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. 2016 Jun;6(1):32.
doi: 10.1007/s13205-015-0326-1. Epub 2016 Jan 14.

Molecular characterization and amplified ribosomal DNA restriction analysis of entomopathogenic bacteria associated with Rhabditis (Oscheius) spp

Balakrishnan Geetha Sangeetha  1 , Cheruvandasseri Arumughan Jayaprakas  2 , Jinachandrannair Vijayakumari Siji  1 , Moochattil Rajitha  1 , Basheerkutty Shyni  1 , Chellappan Mohandas  1
Affiliations

Molecular characterization and amplified ribosomal DNA restriction analysis of entomopathogenic bacteria associated with Rhabditis (Oscheius) spp

Balakrishnan Geetha Sangeetha et al. 3 Biotech. 2016 Jun.

Abstract

Bacterial strains associated with entomopathogenic nematodes (EPNs) Rhabditis (Oscheius) spp. were isolated from infected cadavers of Galleria mellonella. The obtained 18 isolates were subdivided into nine phylogenetically different genera based on comparative sequence analysis of their 16S rRNA genes. The isolates were affiliated to three different class namely γ-proteobacteria (Enterobacter, Proteus, Providencia, Pseudomonas, Stenotrophomonas), β-proteobacteria (Alcaligenes) and Bacilli (Bacillus, Enterococcus, Lysinibacillus). It was observed that Gram-positive strains (Bacilli) were more frequently associated with the EPN, whereas Gram-negative isolates were affiliated to six different genera with more genotypic diversity. Subsequently, all bacterial isolates used in this study were analyzed by amplified ribosomal DNA restriction analysis (ARDRA). Eight restriction endonucleases (CfoI, HinfI, RsaI, DdeI, Sau3AI, AluI, HaeIII, and MspI) were examined and a total of 15 different genotypes were obtained, forming two heterogenous main clusters after analysis by un-weighted pair-group method using arithmetic averages.

Keywords: 16S rDNA; Bacillus; Enterobacter; Entomopathogenic nematode.

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Conflict of interest statement

The authors declare that they have no conflict of interest in the publication.

Figures

Fig. 1
Fig. 1
Phylogenetic tree inferred from 16S rDNA sequences analysis showing the relationships of Rhabditis (Oscheius) spp. associated EPB isolates with several members of the Proteobacteria
Fig. 2
Fig. 2
Phylogenetic tree inferred from 16S rDNA sequences analysis showing the relationships of Rhabditis (Oscheius) spp. associated EPB isolates with several members of the class Bacilli
Fig. 3
Fig. 3
Restriction patterns of PCR-amplified 16S rRNA genes from bacterial strains digested with restriction endonuclease AluI
Fig. 4
Fig. 4
Restriction patterns of PCR-amplified 16S rRNA genes from bacterial strains digested with restriction endonuclease HaeIII. Lane 1–8 Bacillus cereus KPG, BR1, KY1, SBI, KAL, SCI, KK2, HY, 9-Enterococcus faecalis TN5, M-1kbp plus ladder (Bangalore GeNei, India), 10-Lysinibacillus sphaericus 352, 11-Pseudomonas sp. S, 12-Alcaligenes faecalis MM2, 13-Enterobacter sp.MA1, 14-Enterococcus faecalis TAH, 15-A. faecalis KL, 16-Proteus mirabilis D, 17-Providencia sp. F34, 18-Stenotrophomonas sp.MM3
Fig. 5
Fig. 5
Dendrogram based on UPGMA cluster analysis obtained for the combined ARDRA restriction profiles
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