Experimental Program

1) Initial system and method tests

・ Test of reactor system: flow, gas permeability.

・ Test of detection limits and procedure for DNA staining: PicoGreen and SYBR Green.

・ Test of electrochemical sensor detection limits.

2) Reactor inoculation

・ Preparation of an enrichment culture from artificial seawater media inoculated with pipeline scraping solids.

・ Inoculation of reactor system for one day to promote biofilm establishment.

3) Experimental evaluation of single-analyte strategies

・ Regulation of biofilm growth through changes in nutrient load as well as biocide treatment, while determining analyte concentrations in the water phase.

・ Control analyses of planktonic cell growth and microbial biofilm by quantitative polymerase chain reaction (qPCR) technique.

・ Test of correlation between microbial growth and analyte concentrations.

・ Test of detection limits of single-analyte strategies.

・ Selection of 1 or 2 single-analyte strategies suitable for monitoring microbial activity in the system.

Analytes monitored

Analyte

Method

Water

DNA

PicoGreen and SYBR Green assay

ATP

ATP luminometer

H₂S

Electrochemical minisensor

ORP

Electrochemical minisensor

pH

Electrochemical minisensor

Control measures for microbial growth and activity

Analyte

Method

Water

Planktonic population

qPCR and 4,6-diamindino-2-phenylindole (DAPI) staining method

Nutrient consumption

Total Organic Carbon (TOC) and Total Nitrogen (TN) analyses

Coupons

Sessile population

DAPI staining method

Biofilm growth

Wet weight, dry weight, and organic content analyses