Campylobacter jejuni, a major foodborne human pathogen, has become increasingly resistant to fluoroquinolone (FQ) antimicrobials. By using clonally related isolates and genetically defined mutants, we determined the fitness of FQ-resistant Campylobacter in chicken (a natural host and a major reservoir for C. jejuni) in the absence of antibiotic selection pressure. When monoinoculated into the host, FQ-resistant and FQ-susceptible Campylobacter displayed similar levels of colonization and persistence in the absence of FQ antimicrobials. The prolonged colonization in chickens did not result in loss of the FQ resistance and the resistance-conferring point mutation (C257 3 T) in the gyrA gene. Strikingly, when coinoculated into chickens, the FQ-resistant Campylobacter isolates outcompeted the majority of the FQ-susceptible strains, indicating that the resistant Campylobacter was biologically fit in the chicken host. The fitness advantage was not due to compensatory mutations in the genes targeted by FQ and was linked directly to the single point mutation in gyrA, which confers on Campylobacter a high-level resistance to FQ antimicrobials. In certain genetic backgrounds, the same point mutation entailed a biological cost on Campylobacter, as evidenced by its inability to compete with the FQ-susceptible Campylobacter. These findings provide a previously undescribed demonstration of the profound effect of a resistance-conferring point mutation in gyrA on the fitness of a major foodborne pathogen in its natural host and suggest that the rapid emergence of FQ-resistant Campylobacter on a worldwide scale may be attributable partly to the enhanced fitness of the FQ-resistant isolates.colonization ͉ gyrA mutation ͉ poultry A ntimicrobial resistance in bacterial pathogens has become a serious threat to public health. There is a general notion that the acquisition of drug resistance, particularly the resistance mediated by chromosomal mutations, entails a biological cost for pathogens, resulting in reduced fitness in the absence of antibiotic selection pressure (1, 2). However, evidence has accumulated that in vivo-selected or clinically derived isolates may develop compensatory mutations that reduce the fitness cost associated with antimicrobial resistance (3-7). Different environments (in vitro vs. in vivo) may select for different compensatory mutations and varied levels of restoration of fitness (8). Even without compensatory mutations, drug-resistant mutants may show little or no fitness cost (9). One important finding revealed by previous studies is that clinically derived drugresistant isolates display diverse fitness changes, with some showing a biological cost and others showing no cost or even enhanced fitness (4, 10). The retention of ecological fitness in resistant pathogens creates a significant barrier for the elimination of resistant organisms by natural selection.Campylobacter jejuni, a Gram-negative microaerobic bacterium, is a common causative agent of human enterocolitis (11). For antibiotic treatment of ca...

