Campylobacter Enteritis Associated with Unpasteurized Milk
MARTIN J. BLASER, M.D.* JAMES CRAVENS, B.S.t BRADLEY W. POWERS, B.A. F. MARC LAFORCE, M.D. W-L. L. WANG, Ph.D. Denver, Colorudo
Campylobacter fetus ss. jejuni has recently been recognized as a human enteric pathogen. Laboratory isolation has been hindered by its fastidious nature. Methods for recovery of this organism from stool culture and a specific serologic test are described. An outbreak is reported in which three members of the same family became simultaneously ill with fever, severe abdominal cramps and diarrhea. C. fetus ss. jejuni was recovered from stool specimens from all three. A fourfold increase in serum immunoglobulin G (IgG) titer to this organism was demonstrated in each patient. All three patients had been consuming unpasteurized milk from a cow whose feces were infected with C. fetus ss. jejuni. Campylobacter fetus (formerly known as Vibrio fetus] has long been recognized as a commensal and a pathogen of various mammalian species [1,2]. Three subspecies have been identified: fetus, intestinalis and jejuni, the latter two being the more important pathogens in man [3]. Their recognition as agents of human disease has been well documented for 20 years, but only since 1962 have they been routinely differentiated [4]. C. fetus ss. jejuni has been implicated as an enteric pathogen of man; however, isolation has been difficult because of its special growth requirements. Recently, techniques have been devised for recovery of this organism from stool cultures, and investigators in Europe, Africa and North America have isolated this agent in 3 to 8 per cent of cases of diarrhea [S-II]. The epidemiology of infections due to Campylobacter is not well understood. Because of their importance as animal pathogens, transmission of these bacteria from animals to human subjects has been suspected [X2]. This report focuses on a small outbreak of enteritis due to C. fetus ss. jejuni. The affected patients had been drinking unpasteurized milk from an infected cow.
From the Division of Infectious Diseases, Department of Medicine, and Department of Pathology, University of Colorado Medical Center, and the Microbiology Laboratory, Denver V. A. Medical Center, Denver, Colorado. Requests for reprints should be addressed to Dr. Martin Blaser. Manuscript accepted March 19,1979. * Present address: Enteric Disease Branch, Bacterial Diseases Division, Bureau of Epidem. iology, Center for Disease Control, Atlanta, Georgia 30333. + Present address: P.O. Box 144, Anthong, New Mexico 88021.
METHODS Stool specimens were inoculated on a special plate medium to facilitate isolation of Campylobacter. As part of an ongoing study. stools from 80 normal persons were also cultured for C. fetus ss. jejuni and served as controls [ll]. The medium consisted of a Brucella agar base, 10 per cent sheep red blood cells, with vancpmycinl mg/lOO ml, trimethoprim 0.5 mg/lOO ml, polymyxin B 2.5 IU/ml and amphotericin B 2 &ml. Swabs were also placed in thioglycollate broth with the aforementioned antibiotics and 0.16 per cent agar. Plates were incubated at 42% for 48 hours in an atmosphere of 5 per cent oxygen, 10 per cent carbon dioxide and 85 per cent nitrogen whereas broth specimens were refrigerated for 8 hours and then subcultured onto the special medium.
October 1979
The American Journal of Medicine
Volume 67
715
CAMPYLOBACTER
ENTERITIS AND UNPASTEURIZED
MILK-BLASER
Plates were examined at 24 and 48 hours. Colonies which were grey, small or mucoid, nonhemolytic and flat were gram-stained (counter stain with carbol fuchsin) and/or studied under phase microscopy. If the gram stain showed vibrio forms or if phase contrast microscopy revealed a characteristic “darting” motility, further isolation and biochemical testing were performed. Organisms were considered to be C. fetus ss. jejuni if they were motile, oxidase- and catalasepositive, grew in Brucella broth at 37% and 42’C but not at 25’C, did not grow in 3.5 and 6.5 per cent salt, multiplied in 1 per cent glycine, were capable of producing hydrogen sulfide in Kligers media with lead acetate paper and were sensitive to nalidixic acid [3]. All isolates were tested for sensitivity to erythromycin, chloramphenicol, tetracycline, gentamicin, penicillin and cephalothin using the Kirby-Bauer method [l3]. Serums were obtained from patients during the acute and convalescent phase, and serums obtained from hospital personnel served as controls. Anitbody titers were determined using an indirect fluorescent antibody test with autologous isolates as the bacterial antigen. Bacteria were harvested from 24-hour plate cultures, washed twice in phosphate buffered saline solution (pH 7.2), boiled for 1 hour, washed again in phosphate buffered saline solution, and resuspended to a final concentration of 3 X lo7 organisms/ml. Organisms were dropped onto a slide and fixed with acetone. Test serums were diluted in twofold increments, layered onto the slides which were incubated at 37’C for 39 minutes, then washed in phosphate buffered saline solution, and fluorescein isocyanatetagged goat antihuman globulin (1:16 dilution in 0.1 per cent solution of Amido Black) was added. Slides were again incubated, then washed in phosphate buffered saline solution and examined using an epi-illuminated Zeiss fluorescent microscope. Fluorescence was judged at 0 to 4+. Slides were read as unknowns; those which showed 3+ or greater fluorescence were considered positive.
