in Bacterial Pathogens Isolated From Turkeys in Minnesota From 1998 to 2002
Yashpal S. Malik, BVSc, PhD*
Karen Olsen, BS*
Yogesh Chander, PhD
Sagar M. Goyal, BVSc, PhD*
*Departments of Veterinary Diagnostic Medicine and Soils, Water
and Climate, University of
KEY WORDS: antimicrobial resistance, Escherichia
coli, Salmonella spp, Mannheimia hemolytica, Bordetella avium,
surveillance, turkeys, Minnesota
Antimicrobials are commonly used as feed additives in
turkey husbandry either as a growth enhancer or to minimize risks from
bacterial infections. This practice is believed to contribute toward
the development of antimicrobial resistance in bacterial
pathogens. Perusal of the literature revealed the availability of exhaustive
data on antimicrobial resistance in chicken pathogens but a limited
amount of information on the prevalence of antimicrobial resistance
in bacterial pathogens of turkeys. The purpose of this study was to
determine the presence of antimicrobial resistance among bacterial pathogens
isolated from turkeys in Minnesota during 1998 through 2002. The in vitro antimicrobial resistance
of Escherichia coli, Salmonella
spp., Bordetella avium, and Mannheimia
hemolytica was evaluated using the disc diffusion antimicrobial
susceptibility test. Antimicrobial agents tested were amikacin, enrofloxacin,
gentamicin, ampicillin, penicillin, ceftiofur, trimethoprim sulfa, erythromycin,
spectinomycin, tetracycline, clindamycin, sulfadimethoxine, and sulfachloropyridiazine.
The frequency of bacterial isolation from 1998 to 2002 was E. coli (191 or 58.4%) > Salmonella spp.
(87 or 26.6%) >
B. avium (35 or 10.7%)
haemolytica (14 or 4.3%).
A majority of the isolates was susceptible to amikacin, whereas variable
resistance was observed against gentamicin, ampicillin, ceftiofur, and
trimethoprim sulfa. The resistance to erythromycin, spectinomycin, tetracycline,
sulfadimethoxine, and sulfachloropyridiazine was high. In general, antimicrobial
resistance appeared to be increasing from year to year, indicating the
need for continued surveillance to effectively monitor and control the
emergence of resistance in turkey pathogens.
The control of infectious diseases is of basic economic
importance for animal farming to be successful. The use of antibiotics
in feed during the past decades has led to an improvement in the health
and safety of farm animals and poultry. Antibiotics are widely used
for the treatment of infectious diseases or as growth promoters in the
poultry industry,14 which is believed to contribute toward the development
of antibiotic resistance in both the pathogens and normal microflora
Zoonotic pathogens could acquire antibiotic resistance
while inhabiting the gastrointestinal tracts of food animals and could
then transfer this resistance to humans via the food chain.68 Although
transfer of resistant bacteria from turkeys to farmers, caretakers,
slaughterers, and area residents has been reported,9,10 there is no
consensus over the use and/or effects of antimicrobials in food animals.1
The development of antimicrobial resistance is of clinical importance
and requires ongoing surveillance to effectively monitor its spread.
As a part of global surveillance programs, it is important to determine
local patterns of antimicrobial resistance on a regular basis to ascertain
if antimicrobial resistance is increasing or decreasing. Perusal of
literature reveals the existence of exhaustive data on antimicrobial
resistance in chickens but limited information is available on the prevalence
of antimicrobial resistance in turkeys.
This retrospective study was conducted to determine
the prevalence of 4 different pathogens (Escherichia coli, Salmonella spp., Bordetella avium,
and Mannheimia hemolytica) and their associated drug resistance in turkeys submitted to the Minnesota
Veterinary Diagnostic Laboratory during the last 5 years (19982002).
