Clonal emergence of extended-spectrum beta-lactamase (CTX-M-2)-producing Salmonella enterica serovar Virchow isolates with reduced susceptibilities to ciprofloxacin among poultry and humans in Belgium and France (2000 to 2003)

Abstract

Antibiotic treatment is not required in cases of Salmonella enterica gastroenteritis but is essential in cases of enteric fever or invasive salmonellosis or in immunocompromised patients. Although fluoroquinolones and extended-spectrum cephalosporins are the drugs of choice to treat invasive Salmonella, resistance to these antibiotics is increasing worldwide. During the period 2000 to 2003, 90 Salmonella enterica serovar Virchow poultry and poultry product isolates and 11 serovar Virchow human isolates were found to produce an extended-spectrum beta-lactamase, CTX-M-2, concomitantly with a TEM-1 beta-lactamase. The bla(CTX-M-2) gene was located on a large conjugative plasmid (>100 kb). Pulsed-field gel electrophoresis indicated a clonal relationship between the poultry and human isolates. All these isolates displayed additional resistance to trimethoprim-sulfamethoxazole and tetracycline as well as a reduced susceptibility to ciprofloxacin (MICs of between 0.5 and 1 mug/ml). CTX-M-2-producing Salmonella with a reduced susceptibility to fluoroquinolones constitutes a major concern, since such strains could disseminate on a large scale and jeopardize classical antibiotic therapy in immunocompromised patients.

FIG. 1.

Percentage of S. enterica serovar Virchow strains isolated in Belgium from humans, poultry, and poultry products between 1999 and 2003. The percentages of S. enterica serovar Virchow strains isolated from poultry and poultry products show a significant increase over the years (P = 0.011 for poultry and P = 0.055 for poultry products), following similar trends (slopes are not significantly different). The linear regression lines, equations, and r² coefficients are shown.

FIG. 2.

Restriction analysis (EcoRI) (A) and Southern blot hybridization with a bla_CTX-M-2 probe (B) of plasmid DNAs isolated from E. coli transconjugants. Lanes M, markers (SmartLadder; Eurogentec, Seraing, Belgium); 1, E. coli transconjugant p142SA01-1; 2, E. coli transconjugant p1639SA00-1; 3, E. coli transconjugant p02-2928-1; 4, E. coli transconjugant p03-1902-1; 5, E. coli transconjugant p03-664-1; 6, E. coli transconjugant p03-5167-1. All lanes are from the same gel.

FIG. 3.

(A) Representative XbaI PFGE patterns observed among CTX-M-2-producing S. enterica serovar Virchow isolates and among serovar Virchow isolates used as comparison isolates. Lane M, XbaI-digested DNA from S. enterica serovar Braenderup H9812, used as molecular size marker; lane 1, isolate 03-3832; lane 2, isolate 03-5167; lane 3, isolate 03-12787; lane 4, isolate 02-2928; lane 5, isolate 03-664; lane 6, isolate 518SA03; lane 7, comparison isolate 02-4352 (B/2002/H/TEM-52 ESBL, +*); lane 8, isolate 01-2133; lane 9, comparison isolate 02-4366 (B/2002/H/CTX-M-3 ESBL, +*); lane 10, comparison isolate 03-1324 (F/2003/H/nalidixic acid resistant, −*); lane 11, comparison isolate 03-3111 (F/2003/H/nalidixic acid and ampicillin resistant, −*); lane 12, comparison isolate 03-4256 (F/2003/H/pansusceptible, −*). F, France; B, Belgium; V, veterinary isolate; H, human isolate. −*, comparison isolate susceptible to cefotaxime; +*, comparison isolate resistant to cefotaxime. (B) Dendrogram generated by FingerprintingII Informatix software, showing the results of cluster analysis on the basis of PFGE fingerprinting. Similarity analysis was performed using the Dice coefficient, and clustering was by the unweighted-pair group method using average linkages. The different PFGE profiles (asterisks indicate profiles of comparison isolates), numbers of isolates, years of isolation, types of bla genes, and phage types are indicated. ND, not determined.