Background
Methods
Clinical bacterial isolates and drug susceptibility testing
Plasmid digestion, integron cloning and DNA sequencing
Primer | Sequence (5′-3′) | Length (bp) | Reference | |
---|---|---|---|---|
5′-CS | F | GGCATCCAAGCAGCAAGC | Variable | [40] |
3′-CS | R | AAGCAGACTTGACCTGAT | ||
Int1 | F | AGCACCTTGCCGTAGAAGAACAG | 3500 | This study |
R | GTCATAATCGGTTATGGCATCGC | |||
intI1 | F | GGGTCAAGGATCTGGATTTCG | 1250 | [33] |
R | ACATGCGTGTAAATCATCGTCG | |||
sul1 | F | ATGGTGACGGTGTTCGGCATTCTGA | 900 | [34] |
R | TCTGGCTCCCAATCTAGTACGGATC | |||
qacEΔ1 | F | ATCGCAATAGTTGGCGAAGT | 600 | [35] |
R | CAAGCTTTTGCCCATGAAGC | |||
aadA | F | ACATCATTCCGTGGCGTTATC | 1100 | This study |
R | TTATTTGCCGACTACCTTGGTGA | |||
tnpA | F | GGCGGGATCTGCTTGTAGAG | 920 | [36] |
R | CTCCGGAGATGTCTGGCTTACT | |||
aad B | F | TCACAGCCAAACTATCAGG | 857 | This study |
R | TGCTCCACCAATCACAAT | |||
aacA4 | F | TGACCAACAGCAACGATTCC | 800 | [37] |
R | TTAGGCATCACTGCGTGTTC | |||
bla
OXA
| F | ATGAAAAACACAATACATATCAACTTCGC | 900 | This study |
R | GTGTGTTTAGAATGGTGATCGCATT | |||
bla
KPC
| F | TGTCACTGTATCGCCGTC | 1000 | [31] |
R | CTCAGTGCTCTACAGAAAACC | |||
bla
TEM
| F | CTGTCTATTTCGTTCATCC | 1061 | [24] |
R | CTCAGTATTGCCCGCTCC | |||
bla
VIM
| F | CACGAACCCAGTGGACATA | 1512 | This study |
R | TCACAGTAACCAGCAAATCA | |||
bla
IMP
| F | GCACGAACCCAGTTGACATA | 1394 | This study |
R | GTTTCTTCTTCCCACCATCC | |||
tniC | F | GCTCTGGTTGAGTTGGTG | 993 | This study |
R | GGATCCTTCCGCCTGTTG | |||
dfr | F | GTGAAAATATCACTAATGG | 750 | [38] |
R | TTAACCCCTTTGCCAGATTTG | |||
arr-3 | F | GGTGACTTGCTAACCACAG | 450 | This study |
R | ACAGTGACATAGCAAGTTCAG | |||
catB | F | CCTGAAGATTGCCAAGAGTGGT | 980 | [39] |
R | AGTTTGTTCAGGGTGACGAAGG |
Sequence annotation and genome comparison
Carbapenemase activity detection
Nucleotide sequence accession numbers
Results and discussion
Integron cloning experiments and antibiotic susceptibility testing
Strains | Plasmids | Antibiotic susceptibility testing (mg/L) | |||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Cephalosporins | Carbapenems | Aminoglycosides | Fluoroquinolones | ||||||||||
CZ | CAZ | CTX | ETM | MPM | IPM | NET | TOB | AK | NOR | OFL | CIP | ||
Nsa243 | In37
| 256/R | 128/R | 128/R | 16/R | 16/R | 8/R | 64/R | 32/R | 128/R | 0.10/S | 0.05/S | 0.25/S |
In37-TOP10 | 16/R | 8/R | 8/R | 8/R | 8/R | 4/R | 16/R | 16/R | 32/R | 0.05/S | 0.003/S | 0.125/S | |
TOP10 | 1/S | 0.5/S | 0.5/S | 0.5/S | 0.5/S | 0.25/S | 2/S | 0.025/S | 1/S | 0.05/S | 0.003/S | 0.125/S | |
Ecl175 | In843
| 2/S | 1/S | 1/S | 0.5/S | 0.5/S | 0.25/S | 512/R | 128/R | 512/R | 0.10/S | 0.05/S | 0.25/S |
In843-TOP10 | 1/S | 0.5/S | 0.5/S | 0.25/S | 0.25/S | 0.125/S | 128/R | 64/R | 128/R | 0.05/S | 0.003/S | 0.125/S | |
TOP10 | 1/S | 0.5/S | 0.5/S | 0.5/S | 0.5/S | 0.25/S | 64/R | 32/R | 64/R | 0.05/S | 0.003/S | 0.125/S | |
Kpn761 | In62
| 2/S | 1/S | 1/S | 0.5/S | 0.5/S | 0.25/S | 128/R | 64/R | 128/R | 0.10/S | 0.05/S | 0.25/S |
In62-TOP10 | 1/S | 0.5/S | 0.5/S | 0.25/S | 0.25/S | 0.125/S | 64/R | 32/R | 32/R | 0.05/S | 0.003/S | 0.125/S | |
