CYANOCHIP: An Antibody Microarray for High-Taxonomical-Resolution Cyanobacterial Monitoring

Abstract

Cyanobacteria are Gram-negative photosynthetic prokaryotes that are widespread on Earth. Eutrophication and global warming make some aquatic ecosystems behave as bioreactors that trigger rapid and massive cyanobacterial growth with remarkable economic and health consequences. Rapid and efficient early warning systems are required to support decisions by water body authorities. We have produced 17 specific antibodies to the most frequent cyanobacterial strains blooming in freshwater ecosystems, some of which are toxin producers. A sandwich-type antibody microarray immunoassay (CYANOCHIP) was developed for the simultaneous testing of any of the 17 strains, or other closely related strains, in field samples from different habitats (water, rocks, and sediments). We titrated and tested all of the antibodies in succession using a fluorescent sandwich microarray immunoassay. Although most showed high specificity, we applied a deconvolution method based on graph theory to disentangle the few existing cross-reactions. The CYANOCHIP sensitivity ranged from 10(2) to 10(4) cells mL(-1), with most antibodies detecting approximately 10(2) cells mL(-1). We validated the system by testing multiple isolates and crude natural samples from freshwater reservoirs and rocks, both in the laboratory and by in situ testing in the field. The results demonstrated that CYANOCHIP is a valuable tool for the sensitive and reliable detection of cyanobacteria for early warning and research purposes.

Unprecedented rains decimate surface microbial communities in the hyperarid core of the Atacama Desert

“…The water samples were processed and analyzed by multiplex sandwich microarray immunoassay with LDChip as described previously 27 . The anti-cyanobacterial antibodies are specific 52 . A confirmation of this result is that the amplification of 16S rRNA with cyanobacteria-specific primer and the microscopy analysis of the samples in search of cyanobacteria turned both negative.…”

Critical Assessment of Analytical Techniques in the Search for Biomarkers on Mars: A Mummified Microbial Mat from Antarctica as a Best-Case Scenario

“… CYANOCHIP detected cyanobacterial markers in a mummified Antarctic microbial mat. ( A) Schematic of a printing pattern layout (in triplicate) of the anticyanobacterial antibody collection on CYANOCHIP: (o) BSA, bovine serum albumin; (x) only printing buffer; (1–17) each of the antibodies as in Table 1 in the work by Blanco et al 2015 (K1 to K17). From p1 to p17, the corresponding preimmune antibodies as controls.…”

“…Sample A was analyzed with CYANOCHIP, an antibody microarray for detecting the most relevant cyanobacterial genera of both planktonic ( Anabaena , Aphanizomenon , Microcystis , Planktothrix ) and benthic ( Chamaesiphon , Leptolyngbya , Nostoc , Phormidium , Rivularia , Tolypothrix ) habitats (Blanco et al , 2015 ). Aliquots of Sample A were analyzed by a sandwich microarray immunoassay, as described previously (Blanco et al , 2015 ). Briefly, 0.1 g of each transversal section of the mat was ultrasonicated in 1 mL of TBSTRR buffer (0.4 M Tris HCl pH 8, 0.3 M NaCl, 0.1% Tween 20) as above and filtered through a 20-μm filter.…”

Molecular biomarkers in the subsurface of the Salar Grande (Atacama, Chile) evaporitic deposits

“…Antibodies mostly polyclonal and a variety of targets, including biological polymers and strains of extremophile and mesophile microbes covering a wide range of metabolisms and phylogeny, were selected to increase the success of detecting any microbial remain in natural samples, either for environmental monitoring and/or for signs of life detection in planetary exploration (Rivas et al 2008;Parro et al 2008bParro et al , 2011b. Different versions of LDChip have been tested in multiple field campaigns in Rio Tinto (Parro et al 2008a), Atacama Desert (Parro et al 2011a) Deception Island (Blanco et al 2012), or Spanish water reservoirs (Blanco et al 2015a), and it has been implemented for automatic in situ analysis in the instrument SOLID (Signs of Life Detector; Parro et al 2008b), in e.g. Atacama (Parro et al 2011a), or Rio Tinto (Parro et al 2011b).…”