Pathogens were selected on the basis of their direct impact on poultry
and/or humans. It is well known that animals can serve as reservoirs
of E. coli for
humans and transfer of antibiotic resistance genes from poultry to humans
has been documented.10,11 Turkeys have also been reported to be a vehicle
for outbreaks of salmonellosis in humans.12 B. avium is an opportunistic pathogen of chickens and turkeys
and has been isolated from humans.13,14 M. haemolytica is
one of the most prominent pathogens of domestic animals, causing severe
disease and major economic losses in cattle, swine, sheep, and poultry.15
MATERIALS AND METHODS
Source of Samples
Tracheal and sinus swabs and lung tissues from affected
turkeys are routinely submitted to the Minnesota Veterinary Diagnostic
Laboratory for disease diagnosis. These samples were initially inoculated
on sheep blood agar (SBA) followed by incubation at 37˚C for 18
to 24 hours. Suspect colonies of bacteria were then subjected to standard
biochemical tests and were further tested with API-ZYM system (BioMeriuex-France,
Lyon, France) for confirmation.
Antimicrobial susceptibility of bacterial isolates was
determined by methods described in the National Committee for Clinical
Laboratory Standards (NCCLS).1618 Briefly, an isolated colony of bacteria
was inoculated in Mueller Hinton broth followed by overnight incubation
at 37˚C. The overnight culture was than swabbed on the surface
of blood-Mueller Hinton agar followed by the application of antibiotic
discs. In 1998, ampicillin, gentamicin, spectinomycin, sulfadimethoxine,
sulfachloropyridiazine, trimethoprim sulfa, and tetracycline were used
whereas in 1999, amikacin, ceftiofur, clindamycin, erythromycin, penicillin,
and enrofloxacin were also included. Antibiotic resistance was determined
using the criteria for Gram-negative organisms as established by the
A total of 327 isolates of E. coli, Salmonella spp.,
B. avium, and M. hemolytica were isolated from
1998 through 2002 (Table 1). The frequency of bacterial isolation was
coli (58.4%) >
Salmonella spp. (26.6%)
B. avium (10.7%) >
haemolytica (4.3%) (Fig.1).
hemolytica was isolated in 1998. Yearly prevalence of all bacterial
isolates is shown in Figure 1.
As shown in Table 2, a majority of E.
coli isolates was susceptible to amikacin except for one isolate
each in 2000 and 2002. Resistance to ceftiofur (3.88.0%),
enrofloxacin (4.09.7%), and
trimethoprim sulfa (7.519.2%)
was low (Table 2). Resistance to ampicillin (29.042.0%)
and gentamicin (48.469.2%)
was consistent from year to year (Table 2), whereas resistance to spectinomycin,
sulfadimethoxine, and tetracycline was very high (88.7100%).
Amikacin and enrofloxacin were the most effective antibiotics
vitro against Salmonella (Table 3). Thus, none
of the 87 isolates was resistant to amikacin, whereas only one isolate
was resistant to enrofloxacin (Table 3). Resistance to ceftiofur (0.020.8%) and trimethoprim sulfa (0.014.3%) was also low. Resistance to gentamicin (21.461.1%),
ampicillin (7.161.1 %), and tetracycline (20.077.8%) was highly variable from year to year and
resistance to spectinomycin and sulfadimethoxine was almost absolute
varying from 94.4% to 100%.
As shown in Table 4, B. avium isolates were highly susceptible
to amikacin. Although all isolates showed absolute resistance to gentamicin
from 1998 to 2000, none was found resistant in 2001 and 2002. Similarly,
all isolates were susceptible to tetracycline in 2002 while showing
variable resistance (42.9100%)
from 1998 to 2001. An increasing trend of resistance was seen against
erythromycin (from 37.5100%)
and trimethoprim sulfa (from 12.557.1%),
whereas a decreasing trend was observed against ampicillin (from 7540%).
Resistance to sulfachloropyridiazine, clindamycin, and enrofloxacin
was very high (71.4100%).
