Transcriptome and Secretome Analyses of Endophyte Methylobacterium mesophilicum and Pathogen Xylella fastidiosa Interacting Show Nutrient Competition
Abstract
:1. Introduction
2. Materials and Methods
2.1. Xf and Mm Cultivation Conditions
2.2. Bacterial Growth Evaluation
2.3. Transcriptomic Analyses
2.3.1. Experimental Design
2.3.2. qPCR Quantification of Xf and Mm
2.3.3. Total RNA Isolation, cDNA Library Preparation, and Sequencing
2.3.4. Bioinformatic Analyses and Statistics
2.4. Secretomic Analyses
2.4.1. Protein Extraction from Xf, Mm, and Co-Culture Supernatants
2.4.2. Shotgun Proteomics
2.5. Integration of Transcriptomics and Proteomics Analyses
3. Results
3.1. Growth of Mm and Xf in Co-Culture and Xf Biofilm Formation
3.2. Bacteria Quantification by qPCR
3.3. Transcriptomic Analyses
3.4. Transcriptome Analysis of Xf+Mm Co-Culture Compared to Xf Monoculture
3.4.1. Macromolecule Metabolism
3.4.2. Transport
3.4.3. Stress-Related Genes
3.4.4. Pathogenicity
3.5. Transcriptome Analysis of Xf+Mm Co-Culture Compared to Mm Monoculture
3.5.1. DNA, RNA, and Protein Metabolism
3.5.2. Lipid, Sugar, Amino Acid and Nucleotide Metabolism
3.5.3. Transport
3.5.4. Stress-Related Genes
3.5.5. Defense Mechanism-Related Genes
3.6. Secretome Analyses
3.6.1. Total Secretome of Xf and Mm
3.6.2. Xf Secretome
3.6.3. Mm Secretome
3.6.4. Secretome Predictions and Functional Analyses
3.7. Integrating Transcriptome and Secretome Data
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Treatment | Number of Reads | Mapped Reads in Pairs | % Mapped Reads | Unique Fragments |
---|---|---|---|---|
Xf monoculture | 4,400,994 | 3,523,819 | 80.16 | 1,759,574 |
Co-culture mapped in Xf genome | 4,410,865 | 2,284,762 | 51.05 | 1,140,964 |
Co-culture mapped in Mm genome | 4,410,865 | 1,158,502 | 26.41 | 578,282 |
Mm monoculture | 3,981,063 | 2,892,542 | 71.45 | 1,444,234 |
MC | CC | CDS Number IMG Xf * | CDS Number NCBI Xf * | Description and Symbol ** | Predicted Molecular Mass (kDa) | Sequence Coverage (%) | SignalP | SecP Score |
---|---|---|---|---|---|---|---|---|
1 | 3 | XF9a_01687 | XF_1803 | Omp1X | 21 | 26 | 20|21 | |
2 | 1 | XF9a_01416 | XF_1547 | Membrane lipoprotein Lpp | 16 | 73 | 17|18 | |
3 | 2 | XF9a_00315 | XF_0343 | Outer membrane protein mopB | 42 | 28 | 0.94 | |
4 | 11 | XF9a_00326 | XF_0363 | Outer membrane protein OmpA | 26 | 33 | 27|28 | |
5 | 4 | XF9a_00948 | XF_1026 | Serine protease PspB | 95 | 18 | 0.95 | |
6 | 7 | XF9a_01869 | XF_1981 | Adhesin XadA3 | 118 | 23 | 0.95 | |
7 | 12 | XF9a_00900 | XF_0975 | PorinO OprP | 44 | 35 | 21|22 | |
8 | 6 | XF9a_00602 | XF_0668 | Hemolysin toxin protein RTX | 128 | 22 | 0.74 | |
9 | 13 | XF9a_00831 | XF_0898 | Membrane lipoprotein | 13 | 53 | ||
10 | 9 | XF9a_02226 | XF_2349 | Autotransporter beta-domain | 81 | 26 | 0.85 | |
11 | 5 | XF9a_01123 | XF_1219 | Protein of Unknown Function | 9 | 23 | ||
12 | 10 | XF9a_02129 | XF_2237 | TonB-dependent receptor | 103 | 21 | 31|32 | |
13 | - | XF9a_01121 | XF_1217 | Protein of Unknown Function | 9 | 17 | ||
14 | 8 | XF9a_00936 | XF_1011 | Hemolysin toxin protein RTX | 173 | 5 | 0.57 | |
15 | 14 | XF9a_00892 | XF_0964 | Membrane lipoprotein | 19 | 24 | 0.56 | |
