Outbreak of Cryptosporidiosis Among Veterinary Students in a
Bovine Obstetrics Laboratory
Carrie Klumb1,2, Jeff Bender3,Kirk Smith1, Elizabeth Cebelinski1, Joni Scheftel1
1Minnesota Department of Health 2CSTE/CDC Applied Epidemiology Fellowship
3University of Minnesota Veterinary Population Medicine
Background - Cryptosporidiosis• Acute gastrointestinal illness caused by
the protozoan parasite Cryptosporidium• Estimated 748,000 cases and 46 deaths
per year in the United States• Incubation period 1-12 days, mean 7 days• Profuse watery diarrhea, and cramping
over days to weeks. Fever and vomiting also may occur
Background - Cryptosporidiosis (cont.)• 19 species of Cryptosporidium
– C. hominis – human primary reservoir– C. parvum – cattle primary reservoir
• Oocysts (infective stage) are shed in the feces of infected hosts
• In cattle, primarily disease of calves – Shed up to 1010 oocysts per day in
feces • C. parvum is present on 90% of dairy
farms in the United States
Background - Cryptosporidiosis (cont.)• Transmission occurs through water and
food, from person-to-person and through direct animal contact
• 1983: Published report of cryptosporidiosis outbreak due to direct cattle contact among veterinary researchers
• Numerous outbreaks of cryptosporidiosis have been reported among veterinary students
Outbreak Detection• On April 22, 2010 gastrointestinal illness
among second year veterinary students at a college of veterinary medicine (CVM) was reported to the Minnesota Department of Health (MDH)
• Students had participated in a 2-day bovine obstetrics (OB) laboratory prior to becoming ill
Outbreak Detection (cont.)
• Four, 2-day OB laboratory sessions were scheduled at the CVM on:
Lab 1: April 1 and 2Lab 2: April 8 and 9Lab 3: April 15 and 16Lab 4: April 29 and 30
• The first three sessions had already occurred
Methods - Epidemiologic Investigation
• Obtained contact list of all students registered for any of four OB laboratory sessions
• Spoke to professor and employees about laboratory set-up and specific activities
• Developed a questionnaire regarding symptoms and risk factors both in and outside of the laboratory setting
• Interviewed students by phone or in person
Methods - Epidemiologic Investigation (cont.)
• Risk factors assessed– Tracking in small, mixed or large animal
practice– Dates of laboratory attendance – Eating or drinking in the laboratory – Use of personal protective equipment
(PPE)
Methods - Epidemiologic Investigation (cont.)
• Risk factors assessed– Hand hygiene– Being splashed in the face during the
laboratory session– Events or meals in common with other
students• Analyses were performed using Epi-Info
software, version 6.04d and SAS, version 9.2
Methods - Case Definition
A student who attended an OB laboratory session and subsequently had either:
a) A stool sample test positive for Cryptosporidium spp.
ORb) 3 or more days of diarrhea
(≥3 loose stools in 24 hours)
Methods - Laboratory Investigation• Testing was performed on stool samples
– Students and employees with symptoms
– Calves from Dairy Farm A, where animals for class were obtained
• Fecal samples from the calves actually used in the laboratories were unavailable
Methods - Laboratory Investigation (cont.)
• The MDH Public Health Laboratory conducted routine testing for Salmonella, Shigella, Campylobacter, Yersinia and E. coli O157:H7
• PCR for Shiga toxin-producing E. coli virulence factor genes (stx1, stx2, eae and hlyA) performed
Methods - Laboratory Investigation (cont.)
• Testing for Giardia and Cryptosporidium spp. performed using acid fast staining, direct fluorescent antibody tests
• Positive Cryptosporidium specimens identified to species by PCR fragment length polymorphism (PCR-RFLP) – Further identified to subtype by DNA
sequence analysis of the 60-kDa glycoprotein (gp60) gene
• Laboratory gives students hands-on experience assisting cows with difficult labors
• Course is highly physical and occurs in a small room
• Approximately 12 students working in pairs under the supervision of professor or lab technician
Results - OB Laboratory Description
• Mock cows consisting of a wooden box, plastic bag, and cow pelvic bones
• Dead calves are placed inside mock cow• Calves used are generally healthy, 48-hour
old bull calves euthanized by captive bolt
Results - OB Laboratory Description
Mock Cow
Mock Cow
Mock Cow
Mock Cow
• Day 1: practiced re-positioning abnormal fetal presentations and manual assistance of difficult births
Results - OB Laboratory Description
• Day 2: Students practiced re-positioning abnormal presentations and performed a fetotomy– Used when calf has died in utero and too
large to be delivered naturally– Calf is cut up in utero, removed in pieces
• Often results in laceration of the dead calf’s bowels
• Introduces fecal material into the classroom
Results - OB Laboratory Description
• Clean-up procedures– Students hosed down the inside of the
boxes, the plastic linings, and the floor with a garden hose
– No facial protection used
Results - OB Laboratory Description
OB Laboratory
OB Laboratory
Results - OB Laboratory Description
• Calves from Dairy Farm A used during outbreak were:– Mix of male and female – Several weeks old– Some had died of diarrheal illness
• Calves stored together between day 1 and 2 and not labeled– Could not tell who was exposed to which
calves
Results (cont.)
• 42 students had completed a lab at the time of the investigation
• 38 (91%) interviewed• 8 (21%) met the case definition• 22 (58%) reported no symptoms• 8 (21%) reported gastrointestinal symptoms
but did not meet case definition
Results (cont.)
