Natural Transformation as a Mechanism of Horizontal Gene Transfer in Aliarcobacter butzleri
Abstract
1. Introduction
2. Results
2.1. Screening for Aliarcobacter butzleri Isolates with Detectable Levels of Natural Transformation
2.2. Effect of Growth Conditions in the Transformation Frequency of Aliarcobacter butzleri
2.3. Effect of DNA Concentration on Transformation Frequency of Aliarcobacter butzleri
2.4. Transformation of Aliarcobacter butzleri with Different Donor DNA
2.5. Kinetics of Natural Transformation
2.6. Transfer of Antibiotic Resistance Determinants in Aliarcobacter butzleri Co-Cultures
2.7. Identification of Putative Competence Genes in Aliarcobacter butzleri DQ40A1
Campylobacter jejuni | Aliarcobacter butzleri RM4018 a | Aliarcobacter butzleri DQ40A1 (CDS) | ||||
---|---|---|---|---|---|---|
Category and Homolog (Locus ID) | Gene Product | Length (AA) | Homolog (Locus ID) | Homolog (Locus ID) | Length (AA) | Sequence ID (%) [DNA (AA)] b |
DNA Uptake | ||||||
ctsD (cj1474c) | Component of type II secretion/type IV pilus system (potential outer membrane pore) | 472 | ABU_RS09160 | GDI89_RS08280 | 478 | 46.8 (20.9) |
ctsP (cj1473c) | Component of type II secretion/type IV pilus system (putative NTPases/NTP binding protein) | 202 | NA | NA | NA | NA |
ctsX (cj1472c) | Component of type II secretion/type IV pilus system | 195 | NA | NA | NA | NA |
ctsE (cj1471c) | Component of type II secretion/type IV pilus system (putative NTPases/NTP binding protein) | 519 | ABU_RS08225 | GDI89_RS05100 | 453 | 53.5 (36.1) |
pilG c | Competence pseudopilus | 410 | ABU_RS08220 | GDI89_RS05105 | 396 | 38.4 (19.9) |
ctsG (cj1343c) | Putative periplasmic protein | 171 | ABU_RS03485 | GDI89_RS04080 | 132 | 47.5 (28.1) |
comEC (cj1211) | Membrane transporter protein involved in the transfer of DNA across the membrane | 419 | ABU_RS06285 | GDI89_RS02780 | 409 | 56.6 (36.2) |
comE (cj0011c) | Periplasmic DNA-binding competence protein | 79 | ABU_RS11390 | GDI89_RS07505 | 123 | 43.7 (31.7) |
priA c | Helicase | 729 | ABU_RS06960 | GDI89_RS02485 | 599 d | 36.6 (26.6) |
DNA Processing | ||||||
dprA (cj0634) | DNA processing protein | 257 | ABU_RS01180 | GDI89_RS06255 | 258 | 57.3 (41.3) |
recA | DNA recombination protein | 343 | ABU_RS11180 | GDI89_RS02220 | 349 | 73.5 (73.4) |
Others | ||||||
ctsT (cj1077) | Putative periplasmic protein | 100 | NA | NA | NA | NA |
ctsW (cj1028c) | Purine/pyrimidine phosphoribosyltransferase | 191 | ABU_RS02380 | GDI89_RS03920 | 190 | 59.2 (44.0) |
ctsR (cj1475c) | Hypothetical protein | 105 | NA | NA | NA | NA |
proC (cj1076) | Putative PCA reductase | 243 | ABU_RS02955 | NZ_CABVRU010000127.1 + NZ_CABVRU010000230.1 e | 254 | 53.1 (40.5) |
ceuB (cj1352) | Enterochelin uptake permease | 322 | ABU_RS05530 | GDI89_RS07350 | 320 | 49.1 (25.4) |
3. Discussion
4. Materials and Methods
4.1. Bacterial Strains and Growth Conditions
4.2. Genetic Material Preparation
4.3. Natural Transformation Using the Agar Transformation Method
4.4. Natural Transformation Using a Biphasic System
4.5. Effect of Growth Conditions
4.6. Kinetics of Natural Transformation
4.7. Influence of Genetic Material on Natural Transformation
4.8. Exchange of Genetic Material Between Bacterial Co-Cultures
4.9. Bioinformatics Analyses—Identification of Homologs Genes Presence/Absence
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Recipient A. butzleri Strain | Origin/Year of Isolation | ST a | Resistance Profile a | Agar Transformation Method | Biphasic System Method |
---|---|---|---|---|---|
1426_2003 | Diarrheic human stool/2003 | 47 | NAL, CTA b | + + - | - - - |
