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Supplementary table 1 Read Group Treatment Median High Quality Reads Genus level Unique Genera Percent classification High Quality Reads SD classified at reads SD genera identified reads tool Reads the genus identified SD classified at level the genus level BaseSpace RDP All All 47135 22116 44061 21435 110 31 93.53 Infant nTC 47860.5 36374 45598.5 35500 89.5 29 96.07 Infant TC 47135 12649 44760 11849 85 25 95.77 Mother nTC 46976 11743 43256 10655 124 22 91.31 Mother TC 46581 13649 42814 12940 126 25 92.65 DADA2 All All 19390 10589 15498 9411 18 12 84.46 Infant nTC 19266 13606 16043 12781 12 5 94.25 Infant TC 17653 9121 14489.5 8245 12 5 89.4 Mother nTC 20461 9101 15802 7295 34 9 81.65 Mother TC 19858 9594 15139.5 7887 33.5 10 81.81 Supplementary table 2 Household Treatment Group TC Level (pg/mL) Group TC Level (pg/mL) All TC Infant 38.30 (0.1-234.8) Mother 837.05 (63-96664.5) 1008 TC Infant 29 Mother 18502 1053 TC Infant 9 Mother 185 1061 TC Infant 52.2 Mother 96664.5 1084 TC Infant NA Mother 91 2081 TC Infant 33.6 Mother 78.7 2084 TC Infant 0.1 Mother 693.6 2085 TC Infant 63.7 Mother 1070.9 2117 TC Infant 11.3 Mother 916.1 2137 TC Infant NA Mother 118 2169 TC Infant 234.8 Mother 1156.4 2175 TC Infant 9 Mother 63 2211 TC Infant 90 Mother 758 2271 TC Infant 43 Mother 2538 2274 TC Infant 84.6 Mother 6177 2341 TC Infant 52.4 Mother 534.9 2360 TC Infant 13.7 Mother 123.4 2419 TC Infant 66.6 Mother 8681.4 2421 TC Infant 13 Mother 1896 Supplementary table 2 (cont) Household Treatment Group TC Level (pg/mL) Group TC Level (pg/mL) All nTC Infant 10.05 (0 - 574.2) Mother 76.00 (0 - 677.1) 1002 nTC Infant 6 Mother 15 1009 nTC Infant NA Mother 11 1067 nTC Infant 574.2 Mother 48 1092 nTC Infant 2 Mother 399.3 2048 nTC Infant 17.9 Mother 141.5 2050 nTC Infant NA Mother 24 2093 nTC Infant 79 Mother NA 2112 nTC Infant 9 Mother 65 2127 nTC Infant 38 Mother 268 2133 nTC Infant 17.8 Mother 76 2147 nTC Infant 11.1 Mother 76.8 2201 nTC Infant 8 Mother 73 2283 nTC Infant 0.2 Mother 247.6 2284 nTC Infant 6.7 Mother 14.1 2296 nTC Infant 8.4 Mother 166.9 2443 nTC Infant 3 Mother 94 2461 nTC Infant 34.2 Mother 677.1 2463 nTC Infant 30.1 Mother 123.5 2490 nTC Infant 207.7 Mother 0 2534 nTC Infant NA Mother 276 2558 nTC Infant 0 Mother 38.4 2584 nTC Infant NA Mother 8 Supplementary table 3 Group Enrich log2 Fold Adjusted Phylum Class Order Family Genus Species ment Change P-value Infant nTC 2.553 0.0038 Firmicutes Clostridia Clostridiales Clostridiaceae Clostridium paraputrificum 2.620 0.0003 Proteobacteria Gammaproteo - Enterobacteriales Enterobacteriaceae Serratia marcescens bacteria 4.300 0.0003 Bacteroidetes Bacteroidia Bacteroidales Bacteroidaceae Bacteroides fragilis TC 1.702 0.0062 Firmicutes Bacilli Lactobacillales Enterococcaceae Enterococcus 1.716 0.0050 Firmicutes Bacilli Lactobacillales Enterococcaceae Vagococcus teuberi 1.835 0.0084 Firmicutes Clostridia Clostridiales Ruminococcaceae Ruminococcus bromii 2.271 0.0084 Firmicutes Clostridia Coriobacteriales Coriobacteriaceae Eggerthella 2.709 4.61E-06 Actinobacteria Actinobacteria Bifidobacteriales Bifidobacteriaceae Bifidobacterium dentium 2.765 0.0010 Proteobacteria Deltaproteobacteria Desulfovibrionales Desulfovibrionaceae Desulfovibrio 3.233 6.17E-09 Verrucomicrobia Verrucomicrobiae Verrucomicrobiales Verrucomicrobiaceae Akkermansia muciniphila 3.268 0.0008 Firmicutes Clostridia Clostridiales