microcystis strains. When strains were cultivated in two common Microcystis growth media (BG-11 and MA), the qualitative cyanopeptides profiles remained unchanged throughout the growth cycle. For each of the cyanopeptide groups considered, the highest relative cyanopeptide amounts were observed in the mid-exponential growth phase. The outcomes of this study will guide the cultivation of strains producing common and abundant cyanopeptides contaminating freshwater ecosystems. The synchronous production of each cyanopeptide group by Microcystis highlights the need to make more cyanopeptide reference materials available to investigate their distributions and biological functions.

Temporal Dynamics of Cyanobacterial Bloom Community Composition and Toxin Production from Urban Lakes

“…Microcystin-LR identified in our environmental samples had an identical retention time, precursor m / z and MS/MS fragmentation pattern to that of the MC-LR identified in the UTEX positive control (Figure S11). Additional toxins were identified by examining MS/MS networks (e.g., anabaenopeptin B in same cluster as anabaenopeptin A) and by comparing precursor m / z values and MS/MS spectra to previously published work. − Toxin concentrations (as determined by AUC values from mass spec data) coincided with cyanobacterial biomass maxima at all three lakes (Figures and S12A) and toxin production was not constitutive. Additionally, metabolite content in general increased with cyanobacterial biomass (Figure S12B).…”

Cyanopeptolins and Anabaenopeptins Are the Dominant Cyanopeptides from Planktothrix Strains Collected in Canadian Lakes

“…Cluster 11 shows microcystins produced by all strains, excluding P. rubescens CPCC 733. This cluster contains (D-Asp 3 , ( E )-Dhb 7 ) MC-HtyR and (D-Asp 3 , ( E )-Dhb 7 ) MC-HphR, which were previously characterized from P. rubescens CPCC 507 as well as with [Asp 3 , Dha 7 ] MC-LR, which was previously identified from M. aeruginosa CPCC 300 [ 30 , 31 ]. Cluster 12 comprises later-eluting, retention time 5.10–5.58 min, microcystins exclusively produced by P. rubescens CPCC 733.…”

“…All aeruginosins were confirmed by characteristic Choi product ions at m / z 140.1064 and 122.0962 [ 33 ]. Two aeruginosins, 688A and 722, from the seed spectra of M. aeruginosa CPCC 632 aided in the identification of the aeruginosin cluster [ 30 ]. Three additional aeruginosins were identified by manual interpretation of the data: AG 660A, AG 672 and AG 688B.…”

“…Intracellular cyanopeptide extracts were solubilized in 1 mL of 90% aqueous methanol and vortexed for 15 s prior to LC-MS/MS analysis. Non-targeted data were acquired with an Agilent 1290 HPLC (Agilent Technologies, Santa Clara, CA, USA) coupled to a Q-Exactive Orbitrap (Thermo Fisher Scientific, Waltham, MA, USA) mass spectrometer [ 30 ]. Extracts were resolved using an Eclipse Plus C18 RRHD column (2.1 × 100 mm, 1.8 µm; Agilent Technologies) maintained at 35 °C and a 10.5 min gradient program optimized for cyanopeptides.…”