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Avian Pathology
ISSN: 0307-9457 (Print) 1465-3338 (Online) Journal homepage: https://www.tandfonline.com/loi/cavp20
An unusual Clostridium colinum infection in broiler
chickens
B. Perelman , S. Mints , M. Zjut , E. Kuttin & S. Machny
To cite this article: B. Perelman , S. Mints , M. Zjut , E. Kuttin & S. Machny (1991) An unusual
Clostridium�colinum infection in broiler chickens, Avian Pathology, 20:3, 475-480, DOI:
10.1080/03079459108418785
To link to this article: https://doi.org/10.1080/03079459108418785
Published online: 12 Nov 2007.
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2. Avian Pathology (1991) 20, 475-480
An unusual Clostridium colinum infection in broiler
chickens
B. PERELMAN,1
S. MINTS,1
M. ZJUT,1
E. KUTTIN3
& S. MACHNY2
1
Regional Poultry DiseasesLaboratory Experimental Station Gilat, D.N. Negev, Israel,
2
Department of Avian DiseasesKimron Veterinary Institute, Beit Dagan, Israel and
3
Israel Institute for Biological Research, Nes Ziona, Israel
SUMMARY
An unusual syndrome has been observed during the past two years in flocks of 12- to 25-day-old
broiler chickens in Israel. The disease is characterized by enterotoxaemic-like signs including a
high and sudden mortality, apathy, severe localized enteritis and nephrosis. C. colinum which is
the causal agent of ulcerative enteritis has been the only bacteria consistently isolated from the
affected chickens. The clinical and pathological findings in this syndrome are different from
those reported for ulcerative enteritis.
INTRODUCTION
Clostridium colinum is considered the aetiological agent of ulcerative enteritis in several
avian species (Peckham, 1959, 1960, 1978). In chickens detailed descriptions of the
clinical signs and typical pathological changes have been reported (Peckham, 1959,1960,
1978; Kondo et al, 1988). Typically, the affected chickens are between 4 and 15 weeks of
age, and the appearance of ulcerative enteritis is usually secondary to some primary
disease such as coccidiosis, aplastic anaemia, Gumboro disease and stress conditions
(Peckham, 1960, 1978; Davis, 1973).
During the past two years an unusual disease has been observed in several flocks of
very young broiler chickens in the Negev region of Israel. Despite the fact that C.
colinum seems to be related to this problem as a primary pathogen, the clinical and
pathological findings are different from those reported in ulcerative enteritis (Peckham,
1959, 1960, 1978; Kondo et al., 1988).
MATERIALS AND METHODS
Bacteriological tests
Direct swabs taken from the liver, spleen and intestines of dead or sick birds from eight
affected flocks were inoculated onto blood nutrient agar (Difco, Detroit, MI) and
incubated under anaerobic conditions in an anaerobic jar (Gas-Pak, BEL, Cockeysville,
MD) for 24 h. All isolated agents were biochemically characterized according to Berkhoff
et al (1974) and Kondo et al. (1988). Susceptibility tests were carried out on blood
nutrient agar using commercial sensitivity discs (Difco, Detroit, MI) of the following
Received 18 September 1990; Accepted 18 February 1991.
475

3. 476 B. PERELMAN ETAL.
products: oxytetracycline, streptomycin, neomycin, furazolidone, penicillin and ampicil-
lin.
Histological examination
Small pieces of liver, spleen and intestines from recently dead and sick chickens from
eight cases were fixed in 10% buffered formalin, embedded in paraffin and stained with
haematoxylin and eosin.
Chicken embryo inoculation
Six-day-old commercial embryos were inoculated by yolk sac route with a bacterial
suspension prepared by scraping the colonies of 24-h cultures. Embryo inoculation was
performed for all six isolates of Clostridium using six embryos for each isolate. Embryos
were candled daily for mortality for a period of 5 days after inoculation and then
examined for production of gas and presence of Clostridium.
Pathogenicity in quail
Pathogenicity of three isolates was tested by the inoculation of 5-week-old quail. A total
of eight quail for each isolate were infected with 0.2 ml of a bacterial suspension
containing 107
CPU/ml of 24-h cultures in blood agar. Each of four quails was inoculated
intramuscularly or orally.
Pathogenicity in chickens
The same isolates were tested in 12-day-old commercial broiler chickens. Ten chickens
were inoculated with 0.2 ml of a suspension containing 10s
CPU/ml of 24-h culture in
blood agar of each isolate by intramuscular route and ten by oral route. Both groups were
kept in isolation units for a two-week period. Ten chickens, 2 days old, were inoculated
with the bacteria-free supernatant of the intestinal content collected from sick birds. Five
of the chickens were inoculated with 0.2 ml by intramuscular route and the other five
were inoculated with 0.2 ml by oral route.
RESULTS
Clinical and pathological description
All affected chickens were between 12 and 25 days of age. The disease was characterized
by a sharp increase in mortality. Despite the high mortality, which reached 10% within
24 h and 50% if the flock was not treated, the chickens looked apparently healthy, and
usually there were only a few clinically sick birds. Affected chicks showed severe
depression, lack of coordination and slight cyanosis. At post mortem examination the
birds were well-muscled and had food in the crop. The breast muscles were congested,
showed pale streaks and were dry to the touch. Congestion of the liver and kidneys was
frequently found. The most common and characteristic macroscopic lesion was a localized
enteritis of the duodenum, with the accumulation of a dark content in which small white
flakes could be observed (Figure 1). In some chickens the only lesion observed was a
small congested area in the loop of the duodenum. Microscopic examination of this