Ab_1711 | Poultry slaughterhouse equipment surface/2011 | ST_new1 | LEV, CIP, NAL, CTA | - - - | - - - |
Ab_2211 | Slaughterhouse surface/2011 | 460 | AMP, NAL, CTA | - - - | - - - |
Ab_2811 | Poultry carcass neck skin/2011 | 107 | AMP, ERY, LEV, CIP, NAL, CTA | - + + | + - - |
Ab_4211 | Poultry carcass drippings/2011 | ST_new2 | AMP, LEV, CIP, NAL, CTA | + + + | - + + |
Ab_4511 | Poultry carcass drippings/2011 | 510 | AMP, NAL, CTA | + + - | + + + |
CR424 | Poultry meat/2015 | ST_new3 | AMP, ERY, NAL, CTA | - - - | - - - |
CR502 | Poultry meat/2015 | ST_new4 | ERY, LEV, CIP, NAL, CTA | - - - | - - - |
CR604 | Beef meat/2015 | ST_new5 | NAL, CTA | + + + | + + + |
CR641 | Poultry meat/2015 | 108 | ERY, NAL, CTA | + - - | - - - |
CR891 | Poultry meat/2016 | 94 | NAL, CTA | - - - | - - - |
CR892 | Poultry meat/2016 | ST_new6 | AMP, LEV, CIP, NAL, CTA | + + + | - + - |
CR1132 | Ready-to-eat vegetables/2016 | ST_new7 | NAL, CTA | + + + | + + + |
CR1143 | Poultry meat/2016 | ST_new8 | AMP, LEV, CIP, NAL, CTA | + + + | + - + |
DQ20dA1 | Goat milk/2015 | ST_new9 | AMP, LEV, CIP, NAL, CTA | - - - | - - - |
DQ31A1 | Sheep milk/2015 | 172 | AMP, NAL, CTA | + + + | - - - |
DQ40A1 | Dairy plant equipment surface/2015 | ST_new10 | NAL, CTA | + + + | + + + |
Donor DNA (Nature of DNA/Strain) | Resistance Marker | Length of the PCR Fragment | Transformants/CFU |
---|---|---|---|
gDNA/Campylobacter coli 873 | aphA-3 | - | NT |
gyrA PCR fragment/Ab_2811 strain | C254T in gyrA gene | 344 bp | (6.70 ± 2.72) × 10−9 |
gyrA PCR fragment/Ab_2811 strain | C254T in gyrA gene | 1410 pb | (7.85 ± 1.13) × 10−8 |
gDNA/CR1132ΔareB::kanR | aphA-3 (KanR cassette from pUC18-K2) | - | (2.59 ± 0.51) × 10−7 |
PCR fragment/of DQ40A1ΔareB::kanR | aphA-3 (KanR cassette from pUC18-K2), flanked by upstream and downstream regions of about 400 bp | 1638 bp | (3.96 ± 3.54) × 10−7 |
gDNA/Ab_2811 strain | C254T in gyrA gene | - | (6.92 ± 7.67) × 10−7 |
gDNA/DQ40A1ΔareB::kanR | aphA-3 (KanR cassette from pUC18-K2) | - | (7.65 ± 2.25) × 10−6 |
Primers | Sequence | Size of Amplified Fragment (bp) | Reference |
---|---|---|---|
gyrA_Fw | TGGACGTGCATTACCAGATG | 1410 | [50] |
gyrA_Rv | GCAACTTTTCCTTTTCCACCT | ||
F-QRDR | TGGATTAAAGCCAGTTCATAGAAG | 344 | [51] |
R2-QRDR | TCATMGWATCATCATAATTTGGWAC | ||
areB_A1 | TTGAAATAAGGGCTACTTACTCAGG | 1638 | [49] |
areB_B2 | GTTCGCTCTGGCTTGCAAAT |
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Bonifácio, M.; Mateus, C.; Alves, A.R.; Maldonado, E.; Duarte, A.P.; Domingues, F.; Oleastro, M.; Ferreira, S. Natural Transformation as a Mechanism of Horizontal Gene Transfer in Aliarcobacter butzleri. Pathogens 2021, 10, 909. https://doi.org/10.3390/pathogens10070909
Bonifácio M, Mateus C, Alves AR, Maldonado E, Duarte AP, Domingues F, Oleastro M, Ferreira S. Natural Transformation as a Mechanism of Horizontal Gene Transfer in Aliarcobacter butzleri. Pathogens. 2021; 10(7):909. https://doi.org/10.3390/pathogens10070909
Chicago/Turabian StyleBonifácio, Marina, Cristiana Mateus, Ana R. Alves, Emanuel Maldonado, Ana P. Duarte, Fernanda Domingues, Mónica Oleastro, and Susana Ferreira. 2021. "Natural Transformation as a Mechanism of Horizontal Gene Transfer in Aliarcobacter butzleri" Pathogens 10, no. 7: 909. https://doi.org/10.3390/pathogens10070909
APA StyleBonifácio, M., Mateus, C., Alves, A. R., Maldonado, E., Duarte, A. P., Domingues, F., Oleastro, M., & Ferreira, S. (2021). Natural Transformation as a Mechanism of Horizontal Gene Transfer in Aliarcobacter butzleri. Pathogens, 10(7), 909. https://doi.org/10.3390/pathogens10070909