Clostridiaceae Clostridium cadaveris 3.355 0.0074 Firmicutes Clostridia Clostridiales Clostridiaceae Clostridium perfringens 3.548 0.0097 Firmicutes Clostridia Clostridiales Lachnospiraceae Blautia glucerasea 3.575 0.0008 Proteobacteria Betaproteobacteria Burkholderiales Alcaligenaceae Sutterella stercoricanis 4.082 0.0030 Firmicutes Bacilli Lactobacillales Streptococcaceae Streptococcus infantarius 4.416 0.0004 Firmicutes Clostridia Clostridiales Veillonellaceae Megasphaera elsdenii 0.777 0.0095 Firmicutes Bacilli Bacillales Bacillaceae Bacillus Supplementary table 3 (cont) Group Enrich- log2 Adjusted Phylum Class Order Family Genus Species ment Fold P-value Change Mother nTC 1.562 0.0012 Bacteroidetes Bacteroidia Bacteroidales Porphyromonadacea Parabacteroides goldsteinii e 1.640 0.0037 Firmicutes Clostridia Clostridiales Lachnospiraceae Butyrivibrio proteoclasticus 2.065 1.12E-06 Bacteroidetes Bacteroidia Bacteroidales Bacteroidaceae Bacteroides sartorii 2.326 1.52E-05 Bacteroidetes Bacteroidia Bacteroidales Porphyromonadacea Dysgonomonas wimpennyi e 3.519 5.14E-07 Bacteroidetes Bacteroidia Bacteroidales Bacteroidaceae Bacteroides stercoris 4.388 0.0004 Bacteroidetes Bacteroidia Bacteroidales Bacteroidaceae Bacteroides massiliensis 5.206 1.23E-08 Bacteroidetes Bacteroidia Bacteroidales Bacteroidaceae Bacteroides clarus 5.292 1.31E-09 Bacteroidetes Bacteroidia Bacteroidales Bacteroidaceae Bacteroides intestinalis Supplementary table 3 (cont) Mother TC 1.178 0.0011 Bacteroidetes Sphingobacteriia Sphingobacteriales Rhodothermaceae Rhodothermus clarus 1.313 0.0011 Proteobacteria Gammaproteo- Enterobacteriales Enterobacteriaceae Enterobacter nickellidurans bacteria 1.387 0.0039 Bacteroidetes Bacteroidia Bacteroidales 1.540 5.16E-05 Firmicutes Clostridia Clostridiales Veillonellaceae Megasphaera hominis 1.858 0.0072 Firmicutes Clostridia Clostridiales Veillonellaceae Phascolarcto- succinatutens bacterium 1.953 0.0056 Proteobacteria Gammaproteo- Enterobacteriales Enterobacteriaceae Serratia bacteria 2.325 0.0037 Proteobacteria Gammaproteo- Enterobacteriales Enterobacteriaceae Escherichia albertii bacteria 2.334 0.0021 Proteobacteria Gammaproteo- Enterobacteriales Enterobacteriaceae Trabulsiella odontotermitis bacteria 2.405 0.0019 Proteobacteria Gammaproteo- Enterobacteriales Enterobacteriaceae Escherichia bacteria 2.514 0.0001 Proteobacteria Gammaproteo- Enterobacteriales Enterobacteriaceae Enterobacter amnigenus bacteria 2.528 0.0006 Proteobacteria Gammaproteo- Aeromonadales Aeromonadaceae Tolumonas auensis bacteria 2.550 0.0056 Bacteroidetes Bacteroidia Bacteroidales Prevotellaceae Prevotella copri 2.552 0.0087 Proteobacteria Gammaproteo- Enterobacteriales Enterobacteriaceae Escherichia coli bacteria 2.676 1.17E-08 Proteobacteria Gammaproteo- Enterobacteriales Enterobacteriaceae Serratia entomophila bacteria 2.888 0.0004 Proteobacteria Gammaproteo- Enterobacteriales Enterobacteriaceae Enterobacter aceae bacteria 3.011 0.0070 Proteobacteria Gammaproteo- Enterobacteriales Enterobacteriaceae Enterobacter bacteria 3.220 1.03E-11 Proteobacteria Gammaproteo- Enterobacteriales Enterobacteriaceae bacteria 3.608 4.49E-05 Firmicutes Clostridia Clostridiales Veillonellaceae Mitsuokella 4.005 0.0034 Proteobacteria Gammaproteo- Enterobacteriales Enterobacteriaceae Trabulsiella guamensis bacteria 4.611 0.0021 Proteobacteria Gammaproteo- Enterobacteriales Enterobacteriaceae Enterobacter