TOP10 | 1/S | 0.5/S | 0.5/S | 0.5/S | 0.5/S | 0.25/S | 2/S | 0.025/S | 1/S | 0.05/S | 0.003/S | 0.125/S | |
Nsa217 | In1021
| 2/S | 1/S | 1/S | 0.5/S | 0.5/S | 0.25/S | 256/R | 128/R | 256/R | 0.10/S | 0.05/S | 0.25/S |
In1021-TOP10 | 1/S | 0.5/S | 0.5/S | 0.25/S | 0.25/S | 0.125/S | 128/R | 64/R | 128/R | 0.05/S | 0.003/S | 0.125/S | |
TOP10 | 1/S | 0.5/S | 0.5/S | 0.5/S | 0.5/S | 0.25/S | 2/S | 0.025/S | 1/S | 0.05/S | 0.003/S | 0.125/S | |
Eae634 | In846
| 256/R | 128/R | 128/R | 64/R | 64/R | 64/R | 128/R | 64/R | 128/R | 0.10/S | 0.05/S | 0.25/S |
In846-TOP10 | 64/R | 16/R | 16/R | 16/R | 16/R | 16/R | 64/R | 32/R | 64/R | 0.05/S | 0.003/S | 0.125/S | |
TOP10 | 1/S | 0.5/S | 0.5/S | 0.5/S | 0.5/S | 0.25/S | 2/S | 0.025/S | 1/S | 0.05/S | 0.003/S | 0.125/S | |
KP3349 | In1059
| 256/R | 128/R | 128/R | 64/R | 64/R | 128/R | 64/R | 128/R | 64/R | 0.10/S | 0.05/S | 0.25/S |
In1059-TOP10 | 32/R | 16/R | 16/R | 16/R | 16/R | 64/R | 32/R | 16/R | 16/R | 0.05/S | 0.003/S | 0.125/S | |
TOP10 | 1/S | 0.5/S | 0.5/S | 0.5/S | 0.5/S | 0.25/S | 2/S | 0.025/S | 1/S | 0.05/S | 0.003/S | 0.125/S |
Strain carbapenemase activities and genetic features of integrons
Integron | In1059 | In846 | In843 | In37 | In62 | In1021 |
---|---|---|---|---|---|---|
Pc variant | PcH1 | PcH1 | None | None | PcH1 | PcH1 |
“-35”: tggaca | “-35”: tggaca | “-35”: tggaca | “-35”: ggacat | |||
“-35”: taaact | “-10”: taaact | “-10”: ttcgta | “-10”: taaact | |||
P2 promoter | Absent | Absent | Absent | Absent | Absent | Absent |
PintI1 | Yes | Yes | Yes | Yes | Yes | Yes |
“-10”: agtcta | “-10”: agtcta | “-10”: agggcg | “-10”: agtcta | “-10”: agtcta | “-10”: agtcta | |
“-35”: cagcaa | “-35”: cagcaa | “-35”: catcgt | “-35”: cagcaa | “-35”: cagca | “-35”: cagcaa | |
19 bp ORF11 duplication | No | No | No | No | No | No |
intI1
|
IntI1
R32_H39
|
IntI1
R32_H39
| Remant of intI1
| Remant of intI1
|
IntI1
R32_H39
|
IntI1
R32_H39
|
1010 bp-length | 1014 bp-length | 4 bp-length | 720 bp-length | 1014 bp-length | 1014 bp-length | |
attI1
| 63 bp-length | 63 bp-length | 63 bp-length | 63 bp-length | 63 bp-length | 63 bp-length |
Array of gene cassettes | GCbla
VIM-5 - GCaacA4’-37- (tniC-aadA16)- (qacE
30
-aadB) | GCbla
VIM-1 - GCaacA4cr- (qacE
101-catB3) - GCarr-3
| GCaacA4cr - GCarr-3 - GCdfrA27 - (qacE
101-aadA16Δ::IS26) | GCaacA4cr - GCbla
OXA-1 - GCcatB3 - GCarr-3
| GCaacA4cr
| GCaacA4cr - (qacE
109
-arr-3) -
GCdfrA27 - (qacE
30
-aadA16) |
attC
|
attCbla
VIM-5(L;R)
attC
aacA4Δ(LΔ; R);
attC
aadA16/aacA4(L;R);
attC
aadBΔ(L; Del) |
attCbla
VIM-1(L;R);
attC
aacA4cr(L;R);
attC
catB3 (L;R);
attC
aacA4(LΔ; R) |
attC
aacA4cr (L; R);
attC
arr-3 (L;R);
attC
dfrA27 (L;R);
attC
aadA16 (L;R) |
attC
aacA4cr (L;R);
attCbla
OXA-1(L;R);
attC
catB3 (L;R);
attC
arr-3 (L; R) |
attC
aacA4cr (L; R) |
attC
aacA4cr (L;R);
attC
arr-3 (L; R);
attC
dfrA27 (L; R);
attC
aadA16(L;R) mutated |
3′-CS |
qacEΔ1-sul1
|
qacEΔ1 (partial) |
qacEΔ1 (partial) |
qacEΔ1-sul1
|
qacEΔ1-sul1
| IS15-sul1Δ-IS1 |