CASE REPORTS Case I. On June 7, a 20 year old man first noted the onset of intermittent, sharp periumbilical pain which increased in severity. By the following morning, he complained of malaise, anorexia and feverishness. Diarrhea began later that day, and the patient had approximately 10 bowel movements, progressing from loose, to watery, to frankly bloody stools. When seen at a hospital emergency room he was febrile (temperature 38.5%) and had tenderness in the lower right quadrant. Many leukocytes were seen in a wet preparation of his stool. His white blood cell count was 13,700/mm3 with 81 polymorphonuclear leukocytes, 5 juvenile forms, 7 lymphocytes, 6 monocytes and 1 basophil. The patient was treated symptomatically with diphenoxylate hydrochloride. C. fetus ss. jejuni was isolated from a stool specimen which was negative for Salmonellae, Shigellae, Staphylococcus aureus and parasites. Over the next few days there was a gradual diminution in symptoms; however, on June 13 the patient noted a worsening of the abdominal pain and several diarrhea1 stools. The administration of erythromycin (Z g/day] was begun, and within 48 hours the patient was completely well.
716
October 1979
The American Journal of Medicine
ET AL.
Case 2. On June 7, the 47 year old father of the preceding patient [Case 1) noted abdominal bloating and the onset of diarrhea. The following day, he experienced generalized abdominal cramping, malaise, myalgias, and chills, and recorded a temperature of 39.2%. The diarrhea worsened and was associated with severe abdominal cramps. Stools were watery but not bloody. His symptoms gradually diminished over the
next three days, and C. fetus ss. jejuni was isolated from a stool culture obtained on June 12 when he was asymptomatic. Case 3. On the evening of June 6, the 61 year old grandmother of the patient (Case 1) noted malaise and abdominal queasi-
ness. On the following day, she felt weak, complained of myalgias. severe nausea, anorexia and noted a low grade fever (temperature 38’C). Later that day, she experienced intermittent, generalized sharp abdominal cramping and had eight bowel movements within the next 24 hours. The stools were initially loose, and later became mucoid and watery. These symptoms persisted for 48 hours and diminished over the next four days. C. fetus ss. jejuni was isolated from a stool culture taken on June 13. On June 15, the patient noted recurrence of malaise, myalgias and chilly sensations. The following day she was febrile (temperature 39.4%), felt weak and again complained of severe nausea. She did not have abdominal cramps or diarrhea. The administration of erythromycin (2 g/day) was begun, and her symptoms resolved over the next 48 hours.
EPIDEMIOLOGIC INVESTIGATION Stool cultures obtained from the two asymptomatic family members were negative for C. fetus ss. jejuni, as were stool cultures from 80 asymptomatic normal subjects. The three symptomatic patients described in this study were members of a five person family extending over three generations. The index patient (Case 1) and his parents lived in the city. His grandparents (Case 3 and her husband) lived on a small farm 30 miles from the city and raised calves, sheep, swine, chicks and laying hens. In addition, a horse, a milk cow, a cat and a dog were on the farm. A well provided all the water for the farm. The index patient had been a frequent visitor to the farm in the previous few weeks and had contact with many of the animals as part of his routine farm chores. The father of the index patient (Case 1) had not been to the farm in the preceding month. All family members ate eggs from the farm. Cracked eggs were not eaten or used. Rectal culture of the hens did not reveal C. fetus ss. jejuni. The cow provided the family with 1 gallon of milk a day. This was collected in a clean bucket and immediately refrigerated. Four of the five family members consumed the unpasteurized milk. All three patients consumed at least 1 pint of milk a day whereas the fourth family member consumed 1 glass per day. A specimen of milk obtained on June 12 was negative for C. fetus ss. jejuni but repeated fecal specimens from the cow were positive for two months. Treating the cow with tetracycline eradicated the asymptomatic intestinal carriage.