4. CLOSTRIDIUM COLINUM INFECTION 477
content revealed severe desquamation of the epithelium and large numbers of red blood
cells.
Figure 1. Duodenal content of an affected chick. Note the dark fluid with small white flakes.
Gram-stained smears of the duodenal content revealed large numbers of Gram-positive
rods. In chickens with very slight lesions, these Gram-positive rods were localized only in
the area of the lesion. Histological examination revealed congestion of liver and spleen
and enteritis with desquamation of the epithelium, necrosis of the tip of the villi and a
pseudoeosinophilic inflammatory infiltration. Large numbers of rods covering the villi
were observed (Figure 2). Severe nephrosis was observed in all affected chicks.
Treatment of the affected flocks with ampicillin at 1 g/10 1of water reduced mortality
and eliminated the clinical signs within 24 h, and there were no relapses of disease. The
disease was refractory to neomycin, furanes and oxytetracycline.
Microbiological examination
C. colinum was isolated from six out of eight cases reported to the laboratory during a
nine-month period. In most of these cases, C. colinum was isolated as a pure culture from

5. 478 B. PERELMAN£7MZ..
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Figure 2. Desquamation of the intestinal epithelium with large numbers of rods attached to the
surface of the villi. Bar=20 fim.
the area of the lesion in the duodenum. Attempts to isolate C. colinum from the liver and
spleen were unsuccessful.
Characterization of the isolates
All the isolates were Gram-positive motile rods, and their biochemical characteristics
coincided with those reported for C. colinum (Berkhoff et al, 1974; Kondo et al, 1988).
All the isolates were resistant to streptomycin, neomycin, furanes and oxytetracycline and
highly susceptible to penicillin and ampicillin.
Inoculation of 6-day-old chicken embryos caused 100% mortality within 72 h with all
isolates. The embryos showed severe congestion. No gas was produced. Inoculation of 5-
week-old quail by intramuscular route caused 100% mortality within 72 h. Inoculation of
quail by oral route caused 100% mortality within 6 days. Severe haemorrhagic enteritis
was observed in these quail. The bacteria was reisolated from the liver and intestines of
the dead quail.
Inoculation of 12-day-old broilers by intramuscular injection caused 60% mortality

6. CLOSTRIDIUM COLINUM INFECTION 479
within 24 h. The dead birds showed severe muscular necrosis and accumulation of large
amounts of serosanguinous fluid under the skin around the injection site. Wide areas of
necrosis were observed in the liver. The bacteria were reisolated from the subcutaneous
fluid and from the liver. In all surviving birds a severe subcutaneous accumulation ofthe
serosanguinous fluid was observed. Two weeks after inoculation a marked atrophy and
scarring of the muscles and the skin was found in the inoculated area. Chickens
inoculated by oral route did not develop any clinical signs or mortality.
Intramuscular injection or oral administration of the cell-free intestinal content from
sick birds did not cause any clinical or pathological signs in young broiler chicks.
DISCUSSION
Ulcerative enteritis is a well-known disease in light and heavy pullets in Israel (Samberg,
1987-89). All the cases observed until now were quite similar to those previously
reported (Peckham, 1959,I960, 1978; Kondo et al, 1988). The clinical and pathological
signs observed in very young broiler chickens, as reported in this paper, are different
from those observed in ulcerative enteritis (Peckham, 1960, 1978;Kondo et al., 1988).
They resemble an enterotoxaemic syndrome, as observed in some cases of necrotic
enteritis (Parish, 1961; Helmboldt & Bryant, 1971).
The histopathological changes in the intestines of affected chicks are somewhat similar
to those observed in typical cases of ulcerative enteritis (Peckham, 1978), and are
different from the characteristic histological changes of necrotic enteritis (Helmboldt &
Bryant, 1971).
The characterization of the isolates by their biochemical features, and their extreme
pathogenicity for quail, confirms that the clostridium isolated from chickens affected with
this syndrome belongs to the group of C. colinum (Berkhoff et al., 1974;Berkhoff &
Campbell, 1974; Davis et al., 1971; Kondo et al., 1988). The fact that C.colinum was the
only Clostridium consistently isolated strongly supports the idea that this bacteria is
directly involved in the aetiology of this problem.
The absence of other diseases, such as coccidiosis, Gumboro or haemorrhagic syn-
drome, and the immediate response to treatment with only ampicillin, suggests that this is
a primary disease and not a complication such as usually occurs in ulcerative enteritis
(Peckham, 1960,1978; Davis, 1973).
We were not able to demonstrate the presence of toxins in the intestines of sick birds,
as tested by the inoculation of 2-day-old chicks with the cell-free intestinal content from
sick birds, but the nephrosis observed in the affected chicks suggests a toxaemic problem,
and further studies are necessary in order to confirm these preliminary findings.
REFERENCES
BERKHOFF, G.A. & CAMPBELL, S.G. (1974). Etiology and pathogenesis of ulcerative enteritis (Quail disease). The
experimental disease. Avian Diseases, 18,205-212.
BERKHOFF, G.A., CAMPBELL, S.G., NAYLOR, H.B. & SMITH, L.D. (1974). Etiology and pathogenesis of ulcerative
enteritis (Quail disease). Characterization of the causative anaerobe. Avian Diseases, 18,195-204.
DAVIS, R.B. (1973). Ulcerative enteritis in chickens: Coccidiosis and stress as predisposing factors. PoultryScience, 52,
1283-1287.
DAVIS, R.B., BROWN, J. & DAWE, D.L. (1971). Quail—Biological indicators in the differentiation of ulcerative and
necrotic enteritis of chickens. Poultry Science, 50,737-740.
HELMBOLDT, C.F. & BRYANT, E.S.(1971). The pathology of necrotic enteritis in domestic fowl. Avian Diseases, 15,
775-780.

