Carbon : nitrogen : phosphorus ratios influence biofilm formation by Enterobacter cloacae and Citrobacter freundii.
OBJECTIVE
To test the effects of C : N : P ratio modification of a well-known nutrient medium formulation, the Endo formulation on biofilm formation by Enterobacter cloacae Ecl and Citrobacter freundii Cf1 in both single-species and binary species biofilms.


RESULTS
The C : N : P atom : atom ratio of a well-known nutrient medium formulation, the Endo formulation, that has been applied in fermentative biohydrogen studies, was modified to include two different C concentrations, one containing 17.65 g l(-1) and the other 8.84 g l(-1) sucrose, each containing four different C : N : P ratios, two at higher C : N : P ratios (334 : 84 : 16.8 and 334 : 84 : 3) and two at lower C : N : P ratios (334 : 28 : 5.6 and 334 : 28 : 1). Attached cells were enumerated after dislodging the biofilms that had formed on granular activated carbon (GAC). The modified medium containing 17.65 g l(-1) sucrose and having a C : N : P ratio of 334 : 28 : 5.6 resulted in significantly (P < 0.05) higher counts of attached cells for both single-species biofilms at 7.73 log(10) CFU g(-1) GAC and 9.3 log(10)CFU g(-1) GAC for Ent. cloacae Ecl and Cit. freundii Cf1, respectively, and binary species biofilms at 8.2 log(10) CFU g(-1) GAC and 6.34 log(10) CFU g(-1) GAC for Ent. cloacae Ecl and Cit. freundii Cf1, respectively. Scanning electron micrographs showed qualitative evidence that the 334 : 28 : 5.6 ratio encouraged more complex and extensive biofilm growth for both single-species and binary species biofilms.


CONCLUSIONS
The differences in the attachment numbers between the different ratios were found not to be a result of the individual actions of the bacterial isolates involved but rather because of the effects of the various C : N : P ratios. The 334 : 28 : 5.6 ratio showed significantly (P < 0.05) higher counts of attached cells for both single-species and binary species biofilms.


CONCLUSIONS
This study indicates that C : N : P ratios should be a key consideration with regard to maximizing biofilm formation in shake flask and fluidized bed bioreactor studies as well as understanding fundamental factors affecting biofilm growth in natural environments.
Thompson
L J
LJ
School of Molecular and Cell Biology, University of the Witwatersrand, Gauteng, South Africa.
Gray
V
V
Lindsay
D
D
von Holy
A
A
D001422:Bacterial Adhesion / Q000187:drug effects; D018441:Biofilms / Q000187:drug effects* / Q000254:growth & development; D019149:Bioreactors; D002244:Carbon / Q000494:pharmacology; D016971:Citrobacter freundii / Q000187:drug effects / Q000254:growth & development / Q000502:physiology* / Q000648:ultrastructure; D003470:Culture Media / Q000737:chemistry* / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D016972:Enterobacter cloacae / Q000187:drug effects / Q000254:growth & development / Q000502:physiology* / Q000648:ultrastructure; D008855:Microscopy, Electron, Scanning; D009584:Nitrogen / Q000494:pharmacology; D010758:Phosphorus / Q000494:pharmacology; D013395:Sucrose / Q000494:pharmacology
D003470:Culture Media; D010758:Phosphorus; D013395:Sucrose; D002244:Carbon; D009584:Nitrogen