Volume 67
CAMPYLORACTER
C fetus ss. jejuni was also isolated from stool specimens from the swine. Culture of the chicks [whose feed contained tetracycline), sheep, domestic animals and well water were negative. LABORATORY
RESULTS
Serum was collected within five to seven days after the onset of clinical illness (acute serum) and two weeks later (convalescent serum]. Results of antibody studies are presented in Table I. All patients demonstrated a fourfold increase in IgG antibody titer whereas serum from the asymptomatic milk-drinking family member and 14 control subjects had titers from 12 to 1:8.Immunoglobulin M (IgM) antibody titers were initially high and remained so when compared to those in control serums. Convalescent serum from a person who had C. fetus ss. intestinalis bacteremia showed an IgG titer of 1:8to our patients’ isolates of C. fetus ss. jejuni. Our patients’ serums, when tested against C. fetus ss. intestinalis, all showed IgG titers less than 1:8. All stool isolates proved to be sensitive to tetracycline, gentamicin, erythromycin and chloramphenicol, and were resistant to penicillin and cephalothin. Blood cultures were not performed. When this organism was inoculated to milk (to7/ml) and kept at 4’C, survival for three weeks was observed [l4]. COMMENTS The development of better laboratory methods for the isolation of Campylobacter species from coproculture has permitted determination of their roles as enteric pathogens. Recent studies using these techniques have demonstrated a significant rate of isolation of C. fetus ss. jejuni in association with diarrhea1 illness [5-111. Simultaneous isolation of the organism from the stool and blood of several patients with acute enteritis has confirmed pathogenicity in those patients. Lastly, in several human subjects from whom tissue was examined, the acute inflammation of the jejunum and ileum was identical in location and appearance to that seen in animals with enteritis caused by C. fetus ss. jejuni [4,6,151. As a result of these studies, the clinical spectrum of Campylobacter enteritis in human subjects has been defined [4,6]. In a majority of cases, the onset of illness is abrupt, characterized by fever, abdominal pain and diarrhea. The peak of the illness usually lasts for 24 to 72 hours, but longer durations have been reported. Diarrhea is usually the single most prominent symptom and in some cases, occult or gross blood with white blood cells and mucus are present. In the majority of cases, resolution is gradual over a few days to weeks but relapses may occur. There seems little question that the family we describe suffered an acute outbreak of Campylobacter enteritis. Isolation of C fetus ss. jejuni associated with a fourfold increase in IgG antibody titer all support this
ENTERITIS
TABLE I
AND IJNPASTEl~JRIZED
MII,K ~~ULASER ET AI,.
Reciprocal Indirect Fluorescent Antibody Titers to Campylobacter fetus ss. jejuni ___.-W
Case 1 Acute serum Convalescent serum Case 2 Acute serum Convalescent serum Case 3 Acute serum Convalescent serum Asymptomatic family member Acute serum Convalescent serum 14 asymptomatic control subjects, geometric mean
w-l
8 256
8 16
2 128
32 32
128 512
64 128
2 4 5.4
2 2 2.8
NOTE: Acute serum refers to serum collected within five to seven days after the onset of clinical illness and convalescent serum refers to serum collected two weeks later.
contention. Clinical features in patients from our outbreak fall well within the spectrum of disease due to Campylobacter. Of interest, two of our three patients had relapses several days after initial resolution of symptoms. C. fetus ss. jejuni is a frequent commensal in swine [IS] and has been isolated from asymptomatic cattle [v], cats and dogs [6,18]. In sheep and other ungulates it may exist in a commensal state or cause infectious abortion [19]. In fowl, C. fetus ss. jejuni may be a commensal [20] or cause vibrionic hepatitis [21]. The widespread reservoir of C. fetus ss. jejuni in the animal kingdom suggests that contact with animals or animal products would be important in the epidemiology of human discase. Campylobacter enteritis associated with infected dogs [22] and poultry [23] has been reported. A large waterborne outbreak of Campylobacter enteritis illustrated the importance of the oral route of infection [24]. The rate of enteric and vaginal carriage of this organism in asymptomatic subjects is less than 1 per cent [l4]. Current evidence favors an external source for most human infection, but better definition of the specific vectors is necessary. The simultaneous onset of symptoms in three previously healthy persons suggested a common source. Since the patients had different occupations, did not reside together and did not have exposure to the same farm animals, a foodborne source was suspected. In Case 2, the patient never consumed water from the farm. The only sources common to the three patients were the eggs and the raw milk which they consumed. The isolation of C. fetus ss. jejuni from a fecal specimen of the milk cow suggested that the unpasteurized milk was the vehicle for the infection. A fourth family member who drank smaller quantities of the suspect milk had no clinical, cultural or serologic evidence of infection.
October 1979
The American Journal of Medicine
Volume 67
717
CAMPYLOBACTER
ENTERITIS AND UNPASTEURIZED MILK-BLASER
In 1946,Levy [25] described an outbreak of diarrhea1 illness in a closed population, and epidemiologic investigation implicated unpasteurized cow’s milk as the source of infection. No known pathogens were found but gram stain of stool and blood cultures from affected patients demonstrated vibrio forms. However, the organisms did not survive long on routine culture media. In 1977, Taylor [26] reported three cases of septicemia due to C. fetus ss. jejuni. All three patients consumed raw milk from the same dairy. Case-control analysis implicated that this consumption of unpasteurized milk was the most significant risk factor. Recently, in En-
ET AL.
gland, 38 persons who drank unpasteurized milk from the same sources were found to be infected with C. fetus ss. jejuni, and in an outbreak involving another dairy 14 patients were affected. C. fetus ss. jejuni was isolated from the milk on two occasions [27]. The present report provides further evidence for raw milk as a vehicle for the transmission of Campylobacter infections. ACKNOWLEDGMENT We thank Ms. P. Riepe and Mr. A. Tsan for their technical assistance.
