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{"project":"2_test","denotations":[{"id":"23408395-18298829-140428077","span":{"begin":256,"end":258},"obj":"18298829"},{"id":"23408395-21167037-140428078","span":{"begin":259,"end":261},"obj":"21167037"},{"id":"23408395-18298829-140428079","span":{"begin":396,"end":398},"obj":"18298829"},{"id":"23408395-10028268-140428080","span":{"begin":399,"end":401},"obj":"10028268"},{"id":"23408395-21628502-140428081","span":{"begin":402,"end":404},"obj":"21628502"},{"id":"23408395-16558822-140428082","span":{"begin":5637,"end":5639},"obj":"16558822"},{"id":"23408395-12508882-140428083","span":{"begin":7796,"end":7798},"obj":"12508882"},{"id":"23408395-17392223-140428084","span":{"begin":7831,"end":7833},"obj":"17392223"},{"id":"23408395-20308591-140428085","span":{"begin":8057,"end":8059},"obj":"20308591"},{"id":"23408395-18298829-140428086","span":{"begin":8077,"end":8079},"obj":"18298829"}],"text":"Insights from the genome sequence\nMotility of Paenibacillus cells on solid media has been observed with a number of species. P. lautus is reported to spread across plates 69]. P. vortex shows highly unusual organized growth morphologies on solid surfaces [19,20] forming complex patterns on the plate. Another species, P. dendritiformis, also shows unusual growth morphologies on solid surfaces [19,21,22]. Y412MC10 was evaluated for spreading behavior on plates; the results ((Figure 4A Figure 4B Figure 4C) show definite spreading behavior for Y412MC10. The spreading behavior does not, however, appear to be as complex as reported for P. vortex and P. dendritiformis.\nFigure 4A Photograph of Paenibacillus sp. Y412MC10 streaked on YT agar containing 100 mg/l carbenicillin and incubated at 37°C for 18 hours.\nFigure 4B Photograph of Paenibacillus sp. Y412MC10 streaked on YT agar and incubated at 37°C for 50 hours. Note continued clearing of center area and significant spreading of outside edges of culture.\nFigure 4C Photograph of Paenibacillus sp. Y412MC10 streaked on YT agar and incubated at 37°C for 168 hours. Note continued clearing of center area and significant spreading of outside edges of culture. To further understand the phylogenetic relationships of Y412MC10, a separate phylogenetic tree was constructed of Paenibacillus sp. having either complete or draft genome. This was necessary because there is little or no overlap between validly-named Paenibacillus strains and the Paenibacillus strains submitted for whole genome sequencing. The tree was created on the IMG website [54] using the alignment of 16S genes based on the SILVA database and dnadist and neighbor tools from the Phylip package [55]. Paenibacillus sp. JDR-2, Paenibacillus curdlanolyticus YK9, Paenibacillus polymyxa E681, Paenibacillus polymyxa SC2, and Paenibacillus mucilaginosus KNP414 are all species isolated from soil or plant roots. Paenibacillus vortex V453, Paenibacillus sp. HGF5, Paenibacillus sp. HGF7, are oral or intestinal human isolates. Paenibacillus larvae subsp. larvae B-3650, 741161 is a honey bee pathogen that attacks bee larvae. The r16S analysis (Figure 5) shows Y412MC10 is most closely related to Paenibacillus sp. HGF5 (NCBI Taxon ID 908341, Gold ID Gi05716), an organism being sequenced as part of the Human Microbiome Project (HMP) Reference Genomes (http://www.hmpdacc.org/reference_genomes/reference_genomes.php).\nFigure 5 Phylogenetic tree highlighting the position of Paenibacillus sp. Y412MC10 and relative strains within the Bacillales. The strains and their corresponding GenBank taxonomy ID numbers are: Paenibacillus sp. JDR-2, 324057; Paenibacillus curdlanolyticus YK9, A717606; Paenibacillus sp. Y412MC10, 481743; Paenibacillus vortex V453, 715225; Paenibacillus polymyxa E681, 349520; Paenibacillus polymyxa SC2, 886882; Paenibacillus mucilaginosus KNP414, 1036673; Paenibacillus sp. HGF5, 908341; Paenibacillus sp. HGF7, 944559; Paenibacillus larvae subsp. larvae B-3650, 741161; Bacillus subtilis subtilis 168, 2243082. COG [Figure 6] and TIGRfam [Figure 7] whole genome comparisons were carried out between Y412MC10 and draft and finished genomes of closely related organisms using IMG software [56]. The results of the COGs and TIGRfam whole genome comparisons place Y412MC10 clearly among the Paenibacillus species, in agreement with the results from 16S analysis. The 16S analysis shows Y412MC10 is most closely related to P. vortex and P. sp. HGF5, both human isolates, and then to the two P. polymyxa sp. In both whole genome analyses, Y412MC10 is again most closely related to Paenibacillus vortex and Paenibacillus sp. HGF5. In the COG comparison, the other human isolate, P. sp. HGF7, is not closely related to P. vortex and P. sp. HGF5; in the TIGRfam comparison, HGF7 clades closely with Y412MC10, P. vortex and P. sp. HGF5. These results also suggest a mammalian, rather than environmental, ecosystem as the home of Y412MC10.