Comparison of selective media for primary isolation of Campylobacter fetus subsp. jejuni.

Three selective media, Skirrow, Butzler, and a modification of Butzler medium, were compared for the primary isolation of Campylobacter fetus subsp. jejuni. This organism was isolated from 87 of 347 specimens (72 from 240 dogs rectal swabs and 15 from 107 cats rectal swabs). The positive rate for dogs (30%) was twice as high as that for cats (14%). Skirrow and Butzler media were comparable in their isolation of C. fetus subsp. jejuni. A significantly higher rate of positive results was obtained with modified Butzler medium. The best combination of two media was that of modified Butzler and Skirrow media, which detected 98% of the isolates obtained. The percentage of Campylobacter-positive specimens was increased by 9% by holding primary isolation plates 72 h.     

Comparison of basal media for culturing Campylobacter jejuni and Campylobacter coli.

Four strains of Campylobacter jejuni and four strains of Campylobacter coli were used to compare the quantitative growth of Campylobacter cells on blood agar base no. 2 (Oxoid), brucella agar (BBL Microbiology Systems and Difco Laboratories), campylobacter agar base (Difco), Columbia blood agar base (Difco and Oxoid), and Mueller-Hinton agar (Difco and Oxoid). Columbia blood agar base and blood agar base no. 2 were inhibitory to most of the strains tested, as evidenced by reduced (10- to 1,000-fold) colony counts compared with other basal media. One of the brucella agars was inhibitory to two of the C. coli strains. The inhibitory effect of these media could be eliminated by addition of FBP (0.05% each ferrous sulfate hydrate, sodium metabisulfite, and sodium pyruvate) or 7% defibrinated sheep blood. However, addition of FBP or blood to brucella agar, campylobacter agar base, or Mueller-Hinton agar did not significantly affect the count, indicating that supplements are not required in these media for growth of Campylobacter in pure culture.     

A selective medium for isolating Campylobacter jejuni/coli.

Skirrow's medium is effective for isolating Campylobacters from human faeces but is less suitable for animal and environmental specimens owing to the presence of contaminating species. After determining the sensitivity of 104 strains of Campylobacters to several antimicrobial agents, used singly and in various combinations, a selective medium incorporating polymixin, rifampicin, trimethoprim and actidione, was developed. The medium, called Preston medium, was shown to be more selective than Skirrow's medium and suitable for any kind of specimen.     

Inhibition of Campylobacter coli and Campylobacter jejuni by antibiotics used in selective growth media.

The ability of Campylobacter coli and Campylobacter jejuni to grow in the presence of antibiotics used in selective growth media was compared. MIC data for C. coli indicated that some strains were more susceptible to the antibiotics than were the C. jejuni strains tested. A reduction of greater than 1 log cycle in the numbers of cells growing on plates containing antibiotics was considered to be a marked level of inhibition. Only one of nine of the antibiotic combinations studied did not markedly inhibit most of the C. coli strains tested. Although one C. coli strain was not inhibited by any of the antibiotic combinations, the other six strains were inhibited for up to 7 log cycles. The addition of blood or growth supplements reduced but did not eliminate the inhibitory effect. The inhibition of laboratory strains of C. coli on media developed for the isolation of Campylobacter spp. indicates that the incidence of C. coli may be underestimated.     

Trimethoprim activity in media selective for Campylobacter jejuni.

The activity of trimethoprim (TMP) in two selective media used for isolation of Campylobacter jejuni was evaluated. The two selective media, Campy-BAP and Skirrow medium, contain TMP in addition to other antimicrobial agents. The minimal inhibitory concentrations of TMP in blood agar base (basal agar for Skirrow medium) or brucella agar (basal agar for Campy-BAP) for three sensitive control organisms were compared with those in Mueller-Hinton agar, which contains low levels of thymidine. TMP was inactive in both blood agar base and brucella agar, even when lysed horse blood or thymidine phosphorylase was added. TMP had activity when used in combination with the other antimicrobial agents normally included in Skirrow medium and Campy-BAP, probably indicating synergism between TMP and one or more of the other antimicrobial agents. Sheep blood could be substituted for lysed horse blood in Skirrow medium without compromising the activity of TMP.     

