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Bestimmung der minimalen Hemmkonzentration gegenüber antimikrobiellen Wirkstoffen mit dem Verfahren der Bouillon-Mikrodilution bei pathogenen Bakterien von Fischen und molekulare Charakterisierung von Resistenzgenen
| Content Provider | Semantic Scholar |
|---|---|
| Author | Czapiewski, Ellen Von |
| Copyright Year | 2010 |
| Abstract | During the period 2004-2006, fish-pathogenic bacteria from Germany have been investigated for the first time in the national resistance monitoring program GERM-Vet for their in-vitro susceptibility to antimicrobial agents. In total, 206 isolates, obtained from commercially reared fish and from hobby fish, were included in this study. Susceptibility testing was conducted by broth microdilution according to the recommendations given in the CLSI document M49-A. An incubation temperature of 22 ± 2°C and a panel of 22 antimicrobial agents and two combinations of antimicrobial agents were used. The test population comprised 173 motile Aeromonas spp. isolates, 13 Aeromonas salmonicida isolates and 20 Yersinia ruckeri isolates. While the majority of the Aeromonas spp. isolates originated from hobby fish, isolates of the other groups were obtained from commercially reared fish. Aeromonas spp. isolates (n=44), which have been subjected to further molecular analyses based on their high minimum inhibitory concentrations (MICs) of specific antimicrobial agents, were confirmed for their species assignment by sequence analysis of part of the gyrB gene. These 44 isolates consisted of 19 A. hydrophila isolates [hybridisation group 1 (HG1)], 15 A. veronii biovar sobria isolates (HG8), five A. veronii biovar veronii isolates (HG10), two A. sobria (HG7) and A. bestiarum isolates (HG2) each, as well as a single A. caviae isolate (HG4). For the control of bacterial infections in fish, solely the combination trimethoprim/ sulfamethoxazol (1:19; SXT) has been approved in Germany. High SXT MIC values of >32/608mg/L were seen for 30 Aeromonas spp. isolates and two A. salmonicida isolates. All 32 isolates also carried the sulfonamide resistance gene sul1 as confirmed by PCR. Of them, 27 isolates (25 Aeromonas spp. and the two A. salmonicida isolates) harboured a single class 1 integron while four Aeromonas spp. isolates carried two class 1 integons. Among these class 1 integrons, seven different gene cassettes with the trimethoprim resistance genes dfrA1, dfrA12, dfrA14, dfrA28, dfrB1, dfrB3 or dfrB4 were identified by sequence analysis. PCR analysis confirmed the presence of the trimethoprim resistance gene dfrA1 outside of a class 1 integron in three SXT-resistant Aeromonas spp. isolates. Two of these isolates, however, carried class 1 integrons with other gene cassettes. A total of 12 different gene cassette arrangements were detected: dfrA28-orfV (n=1), dfrA12-orfF (n=1), orf-aadA5 (n=1), dfrB3-aadA1 (n=1), dfrA1-aadA1 (n=4), dfrA14-recombined aadA6 (n=1), catB3-aadA2 (n=3), dfrA12-orfF-aadA2 (n=18), dfrB4-catB3-aadA1 (n=1), dfrB1-aadA1-catB2 (n=2), dfrA12-orfF with integrated IS630-aadA2 (n=1) as well as arr-3-aacA4-blaOXA-10-aadA1 (n=1). High MIC values of other antimicrobial agents were also recorded for some of the isolates tested. Eight Aeromonas spp. isolates showed MIC values of ≥ 64 mg/L for the aminocyclitol antibiotic apramycin. In seven of these eight isolates, the apramycin resistance gene aac(3´)-IV was identified by PCR. Eleven Aeromonas spp. isolates exhibited high chloramphenicol MICs of ≥ 256 mg/L (but low MICs of 1 – 4 mg/L for florfenicol) and harboured a catA2 gene. In contrast, two A. salmonicida isolates with chloramphenicol MICs of 64 mg/L were negative by PCR for the chloramphenicol resistance genes catA1, catA2, catA3, cmlA/cmlB and oqxAB. Another 13 Aeromonas spp. isolates exhibited elevated florfenicol MIC values of 16 – > 64 mg/L and carried the resistance gene floR as confirmed by PCR. Seven of these isolates had an additional catA2 gene and chloramphenicol MIC values of ≥ 128 mg/L. All Aeromonas spp. isolates with tetracycline MIC values of ³ 1 mg/L were tested for the presence of the tetracycline resistance genes tet(A)-(E) and tet(Y). The gene tet(E) was seen most frequently in 64 isolates followed by the gene tet(A) (n=13). The genes tet(C) and tet(D) were identified in three and one isolates, respectively. Some isolates carried two different tet genes: tet(A) + tet(E) in four isolates, tet(C) + tet(E) and tet(D) + tet(E) in two isolates each, as well as tet(C) + tet(Y) in a single isolate. Of the four A. salmonicida isolates with tetracycline MICs of 8 or 16 mg/L, two isolates harboured tet(E), and single isolates tet(A) or tet(C), respectively. This study provided for the first time reliable and comparable data on (a) the susceptibility status of fish-pathogenic bacteria from Germany and (b) genes and associated mechanisms for resistance to selected antimicrobial agents. The association of trimethoprim/sulfonamide resistance with class 1 integrons, which in part carried gene cassettes for other resistance properties, bears the risk of co-selection for other resistance genes under the selective pressure imposed by the use of trimethoprim/sulfonamide combinations. |
| File Format | PDF HTM / HTML |
| Alternate Webpage(s) | https://elib.tiho-hannover.de/dissertations/czapiewskie_ss10.pdf |
| Language | English |
| Access Restriction | Open |
| Content Type | Text |
| Resource Type | Article |
