Investigation of community based microbiome and antibiotic resistance
Investigation of community based microbiome and antibiotic resistance Soojin Jang Antibacterial Resistance Research Laboratory We are Pasteurians fighting disease in Korea for all mankind
Seoul Sampling Day (June 29 th) Seoul Subway System • Total 18 lines 9 lines (in Seoul) + 9 lines (outside Seoul but connected to the main lines) • Total 541 stations Line 2 3 4 5 7 9 Total Institut Pasteur Korea - Over 10 Years in Korea Fighting Disease for All Mankind | Confidential The number Area of stations 51 (43) Seoul 44 Seoul 48 Seoul 50 (46) Seoul 51 Seoul 30 Seoul 274 (262) 2
Seoul Sampling Day (June 29 th) Seoul Subway System • 26 people (13 teams) Institut Pasteur Korea - Over 10 Years in Korea Fighting Disease for All Mankind | Confidential 3
Surveillance of antibiotic resistance in community Pilot study : Analysis of culturable bacteria and antibiotic resistance in a small community • Elevator buttons • Door handles • Surface of tables. . . Institut Pasteur Korea - Over 10 Years in Korea Fighting Disease for All Mankind | Confidential 4
The study process Sample collection Cultivation of bacteria Antibiotic susceptibility test Analysis of bacterial species profile (16 s r. RNA sequencing) Identification of resistant strains (whole genome sequencing) Institut Pasteur Korea - Over 10 Years in Korea Fighting Disease for All Mankind | Confidential 5
Analysis of the culturable bacterial community Average composition of the bacterial community Institut Pasteur Korea - Over 10 Years in Korea Fighting Disease for All Mankind | Confidential 6
Analysis of the culturable bacterial community Antibiotic susceptibility test of selected 52 colonies Resistance to at least one antibiotic 17/52 (33%) Multidrug resistance 5/52 (10%) Institut Pasteur Korea - Over 10 Years in Korea Fighting Disease for All Mankind | Confidential 7
Identification of strains TSA 6. 1 whole genome analysis • • Isolated from the elevator button Grew on TSA plate containing Ampicillin (50 ug/ml) or Chloramphenicol (35 ug/ml) Identified as Acinetobacter pittii by whole genome sequencing Total 4, 143, 470 bp (increased genome size by 30 ~300 kb) TSA 6. 1 Institut Pasteur Korea - Over 10 Years in Korea Fighting Disease for All Mankind | Confidential 8
Identification of strains Potential antibiotic resistant genes in TSA 6. 1 : Growth on TSA plates containing Ampicillin (50 ug/ml) or Chloramphenicol (35 ug/ml) Genes TSA 6. 1_00833 TSA 6. 1_00834 TSA 6. 1_00835 TSA 6. 1__00915 TSA 6. 1__00968 TSA 6. 1__01190 Blast. P Hit in CARD Ade. K Ade. J Ade. I multidrug efflux protein Ade. B amp. C protein TSA 6. 1__01190 TSA 6. 1__01348 TSA 6. 1__01349 TSA 6. 1__01350 TSA 6. 1__01356 TSA 6. 1__01696 TSA 6. 1__02202 TSA 6. 1__02732 TSA 6. 1__03368 TSA 6. 1__03369 TSA 6. 1__03386 class C extended-spectrum beta-lactamase ADC-18 putative Outer membrane efflux protein multidrug efflux protein Ade. H multidrug ABC transporter putative multidrug resistance efflux protein Ade. N beta-lactamase OXA-213 penicillin-binding protein 2' Mac. A Mac. B cfr* protein (23 S r. RNA (adenine(2503)-C(2))-methyltransferase Rlm. N ) multidrug efflux pump Abe. M aminoglycoside resistance protein TSA 6. 1__03492 TSA 6. 1__03729 Pathogen Acinetobacter baumannii AYE Acinetobacter baumannii SDF Acinetobacter baumannii AB 0057 Salmonella enterica subsp. enterica serovar Enteritidis Aeromonas dhakensis Acinetobacter baumannii AYE Acinetobacter baumannii SDF Acinetobacter baumannii AYE Acinetobacter baumannii Acinetobacter calcoaceticus Staphylococcus aureus Neisseria gonorrhoeae Staphylococcus sciuri Acinetobacter baumannii AYE Antibiotic Resitance Ontology ade. G ade. B ade. H ade. G ade. F abe. S ade. N OXA-213 mac. A mac. B cfr. A abe. M Bit. Score Percent. Identity 159 31. 1 734 41. 9 154 32. 2 622 37. 91 870 43. 93 280 37. 96 345 859 2007 750 206 364 500 150 264 700 171 47. 13 92. 68 97. 83 95. 07 96. 33 78. 8 91. 19 25. 92 39. 34 52. 58 32. 87 872 150 97. 54 40. 36 Mc. Arthur et al. 2013. The Comprehensive Antibiotic Resistance Database. Antimicrobial Agents and Chemotherapy, 57, 3348 -3357. http: //arpcard. mcmaster. ca Institut Pasteur Korea - Over 10 Years in Korea Fighting Disease for All Mankind | Confidential 9
Identification of strains TSA 15 & TSA 30 • Grew on TSA plate containing ampicillin (50 ug/ml) and kanamycin (50 ug/ml) for TSA 15 and ampicillin, kanamycin and tetracycline (15 ug/ml) for TSA 30 • Identified as Staphylococcus epidermids • TSA 15 has 2, 499, 996 bp genome TSA 15 with 30, 607 bp plasmid • TSA 30 has 2, 501, 949 bp genome TSA 30 with 21, 246 bp plasmid • 97 % of average nucleotide identity by BLASTn Institut Pasteur Korea - Over 10 Years in Korea Fighting Disease for All Mankind | Confidential 10
Identification of strains Comparative genome analysis TSA 30_small (21, 246 bp ) TSA 15_small (30, 607 bp ) • Toxin Yoe. B • GATA zinc finger domain-containing protein 14 • Antitoxin Yef. M • • Toxin RTX-III translocation ATP-binding protein Putative multidrug export ATP-binding/permease protein SA 1683 Lactococcin-G-processing and transport ATP-binding protein Lag. D Histone-lysine N-methyltransferase 2 D Protein rlx Probable replication protein rep Lactococcin-G-processing and transport ATP-binding protein Lag. D Bone sialoprotein-binding protein Institut Pasteur Korea - Over 10 Years in Korea Fighting Disease for All Mankind | Confidential 11
Summary of the pilot study Ø Various culturable bacteria including pathogenic bacteria are present in the sample community Ø 33 % of bacteria were resistant to at least one of tested antibiotics and 10 % of them were resistant to two antibiotics Ø The pilot study demonstrated the potential prevalence of antibiotic resistance in community and the importance of constant surveillance Ø Also, the study clearly showed a number of limitations including culture-dependent analysis and the small sample size Ø We successfully performed the 1 st city sampling! More to come! Institut Pasteur Korea - Over 10 Years in Korea Fighting Disease for All Mankind | Confidential 12
Acknowledgements Antibiotic Resistance Research Laboratory Yunmi Lee Hyungjun Kim Junhyung Park, Yunsun Cha, Takhyun Kim, & Hye-ri Lee (Interns) Dr. Christopher Mason Department of Physiology and Biophysics in Weill Cornell Medical College Meta. SUB consortium Institut Pasteur Korea - Over 10 Years in Korea Fighting Disease for All Mankind | Confidential 13
Thank you for your attention
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