7. 480 B. PERELMAN ET AL.
KONDO, F., TOTTORY, J. & SOKI, K. (1988). Ulcerative enteritis in broiler chickens caused by Clostridium colinumand
in vitro activity of 19 antimicrobial agents in tests on isolates. Poultry Science, 87, 1424-1430.
PARISH, W.E. (1961). Necrotic enteritis in the fowl (Gallus gallus domesticus)1. Histopathology of the disease and
isolation of a strain of Clostridium welchii. Journal of Comparative Pathology, 71,377-393.
PECKHAM, M.C. (1959). An anaerobe the cause of ulcerative enteritis, "Quail disease." Avian Disease, 3, 471-477.
PECKHAM, M.C. (1960). Further studies on the causative organism of ulcerative enteritis. Avian Diseases, 4, 449-458.
PECKHAM, M.C. (1978). Ulcerative enteritis (Quail disease), in: M. S. HOFSTAD, B. W. CALNEK, C. F. HELMBOLDT,
W. M. REID & H. W. YODER (Eds) Diseases of Poultry, 7th Edn, pp. 295-304 (Ames, Iowa State University
Press).
SAMBERG, Y. (1987-89). Annual Reporton Avian Diseases (Bet Dagan, Veterinary Services, Israel).
RESUME
Infection peu commune à Clostridium colinum chez le poulet
Un syndrome inhabituel a été observé, durant les deux dernières années, en Israël, dans des troupeaux de
poulets de chair âgés de 12 à 25 jours. La maladie est caractérisée par des symptômes de pseudo-
entérotoxémie se traduisant par une mortalité élevée et soudaine, de l'apathie, une sévère entérite et de la
néphrose. C. colinum,agent causal de l'entérite ulcérative, a été la seule bactérie principalement isolée des
poulets affectés. Les observations cliniques de ce syndrome sont différentes de celles rapportées pour
l'entérite ulcérative.
ZUSAMMENFASSUNG
Eine ungewöhnliche Clostridium colinum-Infektion bei Mastküken
Ein ungewöhnliches Syndrom ist während der vergangenen zwei Jahre in Israel bei 12- bis 25 tägigen
Mastküen beobachtet worden. Die Krankheit ist durch Enterotoxämie-ähnliche Symptome mit hoher und
plötzlicher Sterblichkeit, Apathie, schwerer herdfömiger Enteritis und Nephrose gekennzeichnet. C.
colinum, der ursächliche Erreger der ulzerativen Enteritis, war das einzige Bakterium, das regelmäßig von
den erkrankten Küken isoliert werden konnte. Die klinischen und pathologischen Befunde bei diesem
Syndrom unterscheiden sich von denen, die für die ulzerative Enteritis beschrieben werden.
RESUMEN
Infección poco común por Clostridium colinum en pollos de engorda
Un sindrome poco comun ha sido observado ultimamente en parvadas de pollitos de engorda de muy
corta edad. La enfermedad se caracteriza por una elavacion subita de la mortalidad, signos de toxemia y
una severa enteritis. Clostridiumcolinum ha sido la unica bacteria aislada en una forma constante de
pollos afectados con este problema. Los signos clinicos y los cambios patologicos observados en estas aves
son diferentes a los reportados como tipicos de la enteritis ulcerativa.