Background
Methods
Bacterial isolates
Antimicrobial susceptibility testing
Biofilm formation
DNA extraction
Detection of biofilm related genes
Target Genes | Primers sequences (5–3) | Annealing Temperature(°c) | DNA amplicon Size (bp) | Reference |
---|---|---|---|---|
epsA
| AGCAAGTGGTTATCCAATCG ACCAGACTCACCCATTACAT | 60 | 451 | [12] |
ompA
| CGCTTCTGCTGGTGCTGAAT CGTGCAGTAGCGTTAGGGTA | 58 | 531 | [12] |
bla
PER-1
| ATGAATGTCATTATAAAAGC AATTTGGGCTTAGGGCAAGAAA | 55 | 927 | [13] |
bap
| TACTTCCAATCCAATGCTAGGGAGGGTACCAATGCAG TTATCCACTTCCAATGATCAGCAACCAAACCGCTAC | 55 | 1225 | [14] |
bfmS
| TTGCTCGAACTTCCAATTTATTATAC TTATGCAGGTGCTTTTTTATTGGTC | 60 | 1428 | [14] |
ptk
| GGCTGAGCATCCTGCAATGCGT ACTTCTGGAGAAGGGCCTGCAA | 57 | 597 | [15] |
pgaB
| AAGAAAATGCCTGTGCCGACCA GCGAGACCTGCAAAGGGCTGAT | 57 | 490 | [15] |
fimH
| TGCAGAACGGATAAGCCGTGG GCAGTCACCTGCCCTCCGGTA | 60 | 870 | [16] |
csgA
| ACTCTGACTTGACTATTACC GATGCAGTCTGGTCAAC | 50 | 200 | [16] |
kpsMII
| GCGCATTTGCTGATACTGTTG CATCCAGACGATAAGCATGAGCA | 58 | 272 | [16] |
csuE
| ATGCATGTTCTCTGGACTGATGTTGAC CGACTTGTACCGTGACCGTATCTTGATAAG | 60 | 976 | [17] |
intI-1
| CAGTGGACATAAGCCTGTTC CCCGAGGCATAGACTGTA | 62 | 160 | [18] |
intI-2
| GTAGCAAACGAGTGACGAAATG CACGGATATGCGACAAAAAGGT | 62 | 788 | [18] |
intI-3
| GCCTCCGGCAGCGACTTTCAG ACGGATCTGCCAAACCTGACT | 62 | 979 | [18] |
Integron characterization
REP-PCR
Statistical analysis
Results
Characteristics of isolates
Antimicrobial susceptibility
No. of antimicrobial agents | Antibiotic resistance patterns | Percent (%) of all isolates |
---|---|---|
5 | Ciprofloxacin-Tobramycin-Gentamicin-Imipenem-Ampicillin/Sulbactam | 2 |
7 | Ceftazidime-Imipenem-Ciprofloxacin-Levofloxacin-Cotrimoxazole-Piperacillin-Cefepime | 2 |
8 | Ceftazidime-Imipenem-Doxycycline-Ciprofloxacin-Levofloxacin-Cotrimoxazole-Piperacillin-Cefepime | 5 |
Levofloxacin-Ampicillin/Sulbactam-Ceftazidime-Imipenem-Ciprofloxacin-Cefepime-Piperacillin-Cotrimoxazole | 8 | |
Ceftazidime-Imipenem-Doxycycline-Ciprofloxacin-Levofloxacin-Cotrimoxazole-Piperacillin-Cefepime | 1 | |
9 | Ampicillin/Sulbactam-Ceftazidime-Imipenem-Doxycycline-Ciprofloxacin-Levofloxacin-Cefepime-Piperacillin-Cotrimoxazole | 20 |
Ampicillin/Sulbactam-Ceftazidim-Imipenem-Gentamycin-Ciprofloxacin-Levofloxacin-Cotromoxazole-Piperacillin-Cefepime | 2 | |
Ceftazidime-Imipenem-Gentamicin-Tobramycin-Ciprofloxacin-Levofloxacin-Cotrimoxazole-Cefepime-Piperacillin | 4 | |
