Enteric Diarrhoeal disease in sheep caused by Campylobacter infection in Alexandria Governorate

Enteric Diarrhoeal disease in sheep caused by Campylobacter infection in Alexandria Governorate

Ola .A. Basha and F. A. El-Shaboury

Animal Health Research Institute

Alexandria branch . Bacteriology Dept.

Summary

    The incidence of Campylobacter jejuni and Campylobacter coli harbouring the intestine of sheep with diarrhea was studied . A total number of 100 rectal swab samples were collected from sheep suffering from diarrhea at Alexandria governorate animal inclosures . The samples were examined bacteriologically . The bacteriological examination revealed the isolation of Campylobacter jejuni from 12 (12 %) out of 100 examined sheep ,while Campylobacter coli could be isolated from 9 cases of diarrheic sheep examined .

Moreover , Campylobacter isolated in this study were tested for susceptibility of 10 antibiotics . None of the isolates were resistant to Chloramphenicol and high percentages of isolates were resistant to Tetracycline , Penicillin and Sulphamethoxzole . These results indicate that Campylobacter colonization in the intestine is very common in sheep and the bacteria play a role in aggravation of diarrhea process . The public health importance of Campylobacter jejuni and Campylobacter coli were discussed .

Introduction

     Diarrhoea is the most common symptom of illness in lambs , which can be caused by many organisms , and more than one causative agent can be present in the one animal like Salmonella ,     E-coli , Viruses such as Rotavirus and Campylobacter which can also cause this problem  ( Radostits et al, . 2000 ) . Campylobacter species are now recognized as one of the most important causes of enteric diarrhoeal disease in domestic animals and humans throughout the world ( Atabay and Corry , 1997) . hence , Campylobacter enteritis constitutes a zoonoses  of major concern in puplic  health and indeed, has been shown to be a greater problem than salmonellosis in several countries (Stanley et al., 1998). Campylobacter is one of the most important foodborn microorganisms leading to gastroenteritis in human . These human cases are most likely associated  with handling or consumption  of  undercooked meat products ( Oosterom  et al .,1984 ) .

     The source of infection for humans in developed countries is though to be the massive reservoir  of Campylobacter jejuni and to a lesser extent Campylobacter coli in the animal population . It is assumed that man may be infected by direct contact with diseased animals suffering from Campylobacter diarrhoea or by consumption of food or water contaminated by secretion or excretion of diseased animals ( Acha and Szyfers . 1991,  Shakespeare , 2002 and Harmut et al ., 2003 ) . Campylobacter is a Gram- negative cylender, curved , and motile rod . It is a  microaerophilic  organism ,as it  requires  a reduced levels  of oxygen for growth . It is relatively fragile , and sensitive to environmental stresses such as 20 % oxygen, drying , heating , disinfectants and acidic conditions . Because of its microaerophilic characteristics , the organism requires 5% oxygen and10 % carbon dioxide for optimal growth  conditions (Betty et al. , 1998 ) . It is evident that Campylobacter jejuni and Campylobacter coli are responsible factor of  diarrhea in sheep ( Diker , 1987 ; Terzolo , 1988 and Khalil et al . , 1993 ) .

   Increasing antimicrobial resistance Campylobacter infection cases is a recognized problem,( Saenz et al . , 2000  and  Jensen and  Aarestrup , 2001 ) . However , Gur et al . ( 1989 ) stated that quinolone antibiotics were very active against Campylobacter species and showed that there was no any marked difference between the susceptibility patterns of C. jejuni and C.coli .

      The present study was designed to investigate the presence of Campylobacter infection in sheep at Alexandria governorate animal inclosures . Moreover , due to the public health hazard of antibiotic resistant for Campylobacter, antibiotic sensitivity patterns of Campylobacter Jejuni and Campylobacter coli were evaluated .

Material and Methods

 Collection of samples :

     Sterile rectal swab samples were collected from 100 diarrheic sheep of different ages and sex suffering from diarrhea , raised in small groups at  Alexandria governorate .

