PMC5627462_01
Phase-contrast micrographs of 10.1601/nm.5116
10.1601/strainfinder?urlappend=%3Fid%3DNRRL+B-23460 and 10.1601/nm.5156
10.1601/strainfinder?urlappend=%3Fid%3DNRRL+B-4156. a Depicts 10.1601/nm.5116
10.1601/strainfinder?urlappend=%3Fid%3DNRRL+B-23460 cells grown in 10.1601/strainfinder?urlappend=%3Fid%3DTSB+for+24 hr. at 30  C. b Depicts 10.1601/nm.5156
10.1601/strainfinder?urlappend=%3Fid%3DNRRL+B-4156 cells grown in 10.1601/strainfinder?urlappend=%3Fid%3DTSB+for+24 hr. at 37  C. Scale bars represent 5 mum
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Fig1
The spore produced by 10.1601/nm.5116 has a rectangular outline, unlike the more ellipsoid shape seen in 10.1601/nm.5156. 10.1601/nm.5116 cells are slightly larger than 10.1601/nm.5156 cells as they range from 3.0-5.0 mum in length and 0.7-0.8 mum in width, while 10.1601/nm.5156 cells are 2.0-3.0 mum long and 0.5-1.0 mum wide (Fig. 1).
Phylogenetic tree of 10.1601/nm.5109 spp. based on 16S rRNA gene sequences. The maximum likelihood tree was inferred from a comparison of sequences from 10.1601/nm.5109 spp., 10.1601/nm.5168
10.1601/strainfinder?urlappend=%3Fid%3DDSM+5760, and 10.1601/nm.10618 168 using FastTree 2.1. The sequences generated from the draft genomes of this study are highlighted, with 10.1601/nm.5156
10.1601/strainfinder?urlappend=%3Fid%3DNRRL+B-4156 in blue font, and 10.1601/nm.5116
10.1601/strainfinder?urlappend=%3Fid%3DNRRL+B-23460 in green font
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Fig2
Our maximum likelihood 16S rRNA gene tree generated by FastTree 2.1 is congruent with these studies (Fig. 2).
Predicted thiamin biosynthesis pathway in 10.1601/nm.5116
10.1601/strainfinder?urlappend=%3Fid%3DNRRL+B-23460. Pathways involved in thiazole biosynthesis are highlighted in blue and pathways involved in pyrimidine biosynthesis are shown in red. The steps in black correspond to the coupling of the thiazole and pyrimidine moieties, and the formation of the active cofactor TPP
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Fig3
THZ-P (Fig. 3); thiG, thiO, thiS, and tenI are located in an operon putatively regulated by a TPP-binding riboswitch. The presence of thiO suggests that 10.1601/strainfinder?urlappend=%3Fid%3DNRRL+B-23460 uses glycine to generate the thiazole moiety.
Putative thiaminase I operons in the three 10.1601/nm.5109 species. Annotations of conserved genes shared amongst the three species are as follows (1) thiM; (2) thiD; (3) thiE; (4) glycosyltransferase; (5) thymidylate synthase; (6) methyltransferase; (7) yzgD NUDIX hydrolase; (8) thiaminase I and (9) HAD hydrolase. Three genes in 10.1601/nm.5116 B23460 that are not conserved include a putative transcriptional regulator and two genes of unknown function. The operons may be under the control of a putative thiamin pyrophosphate binding riboswitch, upstream of thiM. Genes are color-coded based on proposed functions
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Fig4
TMP. In bacteria, these hydrolase genes are either fused to a thiamin biosynthesis gene, like thiD or thiE, or these genes are located in the same operon. The 10.1601/nm.5116 10.1601/strainfinder?urlappend=%3Fid%3DNRRL+B-23460 genome has a HAD subfamily IA hydrolase gene that potentially serves this function, and is located in an operon with other thiamin biosynthesis genes (Fig. 4).   I (Fig. 4). In the 10.1601/nm.5156 10.1601/strainfinder?urlappend=%3Fid%3DNRRL+B-4156 and 10.1601/nm.5127 C454 operons, a HAD subfamily IA hydrolase is located directly after the thiaminase I gene. In 10.1601/nm.5116 10.1601/strainfinder?urlappend=%3Fid%3DNRRL+B-23460, three additional genes are present, which code for a putative transcriptional regulator and two proteins of unknown function (Fig. 4). I, and pyrimidine kinase, all of which are present in the three paenibacilli (Fig. 4), making it likely that they can produce bacimethrin. In the paenibacilli,
Predicted thiamin salvage pathways in all three paenibacilli. Pathways involved in thiazole salvage are highlighted in blue and pathways involved in pyrimidine salvage are shown in red. The dotted red boxes are steps unique to 10.1601/nm.5156
10.1601/strainfinder?urlappend=%3Fid%3DNRRL+B-4156 and 10.1601/nm.5127 C454. Biosynthetic pathways are shown with solid arrows and import pathways are indicated with a striped arrow. The putative importer boxed in green is unique to 10.1601/nm.5116
10.1601/strainfinder?urlappend=%3Fid%3DNRRL+B-23460. In all cases, it is not yet understood how THZ enters the cell
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Fig5
Thiazole kinase (thiM), which phosphorylates THZ (Fig. 5) is the first gene in the operon, followed by thiD and thiE. The thiamin biosynthesis genes are proceeded by a nucleoside 2-deoxyribosyltransferase, a thymidylate synthase, a SAM-dependent methyltransferase, a Nudix-family hydrolase (YzgD), and thiaminase TMP from environmentally acquired THZ and HMP, a strategy used by other bacteria. ThiM can phosphorylate environmentally derived thiazole alcohol, which can be combined with HMP-P by ThiE (Fig. 5). THZ and HMP* could be imported into the cell, phosphorylated by their respective kinases, and combined by ThiE (Fig. 5). TPP is then produced via dephosphorylation by the HAD hydrolase and addition of the pyrophosphate by TPK. The potential pathways available to the three paenibacilli to salvage thiamin are summarized in Fig. 5. TenA appears regulated by a TPP riboswitch. The genomes of all three paenibacilli contain ylmB, which deacetylates base-degraded thiamin forming aminopyrimidine, the preferred substrate for TenA (Fig. 5).