Simple bulk pigment analysis suggests microphytobenthos contributions to food webs may be underestimated due to isotopic contamination by remineralized wetland carbon

Stable isotope studies have revealed the importance of microphytobenthos (MPB) in coastal food webs. Microalgae typically have δ13C values between depleted C3 wetland/terrestrial macrophytes, and enriched C4 macrophytes and seagrasses. However, the challenges of obtaining clean samples of microalgae from sediments means they are often represented by limited sampling in many food web studies; consequently, we have a limited understanding of spatial and temporal variation in their δ13C values. We tested a simple technique to measure the δ13C of bulk pigments extracted from surficial sediments to represent MPB and applied it to quantify fine scale spatial variation in MPB δ13C around salt marshes. The bulk extraction method is logistically simple, and drives substantial but relatively consistent fractionation in δ13C of −3.5 ± 0.13 ‰ (mean ± 1 S.E., range = 2.3–4.4 ‰, n = 18 paired comparisons) compared to whole cell values. The consistency in fractionation suggests that spatial and temporal δ13C patterns measured in field samples should reflect real variation in source values, and that measured values could be corrected and incorporated into isotope mixing models. In 88 MPB samples among four marsh sites over two summers, MPB δ13C in marsh creeks was lower by an average of 4.4 ± 0.72 ‰ and up to 8.4 ‰ compared to sites along the outer marsh-open water fringe 10's of m away. Few food web studies incorporate this magnitude of variation in their MPB source estimates into mixing models. Over three weekly samplings at one marsh creek site, low tide dissolved inorganic carbon (DIC) δ13C was similarly lower by 4.8 ± 0.36 ‰ and up to 6.2 ‰ compared to high tide and adjacent open water DIC values. The significant small-scale variability in MPB δ13C appears to be driven by remineralized marsh carbon which depletes the DIC δ13C in the marsh creeks, a phenomenon that has long been recognized but is rarely considered in food web studies. Mixing models that assume a narrower range in MPB source values will erroneously attribute isotopically variable MPB contributions to end-member production sources thereby clouding our understanding of energy flows through coastal seascapes. •Microphytobenthos is important in foodwebs but hard to sample for isotope analysis.•We tested for fractionation in δ13C with a simple pigment extraction method.•Bulk pigments were consistently depleted by 3.5‰ compared to whole cells.•Remineralized marsh carbon drove >4‰ variation in MPB δ13C around