Abstract
Copepods are small crustaceans capable to survive in various aquatic environments. Their responses to changes in different external factors such as salinity and temperature can be observed at different integration levels from copepod genes to copepod communities. Until now, no thorough observation of the temperature or salinity effect stresses on copepods has been done by optical microscopy. In this study, we used autofluorescence to visualize these effects on the morphology of the calanoid copepod Pseudodiaptomus marinus maintained during several generations in the laboratory at favorable and stable conditions of salinity (30 psu) and temperature (18°C). Four different stress experiments were conducted: at a sharp decrease in temperature (18 to 4°C), a moderate decrease in salinity (from 30 to 15 psu), a major decrease in salinity (from 30 to 0 psu), and finally a combined stress with a decrease in both temperature and salinity (from 18°C and 30 psu to 4°C and 0 psu). After these stresses, images acquired by confocal laser scanning microscopy (CLSM) revealed changes in copepod cuticle and muscle structure. Low salinity and/or temperature stresses affected both the detection of fluorescence emitted by muscle sarcomeres and the distance between them. In the remaining paper we will use the term sarcomeres to describe the elements located within sarcomeres and emitted autofluorescence signals. Quantitative study showed an increase in the average distance between two consecutive sarcomeres from 2.06 +/- 0.11 μm to 2.44 +/- 0.42 μm and 2.88 +/- 0.45μm after the exposure to major haline stress (18°C, 0 psu) and the combined stress (4°C, 0 psu), respectively. These stresses also caused cuticle cracks which often occurred at the same location, suggesting the cuticle as a sensitive area for osmoregulation. Our results suggest the use of cuticular and muscle autofluorescence as new biomarkers of stress detectable in formalin-preserved P. marinus individuals. Our label-free method can be easily applied to a large number of other copepod species or invertebrates with striated musculature.
Highlights
Copepods are small crustaceans and a major component of zooplankton, which represents the main food for higher trophic levels in aquatic systems including fish larvae
The effects of thermal or haline stressors on copepods have been observed at the levels of gene expression [14], protein expression [27] and enzymatic activity [28]
For the first time, the effects of these stressors on the muscular and cuticle structures of the copepod Pseudodiaptomus marinus adult females by using the confocal laser scanning microscopy (CLSM) tool
Summary
Copepods are small crustaceans and a major component of zooplankton, which represents the main food for higher trophic levels in aquatic systems including fish larvae. Copepods occur in most aquatic environments, from large oceans to small ponds. They are even found in extended transient water so after heavy rains in wet plant detritus [1]. A rich literature on the biology and ecology of copepods focuses on the effects of temperature on their development, growth, survival and reproduction [9,10,11]. Regarding their salinity tolerance, two evolutionary pathways are characterizing marine and freshwater copepods, even though copepods can be found in hypersaline habitats [12]. When the environmental conditions are not optimal, these copepods experience stress that can be observed at different levels of organization from cells to individuals
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