Bacteriological examination :

    Rectal swab samples were inoculated into sterile tube containing brucella broth (Retting , 1979) to preserve the viability of the organism during transportation to the laboratory . The samples were examined for Campylobacter jejuni and Campylobacter Coli by streaking on Skirrow medium (Skirrow , 1977 )containing 5-7 % horse blood  and Campylobacter selective supplement (Oxoid ) . The plates were incubated at 42 Cْ for 48 hours in an atmosphere of 5 % oxygen , 10 % carbon dioxide and 85% nitrogen in standard anaerobic jars . The bacterial isolates were purified and identified by using colonial , morphological and biochemical characteristics ( Koneman et al . 1988 and                   Quinn et al . 2002 ) .

In – vetro antibiotic sensitivity test :

      Different types of antibacterial sensitivity discs  were  used in the sensitivity tests to determine  the sensitivity  and  resistance of  different  isolated  strains  of Campylobacter jejuni and Campylobacter coli . The antibiotic discs were obtained from Oxoid . The  technique was  carried out  using  disc  diffusion  method according to Bopp et al . ( 1985).Three colonies of Campylobacter organisms were  inoculated  into tubes containing 5 ml Muller Hinton broth ( Oxoid ) , then incubated for eight hours under reduced  oxygen tension at 37°C until  the turbidity exceeded that of the standard McFarland 0.5 barium sulphate tube ( 0.5 ml of 1.175 % barium chloride hydrate to 99 ml of 1 % sulphuric acid ) . The turbidity was adjusted to match that of the McFarland 0.5 barium sulphate standard tube by adding sterile saline solution .  The suspension was then inoculated evenly on 150 mm Muller – Hinton agar plates supplemented with 5 % defebrinated sheep blood . Antibiotic discs were placed on the surface  of agar plate in a radial  pattern  with the lowest concentration toward  the center . The plates were incubated for 72 hours at 37 °C under the microaerophilic conditions and the inhibitory zone diameters were measured .

Results

Incidence of C. jejuni  and C. coli in rectal  swab samples collected  from  diarrheic  sheep:

      Isolates of Campylobacter jejuni  were recovered from 12 ( 12 % ) out of 100 rectal swab fecal samples examined while Campylobacter coli was recovered in 9 ( 9 % )of diarrheic sheep .

Totally C. jejuni  and C. coli could be isolated from 21 out of 100 sheep examined Table (1 ) .

The biochemical characteristics of Campylobacter isolates are illustrated in Table (2 ) .

In – Vitro antimicrobial sensitivity

       As shown in Table (3 ) , C. jejuni isolated were sensitive to Chloramphenicol ( 100 % ) followed by Neomycin and Gentamycin ( 80 % ) , while 60 % only for Ampicillin  . On the other hand , all

isolates of C. jejuni were resistant to Tetracyclin ( 100 % ) and  80 % of  isolates  were resistant to Erythromycin , Sulphamethoxazole and Pencillin .

     As shown in table ( 4 ) , C. coli isolated  from sheep rectal swab samples were sensitive to

Ampicillin and Neomycin  ( 100 % ) followed by Chloramphenicol ( 80 % ) but they showed high resistance to Penicillin , Tetracyclin and Naladixic acid .

Discussion

       Campylobacter species are microaerobic inhabitant of the gastrointestinal tract of various animals including  sheep ( Betty et al . 1998 ) . It have been recognized as a cause of diarrhea in sheep ( Radostitis  et al . 2000 ) . In this study , rectal swab samples were collected from sheep with diarrhea  and used  for  isolation  of  Campylobacter  jejuni  and  Campylobacter  coli . As shown in     table 1 Campylobacter jejuni was isolated from examined  rectal swab sheep samples  at a percentage  of  12 %.This  bacteria  was  isolated  from  sheep faeces  by  many  various studies ( Turkoson et al . 1988  and Adesiyun et al . 1992 ). Nearly similar percentage was recorded by Rosef et al . ( 1983 ) who isolated 8.1 % . Higher recovery rates were recorded by Manser and Dalziel ( 1985 ) and Adesiyun et al . ( 1992 ) who isolated 22 and 17.9 % respectively .