• 8 of 8 (100%) and 4 of 22 (18%) controls attended Lab 2 on April 8 and 9 – Fisher’s exact p < 0.001
• There were no cases among Lab 1 or Lab 3 attendees
Case Symptoms (n = 8)
Symptom No. (%)Diarrhea 8 (100)Cramps 8 (100)Gas 7 (88)Weight Loss 4 (50)Fever 3 (38)Vomiting 1 (13)Bloody Diarrhea 1 (13)
Results (cont.)
• Median incubation period from Lab 2 was 7 days (range, 5 to 12 days)
• Median duration of illness for the 6 cases that had recovered at the time of the interview was 8.5 days (range, 7 to 14 days)
• 1 case was seen by their physician and 1 case was seen in an Emergency Department
Results (cont.)
• A CVM employee also became ill with bloody diarrhea, gas and cramps with illness onset on April 23
• Employee still ill at time of interview• Set up and cleaned up the OB laboratories on
several different dates• Incubation period calculated from the date of
Lab 2 was 15 days
• 4 of 8 human samples (3 student cases and CVM employee) positive for C. parvum of the same gp60 subtype (IIaA15G2R2)
• All human stool samples negative for Salmonella, Shigella, Campylobacter, Yersinia, E. coli O157:H7, Shiga toxin-producing E. coli virulence factors, and Giardia
Results - Human Testing
No.
of C
ases
16 17 18 19 20 21 2212 13 14 153129 30
March
1
4
23
1 1110987
April
62 43
First Obstetrics Lab
5
Second Obstetrics Lab
Third Obstetrics Lab
23 24 25 2628
Second Obstetrics Lab Attendee
Laboratory-confirmedVeterinary School Employee
Onset Date
Cryptosporidium parvum Cases Associated with a Veterinary School Bovine Obstetrics Laboratory by
Illness Onset Date, Minnesota, 2010
Results (cont.)• Among the Lab 2 attendees, the only risk
factor that approached significance was being splashed in the face at any point during day 2 of the lab– 7 of 7 cases vs. 1 of 3 controls*– Fisher’s exact p = 0.07
*1 case and 1 control were not able to recall and could not answer the question
Results (cont.)
• Students in Lab 2 were significantly more likely to have been splashed in the face at any point during the lab session than students in Labs 1 and 3– Odds ratio, 13.0– 95% confidence interval, 1.44 to 117.20 – p = 0.01
Self-Reported Personal Protective Equipment (PPE) Use Among Students
PPE No. (%) PPE No. (%)
Day 1 Day 2Sleeves 42 (100) Sleeves 42 (100)
Coveralls 28 ( 93) Coveralls 27 ( 96)
Boots 29 ( 97) Boots 29 (100)Rubber Apron 0 ( 0) Rubber
Apron 0 ( 0)
Mask/Face Shield 0 ( 0) Mask/Face
Shield 0 ( 0)
• 3 of 9 calf fecal samples collected at Dairy Farm A positive for C. parvum gp60 subtype (IIaA15G2R2)– Matched C. parvum in human samples– Subtype commonly found in Minnesota
• 3 fecal samples were positive for the E. coli virulence factor gene stx1
• Calf fecal samples negative for Salmonella, Shigella, Campylobacter, Yersinia, E. coli O157:H7 and Giardia
Results - Calf Testing
Conclusion• This was an outbreak of cryptosporidiosis
among veterinary students in an OB laboratory
• Calves that had died of a diarrheal illness and then used in the laboratories were the source of infection
Conclusion (cont.)
• Not surprising that stool samples from calves at Dairy Farm A tested positive for C. parvum
– C. parvum is present on 90% of dairy farms in the U.S.
– Higher prevalence in calves • Highlights the zoonotic risk inherent in
certain veterinary school activities
RecommendationsMDH epidemiologists and infection control specialists from CVM made recommendations
• Have students read provided information about zoonoses and prevention measures
• Remind students about proper hand hygiene
• Restrict calves used to euthanized 48-hour old healthy bull calves
Recommendations (cont.)
• Provide face shields/masks and rubber aprons to students
• Offer laundry service at school• Use an EPA-approved disinfectant to
clean laboratory
No illnesses were identified during the last laboratory on April 29 and 30
Follow Up• Visited laboratory on May 12, 2011 to observe• Warning was added to this year’s syllabus
Follow Up (cont.)
• Observations made– All students wore boots, coveralls,
sleeves – No face shields/masks offered or worn
by students– Laundry service not being provided by
the school– Near-term fetuses were being obtained
from slaughter facilities
Follow Up (cont.)
Conclusion• Cryptosporidiosis is sometimes called a
“rite of passage” in veterinary school– Serious disease– Infections should be avoided to the
extent possible
Conclusion• Prevention of future outbreaks
– Educating students about the risks– Ensuring calves used in laboratories
are healthy– Providing face shields or a mask and
goggles• Compendium of Veterinary Standard
Precautions for Zoonotic Disease Prevention in Veterinary Personnel offers infection control guidance
Acknowledgments
Minnesota Department of Health
Team Diarrhea
Foodborne, Vectorborne, and Zoonotic Disease Unit
University of Minnesota
Dr. Ricardo Chebel Amanda Doran
Dr. Tom Molitor Leslie Hiber