for bacterivory, and acid P-galactosidase activity for herbivory. Samples will also be preserved 

 for later microscopic inspection of the protistan community in the nanoplankton and 

 microplankton size ranges. 



STRENGTHS AND LIMITATIONS OF PROPOSED RESEARCH 



Protistan grazing on bacteria and on phytoplankton is a major process in marine food 

 webs. Thus, in order to properly evaluate pathways of carbon flow in marine systems, it is 

 essential to have a good understanding of the trophic activities of heterotrophic protists. 

 Unfortunately, existing assays for protist grazing are so cumbersome to carry out that not very 

 many measurements can be done within short time periods (days). The DEA approach that we 

 are planning to use will be the first method which allows high-resolution sampling for protist 

 grazing, so that vertical and horizontal variability in grazing activity can be assessed. The 

 weakness of the method is that this is a new approach, and we are just now working out a 

 calibration curve to allow quantification of herbivory (chl-a mortality) based on measured in situ 

 p-galactosidase activity. 



STATUS OF RESEARCH 



Under the auspices of the OMP, the first application of the DEA approach was the acid 

 lysozyme method developed by our post-doctoral colleague, Dr. Juan Gonzalez (1993, MEPS 

 100:197-206). Rate of cleavage of MUF from a fluorochrome linked substrate analogue of 

 peptidoglycan, a major structural compound in bacterial cell walls, was calibrated as a 

 quantitative indicator of bacterivory via rates of uptake of FLB. We are now evaluating 

 measurement of p-galactosidase-like activity at acid pH as an indicator of herbivory. All 

 phototrophic organisms, including cyanobacteria, have sulfo-lipid compounds associated with 

 their chloroplasts (thylakoid membranes) that can be cleaved by the enzyme p-galactosidase. 

 Thus, herbivorous protists should elaborate digestive enzymes for cleaving the sulfo-lipids of 

 phototrophs, but which also would be able to cleave MUF from the substrate MUF-galactoside. 

 Preliminary results using this substrate are promising. Herbivorous protists in lab culture and in 

 natural seawater assemblages have high P-galactosidase-like activity at acid pH, comparable in 

 magnitude to P-glucosidase activity. Bacterivorous protists have negligible P-galactosidase-like 

 activity. The challenge now is to calibrate measured P-galactosidase activity by simultaneous 

 determination of actual chlorophyll loss due to grazing. We are attempting to do this via 

 comparison of MUF- p-galactosidase-like activity with chlorophyll grazing loss estimated via 

 dilution experiments using coastal seawater. 



We plan to use DEA methods for both bactivory and herbivory on the upcoming cruise 

 in June. We will cany out on-board FLB-uptake experiments to calibrate the acid lysozyme 

 methods, and will use a laboratory-based calibration factor to convert acid P-galactosidase 

 activities to phytoplankton grazing rates. 



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