Enrofloxacin treatment of chickens infected with fluoroquinolone(FQ)-sensitiveCampylobacter promoted the emergence of FQ-resistant Campylobacter mutants which propagated in the intestinal tract and recolonized the chickens. The recovered isolates were highly resistant to quinolone antibiotics but remained susceptible to non-FQ antimicrobial agents. Specific single-point mutations in the gyrA gene and the function of the CmeABC efflux pump were linked to the acquired FQ resistance. These results reveal that Campylobacter is hypermutable in vivo under the selection pressure of FQ and highlight the need for the prudent use of FQ antibiotics. Fluoroquinolone (FQ)-resistantCampylobacter jejuni strains are rapidly increasing throughout the world, which has posed a serious threat to public health (19,20). Although FQ resistance in Campylobacter can occur following the treatment of humans with the antibiotic (4, 17, 22), poultry are considered a significant source for FQ-resistant Campylobacter (1,5,7,18). Laboratory studies have demonstrated the emergence of FQ-resistant Campylobacter in experimental chickens treated with FQ antibiotics (10, 12). However, the previously published works revealed little information on the in vivo dynamics of the emergence of FQ-resistant Campylobacter in chickens. Specifically, it is unknown how Campylobacter populations shift in individual birds in response to the antibiotic treatment and how extensively the infected chickens are colonized by the resistant organisms. In addition, the genetic mechanisms responsible for the in vivo-acquired resistance to FQ in Campylobacter strains are not known. Answering these questions will greatly improve our understanding of the development and mechanisms of FQ resistance in Campylobacter and may facilitate the design of means to prevent the occurrence of FQresistant Campylobacter in vivo.In this study, we examined the dynamics of changes of Campylobacter populations in chickens treated with enrofloxacin and determined the molecular mechanisms associated with the acquired FQ resistance in the in vivo-selected resistant isolates. C. jejuni strain S3B was originally isolated from chicken feces in our laboratory. Bacterial cultures were routinely grown in Mueller-Hinton (MH) broth or plates (Becton Dickinson and Company, Sparks, Md.) at 42°C under microaerophilic conditions generated by the CampyPak Microaerophilic System (BBL). Day-old broiler chickens were obtained from a commercial hatchery. Prior to use, the birds were tested negative for Campylobacter by conventional culture methods. Two independent experiments (A and B) were conducted whose designs are detailed in Table 1. Each group of chickens was maintained in a sanitized wire-floored cage. Feed and water were provided ad libitum. Infection of the chickens with C. jejuni strain S3B (ciprofloxacin MIC ϭ 0.125 g/ml) and treatment with enrofloxacin (Baytril; Bayer Corporation) are detailed in Table 1. Cloacal swabs were collected periodically, resuspended in MH broth, and plated onto MH plates con...

Using laboratory challenge experiments, we examined whether Campylobacter-specific maternal antibody (MAB) plays a protective role in young chickens, which are usually free of Campylobacter under natural production conditions. Kinetics of C. jejuni colonization were compared by infecting 3-day-old broiler chicks, which were naturally positive for Campylobacter-specific MAB, and 21-day-old broilers, which were negative for Campylobacter-specific MAB. The onset of colonization occurred much sooner in birds challenged at the age of 21 days than it did in the birds inoculated at 3 days of age, which suggested a possible involvement of specific MAB in the delay of colonization. To further examine this possibility, specific-pathogen-free layer chickens were raised under laboratory conditions with or without Campylobacter infection, and their 3-day-old progenies with (MAB ؉ ) or without (MAB ؊ ) Campylobacter-specific MAB were orally challenged with C. jejuni. Significant decreases in the percentage of colonized chickens were observed in the MAB ؉ group during the first week compared with the MAB ؊ group. These results indicate that Campylobacter-specific MAB plays a partial role in protecting young chickens against colonization by C. jejuni. Presence of MAB in young chickens did not seem to affect the development of systemic immune response following infection with C. jejuni. However, active immune responses to Campylobacter occurred earlier and more strongly in birds infected at 21 days of age than those infected at 3 days of age. Clearance of Campylobacter infection was also observed in chickens infected at 21 days of age. Taken together, these findings (i) indicate that anti-Campylobacter MAB contributes to the lack of Campylobacter infection in young broiler chickens in natural environments and (ii) provide further evidence supporting the feasibility of development of immunization-based approaches for control of Campylobacter infection in poultry.

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Thirty-eight homozygous sickler sera were compared with a large pool of serum from healthy African non-sicklers with regard to bactericidal and phagocytic indices. One third of the sera showed reduced bactericidal activity against Salmonella enteritidis which was restored by the addition of 4% control serum; control serum provided both heat-labile (HL) and absorbable (ABS) serum factors. 76% of test sera showed greatly defective opsonization as indicated by ingestion by normal human neutrophils. Activity was not readily restored by the addition of control serum which provided only HL factors. Intracellular survival was increased when bacteria were ingested from sickler serum; activity was readily restored by control serum which provided both HL and ABS factors. In the presence of both serum and neutrophils 84% of test sera permitted increased bacterial survival; the defect was not readily reversed by the addition of control serum which provided both HL and ABS factors.