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CAMPYLOBACTER SELECTIVE MEDIA
The revelation in the 1970's that campylobacters are important human pathogens marked the beginning of an upsurge of interest in these organisms which has continued unabated. New species are being identified and some existing species have been assigned to new genera.
In 1977, enteritis-causing campylobacters were still in the province of a few experts but with improved media and isolation procedures, clinical laboratories can isolate thermophilic campylobacters routinely. A considerable body of literature on the genus has arisen; reference to this work and an up-date on developments has been made in a review by Penner1. The table below has been adapted from this review.
Differential reaction and characteristics for species of the genera Campylobacter, Arcobacter and Helicobacter pylori. (Adapted from Penner1)
Growth
|
Susceptibility
|
||||||||||
Species |
Catalase
|
Nitrate
|
H2S (TSI)
|
Hippurate
|
Urease
|
25°C
|
37°C
|
42°C
|
Nali dixic acid
|
Cepho-lothin
|
G+C content (mol%) |
Campylobacter fetus subsp. fetus |
+
|
+
|
-
|
-
|
-
|
+
|
+
|
(-)
|
R
|
S
|
33-34 |
C. fetus subsp. venerealis |
+
|
+
|
-
|
-
|
-
|
+
|
+
|
-
|
R
|
S
|
33-34 |
C. hyointestinalis |
+
|
+
|
+
|
-
|
-
|
(+)
|
+
|
+
|
R
|
S
|
35-36 |
C. jejuni |
+
|
+
|
-
|
+
|
-
|
-
|
+
|
+
|
S
|
R
|
30-32 |
C. coli |
+
|
+
|
-
|
-
|
-
|
-
|
+
|
+
|
S
|
R
|
31-33 |
C. lari |
+
|
+
|
-
|
-
|
-
|
-
|
+
|
+
|
R
|
R
|
31-33 |
C. upsaliensis |
(-)
|
+
|
-
|
-
|
-
|
-
|
+
|
+
|
S
|
S
|
35-36 |
‘C. cinaedi’ |
+
|
+
|
-
|
-
|
-
|
-
|
+
|
-
|
S
|
I
|
37-38 |
‘C. fennelliae’ |
+
|
- |
-
|
-
|
-
|
-
|
+
|
-
|
S |
S
|
37-38 |
C. sputorum |
-
|
+
|
(+)
|
-
|
-
|
-
|
+
|
+
|
(S)
|
S
|
31-32 |
Biovar sputorum | |||||||||||
Biovar bubulus |
-
|
+
|
+
|
-
|
-
|
-
|
+
|
+
|
R
|
S
|
31-32 |
Biovar fecalis |
+
|
+
|
+
|
-
|
-
|
-
|
+
|
+
|
R
|
S
|
32-33 |
C. mucosalis |
-
|
+
|
+
|
-
|
-
|
-
|
+
|
+
|
R
|
S
|
38-39 |
C. concisus |
-
|
+
|
+
|
-
|
-
|
-
|
+
|
+
|
R
|
R
|
38-39 |
Arcobacter cryaerophilia |
+
|
+
|
-
|
-
|
-
|
+
|
+
|
-
|
d
|
R
|
29-30 |
A. nitrofigilis |
+
|
+
|
ND
|
-
|
+
|
+
|
+
|
-
|
S
|
S
|
28-29 |
Helicobacter pylori |
+
|
d
|
-
|
-
|
+
|
-
|
+
|
+
|
R
|
S
|
36-37 |
+ Positive reaction; - negative reaction; ND no test results found; (+) most
strains positive but a low percentage negative: (-) most strains negative but
some positive or weakly positive; d different reactions; R resistant; S suscpetible;
I intermediate zones of inhibitions.
aSusceptbility to antibiotics was determined with 30m
g disks.
The major step forward in recognising the importance of campylobacters in human
disease was the development of isolation media which contain antibiotics. These
media suppress competing faecal flora and allowed the campylobacters to grow
into easily detected colonies.
The first selective supplement was developed by Skirrow1 and other
workers followed with other antibiotic combinations.
Differential reactions and characteristics for species of the genera Campylobacter,
Arcobacter and Helicobacter pylori. (Adapted from Penner1)Gun-Monro
et al.2 carried out a laboratory and clinical evaluation of the various
selective isolation media for thermophilic campylobacters. Summary tables of
their findings for the above antibiotic supplements are shown.
Selective Antibiotic Supplements for the isolation of Campylobacters
Antibiotics (mg/litre) |
Butzler SR85 |
Blaser-Wang SR98 |
CCDA SR155 |
Preston SR117 |
Skirrow SR69 |
Karmali SR167 |
CAT SR174 |
Amphotericin B |
- |
2 |
10 |
- |
- |
- |
10 |
Bacitracin |
25,000* |
- |
- |
- |
- |
- |
- |
Cephalothin |
- |
15 |
- |
- |
- |
- |
- |
Cefazolin |
15 |
- |
- |
- |
- |
32 |
- |
Cefoperazone |
- |
- |
32 |
- |
- |
- |
8 |
Colistin |
10,000* |
- |
- |
5,000* |
- |
- |
- |
Cycloheximide |
50 |
- |
- |
100 |
- |
100 |
- |
Novobiocin |
5 |
- |
- |
- |
- |
- |
- |
Polymyxin |
- |
2,500* |
- |
- |
2,500 |
- |
- |
Rifampicin |
- |
- |
- |
10 |
- |
- |
- |
Trimethoprim |
- |
5 |
- |
10 |
5 |
- |
- |
Vancomycin |
- |
10 |
- |
- |
10 |
20 |
- |
Teicoplanin |
- |
- |
- |
- |
- |
- |
4 |
*IU/L
CCDA – Modified Charcoal Cefoperazone Desoxycholate Agar (Blood-free medium)
Table 1. Recovery of 70 C. jejuni strains on five selective media.
