Screening is a critical step in the discovery of microbial agents that can exert biological control of Fusarium verticillioides at the root level. The objectives of this research were to determine the utility of a niche overlap index to realise the first screening of maize rhizobacterial isolates during different water activities. Studies were conducted to evaluate various methods for second screening with different modes of action. The antifungal activity of bacterial isolates through antibiosis assay was checked and the influence of different isolates on Fusarium verticilliodes growth and fumonisin B(1) was studied. Eleven competitive rhizobacterial isolates (Arthrobacter globiformis RC1, Azotobacter armeniacus RC2, A. armeniacus RC3, A. globiformis RC4, A. globiformis RC5, A. armeniacus RC6, Pseudomonas solanacearum RC7, Bacillus subtilis RC8, B. subtilis RC9, P. solanacearum RC10, B. subtilis RC11) were selected for the studies which followed. All bacteria were able to utilise the widest range of carbon sources and showed the highest niche overlap indices at the water activities tested. All bacterial antagonists reduced fumonisin B(1) production at all levels tested. Isolates belonging to Pseudomonas and Bacillus genera significantly inhibited fumonisin B(1) production, which ranged between 70 and 100%. Also, A. armeniacus RC2 caused important fumonisin B(1) reduction. The results of the present work suggest that A. armeniacus RC2, A. armeniacus RC3, B. subtilis RC8, B. subtilis RC9, B. subtilis RC11, P. solanacearum RC7, and P. solanacearum RC10 could have practical value in the control of F. verticillioides root colonisation. This paper is part of an on-going study to determine their application at the field level.