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Plant defensins are small, basic, cysteine-rich proteins with antimicrobial activity. In most plants, they are encoded by small multigene families and have been shown to have variable expression in different plant tissues. A search of the Nicotiana benthamiana Gene Index (http://www.tigr.org) revealed five defensin genes belonging to two types. The first type are short defensins of about 78 amino acids, while the second type are about 105 amino acids with an additional 27 to 33 acidic amino acid C-terminal domain. To study the expression of two of these genes, relative reverse transcriptase polymerase chain reaction was used with a translation elongation factor-1 gene as an internal control. In a compatible interaction with Pseudomonas syringae pv. tabaci, expression of both defensin genes were similarly down-regulated in leaves at 48 HPI. In contrast, no change in expression compared to the controls was observed in compatible interactions with Colletotrichum destructivum and Colletotrichum orbiculare, the causal agents of anthracnose, or in an incompatible interaction of N. benthamiana expressing Pto with P. syringae pv. tabaci expressing AvrPto. Both these defensin genes appear to be co-regulated being suppressed during disease development by P. syringae pv. tabaci. This suppression during a compatible interaction may be related to the action of bacterial effectors released through the type III secretion system.