Medium |
Colony Count |
P valueb |
Blood agar control |
7.95 ± 0.36 |
|
Skirrow |
7.82 ± 0.48 |
Nsc |
Butzler |
7.77 ± 0.51 |
<0.05 |
Blaser-Wang |
7.70 ± 0.56 |
<0.05 |
Preston |
7.76 ± 0.52 |
<0.05 |
Modified CCDA |
7.91 ± 0.36 |
NS |
Table 2. Isolation of C. jejuni from 70 simulated positive faeces samples.
Medium |
24 Hours |
48 Hours |
Blood agar control |
69 (99) |
70 (100) |
Skirrow |
39 (56) |
67 (96) |
Butzler |
38 (54) |
60 (86) |
Blaser-Wang |
17 (24) |
31 (41) |
Preston |
32 (46) |
64 (91 |
Modified CCDA |
61 (87) |
69 (99) |
aLog10 mean colony counts ±
standard deviation.
bsignificance determined by Student's t test for unpaired samples.
cNS, Not significant.
Table 3. Suppression of faecal flora from 70 simulated positive faecal samples.
No. (%) of plates with 75% reduction of faecal flora compared with control
Medium number (%) of strains isolated after incubation for: |
||
Medium |
24 Hours |
48 Hours |
Skirrow |
46 (66) |
38 (54) |
Butzler |
56 (80) |
47 (67) |
Blaser-Wang |
28 (40) |
15 (21) |
Preston |
58 (83) |
50 (71) |
Modified CCDA |
64 (91) |
59 (84) |
The general conclusions reached by Gun-Munro et al. were confirmed by Griffiths and Ribeiro3.
Enrichment broth cultures - the value of enrichment media for campylobacters
is controversial1 but in food and environmental studies enrichment
may be essential4. Enrichment at 42°
C4,5 and cold enrichment at 4° C6
have been reported. Where enrichment increases competitive flora, the use of
membrane filters on the surface of the agar can help select campylobacters7.
A comprehensive review of selective media for Campylobacter and Arcobacter
species has been published in a special issue of International Journal of Food
Microbiology.
Reference
Corry J.E.L., Post D.E., Colin P. and Laisney M.J. (1995). Int. J. Food
Microbiol. 26. 43-76.
Laboratory growth environment
Atmosphere - members of the Campylobacter genus require a wide spectrum
of atmospheres for optimum growth, ranging from complete anaerobiosis to ambient
air tolerance8. Most species, however, lie between these extremes
and are micro-aerophilic. The Oxoid Campylobacter Gas Generating Kits (BR56
and BR60) ensure that the correct oxygen and carbon dioxide levels are produced
for the optimum growth of these micro-aerophilic organisms. Alternatively use
CampyGen CN025A or CN035A. CampyGen does not require the addition of water or
a catalyst.
Temperature - the temperature range for incubation of Campylobacter
species and related organisms varies from 15°
C for Arcobacter cryaerophilea to 42°
C for the thermophilic species. However, most strains have a considerable tolerance
of growth temperatures around those required for optimum growth.
Helicobacter pylori - this organism is implicated as a cause of gastritis
and peptic ulceration9,10,11.
A specific selective culture medium, prepared from Helicobacter pylori Selective
Supplement (Dent) SR147 and Columbia Blood Agar Base CM331 is required to isolate
this organism from gastric biopsy specimens.
References
1 Penner J. L. (1988) Clin. Microbiol. Reviews. 1. 157-172.
2 Gun-Munro J., Rennie R. P. , Thornley J. H., Richardson H.L., Hodge D. and
Lynch J. (1987) J. Clin. Microbiol. 25. 2274-2277.
3 Griffiths A. and Ribeiro C. D. (1988) J. Clin. Path. 41. 704-705.
4 Marinescu M., Festy B., Derimay R. and Megraud F. (1987) Eur. J. Clin.
Microbiol. 6. 693-694.
5 Bolton F. J., Coates D., Hinchliffe P. M. and Robertson L. (1983) J. Clin.
Path. 36. 78-83.
6 Rubin S. J. and Woodward N. (1983) J. Clin. Microbiol. 18. 1008-1010.
7 Steele T. W. and McDermott S. N. (1984) Pathology 16. 263-265.
8 Neill S. D., Campbell J. N., O'Brien J. J., Weatherup S. T. and Ellis W. A.
(1985) Int. J. Sys. Bacteriol. 35. 342-356.
9 Marshall B. K., Warren J. R., Blincow E. D., Phillips M., Goodwin C. S., Murray
R., Blackbourne S. J. and Waters T. E. (1988) Lancet ii. 626-627.
10 Dent J. C. and McNulty C. A. M. (1988) Eur. J. Clin. Microbiol. Infect.
Dis. 7. 555-568.
11 Buck G. E. (1988) Lab. Managemt. 26. 9.