REFERENCES 1. Smith T: Spirilla associated
with disease of the fetal membranes in cattle (infectious abortionl. ,, 1 EXD . Med 28: 701. 1918. 2. MacFavdean F. Stockman S: Final renort of the Deuartmental Com”mittee appointed by the Board of Agriculture and Fisheries to Enquire into Epizootic Abortion. III. Abortion in Sheep, London, His Majesty’s Stationary Office, 1913. 3. Smibert RM: Cam ylobacter. Spiral and curved bacteria. Bergey’s Manua P of Determinative Bacteriology, 8th ed. (Buchanan RE, Gibbons NE, eds], Baltimore, Williams & Wilkins, 1974. p 207. 4. Kine EO: The laboratorv recognition of Vibrio fetus and a closely related vibrio isolated from cases of human vibriosis. Ann NY Acad Sci 98: 700,1962. 5. Dekeyser, P, Gossvin-Detrain M, Butzler JP, et al.: Acute enteritis due to related vibrio: first positive stool cultures. J Infect Dis 125: 390, 1972. 6. Skirrow MB: Campylobacter enteritis: a “new” disease. Br Med 12: 9.1977. 7. Pearson’ AD; Suckling WG. Ricciardi ID, et al.: Campylobatter associated diarrhea in Southamuton. Br Med 12: 955. MY
1977.
8. 9. 10. 11. 12. 13.
718
Lauwers S. DeBoek M. Butzler IP: Camovlobacter enteritis _” in Brussels. Lancet i: 604.1978. Demo1 P, Bosmans E: Campylobacter enteritis in Central Africa. Lancet 1: 604.1978. Dixon JMS. Mueller L: Campylobacter enteritis-Alberta. Canada Diseases Weeklv Reoort ~011. o 6.1978. Blaser MJ, Berkowitz ID. LaFor& FM, et 2,:.Campylobacter enteritis: clinical and epidemiologic features. Ann Intern Med (in press). Bokkenhauser V: Vibrio fetus infection in man. I. Ten new cases and some epidemiologic observations. Am J Epidemiol91: 400.1970. Bauer AW, Kirby WM, Sherris JC. et al.: Antibiotic susceptibility testing by a standardized single disc method. Am J Clin Path01 45: 493.1966.
October 1979
The American Journal of Medicine
14.
15. 16. 17. 18. 19. 20. 21. 22. 23. 24.
Blaser MJ, Hardesty HL, Wang WLL: Survival of Campylobatter fetus ss. ieiuni in biolonical milieus labstractl. Presented at the 79th’Annual Mee%ng of the American Society for Microbiology, Los Angeles, California, 1979. Evans RG, Dadswell JV: Human vibriosis. Br Med J 3: 240. 1967. Deas DW: Observations on swine dysentery and associated vibrios. Vet Ret 72: 65.1960. Bryner JH, O’Berry PA, Estes PC, et al.: Studies of vibrios from gallbladder of market sheep and cattle. Am J Vet Res 33: 1439.1972. Blaser MJ, LaForce FM, Wang WLL: Reservoirs for human campylobacteriosis. Clin Res 27: 40A, 1979. Firehammer BD: The isolation of vibrios from ovine feces. Cornell Vet 55: 482.1965. Smibert RM: Vibrio fetus var. intestinalis isolated from the intestinal contents of birds. Am J Vet Res 30: 1437,1969. Peckham MC: Avian vibrio infections. Diseases of Poultry, 6th ed, (Hofstad MS, ed). Ames, Iowa, State University Press, 1972, p 332. Blaser MJ, Cravens J, Powers BW. et al.: Campylobacter enteritis associated with canine infection. Lancet 2: 979, 1978. Hayek LF, Cruikshank JG: Campylobacter enteritis. Br Med 12: 1219.1977. Tiohan W, Vogt Rt: Waterborne campylobacter gastroenteritis-Vermont. Morbiditv Mortalitv Weekly. Rep. 27: 207, 1978.
25.
Levy AJ: Gastro-enteritis outbreak probably due to a bovine strain of vibrio. Yale J Biol Med 18: 243.1946. 26. Taylor PR, Weinstein WM. Bryner JH: Campylobacter fetus infection in human subjects: association with raw milk. Am 1Med 66: 779.1979. 27. Communicable Diseases Surveillance Center (United Kingdom): An outbreak of Campylobacter infection, presumed milkborne. Communicable Disease Report 78: 47, 1978.
