Molecular detection of some Gram-negative bacterial species using folp gene sequences
In this study, the folp gene encoding DHPS was tested as a possible alternative phylogenetic marker for more closely related Gram-negative bacterial species. In this new method, 854 bp were implemented for classification instead of 1435 mostly used in 16S gene detection. Phylogenetic analysis was performed based on DNA sequences obtained from the GenBank including the most important Gram-negative bacterial species. The 16S rRNA-based tree openly portrayed three distinct clusters where cluster 1 is a mixed species cluster including E.coli, Shigella sp., Proteus sp., Enterobacter sp. Citrobacter sp., Salmonella sp., Serratia sp., and Klebsiella sp., while clusters 2 and 3 contained P. aeruginosa and Haemophilus sp. respectively. Comparatively, the folp gene-based tree yielded 9 clusters in which, E.coli and Shigella sp. were identified in one mixed cluster. However, other Gram-negative species such as Klebsiella sp., Enterobacter sp., Salmonella sp., Citrobacter sp., Serratia sp., Proteus sp., Haemophilus sp., and P. aeruginosa were found each in a separate cluster. In addition, DNA-DNA relatedness studies indicated high sequence divergence of folp gene exhibiting 47.54- 98.37% interspecies homology compared to 16S rRNA with sequence similarities of 79.58-98.11%. In addition, 71.9-100 % intraspecific similarities were obtained for the folp gene which indicates the possibility for use of the folp gene as a possible efficient target for Gram-negative bacterial group taxonomic analysis. Moreover, in blind tests, this method was able for the correct identification of 10 Gram-negative bacteria isolates. In conclusion, folp gene sequences provide better analysis of Gram-negative bacteria.
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