16 | 15 | XF9a_01736 | XF_1851 | Serine protease | 105 | 10 | 0.95 | |
17 | - | XF9a_01786 | XF_1896 | Outer membrane protein OmpA | 20 | 10 | 27|28 | |
18 | 21 | XF9a_00582 | XF_0644 | Peptidylprolyl isomerase Fkbp | 25 | 18 | 19|20 | |
19 | 19 | XF9a_00514 | XF_0565 | Lipid-binding SYLF | 32 | 13 | 22|23 | |
20 | 16 | XF9a_02272 | XF_2407 | Hemolysin toxin protein RTX | 219 | 5 | 0.69 | |
21 | 18 | XF9a_02555 | XF_2713 | Porin CirA | 96 | 15 | 24|25 | |
22 | 29 | XF9a_02447 | XF_2586 | Outer membrane export factor TolC | 49 | 23 | 21|22 | |
23 | 17 | XF9a_00557 | XF_0615 | Chaperonin GroEL | 58 | 16 | ||
24 | 20 | XF9a_01776 | XF_1887 | Cysteine/serine peptidase PS-46 | 79 | 16 | 22|23 | |
25 | 26 | XF9a_01787 | XF_1897 | protein TolB | 48 | 17 | 0.78 | |
26 | - | XF9a_02412 | XF_2548 | Succinyl-CoA ligase SucD | 30 | 21 | 0.64 | |
27 | 30 | XF9a_00323 | XF_0357 | Lipase/Esterase LesA | 42 | 15 | 0.90 | |
28 | - | XF9a_01186 | XF_1297 | Gluconolactonase | 37 | 5 | 25|26 | |
29 | 23 | XF9a_01725 | XF_1840 | Protein of Unknown Function | 25 | 7 | 0.90 | |
30 | - | XF9a_01712 | XF_1827 | Organic hydroperoxide reductase OsmC/OhrA | 15 | 20 | ||
31 | - | XF9a_00558 | XF_0616 | Chaperonin GroES | 10 | 39 | ||
32 | - | XF9a_01475 | XF_1604 | Glutathione peroxidase | 21 | 16 | 27|28 | |
33 | - | XF9a_00607 | XF_0672 | Acyl carrier protein AcpP | 9 | 24 | ||
34 | 22 | XF9a_00073 | XF_0082 | Fimbrial chaperone protein PapD | 29 | 15 | 30|31 | |
35 | - | XF9a_00777 | XF_0855 | Lipoprotein NlpD | 26 | 8 | 26|27 | |
36 | - | XF9a_00319 | XF_0353 | Translation initiation inhibitor | 14 | 18 | ||
37 | 25 | XF9a_02217 | XF_2340 | Chaperone protein DnaK | 68 | 15 | ||
38 | - | XF9a_01697 | XF_1811 | Outer membrane protein Slp | 18 | 17 | 0.91 | |
39 | - | XF9a_01043 | XF_1133 | Tryptophan repressor binding protein WrbA | 20 | 10 | 0.92 | |
40 | 24 | XF9a_01514 | XF_1649 | Protein of Unknown Function | 30 | 11 | ||
41 | - | XF9a_00742 | XF_0816 | Zn-dependent peptidase | 108 | 4 | ||
42 | - | XF9a_00954 | XF_1036 | Porin | 111 | 9 | 33|34 | |
43 | 27 | XF9a_00221 | XF_0239 | Polyribonucleotide nucleotidyltransferase Pnp | 76 | 7 | ||
44 | 28 | XF9a_00502 | XF_0550 | TonB-dependent receptor | 114 | 4 | 34|35 | |
45 | - | XF9a_02478 | XF_2622 | Cold shock protein, CspA | 9 | 21 | 0.89 | |
46 | - | XF9a_02496 | XF_2640 | Elongation factor Tu tufA | 43 | 16 | ||
47 | - | XF9a_01153 | XF_1253 | Acetyl esterase/lipase | 35 | 6 | 0.59 | |
48 | - | XF9a_00069 | XF_0078 | Fimbrial protein MrkD | 37 | 6 | 37|38 | |
49 | - | XF9a_02168 | XF_2283 | Beta-lactamase-like | 34 | 8 | 23|24 | |
50 | - | XF9a_01476 | XF_1605 | Fkbp-type peptidyl-prolyl cis-trans isomerase FkpA | 32 | 7 | 0.74 | |
51 | - | XF9a_00181 | XF_0196 | DUF2059 | 20 | 20 | 26|27 | |
52 | - | XF9a_00120 | XF_0138 | Aminopeptidase PepA | 52 | 12 | ||
53 | - | XF9a_00760 | XF_0838 | Chaperone SurA | 51 | 5 | 25|26 | |
54 | 32 | XF9a_00744 | XF_0820 | Zn-dependent amino- or carboxypeptidase | 58 | 9 | 0.52 | |
55 | - | XF9a_01399 | XF_1527 | Type II protein GspD/PulD | 81 | 3 | 0.95 | |
56 | - | XF9a_02411 | XF_2547 | Succinate--CoA ligase subunit beta SucC | 41 | 7 | ||