Volume 67
MARTIN J. BLASER, M.D.* JAMES CRAVENS, B.S.t BRADLEY W. POWERS, B.A. F. MARC LAFORCE, M.D. W-L. L. WANG, Ph.D. Denver, Colorudo
Campylobacter fetus ss. jejuni has recently been recognized as a human enteric pathogen. Laboratory isolation has been hindered by its fastidious nature. Methods for recovery of this organism from stool culture and a specific serologic test are described. An outbreak is reported in which three members of the same family became simultaneously ill with fever, severe abdominal cramps and diarrhea. C. fetus ss. jejuni was recovered from stool specimens from all three. A fourfold increase in serum immunoglobulin G (IgG) titer to this organism was demonstrated in each patient. All three patients had been consuming unpasteurized milk from a cow whose feces were infected with C. fetus ss. jejuni. Campylobacter fetus (formerly known as Vibrio fetus] has long been recognized as a commensal and a pathogen of various mammalian species [1,2]. Three subspecies have been identified: fetus, intestinalis and jejuni, the latter two being the more important pathogens in man [3]. Their recognition as agents of human disease has been well documented for 20 years, but only since 1962 have they been routinely differentiated [4]. C. fetus ss. jejuni has been implicated as an enteric pathogen of man; however, isolation has been difficult because of its special growth requirements. Recently, techniques have been devised for recovery of this organism from stool cultures, and investigators in Europe, Africa and North America have isolated this agent in 3 to 8 per cent of cases of diarrhea [S-II]. The epidemiology of infections due to Campylobacter is not well understood. Because of their importance as animal pathogens, transmission of these bacteria from animals to human subjects has been suspected [X2]. This report focuses on a small outbreak of enteritis due to C. fetus ss. jejuni. The affected patients had been drinking unpasteurized milk from an infected cow.
From the Division of Infectious Diseases, Department of Medicine, and Department of Pathology, University of Colorado Medical Center, and the Microbiology Laboratory, Denver V. A. Medical Center, Denver, Colorado. Requests for reprints should be addressed to Dr. Martin Blaser. Manuscript accepted March 19,1979. * Present address: Enteric Disease Branch, Bacterial Diseases Division, Bureau of Epidem. iology, Center for Disease Control, Atlanta, Georgia 30333. + Present address: P.O. Box 144, Anthong, New Mexico 88021.
METHODS Stool specimens were inoculated on a special plate medium to facilitate isolation of Campylobacter. As part of an ongoing study. stools from 80 normal persons were also cultured for C. fetus ss. jejuni and served as controls [ll]. The medium consisted of a Brucella agar base, 10 per cent sheep red blood cells, with vancpmycinl mg/lOO ml, trimethoprim 0.5 mg/lOO ml, polymyxin B 2.5 IU/ml and amphotericin B 2 &ml. Swabs were also placed in thioglycollate broth with the aforementioned antibiotics and 0.16 per cent agar. Plates were incubated at 42% for 48 hours in an atmosphere of 5 per cent oxygen, 10 per cent carbon dioxide and 85 per cent nitrogen whereas broth specimens were refrigerated for 8 hours and then subcultured onto the special medium.
October 1979
The American Journal of Medicine
Volume 67
715
CAMPYLOBACTER
ENTERITIS AND UNPASTEURIZED
MILK-BLASER
Plates were examined at 24 and 48 hours. Colonies which were grey, small or mucoid, nonhemolytic and flat were gram-stained (counter stain with carbol fuchsin) and/or studied under phase microscopy. If the gram stain showed vibrio forms or if phase contrast microscopy revealed a characteristic “darting” motility, further isolation and biochemical testing were performed. Organisms were considered to be C. fetus ss. jejuni if they were motile, oxidase- and catalasepositive, grew in Brucella broth at 37% and 42’C but not at 25’C, did not grow in 3.5 and 6.5 per cent salt, multiplied in 1 per cent glycine, were capable of producing hydrogen sulfide in Kligers media with lead acetate paper and were sensitive to nalidixic acid [3]. All isolates were tested for sensitivity to erythromycin, chloramphenicol, tetracycline, gentamicin, penicillin and cephalothin using the Kirby-Bauer method [l3]. Serums were obtained from patients during the acute and convalescent phase, and serums obtained from hospital personnel served as controls. Anitbody titers were determined using an indirect fluorescent antibody test with autologous isolates as the bacterial antigen. Bacteria were harvested from 24-hour plate cultures, washed twice in phosphate buffered saline solution (pH 7.2), boiled for 1 hour, washed again in phosphate buffered saline solution, and resuspended to a final concentration of 3 X lo7 organisms/ml. Organisms were dropped onto a slide and fixed with acetone. Test serums were diluted in twofold increments, layered onto the slides which were incubated at 37’C for 39 minutes, then washed in phosphate buffered saline solution, and fluorescein isocyanatetagged goat antihuman globulin (1:16 dilution in 0.1 per cent solution of Amido Black) was added. Slides were again incubated, then washed in phosphate buffered saline solution and examined using an epi-illuminated Zeiss fluorescent microscope. Fluorescence was judged at 0 to 4+. Slides were read as unknowns; those which showed 3+ or greater fluorescence were considered positive.
