Efficient Isolation of Campylobacteria from Stools

Engberg et al. (2) compared selective media and filtration for the efficient isolation of campylobacteria from stools. Since 1977, we have routinely isolated campylobacters from the diarrhetic stools of pediatric patients at the Red Cross Children's Hospital in Cape Town, South Africa. In 1990, the use of antibiotic-containing plates was discontinued, and the Cape Town protocol, the first to combine both membrane filtration onto antibiotic-free blood agar plates and incubation in an H₂-enhanced microaerobic atmosphere (3), was introduced.

With the use of this protocol, stool cultures positive for campylobacteria rose to 21.8% from the 7.1% previously obtained with Skirrow's medium and other selective media used in conjunction with conventional microaerobic incubation (3). Our laboratory could only begin to isolate Campylobacter upsaliensis, Campylobacter concisus, Campylobacter curvus, Campylobacter rectus, Campylobacter sputorum biovar sputorum, Helicobacter fennelliae, Helicobacter cinaedi, “Helicobacter rappini”, and Arcobacter butzleri from stool samples with the use of the Cape Town protocol. Strains of these species are sensitive to antibiotics commonly used in selective media or have an essential requirement for an H₂-enhanced microaerobic atmosphere.

With a combination of filtration and selective media, Engberg et al. (2) documented an isolation rate of 4.8% for eight species of campylobacteria from diarrhetic stools. The Cape Town protocol, utilizing only membrane filtration, yielded an isolation rate of 21.1% for 16 species or subspecies of Campylobacter and related organisms (Table 1). Differences in colonial morphology on primary isolation and subsequent biochemical, serological, or molecular confirmation indicated that 16.2% of South African children suffering gastroenteritis had coinfections with up to five different species of campylobacteria (3).

TABLE 1.

Distribution of Campylobacter and related species isolated from 19,535 diarrhetic stools of pediatric patients at the Red Cross Children's Hospital, Cape Town, South Africa, from 1 October 1990 to 29 February 2000

Species and subspecies	Specimens
	No. positive	% Positive
C. jejuni subsp. jejuni, biotype 1a	1,166	28.29
C. concisus	971	23.55
C. upsaliensis	948	23.00
C. jejuni subsp. doylei	378	9.17
H. fennelliae	260	6.31
C. coli	119	2.87
C. jejuni subsp. jejuni, biotype 2a	115	2.79
C. hyointestinalis	53	1.29
H. cinaedi	42	1.01
CLO/HLOb	35	0.85
Arcobacter butzleri	16	0.39
C. fetus subsp. fetus	7	0.17
“H. rappini”	4	0.10
C. lari	2	0.05
C. curvus	2	0.05
C. rectus	2	0.05
C. sputorum biovar sputorum	2	0.05
Total	4,122	100.00

^aBiotype of Skirrow and Benjamin (4). 

^bCLO/HLO, Campylobacter or Helicobacter organisms that could not be fully characterized. 

Discrepancy in the isolation rate and numbers of species detected between the Danish and South Africa studies may reflect geographical differences in the prevalence of various campylobacteria, the nature of infection in these countries, or other factors. A final hydrogen concentration of 3% was advocated by Engberg et al. (2) as being optimal. Use of Oxoid BR 38 GasPaks with no catalyst in the Cape Town protocol generates a hydrogen concentration of greater than 40%. This increased level of hydrogen, combined with strain variation in hydrogen requirements, may contribute to the higher isolation rate of the Cape Town protocol.

Campylobacter jejuni and Campylobacter coli comprise about one-third of the campylobacteria routinely isolated in Cape Town (Table 1). We concur with Engberg et al. (2) that campylobacteria other than C. jejuni and C. coli are responsible for gastroenteritis and are undetected by inappropriate techniques. We also agree that C. concisus is an important opportunistic pathogen of very young children (1). The pathogenic potential, reservoirs, and modes of transmission of these non-C. jejuni or non-C. coli species have yet to be fully determined (1).

Engberg et al. (2) recommend a combination of both membrane filtration and selective media for optimal isolation of campylobacteria from stool. They also state that membrane filtration is costly, labor-intensive, and less sensitive than selective media. We do not agree, as the Cape Town protocol in our laboratory over the last decade has proved to be a simple, efficient, and cost-effective alternative to the use of selective media.