All isolates were susceptible to amikacin, ampicillin,
ceftiofur, trimethoprim sulfa, and gentamicin (Table 5). During 2001
and 2002, a sudden increase in resistance to enrofloxacin (42.9100%) and complete resistance to tetracycline was
observed, although these antibiotics were very effective during 1999
and 2000 (Table 5). Resistance to sulfadimethoxine, spectinomycin, erythromycin,
and penicillin was very high (66.6100%).
general, isolation rate of E. coli, Salmonella, B. avium,
and M. haemolytica continued to increase
from 1998 through 2001 with a decreasing trend in 2002. It would be
interesting to determine if this decreasing trend continues in the coming
years. Limited information is available in the literature on antimicrobial
resistance in bacterial pathogens of turkeys whereas exhaustive information
has been published on antimicrobial resistance in chickens. Therefore,
in this study, the results for some of the antimicrobials against bacterial
isolates of turkeys are compared with resistance data available for
High resistance to
spectinomycin and tetracycline seen in E. coli isolates is in accordance with earlier reports11,19
in which high resistance to these antibiotics (57.099.0%) was reported in chicken isolates. Low resistance
to trimethoprim sulfa and amikacin observed in the present study is
in contrast to the reports by Al-Ghamdi et al.19 and Over et al.,20
respectively, in which high resistance to these antibiotics was reported
in turkey isolates. Our results on resistance of E. coli isolates
to gentamicin (48.469.2%) are similar
to those observed in an earlier report21 in which 86% of the E. coli isolates from turkeys were resistant. High resistance
to sulfadimethoxine (93.5100%) is in accordance
with the report of Takahashi et al.22 in which high resistance to this
antimicrobial was observed in chicken isolates. Results on resistance
to ampicillin (2942.3%) and enrofloxacin (4.09.7%) are in accordance with a report by Amara et al.23
in which low to medium resistance (1540%) was
reported in E. coli isolates of chickens. A search of the literature
revealed no data on ceftiofur resistance in E. coli isolates
from chickens and turkeys, although very low resistance (3.88.0%) was seen in this study.
of the Salmonella isolates showed high
resistance to sulfadimethoxine and spectinomycin, which is in accordance
with the findings of Rajashekera et al.24 in which high resistance to
sulfonamides was reported. However, results on ampicillin resistance
are in contrast to their findings because a decreasing trend of resistance
was seen in the present study. The resistance to gentamicin and ampicillin
decreased from year to year, which is in contrast to the findings of
Hirsh et al.25
who reported an increasing trend of resistance to gentamicin and ampicillin
in turkeys. Our results on tetracycline resistance are in contrast to
the findings of Ekperigin et al.26 in which tetracycline was reported
to be the most effective drug in vitro for Salmonella
in chickens. In the present study, Salmonella isolates appeared to
show higher resistance against tetracycline. These results are more
close to those of Poppe et al.27 in which resistance to amikacin, gentamicin,
and spectinomycin was low, medium, and high, respectively. Low resistance
(0.014.3%) against trimethoprim sulfa is similar to
that reported in a study by Nayak and Kenney28 in which 25% of the Salmonella isolates from turkeys
were resistant. Low resistance to enrofloxacin (04.8%) and ceftiofur (020.8%)
is also in accordance with a report by Pedersen et al.29 in which Salmonella
isolates from Danish turkeys were reported to be highly susceptible
to ceftiofur and enrofloxacin.
avium isolates showed high susceptibility to amikacin, which
is in agreement with Mortensen et al.30 who found B. avium to be highly sensitive
to amikacin. The organism was highly resistant to clindamycin (72.7100%), enrofloxacin (71.4100%), and erythromycin (37.5100%). None of the isolates during 2001 and 2002
showed resistance to gentamicin, whereas complete resistance was seen
during 1998 through 2000. Similarly high resistance (42.9100%) against tetracycline was seen from 1998 through 2001 but not
in 2002. These results are in contrast to those reported by Blackall
et al.31 in which tetracycline was found to be effective. In the present
avium isolates showed a decreasing trend of resistance to ampicillin
from 1998 to 2002 (75.040.0%).
These results are in agreement with those of Blackall et al.31 in which
avium isolates were reported to be sensitive to these 2 antimicrobials.