CASE REPORTS Case I. On June 7, a 20 year old man first noted the onset of intermittent, sharp periumbilical pain which increased in severity. By the following morning, he complained of malaise, anorexia and feverishness. Diarrhea began later that day, and the patient had approximately 10 bowel movements, progressing from loose, to watery, to frankly bloody stools. When seen at a hospital emergency room he was febrile (temperature 38.5%) and had tenderness in the lower right quadrant. Many leukocytes were seen in a wet preparation of his stool. His white blood cell count was 13,700/mm3 with 81 polymorphonuclear leukocytes, 5 juvenile forms, 7 lymphocytes, 6 monocytes and 1 basophil. The patient was treated symptomatically with diphenoxylate hydrochloride. C. fetus ss. jejuni was isolated from a stool specimen which was negative for Salmonellae, Shigellae, Staphylococcus aureus and parasites. Over the next few days there was a gradual diminution in symptoms; however, on June 13 the patient noted a worsening of the abdominal pain and several diarrhea1 stools. The administration of erythromycin (Z g/day] was begun, and within 48 hours the patient was completely well.
716
October 1979
The American Journal of Medicine
ET AL.
Case 2. On June 7, the 47 year old father of the preceding patient [Case 1) noted abdominal bloating and the onset of diarrhea. The following day, he experienced generalized abdominal cramping, malaise, myalgias, and chills, and recorded a temperature of 39.2%. The diarrhea worsened and was associated with severe abdominal cramps. Stools were watery but not bloody. His symptoms gradually diminished over the
next three days, and C. fetus ss. jejuni was isolated from a stool culture obtained on June 12 when he was asymptomatic. Case 3. On the evening of June 6, the 61 year old grandmother of the patient (Case 1) noted malaise and abdominal queasi-
ness. On the following day, she felt weak, complained of myalgias. severe nausea, anorexia and noted a low grade fever (temperature 38’C). Later that day, she experienced intermittent, generalized sharp abdominal cramping and had eight bowel movements within the next 24 hours. The stools were initially loose, and later became mucoid and watery. These symptoms persisted for 48 hours and diminished over the next four days. C. fetus ss. jejuni was isolated from a stool culture taken on June 13. On June 15, the patient noted recurrence of malaise, myalgias and chilly sensations. The following day she was febrile (temperature 39.4%), felt weak and again complained of severe nausea. She did not have abdominal cramps or diarrhea. The administration of erythromycin (2 g/day) was begun, and her symptoms resolved over the next 48 hours.
EPIDEMIOLOGIC INVESTIGATION Stool cultures obtained from the two asymptomatic family members were negative for C. fetus ss. jejuni, as were stool cultures from 80 asymptomatic normal subjects. The three symptomatic patients described in this study were members of a five person family extending over three generations. The index patient (Case 1) and his parents lived in the city. His grandparents (Case 3 and her husband) lived on a small farm 30 miles from the city and raised calves, sheep, swine, chicks and laying hens. In addition, a horse, a milk cow, a cat and a dog were on the farm. A well provided all the water for the farm. The index patient had been a frequent visitor to the farm in the previous few weeks and had contact with many of the animals as part of his routine farm chores. The father of the index patient (Case 1) had not been to the farm in the preceding month. All family members ate eggs from the farm. Cracked eggs were not eaten or used. Rectal culture of the hens did not reveal C. fetus ss. jejuni. The cow provided the family with 1 gallon of milk a day. This was collected in a clean bucket and immediately refrigerated. Four of the five family members consumed the unpasteurized milk. All three patients consumed at least 1 pint of milk a day whereas the fourth family member consumed 1 glass per day. A specimen of milk obtained on June 12 was negative for C. fetus ss. jejuni but repeated fecal specimens from the cow were positive for two months. Treating the cow with tetracycline eradicated the asymptomatic intestinal carriage.