57 | - | XF9a_01116 | XF_1211 | Malate dehydrogenase Mdh | 35 | 8 | ||
58 | - | XF9a_01785 | XF_1895 | Tol-pal System protein YbgF | 30 | 8 | 23|24 | |
59 | - | XF9a_00964 | XF_1046 | Outer membrane protein BamA | 88 | 6 | 26|27 | |
60 | - | XF9a_01449 | XF_1577 | DUF2184 | 37 | 6 | 0.53 | |
61 | 31 | XF9a_02619 | XF_2773 | Hemagglutinin HxfA | 361 | 1 | 0.96 | |
62 | - | XF9a_00008 | XF_0007 | Protein of Unknown Function | 44 | 6 | 0.75 | |
63 | - | XF9a_00264 | XF_0290 | Aconitase | 98 | 3 | 0.78 | |
64 | - | XF9a_00072 | XF_0081 | Outer membrane FimD | 98 | 2 | 0.91 |
MC | CC | CDS Number Mb * | CDS Number “Old NCBI Locustag” Mb * | Description and Symbol ** | Predicted Molecular Mass kDa | Sequence Coverage (%) | SignalP | SecP Score |
---|---|---|---|---|---|---|---|---|
1 | - | MMSR116_RS18845 | MMSR116_19085 | Flagellin | 41 | 16 | 0.97 | |
2 | 2 | MMSR116_RS23965 | MMSR116_24295 | Cytochrome c class I | 13 | 15 | 22|23 | |
3 | - | MMSR116_RS28515 | MMSR116_28855 | Uncharacterized protein | 12 | 10 | 24|25 | |
4 | 3 | MMSR116_RS16505 | MMSR116_16705 | Formaldehyde-activating enzyme | 18 | 8 | 0.82 | |
5 | 10 | MMSR116_RS16515 | MMSR116_16715 | Chaperonin GroES | 11 | 21 | ||
6 | - | MMSR116_RS22410 | MMSR116_22715 | Uncharacterized protein | 17 | 19 | 25|26 | |
7 | - | MMSR116_RS23130 | MMSR116_23445 | Peptidyl-prolyl cis-trans isomerase | 19 | 19 | 22|23 | |
8 | 9 | MMSR116_RS12040 | MMSR116_12205 | Thioredoxin Trx | 12 | 20 | 0.73 | |
9 | 5 | MMSR116_RS00465 | MMSR116_00470 | Porin | 29 | 19 | 20|21 | |
10 | - | MMSR116_RS24205 | MMSR116_24535 | Uncharacterized protein | 15 | 14 | 23|24 | |
11 | 8 | MMSR116_RS12140 | MMSR116_12305 | Uncharacterized protein | 10 | 12 | 23|24 | |
12 | 22 | MMSR116_RS25265 | MMSR116_25605 | Elongation factor Tuf1 | 43 | 6 | ||
13 | - | MMSR116_RS13860 | MMSR116_14035 | Signal peptide protein | 19 | 20 | 21|22 | |
14 | 6 | MMSR116_RS20185 | MMSR116_20450 | Extracellular solute-binding protein | 33 | 23 | 27|28 | |
15 | 12 | MMSR116_RS27490 | MMSR116_27830 | Multiple sugar-binding periplasmic receptor ChvE | 38 | 15 | 27|28 | |
16 | 7 | MMSR116_RS07030 | MMSR116_07145 | Glyceraldehyde-3-phosphate dehydrogenase | 36 | 13 | ||
17 | 4 | MMSR116_RS11970 | MMSR116_12135 | Cysteine synthase A | 34 | 14 | ||
18 | - | MMSR116_RS17175 | MMSR116_17365 | Peptidase PepSY | 20 | 21 | 21|22 | |
19 | 13 | MMSR116_RS09940 | MMSR116_10105 | Chaperone protein DnaK | 69 | 10 | ||
20 | - | MMSR116_RS16510 | MMSR116_16710 | Chaperone protein GroEL | 58 | 20 | ||
21 | 1 | MMSR116_RS07550 | MMSR116_07665 | Extracellular ligand-binding receptor | 39 | 26 | 20|21 | |
22 | 14 | MMSR116_RS09540 | MMSR116_09680 | NAD-binding 6-phosphogluconate dehydrogenase | 30 | 14 | ||
23 | - | MMSR116_RS26210 | MMSR116_26565 | Catalase-related peroxidase | 35 | 8 | 19|20 | |
24 | - | MMSR116_RS09370 | MMSR116_09510 | Superoxide dismutase | 23 | 9 | 0.51 | |
25 | - | MMSR116_RS25335 | MMSR116_25675 | Transcription elongation factor GreA | 16 | 13 | ||
26 | - | MMSR116_RS18790 | MMSR116_19035 | Uncharacterized protein | 8 | 28 | 23|24 | |
27 | - | MMSR116_RS03745 | MMSR116_03785 | Uncharacterized protein | 23 | 7 | 23|24 | |