Blood-free selective medium for isolation of Campylobacter jejuni from feces

A blood-free selective agar is described which contains charcoal, ferrous sulfate, sodium pyruvate, casein hydrolysates, cefazolin, and sodium deoxycholate (CCD agar). CCD agar was compared with Preston medium for isolation of Campylobacter jejuni from human feces, and isolation rates were similar on both media, but CCD agar was less selective. Temperature studies at 37 and 42 degrees C confirmed that incubation of direct plates at 42 degrees C for 48 h was necessary for maximum isolation of C. jejuni.     

Evaluation of four enrichment media for isolation of Campylobacter jejuni.

Diarrheal stool specimens were inoculated into the following media: alkaline peptone water (APW), Bruce-Zochowsky medium (BZ), Campylobacter enrichment broth (CEB), Campy-thio broth (CT), and Skirrow blood-agar (SK) plate. All media were incubated at 42 degrees C in microaerophilic conditions for 24 h. Afterwards, a new SK plate was inoculated from every liquid medium. Campylobacter jejuni was isolated from 43 of the 259 specimens when CT was used, from 45 when APW was used, from 46 when BZ was used, and from 46 when CEB was used; these totals include specimens that grew after enrichment only, on SK plates only, and both after enrichment and on SK plates. No significant differences were found between the isolates obtained with and without enrichment procedures.     

Laboratory and clinical evaluation of isolation media for Campylobacter jejuni.

Six selective isolation media were evaluated for their ability to support the growth of Campylobacter jejuni. Colony counts of 70 isolated strains of C. jejuni and recovery studies on these strains in simulated positive feces samples demonstrated that Bolton and Hutchinson' charcoal, cefoperazone, deoxycholate agar and Karmali's charcoal-based selective medium produced the highest recovery rates with the greatest suppression of other fecal flora. C. jejuni colonies were more easily recognized on charcoal-based selective medium. A clinical evaluation performed on 2,780 human, animal, and avian feces specimens confirmed the results of the laboratory investigation. From human samples, 4 more strains of C. jejuni were isolated on charcoal-based selective medium than were isolated on Skirrow medium, and 19 more strains of C. jejuni or C. coli were isolated on charcoal-based selective medium from animal specimens. Suppression of normal fecal flora was also greater on charcoal-based selective medium.     

Comparative efficacy of seven selective media for isolating Campylobacter jejuni.

Diarrheal stools from 263 patients were inoculated on seven selective media: Butzler selective medium, Blaser medium, Skirrow blood agar, Preston campylobacter selective medium, Preston campylobacter blood-free medium, Butzler Virion medium, and modified Preston medium (with amphotericin B [2 mg/liter]). A similar number of Campylobacter jejuni strains were isolated from all the media studied; nevertheless, the presence of competing fecal flora (FF) made the detection of suspect colonies difficult. Preston campylobacter blood-free medium with cefoperazone yielded the greatest number of C. jejuni isolations, and contaminating FF grew in only 9% of the plates showing C. jejuni growth; all the other media allowed the abundant growth of other FF, regardless of whether C. jejuni was isolated from them or not.     

Epidemiology of Campylobacter jejuni/coli

The authors describe the epidemic incidence of diseases caused by Campylobacter jejuni/coli. During the 5-week follow up period (June 1-July 1, 1988) a total of 74 subjects fell sick. In 31 instances the suspect factor of transmission was non-pasteurized cheese prepared from sheep's milk. In this group of patients a significant shift to higher age categories was noted, contrary to the other 43 diseases. It did not prove possible to isolate Campylobacter from cheese, smears from the cottage or from rectal swabs of the workers from the cottage. In an investigation focused on assessment of survival of Campylobacter jejuni/coli in sheep's milk and cheese long-term survival of the microorganism only in non-pasteurized milk was found. The authors assume that cheese becomes contaminated secondarily, by lack of adherence to hygienic rules.     

Serotyping of Campylobacter jejuni/coli.