Volume 67
CAMPYLORACTER
C fetus ss. jejuni was also isolated from stool specimens from the swine. Culture of the chicks [whose feed contained tetracycline), sheep, domestic animals and well water were negative. LABORATORY
RESULTS
Serum was collected within five to seven days after the onset of clinical illness (acute serum) and two weeks later (convalescent serum]. Results of antibody studies are presented in Table I. All patients demonstrated a fourfold increase in IgG antibody titer whereas serum from the asymptomatic milk-drinking family member and 14 control subjects had titers from 12 to 1:8.Immunoglobulin M (IgM) antibody titers were initially high and remained so when compared to those in control serums. Convalescent serum from a person who had C. fetus ss. intestinalis bacteremia showed an IgG titer of 1:8to our patients’ isolates of C. fetus ss. jejuni. Our patients’ serums, when tested against C. fetus ss. intestinalis, all showed IgG titers less than 1:8. All stool isolates proved to be sensitive to tetracycline, gentamicin, erythromycin and chloramphenicol, and were resistant to penicillin and cephalothin. Blood cultures were not performed. When this organism was inoculated to milk (to7/ml) and kept at 4’C, survival for three weeks was observed [l4]. COMMENTS The development of better laboratory methods for the isolation of Campylobacter species from coproculture has permitted determination of their roles as enteric pathogens. Recent studies using these techniques have demonstrated a significant rate of isolation of C. fetus ss. jejuni in association with diarrhea1 illness [5-111. Simultaneous isolation of the organism from the stool and blood of several patients with acute enteritis has confirmed pathogenicity in those patients. Lastly, in several human subjects from whom tissue was examined, the acute inflammation of the jejunum and ileum was identical in location and appearance to that seen in animals with enteritis caused by C. fetus ss. jejuni [4,6,151. As a result of these studies, the clinical spectrum of Campylobacter enteritis in human subjects has been defined [4,6]. In a majority of cases, the onset of illness is abrupt, characterized by fever, abdominal pain and diarrhea. The peak of the illness usually lasts for 24 to 72 hours, but longer durations have been reported. Diarrhea is usually the single most prominent symptom and in some cases, occult or gross blood with white blood cells and mucus are present. In the majority of cases, resolution is gradual over a few days to weeks but relapses may occur. There seems little question that the family we describe suffered an acute outbreak of Campylobacter enteritis. Isolation of C fetus ss. jejuni associated with a fourfold increase in IgG antibody titer all support this
ENTERITIS
TABLE I
AND IJNPASTEl~JRIZED
MII,K ~~ULASER ET AI,.
Reciprocal Indirect Fluorescent Antibody Titers to Campylobacter fetus ss. jejuni ___.-W
Case 1 Acute serum Convalescent serum Case 2 Acute serum Convalescent serum Case 3 Acute serum Convalescent serum Asymptomatic family member Acute serum Convalescent serum 14 asymptomatic control subjects, geometric mean
w-l
8 256
8 16
2 128
32 32
128 512
64 128
2 4 5.4
2 2 2.8
NOTE: Acute serum refers to serum collected within five to seven days after the onset of clinical illness and convalescent serum refers to serum collected two weeks later.
contention. Clinical features in patients from our outbreak fall well within the spectrum of disease due to Campylobacter. Of interest, two of our three patients had relapses several days after initial resolution of symptoms. C. fetus ss. jejuni is a frequent commensal in swine [IS] and has been isolated from asymptomatic cattle [v], cats and dogs [6,18]. In sheep and other ungulates it may exist in a commensal state or cause infectious abortion [19]. In fowl, C. fetus ss. jejuni may be a commensal [20] or cause vibrionic hepatitis [21]. The widespread reservoir of C. fetus ss. jejuni in the animal kingdom suggests that contact with animals or animal products would be important in the epidemiology of human discase. Campylobacter enteritis associated with infected dogs [22] and poultry [23] has been reported. A large waterborne outbreak of Campylobacter enteritis illustrated the importance of the oral route of infection [24]. The rate of enteric and vaginal carriage of this organism in asymptomatic subjects is less than 1 per cent [l4]. Current evidence favors an external source for most human infection, but better definition of the specific vectors is necessary. The simultaneous onset of symptoms in three previously healthy persons suggested a common source. Since the patients had different occupations, did not reside together and did not have exposure to the same farm animals, a foodborne source was suspected. In Case 2, the patient never consumed water from the farm. The only sources common to the three patients were the eggs and the raw milk which they consumed. The isolation of C. fetus ss. jejuni from a fecal specimen of the milk cow suggested that the unpasteurized milk was the vehicle for the infection. A fourth family member who drank smaller quantities of the suspect milk had no clinical, cultural or serologic evidence of infection.
October 1979
The American Journal of Medicine
Volume 67
717
CAMPYLOBACTER
ENTERITIS AND UNPASTEURIZED MILK-BLASER
In 1946,Levy [25] described an outbreak of diarrhea1 illness in a closed population, and epidemiologic investigation implicated unpasteurized cow’s milk as the source of infection. No known pathogens were found but gram stain of stool and blood cultures from affected patients demonstrated vibrio forms. However, the organisms did not survive long on routine culture media. In 1977, Taylor [26] reported three cases of septicemia due to C. fetus ss. jejuni. All three patients consumed raw milk from the same dairy. Case-control analysis implicated that this consumption of unpasteurized milk was the most significant risk factor. Recently, in En-
ET AL.
gland, 38 persons who drank unpasteurized milk from the same sources were found to be infected with C. fetus ss. jejuni, and in an outbreak involving another dairy 14 patients were affected. C. fetus ss. jejuni was isolated from the milk on two occasions [27]. The present report provides further evidence for raw milk as a vehicle for the transmission of Campylobacter infections. ACKNOWLEDGMENT We thank Ms. P. Riepe and Mr. A. Tsan for their technical assistance.