cowanii bacteria 4.896 0.0011 Proteobacteria Gammaproteo- Enterobacteriales Enterobacteriaceae Yersinia bacteria 5.158 1.12E-06 Bacteroidetes Bacteroidia Bacteroidales Prevotellaceae Prevotella multisaccharivorax Supplementary table 4 Group Visit Enrichment Log2 Fold Adjuste Phylum Class Order Family Genus Species (Mon Change d p- ths) value Infant 2 nTC 2.304 0.0235 Bacteroidetes Bacteroidia Bacteroidales Bacteroidaceae Bacteroides xylanisolven s 3.794 0.0196 Proteobacteria Gammaproteo- Enterobacteriales Enterobacteriaceae Citrobacter freundii bacteria 5.141 0.0196 Firmicutes Clostridia Clostridiales Clostridiaceae Clostridium neonatale 5.358 0.0196 Firmicutes Erysipelotrichi Erysipelotrichales Erysipelotrichaceae Eubacterium dolichum TC 2.853 0.0241 Firmicutes Clostridia Clostridiales Clostridiaceae Clostridium cavendishii 3.232 0.0003 Bacteroidetes Bacteroidia Bacteroidales Bacteroidaceae Bacteroides dorei 3.241 0.0092 Proteobacteria Gammaproteo- Pasteurellales Pasteurellaceae Mannheimia caviae bacteria 5.500 0.0003 Firmicutes Clostridia Coriobacteriales Coriobacteriaceae Collinsella aerofaciens 5.530 0.0196 Firmicutes Clostridia Clostridiales Clostridiaceae Clostridium perfringens 5.626 1.78E-09 Actinobacteria Actinobacteria Bifidobacteriales Bifidobacteriaceae Bifidobacterium dentium Infant 6 nTC 5.070 0.0094 Bacteroidetes Bacteroidia Bacteroidales Bacteroidaceae Bacteroides fragilis 5.305 3.73E-05 Bacteroidetes Bacteroidia Bacteroidales Porphyromonadaceae Parabacteroides merdae TC 1.766 0.0279 Firmicutes Bacilli Lactobacillales 4.048 0.0405 Proteobacteria Betaproteo- Burkholderiales Alcaligenaceae Sutterella stercoricanis bacteria 4.728 0.0005 Firmicutes Clostridia Clostridiales Veillonellaceae Veillonella ratti Infant 10 nTC 2.000 0.0068 Firmicutes Clostridia Clostridiales Clostridiaceae Clostridium 2.049 0.0139 Firmicutes Clostridia Clostridiales Clostridiaceae Clostridium cavendishii 2.295 0.0268 Firmicutes Clostridia Clostridiales Clostridiaceae Clostridium taeniosporu m 2.982 0.0350 Proteobacteria Betaproteo- bacteria 3.036 0.0481 Firmicutes Bacilli Lactobacillales Lactobacillaceae Lactobacillus siliginis TC 3.028 0.0025 Firmicutes Bacilli Lactobacillales Streptococcaceae Streptococcus bovis 4.037 0.0007 Bacteroidetes Bacteroidia Bacteroidales Prevotellaceae Prevotella copri 4.073 0.0430 Verrucomicrobia Verrucomicrobiae Verrucomicrobiale Verrucomicrobiaceae Luteolibacter algae s 4.753 0.0350 Firmicutes Bacilli Lactobacillales Streptococcaceae Streptococcus luteciae 4.979 0.0095 Firmicutes Bacilli Lactobacillales Streptococcaceae Streptococcus infantarius 5.900 1.02E-07 Verrucomicrobia Verrucomicrobiae Verrucomicrobiale Verrucomicrobiaceae Akkermansia muciniphila s 6.420 2.05E-07 Bacteroidetes Bacteroidia Bacteroidales
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    Phylogeny of Selenophosphate synthetases (SPS) Supplementary Material S1 ! SelD in prokaryotes! ! ! SelD gene finding in sequenced prokaryotes! We downloaded a total of 8263 prokaryotic genomes from NCBI (see Supplementary Material S7). We scanned them with the program selenoprofiles (Mariotti 2010, http:// big.crg.cat/services/selenoprofiles) using two SPS-family profiles, one prokaryotic (seld) and one mixed eukaryotic-prokaryotic (SPS). Selenoprofiles removes overlapping predictions from different profiles, keeping only the prediction from the profile that seems closer to the candidate