Crayfish are known to benefit from branchiobdellidan worms as these worms consume bacteria on crayfish gills, providing a cleaning symbiosis. In our first experiment, we inoculated crayfish with four branchiobdellidans and others had all branchiobdellidans removed. It was expected that the presence of worms would decrease the amount of bacteria in the gill chamber, thus allowing an increase in both growth rates and dissolved oxygen (DO) uptake. Crayfish were then observed for DO uptake and microbial swabs were collected to identify colony forming units (CFUs/g crayfish). Our hypothesis was that the branchiobdellidan presence would result in an increase in DO uptake due to the cleaning effect of the worms on the gills. Based on the results from this study, we then began collecting microbial samples from crayfish in a stream with and without branchiobdellidans. Aseptic technique was used to obtain microbial swabs of crayfish and environment from five collection sites during the months of May, June, July, and August (2018). DNA was extracted from the microbial swabs and the 16S gene was amplified for DNA sequencing. Sequenced microbes were then compared to stream order, stream environment (water and substrate), and the number of branchiobdellidan worms present on the crayfish.
Results/Conclusions
From the tank study, the observed percent change in blotted wet mass (BWM) over eight weeks showed that the control crayfish group grew 26.2% more than the experimental group (with four worms/crayfish). No significant difference was found in the mean DO consumption between control (0.094 mg/l/g*2hr) and experimental (0.090 mg/l/g*2hr) groups. A gill chamber bacterial analysis and characterization showed that the logged mean number of colony forming units/gram crayfish (log10 CFUs/g) was greater for the experimental group (3.80 log10 CFUs/g) than the control group (2.84 log10 CFUs/g). The bacteria identified revealed two unique microbial assemblages which was a basis for looking for distinct microbial assemblages on crayfish with and without branchiobdellidans in a stream. From the field based study, preliminary Qiime2 analysis of the raw sequencing data from crayfish with (N=19) and without(N=14) branchiobdellidans revealed significant differences in alpha diversity. However, the limited sample size warrants further investigation into how branchiobdellidan symbionts alter microbial assembly.