Antisera were prepared from strains of Campylobacter jejuni/coli isolated from patients in six outbreaks of enteritis. Bactericidal antibodies, and agglutinating antibodies to heat-labile and heat-stable antigens, were demonstrated. These reactions were used to type a number of strains isolated from patients in each outbreak, and to distinguish 'epidemic' from 'non-epidemic' strains.     

Comparison of selective media for the isolation of salmonellae

A study of the selective action of four fluid media used for the isolation of salmonellae is described. When incubated in the presence of three competing organisms, test strains were recovered most frequently from Rappaport medium but mannitol selenite broth proved superior for the isolation of S. typhi.     

Comparison of Characteristics of Patients Infected by Campylobacter jejuni,Campylobacter coli,and Campylobacter fetus

A large database of Campylobacter isolates precisely identified at the species level was used to compare patients'' characteristics. In a multivariate analysis, Campylobacter coli was found more often in older patients and in patients having traveled abroad and less often in summertime than Campylobacter jejuni. Campylobacter fetus infection occurred in much older patients and in hospitalized patients with a systemic disease.     

Evaluation of media for isolation of Campylobacter fetus subsp. jejuni from fecal specimens.

Isolation rates of Campylobacter fetus subsp. jejuni from human fecal specimens were equivalent after broth enrichment (thioglycolate medium containing antibiotics) and direct inoculation on two brucella blood agar media containing ferrous sulfate, sodium metabisulfite, and sodium pyruvate, identical concentrations of vancomycin and trimethoprim, and different concentrations of polymyxin B and cephalothin. Studies with clinical isolates of C. fetus subsp. jejuni demonstrated temperature-dependent activity of polymyxin E (colistin) and substantial inhibition of growth on Thayer-Martin and Martin-Lewis media.     

Improved biotyping schemes for Campylobacter jejuni and Campylobacter coli.

Campylobacter jejuni (20 strains) and Campylobacter coli (12 strains) were assigned to four biovars for each species based on phenotypic tests that were easy to perform and interpret. The resulting biotyping schemes offer a greater degree of distinction among C. jejuni and C. coli strains than any of the other biotyping schemes previously described for these organisms.     

Serological response to Campylobacter jejuni/coli infection.

The antibody response to Campylobacter jejuni/coli infection was investigated in 59 patients involved in two outbreaks of milk-borne infection and in sporadic infections in the community. Agglutinins and complement-fixing (CF) antibodies were detected in nearly all these patients. Agglutinins were present in 25% of normal sera at low titres (not greater than 1/160) but CF antibody titres of 1/4 or 1/8 were present in only 2.0%. The agglutination reactions in convalescent sera were best developed with the homologous or an antigenically similar strain whereas the CF test, with sonicated organisms as antigen, was less strain-specific and was more suitable as a routine test. Antibody was present seven to 10 days after the onset of symptoms.     

Fitness of macrolide resistant Campylobacter coli and Campylobacter jejuni

The aim of this study was to investigate the fitness of macrolide resistant Campylobacter coli and Campylobacter jejuni. The in vitro growth, the survival on food matrix, and the in vivo colonization of C. jejuni and C. coli susceptible isolates and their isogenic resistant mutants were studied. In vitro experiments demonstrated that macrolide resistance imposed a fitness cost when the susceptible strains and their isogenic resistant mutants were cultured in competition. When inoculated in food matrix, the resistant C. jejuni mutant was no longer detectable after 3 to 5 days but the susceptible strain remained detectable for over 18 days. No difference in survival in food matrix was observed between susceptible and resistant C. coli. When inoculated in vivo in chickens, the macrolide susceptible and resistant C. coli displayed similar levels of colonization, both in separated inoculations and during competitive assays. Strikingly, when mono-inoculated or co-inoculated into chickens, macrolide susceptible C. jejuni outcompeted the macrolide resistant population. However, a spontaneous mutant that evolved in vivo showed a colonization capacity similar to the susceptible strain. Our findings demonstrate the effect of macrolide resistance on the fitness of Campylobacter but suggest that evolved mutants may be as fit as susceptible strains.