sequence. As expected, the great majority of output predictions in prokaryotic genomes were from the seld profile. We will refer to the prokaryotic SPS/SelD !genes as SelD, following the most common nomenclature in literature.! To be able to inspect results by hand, and also to focus on good-quality genomes, we considered a reduced set of species. We took the prok_reference_genomes.txt list from ftp://ftp.ncbi.nlm.nih.gov/genomes/GENOME_REPORTS/, which NCBI claims to be a "small curated subset of really good and scientifically important prokaryotic genomes". We named this the prokaryotic reference set (223 species - see Supplementary Material S8). We manually curated most of the analysis in this set, while we kept automatized the !analysis on the full set.! We detected SelD proteins in 58 genomes (26.0%) in the prokaryotic reference set (figure 1 in main paper), which become 2805 (33.9%) when considering the prokaryotic full set (figure SM1.1). The difference in proportion between the two sets is due largely to the presence of genomes of very close strains in the full set, which we consider redundant.
  • Rational Construction of Genome-Reduced Burkholderiales Chassis Facilitates Efficient Heterologous Production of Natural Products from Proteobacteria

    Rational Construction of Genome-Reduced Burkholderiales Chassis Facilitates Efficient Heterologous Production of Natural Products from Proteobacteria

    ARTICLE https://doi.org/10.1038/s41467-021-24645-0 OPEN Rational construction of genome-reduced Burkholderiales chassis facilitates efficient heterologous production of natural products from proteobacteria Jiaqi Liu1, Haibo Zhou1, Zhiyu Yang1, Xue Wang1, Hanna Chen1, Lin Zhong1, Wentao Zheng1, Weijing Niu1, Sen Wang2, Xiangmei Ren2, Guannan Zhong1, Yan Wang3, Xiaoming Ding4, Rolf Müller 5, Youming Zhang1 & ✉ Xiaoying Bian 1 1234567890():,; Heterologous expression of biosynthetic gene clusters (BGCs) avails yield improvements and mining of natural products, but it is limited by lacking of more efficient Gram-negative chassis. The proteobacterium Schlegelella brevitalea DSM 7029 exhibits potential for het- erologous BGC expression, but its cells undergo early autolysis, hindering further applica- tions. Herein, we rationally construct DC and DT series genome-reduced S. brevitalea mutants by sequential deletions of endogenous BGCs and the nonessential genomic regions, respectively. The DC5 to DC7 mutants affect growth, while the DT series mutants show improved growth characteristics with alleviated cell autolysis. The yield improvements of six proteobacterial natural products and successful identification of chitinimides from Chit- inimonas koreensis via heterologous expression in DT mutants demonstrate their superiority to wild-type DSM 7029 and two commonly used Gram-negative chassis Escherichia coli and Pseudomonas putida. Our study expands the panel of Gram-negative chassis and facilitates the discovery of natural products by heterologous expression. 1 Helmholtz International Lab for Anti-Infectives, Shandong University-Helmholtz Institute of Biotechnology, State Key Laboratory of Microbial Technology, Shandong University, Qingdao, Shandong, China. 2 Core Facilities for Life and Environmental Sciences, Shandong University, Qingdao, Shandong, China. 3 College of Marine Life Sciences, and Institute of Evolution & Marine Biodiversity, Ocean University of China, Qingdao, China.