The results on erythromycin resistance (37.5100%)
are similar to those reported by Blackall et al.31 in which increasing
resistance to erythromycin was reported. Data on amikacin and sulfachloropyridiazine
resistance in turkey and chicken isolates have not been reported. In
the present study, however, none of the isolates were resistant to amikacin,
whereas 81.8% to 100% were resistant to sulfachloropyridiazine.
haemolytica does not appear to be an important pathogen of turkeys
because only 14 isolates were obtained during the 5 years of the study.
The presence of high resistance in M. haemolytica against erythromycin
and tetracycline is in accordance with the findings of Watt et al.32
who reported high tetracycline and erythromycin resistance in M.
haemolytica isolates. However, the absence of resistance to ampicillin
is in contrast to the findings of Watt et al.32 in which high resistance to
ampicillin was found. The results on ceftiofur resistance are in agreement
with those of Watt et al.32 who reported M. haemolytica isolates to be highly
sensitive to ceftiofur. Our results are in agreement with those of Hormandorfer
and Bauer33 as far as resistance to sulfadimethoxine, tetracycline,
and enrofloxacin is concerned. Our results are also in agreement with
those of Diker et al.34 for sensitivity to ampicillin but not for erythromycin.
No data are available on amikacin and penicillin resistance in turkey
and chicken isolates, although none of the isolate was resistant to
amikacin in his study and none was sensitive to penicillin.
These results clearly indicate the
in vitro efficacy of amikacin, ceftiofur, and trimethoprim sulfa
against turkey pathogens whereas spectinomycin and sulfadimethoxine
appeared to be useless in vitro. It is rather easier to
explain an increasing trend of resistance to tetracycline and erythromycin
avium, but why this organism became susceptible to gentamicin
in 2001 and 2002 is not clear. These data on antimicrobial resistance
patterns and distribution frequency of bacterial pathogens in turkeys
could prove to be useful in developing strategies for the control of
bacterial infections in the turkey industry. In summary, E. coli and Salmonella
appeared to be the major pathogens of turkeys in Minnesota. A
majority of the isolates showed resistance to erythromycin, spectinomycin,
tetracycline, sulfadimethoxine, and sulfachloropyridiazine, whereas
variable resistance was seen against gentamicin, ampicillin, ceftiofur,
and trimethoprim sulfa. Amikacin appeared to be the most effective antibiotic
vitro. In general, antimicrobial resistance appeared to be increasing
from year to year.
1. McEwen SA, Fedorka-Cray PJ: Antimicrobial use
and resistance in animals. Clin Infect Dis 34:93106, 2002.
2. Swezey JL, Baldwin BB, Bromel MC: Effects of oxytetracycline
as a turkey feed additive. Poult Sci 60:738743, 1981.
3. Devriese LA, Hommez J, Vandamme P, et al: In
vitro antibiotic sensitivity of Ornithobacterium rhinotracheale strains
from poultry and wild birds. Vet Rec 137:435436, 1995.
4. van Veen L, Hartman E, Fabri T: In
vitro antibiotic sensitivity of strains of Ornithobacterium
rhinotracheale isolated in the Netherlands between 1996 and 1999.
Rec 149:611613, 2001.
5. Manie T, Khan S, Brozel VS, et al: Antimicrobial
resistance of bacteria isolated from slaughtered and retail chickens
in South Africa. Lett Appl Microbiol 26:253258,
6. van den Bogaard AE, London N, Driessen C, et al:
Antibiotic resistance of fecal Escherichia coli in poultry, poultry
farmers and poultry slaughterers. J Antimicrob Chemother 47:763771,
7. van den Bogaard AE, Stobberingh E: Antibiotic
usage in animals: impact on bacterial resistance and public health.
8. Threlfall EJ: Antimicrobial drug resistance in
problem and perspectives in food and waterborne infections. FEMS
Microbiol Rev 26:141148, 2002.
9. Stobberingh E, van den Bogaard AE, London N, et
al: Enterococci with glycopeptide resistance in turkeys, turkeys farmers,
turkey slaughterers and (sub) urban residents in South of the Netherlands:
evidence for transmission of vancomycin resistance from animals to humans?
