Computational protocol: Draft Genome Sequences of Paenibacillus polymyxa NRRL B-30509 and Paenibacillus terrae NRRL B-30644, Strains from a Poultry Environment That Produce Tridecaptin A and Paenicidins

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Protocol publication

[…] Campylobacter jejuni is a leading cause of gastroenteritis (). Infection with Campylobacter occurs primarily through the consumption and handling of poultry. Antibiotic resistance of Campylobacter strains is a concern, and recently, attention has focused on the isolation of peptides that are active against Campylobacter species. Paenibacillus polymyxa NRRL B-30509 and Bacillus circulans NRRL B-30644, isolated from a poultry production environment, were reported to produce bacteriocins inhibitory to C. jejuni (). However, subsequent studies showed no evidence that these bacteriocins were produced, and instead the activity in both strains against C. jejuni was attributed to the nonribosomal lipopeptide tridecaptin A (, ). In addition, NRRL B-30509 and NRRL B-30644 were found to produce the novel lantibiotics paenicidin A and B, respectively (, ). Based on 16S rRNA analysis, B. circulans NRRL B-30644 was renamed Paenibacillus terrae NRRL B-30644 ().Genomic DNA of P. polymyxa NRRL B-30509 and P. terrae NRRL B-30644 were isolated using the DNeasy blood & tissue kit (Qiagen) and sequenced by 454 GS FLX Titanium pyrosequencing (Roche) at GenoSeq (UCLA Genotyping and Sequencing Core, Los Angeles, CA). The reads were assembled into contigs using the GS De Novo Assembler software (Roche). The assembly of the draft genome sequence of P. polymyxa NRRL B-30509 yielded 75 contigs (>200 bp) and contains 5,948,280 bases with a G+C content of 45.1%. For P. terrae NRRL B-30644, the assembled draft genome sequence consists of 234 contigs (>200 bp) and 6,514,862 bases with a G+C content of 45.7%. One of these 234 contigs carries the genes for tridecaptin A production and was assembled from different contigs previously (). Genome annotation by the NCBI Prokaryotic Genomes Automatic Annotation Pipeline (PGAAP) predicted 5,169 genes, including 4,839 coding sequences (CDS), 33 rRNAs, and 109 tRNAs in the genome of NRRL B-30509, and 5,988 genes, including 5,482 CDS, 14 rRNAs, and 95 tRNAs in the genome of NRRL B-30644.The average nucleotide identity (ANI) between P. polymyxa NRRL B-30509, P. terrae NRRL B-30644, P. polymyxa SC2 (), and P. terrae HPL-003 () was calculated using JSpecies software (). NRRL B-30509 has a higher identity to SC2 (94.71% ANI) than to NRRL B-30644 and HPL-003 (84.82% and 84.68% ANI, respectively), whereas NRRL B-30644 has a higher identity to HPL-003 (93.28% ANI) than to NRRL B-30509 and SC2 (84.75% and 84.54% ANI, respectively). This is in agreement with the previous finding that NRRL B-30644 is a P. terrae species ().P. polymyxa NRRL B-30509 carries gene clusters for several nonribosomal lipopeptides, including tridecaptin A, polymyxin, and fusaricidin. Indeed, it was previously observed that NRRL B-30509 also produces polymyxins E1 and E2 (). The fusaricidin synthetase gene in NRRL B-30509 contains a frameshift mutation, which might prevent the production of fusaricidin. In addition to the paenicidin A gene cluster, NRRL B-30509 also harbors a lantibiotic gene cluster that might produce a novel subtilin-like bacteriocin. The genome of P. terrae NRRL B-30644 carries the gene clusters for tridecaptin A and fusaricidin, as well as for paenicidin B. […]

Pipeline specifications

Software tools Newbler, PGAP, JSpeciesWS
Application Nucleotide sequence alignment
Organisms Paenibacillus polymyxa, Paenibacillus terrae, Campylobacter jejuni