CellTracker Green labelling vs. Rose Bengal staining: CTG wins by points in distinguishing living from dead anoxia-impacted copepods and nematodes

Abstract

Hypoxia and anoxia have become a key threat to shallow coastal seas. Much is known about their impact on macrofauna, less on meiofauna. In an attempt to shed more light on the latter group, in particular from a process-oriented view, we experimentally induced short-term anoxia (1 week) in the Northern Adriatic Sea, Mediterranean, and examined the two most abundant meiofauna taxa – harpacticoid copepods and nematodes. Both taxa also represent different ends of the tolerance spectrum, with copepods being the most sensitive and nematodes among the most tolerant. We compared two methods: CellTracker Green (CTG) – new labelling approach for meiofauna – with the traditional Rose Bengal (RB) staining method. CTG binds to active enzymes and therefore colours live organisms only. The two methods show considerable differences in the number of living and dead individuals of both meiofauna taxa. Generally, RB will stain dead but not yet decomposed copepods and nematodes equally as live ones. Specifically, RB significantly overestimated the number of living copepods in all sediment layers in anoxic samples, but not in any normoxic samples. In contrast, for nematodes, the methods did not show such a clear difference between anoxia and normoxia. Surprisingly, RB overestimated the number of living nematodes in the top sediment layer of normoxic samples, which implies an overestimation of the overall live nematofauna. For monitoring and biodiversity studies, the RB method might be sufficient, but for more fine-scaled (days, hours, tipping points) studies, especially on hypoxia and anoxia where it is necessary to resolve the course of events, CTG labelling is a better tool. Moreover, it clearly highlights the surviving species within the copepod or nematode community. As already accepted for foraminiferal research, we demonstrate that the CTG labelling is also valid for other meiofauna groups

Structural and functional responses of harpacticoid copepods to anoxia in the Northern Adriatic: an experimental approach

“…Moreover, novel approaches, such as the use of vital stains, may allow a more accurate assessment of alive/dead individuals following short-term impact by hypoxia and anoxia (e.g. see Grego et al, 2013a, for copepods and nematodes). Harpacticoid copepods were the most sensitive meiofauna taxa to low oxygen concentrations.…”

“…As there was no significant difference in overall density between the natural (normoxic) and the treated cores (anoxia), we conclude that the impacted meiofauna community corresponded to a "natural, pristine" meiofauna community. Note that the meiofauna counts for this study were based on rose-bengalcoloured individuals and may therefore be an overestimation of the actual number of life animals (see Grego et al, 2013a).…”

Is the meiofauna a good indicator for climate change and anthropogenic impacts?

“…Harpacticoid copepods are the most sensitive meiofauna taxa to low oxygen concentrations . Several studies have shown that nematodes are generally more resistant to hypoxia than copepods (Murrell and Fleeger 1989;Modig and Olafsson 1998;Grego et al 2013;Sergeeva and Zaika 2013). Copepods may stop feeding under anoxic conditions, yielding increased primary producer levels .…”

Characteristics of meiofauna in extreme marine ecosystems: a review

“…Several studies showed that nematodes are generally more resistant to hypoxia than copepods (Murrell and Fleeger 1989;Modig and Olafsson 1998;Grego et al 2013;Sergeeva and Zaika 2013). Nematodes are adapted to these extreme conditions (e.g.…”