28 | - | MMSR116_RS24405 | MMSR116_24735 | Transcriptional regulator, MucR family | 18 | 10 | 0.96 | |
29 | - | MMSR116_RS12930 | MMSR116_13100 | Uncharacterized protein | 22 | 9 | 27|28 | |
30 | - | MMSR116_RS09955 | MMSR116_10120 | Redoxin | 17 | 11 | ||
31 | - | MMSR116_RS23810 | MMSR116_24135 | Citrate synthase | 48 | 3 | ||
32 | - | MMSR116_RS14870 | MMSR116_15050 | Uncharacterized protein | 17 | 16 | 36|37 | |
33 | - | MMSR116_RS01755 | MMSR116_01770 | Alanine racemase domain-containing protein | 40 | 12 | ||
34 | 15 | MMSR116_RS14980 | MMSR116_15160 | Phosphoenolpyruvate carboxykinase PckA | 59 | 6 | ||
35 | 18 | MMSR116_RS20370 | MMSR116_20635 | Malate dehydrogenase Mdh | 34 | 7 | ||
36 | - | MMSR116_RS20190 | MMSR116_20455 | branched-chain amino acid ABC transporter substrate-binding protein | 43 | 9 | 25|26 | |
37 | 20 | MMSR116_RS00865 | MMSR116_00880 | Methanol/ethanol family PQQ-dependent dehydrogenase | 69 | 3 | 24|25 | |
38 | 19 | MMSR116_RS20480 | MMSR116_20745 | Polyribonucleotide nucleotidyltransferase Pnp | 80 | 3 | ||
39 | 23 | MMSR116_RS29630 | MMSR116_29970 | Ketol-acid reductoisomerase IlvC | 37 | 5 | ||
40 | - | MMSR116_RS11180 | MMSR116_11330 | Adenylosuccinate synthetase PurA | 48 | 6 | ||
41 | - | MMSR116_RS21010 | MMSR116_21290 | Methylenetetrahydrofolate dehydrogenase | 29 | 8 | 0.81 | |
42 | - | MMSR116_RS05635 | MMSR116_05745 | 30S ribosomal protein S1 RpsA | 63 | 3 | ||
43 | - | MMSR116_RS03750 | MMSR116_03790 | Aconitate hydratase | 97 | 3 | 0.60 | |
- | 11 | MMSR116_RS21075 *** | MMSR116_21360 | Inorganic pyrophosphatase Ppa | 20 | 8 | ||
44 | - | MMSR116_RS13495 | MMSR116_13665 | Pyruvate, phosphate dikinase | 97 | 2 | ||
- | 16 | MMSR116_RS19390 *** | MMSR116_19640 | Adenosylhomocysteinase AhcY | 51 | 5 | ||
- | 17 | MMSR116_RS19735 *** | MMSR116_19990 | Phosphate-binding protein PstS | 37 | 6 | 23|24 | |
- | 22 | **** | MMSR116_18145 *** | Porin | 59 | 3 | 0.89 |
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Dourado, M.N.; Pierry, P.M.; Feitosa-Junior, O.R.; Uceda-Campos, G.; Barbosa, D.; Zaini, P.A.; Dandekar, A.M.; da Silva, A.M.; Araújo, W.L. Transcriptome and Secretome Analyses of Endophyte Methylobacterium mesophilicum and Pathogen Xylella fastidiosa Interacting Show Nutrient Competition. Microorganisms 2023, 11, 2755. https://doi.org/10.3390/microorganisms11112755
Dourado MN, Pierry PM, Feitosa-Junior OR, Uceda-Campos G, Barbosa D, Zaini PA, Dandekar AM, da Silva AM, Araújo WL. Transcriptome and Secretome Analyses of Endophyte Methylobacterium mesophilicum and Pathogen Xylella fastidiosa Interacting Show Nutrient Competition. Microorganisms. 2023; 11(11):2755. https://doi.org/10.3390/microorganisms11112755
Chicago/Turabian StyleDourado, Manuella Nobrega, Paulo Marques Pierry, Oseias Rodrigues Feitosa-Junior, Guillermo Uceda-Campos, Deibs Barbosa, Paulo A. Zaini, Abhaya M. Dandekar, Aline Maria da Silva, and Welington Luiz Araújo. 2023. "Transcriptome and Secretome Analyses of Endophyte Methylobacterium mesophilicum and Pathogen Xylella fastidiosa Interacting Show Nutrient Competition" Microorganisms 11, no. 11: 2755. https://doi.org/10.3390/microorganisms11112755