10 | Ampicillin/Sulbactam-Ceftazidime-Imipenem-Gentamicin-Tobramycin-Ciprofloxacin-Levofloxacin-Cotrimoxazole-Piperacillin-Cefepime | 20 |
Ampicillin/Sulbactam-Ceftazidime-Imipenem-Gentamicin-Tobramycin-Doxycycline-Ciprofloxacin-Levofloxacin-Piperacillin-Cefepime | 2 | |
Ceftazidime-Imipenem-Gentamicin-Tobramycin-Doxycycline-Ciprofloxacin-Levofloxacin-Cotrimoxazole-Piperacillin-Cefepime | 2 | |
11 | Ampicillin/Sulbactam-Ceftazidime-Imipenem-Gentamicin-Tobramycin-Doxycycline-Ciprofloxacin-Levofloxacin-Cotrimoxazole-Pipercillin-Cefepime | 32 |
Biofilm formation assay
Distribution of biofilm related genes
Biofilm related genes | No = (%) of isolates |
---|---|
pgaB | 98 |
ompA
| 81 |
Bap
| 43 |
bfmS
| 92 |
ptk
| 95 |
epsA
| 95 |
kpsMII
| 57 |
bla
PER-1
| 2 |
fimH
| 0 |
csgA
| 0 |
csuE
| 100 |
No. of biofilm related genes | Patterns of biofilm related genes | Percent (%) of A. baumannii |
---|---|---|
4 gene |
bfmS-epsA-pgaB-csuE
| 2 |
5 gene |
bfmS- csuE- ompA- ptk- pgaB
| 5 |
bfmS- csuE- epsA - ptk- pgaB
| 13 | |
csuE- epsA - ompA- ptk- pgaB
| 3 | |
csuE- epsA- bap - ompA- ptk
| 2 | |
6 gene |
bfmS- csuE- epsA - ompA- ptk- pgaB
| 8 |
csuE- epsA- bap - ompA- ptk-pgaB
| 3 | |
bfmS- csuE- epsA -kpsMT- ompA- pgaB
| 3 | |
bfmS- csuE- epsA- bap- ptk- pgaB
| 3 | |
7 gene |
csuE- epsA- bap -kpsMT- ompA- ptk-pgaB
| 2 |
bfmS- csuE - bap -kpsMT- ompA- ptk- pgaB
| 2 | |
bfmS- csuE- epsA -kpsMT- ompA- ptk- pgaB
| 21 | |
bfmS- csuE- epsA- bap - ompA- ptk- pgaB
| 3 | |
bfmS- csuE- epsA- bap -kpsMT- ptk - pgaB
| 2 | |
8 gene |
bfmS- csuE- epsA- bap -kpsMT- ompA- ptk- pgaB
| 26 |
9 gene |
bfmS- csuE- epsA-bla
PER-1
-bap -kpsMT- ompA- ptk- pgaB
| 2 |
Antibiotic resistance | Cefepime (n = 97) | Piperacillin (n = 99) | Cotrimoxazole (n = 95) | Levofloxacin (n = 97) | Ciprofloxacin (n = 100) | Doxycycline (n = 62) | Tobramycin (n = 63) | Gentamicin (n = 67) | Imipenem (n = 100) | Ceftazidime (n = 97) | Ampicillin-sulbactam (n = 87) |
---|---|---|---|---|---|---|---|---|---|---|---|
Biofilm related genes | |||||||||||
csuE
| 95 | 99 | 95 | 96 | 100 | 62 | 63 | 65 | 100 | 96 | 87 |
pgaB
| 95 | 98 | 92 | 95 | 98 | 61 | 62 | 63 | 98 | 95 | 86 |
ompA
| 76 | 79 | 74 | 76 | 79 | 62 | 44 | 44 | 79 | 76 | 70 |
bap
| 41 | 43 | 41 | 43 | 42 | 31 | 27 | 27 | 43 | 43 | 35 |
bfmS