REFERENCES 1. Smith T: Spirilla associated
with disease of the fetal membranes in cattle (infectious abortionl. ,, 1 EXD . Med 28: 701. 1918. 2. MacFavdean F. Stockman S: Final renort of the Deuartmental Com”mittee appointed by the Board of Agriculture and Fisheries to Enquire into Epizootic Abortion. III. Abortion in Sheep, London, His Majesty’s Stationary Office, 1913. 3. Smibert RM: Cam ylobacter. Spiral and curved bacteria. Bergey’s Manua P of Determinative Bacteriology, 8th ed. (Buchanan RE, Gibbons NE, eds], Baltimore, Williams & Wilkins, 1974. p 207. 4. Kine EO: The laboratorv recognition of Vibrio fetus and a closely related vibrio isolated from cases of human vibriosis. Ann NY Acad Sci 98: 700,1962. 5. Dekeyser, P, Gossvin-Detrain M, Butzler JP, et al.: Acute enteritis due to related vibrio: first positive stool cultures. J Infect Dis 125: 390, 1972. 6. Skirrow MB: Campylobacter enteritis: a “new” disease. Br Med 12: 9.1977. 7. Pearson’ AD; Suckling WG. Ricciardi ID, et al.: Campylobatter associated diarrhea in Southamuton. Br Med 12: 955. MY
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Lauwers S. DeBoek M. Butzler IP: Camovlobacter enteritis _” in Brussels. Lancet i: 604.1978. Demo1 P, Bosmans E: Campylobacter enteritis in Central Africa. Lancet 1: 604.1978. Dixon JMS. Mueller L: Campylobacter enteritis-Alberta. Canada Diseases Weeklv Reoort ~011. o 6.1978. Blaser MJ, Berkowitz ID. LaFor& FM, et 2,:.Campylobacter enteritis: clinical and epidemiologic features. Ann Intern Med (in press). Bokkenhauser V: Vibrio fetus infection in man. I. Ten new cases and some epidemiologic observations. Am J Epidemiol91: 400.1970. Bauer AW, Kirby WM, Sherris JC. et al.: Antibiotic susceptibility testing by a standardized single disc method. Am J Clin Path01 45: 493.1966.
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The American Journal of Medicine
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Blaser MJ, Hardesty HL, Wang WLL: Survival of Campylobatter fetus ss. ieiuni in biolonical milieus labstractl. Presented at the 79th’Annual Mee%ng of the American Society for Microbiology, Los Angeles, California, 1979. Evans RG, Dadswell JV: Human vibriosis. Br Med J 3: 240. 1967. Deas DW: Observations on swine dysentery and associated vibrios. Vet Ret 72: 65.1960. Bryner JH, O’Berry PA, Estes PC, et al.: Studies of vibrios from gallbladder of market sheep and cattle. Am J Vet Res 33: 1439.1972. Blaser MJ, LaForce FM, Wang WLL: Reservoirs for human campylobacteriosis. Clin Res 27: 40A, 1979. Firehammer BD: The isolation of vibrios from ovine feces. Cornell Vet 55: 482.1965. Smibert RM: Vibrio fetus var. intestinalis isolated from the intestinal contents of birds. Am J Vet Res 30: 1437,1969. Peckham MC: Avian vibrio infections. Diseases of Poultry, 6th ed, (Hofstad MS, ed). Ames, Iowa, State University Press, 1972, p 332. Blaser MJ, Cravens J, Powers BW. et al.: Campylobacter enteritis associated with canine infection. Lancet 2: 979, 1978. Hayek LF, Cruikshank JG: Campylobacter enteritis. Br Med 12: 1219.1977. Tiohan W, Vogt Rt: Waterborne campylobacter gastroenteritis-Vermont. Morbiditv Mortalitv Weekly. Rep. 27: 207, 1978.
25.
Levy AJ: Gastro-enteritis outbreak probably due to a bovine strain of vibrio. Yale J Biol Med 18: 243.1946. 26. Taylor PR, Weinstein WM. Bryner JH: Campylobacter fetus infection in human subjects: association with raw milk. Am 1Med 66: 779.1979. 27. Communicable Diseases Surveillance Center (United Kingdom): An outbreak of Campylobacter infection, presumed milkborne. Communicable Disease Report 78: 47, 1978.
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