\nFigure 6 COGs whole genome comparison of selected strains. Comparison was performed as described in text; organisms and GenBank accession numbers are described in Figure 5.\nFigure 7 TIGRfam whole genome comparison of selected strains. Comparison was performed as described in text; organisms and GenBank accession numbers are described in Figure 5. To further understand the relationship between these organisms, whole genome alignments were performed using Mummer software to generate dot plot diagrams comparing pairs of genomes on the IMG website [57] using input DNA sequences directly (NUCmer). The close relationship between the genome of Y412MC10 and the genomes of HGF5 and P. vortex is reflected in the high levels of homology and synteny seen (Figure 8, Figure 9) with these two human isolates.\nFigure 8 Dot plot comparison of Y412MC10 and HGF5 performed on IMG website. HGF5 draft genome contains 185 scaffolds.\nFigure 9 Dot plot comparison of Y412MC10 and P. vortex performed on IMG website. P. vortex draft genome contains 56 scaffolds. In comparison, whole genome alignment of Y412MC10 with the genomes of P. polymyxa and P. mucilaginosus show little homology or synteny between Y412MC10 and the two soil organisms (Figure 10, Figure11).\nFigure 10 Dot plot comparison of Y412MC10 and P. polymyxa performed on IMG website.\nFigure 11 Dot plot comparison of Y412MC10 and P mucilaginosus performed on IMG website. The similarity between the r16S sequences of P. lautus and Y412MC10 led us to examine if biochemical evidence suggested a similar habit for both. Bacillus lautus was first isolated from the intestinal tract of children [58]; later, the identity of the organism was re-confirmed and the organism was reclassified and renamed Paenibacillus lautus. Examination of the genome of Y412MC10 lends support to the hypothesis that Y412MC10 also has an intestinal origin. An analysis of the carbohydrate active enzymes (CAZY [59]) shows very low levels of GH family 5, 6, 8, 9, 10, 11, and 48 as well as no CBM 2 or 3 members, suggesting an inability to significantly degrade cellulose and hemicellulose components of biomass. CAZy analysis shows a genome enriched in GH29 and GH95 α-fucosidases; the genome is also enriched in GH38 and GH125 α-mannosidases and GH78 α-L-rhamnosidases. All these enzyme groups attack carbohydrate sidechains attached to eukaryotic glycoproteins; such glycoproteins are found in abundance in intestinal cell walls. CAZy analysis also shows a genome enriched in GH18 chitinases, GH28 polygalacturonases, GH88 unsaturated glucuronyl hydrolases, GH105 unsaturated rhamnogalacturonyl hydrolases and pectate lyase (PL) family members. These enzymes attack dietary fiber components that would be resistant to digestion by most ruminant bacteria, allowing the organism to scavenge sugars from pre-digested dietary sources. The enzymes required for bacillibactin production appear to be present in the genome of Y412MC10; bacillibactin is involved in iron acquisition. Iron is in limited supply in intestinal environments, but is present in large excess (approximately 2 μM Fe2+) in Obsidian hot spring. This again argues for an intestinal origin for the organism. Y4112MC10 does not possess genes usually involved in detoxification of heavy metals and sulfide found in other hot springs organisms (unpublished results). The organism also lacks antibiotic production genes, indicating it comes from an environment with excess resources, typical of the intestine. The growth temperature range and optimum of Y412MC10 is an excellent match for intestinal conditions, but a poor fit for the conditions of Obsidian hot spring, where temperatures average 79±4°C. Nitrogen fixing Paenibacillus have been isolated from the rhizosphere, including Paenibacillus brasilensis [60], and Paenibacillus zanthoxyli [61]. Paenibacillus lautus Y412MC10 has no nitrogen-fixing genes; these would be of no advantage for a free-living organism in an intestinal environment.\nComplex cooperative behaviors such as those seen with P. dendritiformis [62], and P. vortex [19] are not observed with Y412MC10; again, these behaviors may be unnecessary for survival in the intestine. Formation of external matrices in liquid and solid cultures may be beneficial to Y412MC10 for survival; the matrix may allow attachment of the bacteria to intestinal mucosa."}

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PMC:3558958 / 13685-22044 JSON TXT