Agents Chemother 43:22152221, 1999.
10. Levy SB, Fitzgerald GB, Macone AB: Spread of antibiotic
resistant plasmids from chicken to chicken and from chicken to man.
11. Ojeniyi AA: Direct transmission of Escherichia
coli from poultry to humans. Epidemiol Infect 103:513522, 1989.
12. Grein TO, Flanagan D, McCarthy T, et al: An outbreak
of multidrug resistant Salmonella typhimurium food poisoning
at a wedding reception. Irish Med J 92:238241, 1999.
13. Jackwood MW, McCarter SM, Brown TP: Bordetella
avium: an opportunistic pathogen in Leghorn chickens. Avian
Dis 39:360367, 1995.
14. Dorittke C, Vandamme P, Hizaz KH, et al: Isolation
of a Bordetella avium-like organism
from human specimen. Eur J Clin Microbiol Infect Dis
15. Confer AW: Immunogens of Pasteurella. Vet
Microbiol 37:353368, 1993.
16. Performance Standards for Antimicrobial
Susceptibility Testing: Approved Standards M100-S6. Wayne, PA:
National Committee for Clinical Laboratory Standards; 1995.
17. Performance Standards for Antimicrobial
Disc Susceptibility Tests. Approved Standards M2-A6. Wayne, PA:
National Committee for Clinical Laboratory Standards; 1997.
18. Performance Standards for Antimicrobial
Disk Dilution Susceptibility Tests for Bacteria Isolated From Animals.
Approved Standards, M31- A2. Wayne, PA: National Committee for
Clinical Laboratory Standards; 2002.
19. Al-Ghamdi MS, El-Morsy F, Al-Mustafa ZH, et al:
Antibiotic resistance of Escherichia coli isolated from
poultry workers, parents and chicken in the eastern province of Saudi
Med Int Health 4:278283, 1999.
20. Over U, Gur D, Unal S, et al: The changing nature
of aminoglycoside resistance mechanisms and prevalence of newly recognized
resistance mechanisms in turkey. Clin Microbiol Infect 7:470478,
21. Dubel JR, Zink DL, Kelley LM, et al: Bacterial
antibiotic resistance: frequency of gentamicin resistant strains of
coli in the fecal microflora of commercial turkeys. Am
J Vet Res 43:17861789, 1982.
22. Takahashi I, Yoshida T, Higashide Y, et al: Susceptibilities
of Escherichia coli, Salmonella and
aureus isolated from animals to ofloxacin and commonly used antimicrobial
agents. Jpn J Antibiot 43:8999, 1990.
23. Amara A, Ziani Z, Bouzoubaa K: Antibioresistance
of Escherichia coli strains isolated
in Morocco from chickens with colibacillosis. Vet
Microbiol 43:325330, 1995.
24. Rajashekara G, Haverly E, Halvorson DA, et al:
Multidrug resistant Salmonella typhimurium DT104 in
poultry. J Food Prot 63:155161, 2000.
25. Hirsh DC, Ikeda JS, Martin LD, et al: R plasmid-mediated
gentamicin resistance in Salmonella isolated from turkeys and their
environment. Avian Dis 27:766772, 1983.
26. Ekperigin HE, Jang S, McCapes RH: Effective control
of a gentamicin resistant Salmonella arizona infection in
turkeys poults. Avian Dis 27:822829, 1983.
27. Poppe C, Koalr JJ, Demczuk WH, et al: Drug resistance
and biochemical characteristics of Salmonella from turkeys. Can
J Vet Res 59:241248, 1995.
28. Nayak R, Kenney PB: Screening of Salmonella
isolates from a turkey production facility for antibiotic resistance.
Sci 81:14961500, 2002.
29. Pedersen K, Hansen HC, Jorgensen JC, et al: Serovars
of Salmonella isolated from Danish
turkeys between 19952000 and their antimicrobial resistance.
Vet Rec 150:471474, 2002.