       On the other hand , low  rate  of  2 % was recorded by Turkoson et al . (1988 ) . These differences in isolation rates may be attributed to sample collection in different seasons as high temperature in summer months , while lower recovery rates occurred during winter and spring and this was reported by Hanninen et al . ( 2000 ) . The result illustrated in table 1 showed that Campylobacter coli was isolated from examined rectal swab sheep samples at percentage of 9 % . Nearly similar results were recorded by Terzolo ( 1988 ) and Kakkar and Dogra ( 1990 ) , and lower result was reported by Rosef et al . ( 1983 ) but higher result was recorded by Giacoboni et al.(1993 )

       The rate of  isolation  for Campylobacter jejuni was higher than Campylobacter coli in rectal swab samples of sheep . Table ( 1 ) . This support the observation of Rosef et al ., ( 1983 ) and Busato et al.(1999) who reported that Campylobacter coli is less frequently involved in animal than Campylobacter jejuni . However , the epidemiological features of both are similar and sometimes not further differentiated in literature ( Busato et al . 1999 ) .

     Campylobacteriosis is acute or chronic infection of humans and animals , Transmission from animals to human has been known for Campylobacter jejuni and Campylobacter coli . The infection in man is manifested generally by acute enteritis , abdominal pain , diarrhea in most cases with stool contain blood , pus or mucous , fever up to 40 °C and in few cases vomiting . Abdominal symptoms may  lead to laparotomy or appendectomy ( Acha and Szyfers , 1991 and Hartmut et al . 2003 ) .

    In- vetro antibiotic sensitivity  test  was done against  isolated  strains of  Campylobacter  using

a panel of 10 antibiotics ( Table ,3 and 4 ) . The most effective antibiotics for all isolates of Campylobacter jejuni and Campylobacter coli were Chloramphenicol . This support the data reported by Diker et al . ( 1990 ) The isolates of Campylobacter species at this study were resistant to Tetracycline  Penicillin and Sulphamethoxazole . These findings  agree with that  recorded  by

Erdger and Diker ( 1995 ) .

 In conclusion , due to the presence of Campylobacter in the small intestine of sheep   represent         a public health hazard . Moreover , because of  multidrug resistance among Campylobacters

hygienic measures should be undertaken such as cleaning and disinfection of animal houses , avoidance  of  improper use of antibiotics , hygienic disposal of animal excreta and proper cooking and hygienic handling of meat befor consumption .

Table ( 1 ) : Incidence of C . jejuni and C.coli in rectal swab samples collected from diarrheic sheep.

Table ( 2 ) : Biochemical reactions of Campylobacter jejuni and Campylobacter coli isolated from sheep suffering from diarrhea .

Table (3) : Antimicrobial sensitivity of Campylobacter jejuni isolated from sheep ( n = 5 )

Table (4) : Antimicrobial sensitivity of Campylobacter coli isolated from sheep ( n = 5 )

Refrences

Acha , P. N. and Szyfers, B. (1991 ) :

Zoonoses and Communicable Diseases Common to Man and Animals . 2^nd Ed . Pan American Health Organization , Washington D. C.

Adesiyun , A. A. ; Kaminojolo , J. S. ; Loregnard , R. and Kitson , P. W. ( 1992 ) :

Campylobacter infection in calves , piglets , lambs and kids in Trenedad . Br. Vet. J., 148 (6):547-556 .

Atabay , H. I. and Corry, J. E. (1997) :

The prevalence of Campylobacters and Arcobacters in broiler chickens . J. App. Miicrobiol. , 83(5):619-626 .

Betty, A. ; Dniel , F. and Alice , S. W. (1998):

Bailey and scott's Diagnostic microbiology . 10^th Ed., Mosby .

Bopp , F. L. Bikness, K. A. ; Washsmuth, I. K. and Barrett , T. J. (1985):

In vetro antimicrobial susceptibility , plasmid analysis and serology of epidemic Campylobacter jejuni . J. Clin. Microbiol . , 21 (1) : 4-7 .

Busato , A. ; Hofer, D.; Lentze, T. ; Gaillard, C. and Burnens , A. (1999) :          Prevalence and infection risk of zoonotic enteropathogenic bacteria in swiss cow – calf   farms .     Vet . Microbiol . , 69: 251-263 .

Diker , K. S (1987):

Isolation of  Campylobacter species from various animals and evaluation  of zoonotic  species

Microbiol . Bull . , 21(4):957 – 962 .

Diker , K. S.  Diker, S. and Ozlem , M. B. (1990)

Bovine diarrhea associated with Campylobacter hyointestinalis . Zentralbl Veterinarmed,       B, Mar . ; 37 (2) : 158-160 .