| 87 | 90 | 85 | 87 | 90 | 41 | 35 | 33 | 90 | 87 | 77 |
ptk
| 92 | 95 | 90 | 92 | 95 | 60 | 61 | 62 | 95 | 93 | 84 |
epsA
| 92 | 93 | 90 | 92 | 93 | 41 | 35 | 35 | 93 | 92 | 79 |
kpsMT
| 55 | 57 | 54 | 55 | 57 | 49 | 30 | 33 | 57 | 55 | 49 |
bla
PER-1
| 1 | 2 | 1 | 1 | 1 | 1 | 0 | 0 | 2 | 1 | 1 |
fimH
| 0 | 0 | 0 | 0 | 0 | 0 | 0 | 0 | 0 | 0 | 0 |
csgA
| 0 | 0 | 0 | 0 | 0 | 0 | 0 | 0 | 0 | 0 | 0 |
Integron characterization
REP-PCR
REP Types | Subtypes | Size of REP-PCR products (bp) | Total isolates number (%) |
---|---|---|---|
A (3 bands) (24%) | A1 | 800, 1000, 3000 | 23 |
A2 | 700, 1500, 2500 | 1 | |
B (4 bands) (4%) | B1 | 700, 850, 1000, 1600 | 3 |
B2 | 900, 1400, 1800, 3000 | 1 | |
C (5 bands) (27%) | C1 | 900, 950, 1000, 1500, 2000 | 15 |
C2 | 800, 1000, 1500, 2500, 3000 | 9 | |
C3 | 700, 1600, 2000, 3000, 4000 | 1 | |
C4 | 550, 700, 800, 1000, 3100 | 1 | |
C5 | 750, 1000, 1900, 2900, 3000 | 1 | |
D (6 bands) (14%) | D1 | 800, 1300, 2200, 2500, 3000, 3400 | 4 |
D2 | 700, 1000, 1900, 2300, 3000, 3300 | 4 | |
D3 | 650, 900, 1900, 2700, 3000, 4750 | 3 | |
D4 | 900, 1000, 1200, 1500, 2000, 3000 | 3 | |
E (7 bands) (16%) | E1 | 750,800,1000,1300,1500,2500,3000 | 6 |
E2 | 650,850,900,1300,2000,2800,3000 | 9 | |
E3 | 250,450,900,1400,2100,2700,3000 | 1 | |
F (8 bands) (7%) | F1 | 600,700,900,1200,1600,2000,2800,3200 | 3 |
F2 | 100,300,400,800,1000,1500,2800,3000 | 4 | |
G (9 bands) (7%) | G1 | 300,500,800,900,1600,2300,2800,3200,4300 | 4 |
G2 | 100,300,500,650,700,900,1100,1700,3000 | 3 | |
H (10 bands) (1%) | H | 100,250,500,700,800,1000,1500,2000,2700,3000 | 1 |
REP Types | pgaB (n = 98) |
ompA
(n = 81) |
Bap
(n = 43) |
bfmS
(n = 92) |
Ptk
(n = 95) |
epsA
(n = 95) |
kpsMII
(n = 57) |
bla
PER-1
(n = 2) |
fimH
(n = 0) |
csgA
(n = 0) |
csuE
(n = 100) |
---|---|---|---|---|---|---|---|---|---|---|---|
A (3 bands) (24%) | 24 | 23 | 19* | 22 | 24 | 23 | 18 | 2 | 0 | 0 | 24 |
B (4 bands) (4%) | 2 | 3 | 1 | 4 | 4 | 4 | 2 | 0 | 0 | 0 | 4 |
C (5 bands) (27%) | 27 | 22 | 8 | 25 | 25 | 26 | 17 | 0 | 0 | 0 | 27 |
D (6 bands) (14%) | 14 | 10 | 4 | 12 | 14 | 13 | 6 | 0 | 0 | 0 | 14 |
E (7 bands) (16%) | 16 | 12 | 6 | 16 | 15 | 15 | 8 | 0 | 0 | 0 | 16 |
F (8 bands) (7%) | 7 | 6 | 2 | 5 | 7 | 6 | 3 | 0 | 0 | 0 | 7 |
G (9 bands) (7%) | 7 | 5 | 2 | 7 | 6 | 7 | 2 | 0 | 0 | 0 | 7 |
H (10 bands) (1%) | 1 | 0 | 1 | 1 | 0 | 1 | 1 | 0 | 0 | 0 | 1 |