30. Mortensen JE, Brumbach A, Shryock TR: Antimicrobial
susceptibility of Bordetella avium and Bordetella
bronchiseptica isolates. Antimicrob Agents Chemother 33:771772,
31. Blackall PJ, Eaves LE, Fegan M: Antimicrobial
sensitivity testing of Australian isolates of Bordetella avium and Bordetella
avium- like organisms. Aust Vet J 72:97100, 1995.
32. Watt JL, Yancey RJ Jr, Salmon SA, et al: A 4-year
survey of antimicrobial susceptibility trends for isolates from cattle
with bovine respiratory disease in North America. J
Clin Microbiol 32:725731, 1994.
33. Hormandorfer S, Bauer J: Resistance pattern of
bovine Pasteurella. Berl Munch Tierarztt Wochenschr 109:168171,
34. Diker KS, Akam M, Hazirouglu R: Antimicrobial
susceptibility of Pasteurella hemolytica and Pasteurella
multocida isolated from pneumonic ovine lungs. Vet
Rec 134:597598, 1994.
Figure 1. Occurrence of Escherichia coli, Salmonella
spp., Bordetella avium, and Mannheimia hemolytica in turkeys from 1998
Isolation of Bacterial Pathogens From
Turkeys From 1998 to 2002
Number isolated in
1998 1999 2000 2001 2002 Total
coli 31 26 50 53
spp. 18 14
avium 4 8 11 7
hemolytica 0 2
Total Total = 327
Table 2. Percent Antibiotic Resistance in Escherichia
coli Isolated From Turkeys
Percent resistant during the indicated year
(number of isolates tested)
1998 1999 2000
Antimicrobial agents (31)
(26) (50) (53)
Amikacin NT 0.0 2.0
NT 3.8 8.0 7.5
NT 7.7 4.0 7.5
sulfa 12.9 19.2 10.0
Ampicillin 29.0 42.3 34.0
Gentamicin 48.4 69.2 62.0
Spectinomycin 100.0 100.0 100.0
Sulfadimethoxine 93.5 100.0
98.0 100.0 100.0
Tetracycline 93.5 96.1 96.0
Percent Antibiotic Resistance in Salmonella
spp. Isolated From Turkeys
resistant during the indicated year
(number of isolates tested)
1998 1999 2000
Antimicrobial agents (18)
(14) (24) (21)
NT 0.0 0.0 0.0
NT 0.0 0.0 4.8
NT 7.1 20.8 4.8
sulfa 11.1 14.4 0.0
Gentamicin 61.1 21.5 54.2
Ampicillin 61.1 7.1 37.5
Tetracycline 77.8 21.4 70.8
Spectinomycin 94.4 100.0 100.0
Sulfadimethoxine 100.0 100.0
95.8 100.0 100.0
Percent Antibiotic Resistance in Bordetella
avium Isolated From Turkeys
resistant during the indicated year
(number of isolates tested)
1998 1999 2000
(4) (8) (11)
NT 0.0 0.0 0.0
Gentamicin 100.0 100.0 100.0
Tetracycline 50.0 100.0 81.8
NT 37.5 45.5 100.0
sulfa 25.0 12.5 36.4
Ampicillin 75.0 50.0 27.3
100.0 81.8 NT
NT 100.0 72.7 100.0
NT 100.0 100.0 71.4
Table 5. Percent Antibiotic Resistance in Mannheimia
during the indicated year
of isolates tested)
Antibiotics (2) (3)
Amikacin 0.0 0.0
Ampicillin 0.0 0.0
Ceftiofur 0.0 0.0
sulfa 0.0 0.0 0.0 0.0
Gentamicin 0.0 0.0 0.0 0.0
Enrofloxacin 0.0 0.0 42.9 100.0
Tetracycline 0.0 0.0 100.0 100.0
Spectinomycin 100.0 66.7 85.7 100.0
Erythromycin 100.0 100.0 85.7 100.0
Penicillin 100.0 100.0
M. hemolytica was
isolated in 1998.