Erdger, J. and Diker , K. S. (1995) :

Multiple antibiotic resistance in poultry isolates of Campylobacter . Veterinar Faku  Itesi Dergisi , Ankara universitesi , Ankara , Turkey .

Giacoboni , G. L. ; Itoh , K. ; Hirayama K.; Takahashi, E . and Mitsuoka , T. (1993) :

Comparison of  faecal Campylobacter in calves and  cattle  of  different  ages and  areas  in Japan .  J. vet . Med . sci . , 55 (4): 555 – 559 .

Gur , D. ; Harscelik , G., Akyon , Y. ; Akalin , H. E. and Diker , S. (1989) :                            In vetro susceptibility of Campylobacter coli to quinolone antibiotics . Mikrobiyol. Bul. , 23 (3) : 185-189 .

Hanninen , M. L.  ; perk, O.  Makela , P. ; pitkala , A. and Rautelin , H. (2000) :

A three year study of Campylobacter jejuni genotypes in humans with domestically acquired infections and in chicken samples from the Helsinki area . J. clin . Microbiol., 38 (5) :      1998-2000  .

Hartmut , K. ; Albert , W. ; Max , A., Burkhard , E. ; Henry , D. ; HansGerd, S. ; Werner , S. ; Alexander , V. and Horst , Z ., ( 2003 ) :

Zoonoses , Infectious Diseases Transmissible from Animals to Humans . 3^rd Ed. American Society for Mirobiology , Washington D. C.

Jensen , L. B. and Aarestrup , F. M. (2001) :

Macrolid resistance in Campylobacter coli of animals origin in Denmark . Antimicrob . agents chemother . , 45 : 371-372 .

Kakkar , M . and Dogra , S. C. (1990) :

Prevalence of Campylobacter infection in animals and children in Haryana , India .

J . Diarrhoeal Dis . Res. , 8 ( 1-2 )  34-36 .

Khalil , k.; Lindblom , G. B. ; Mazhar , k. ; Sjogren, E . and Kaijser, B. (1993) :

Frequency and enterotoxicity of Campylobacter jejuni and Campylibacter coli in domestic animals in Pakistan as compared to Sweden .J. Trop. Med. Hyg., 96(1) : 35-40 .

Koneman , E. W. ; Allen , S. D. ; Dowell, V. R. ;Janda , W. M. ; Sommers, H. M. and Winn , W. C. W. C. (1988) :

Colour Atlas and Textbook of Diagnostic Microbiology . 3^rd Ed. T. B. Lippencott, NewYork , London .

Manser , P. A. and Dalziel , R. W. ( 1985 ) :                                                                           A survey of Campylobacter in animals . J. Hyg ., 95 (1) : 15-21 .

Oostrom , J. ; Uyl, C. H. Den ; Banfter, J. R. J. and Huisman , J. (1984) :

Epidemiological investigations on Campylobacter jejuni in households with a primary infection . J. Hyg. Camb . 92  325-332 .

Quinin , P. T. ; Markey , B. K. ; Carter , M. E. ; Donnelly , W. J. and Leonard , F. C. (2002) :

Veterinary Microbiology and Microbial Diseases . Blackwell science L T D .

Radostits , O. M. ; Gay , C. C. ; Blood , D. C. ; Hinchcliff, K. W. and Arundel , J. H. (2000):

Diseases caused by bacteria , P. 967. In veterinary medicine , 9^th E.d. The W.B. sunders Co., Philadelphia .

Retting , P. J. (1979) :

Campylobacter infections in human being J. Ped., 94 (6) : 855-864 .

Rosef , O ; Gondrosen , B.  ; Kapperud , G  and Underdal , B. (1983) :

Isolation and characterization of Campylobacter jejuni and Campylobacter coli from domestic and wild mammals in Norway . Appl. Envirnm. Microbiol. , 46 (4) : 855-859 .

Saenz , Y.; Zarazoga , M.; Lantero , M. ; Jose  Gastanares  F ;  Baquero ,  M.   and     Torres , C. (2000) :

Antibiotic resistance in Campylobacter strains isolated from animals , foods , and humans in spain in 1997-1998 . Antimicrob . Agents Chemother . 44 : 267-271 .

Shakespeare , M. (2002) :

Zoonoses . Pharmaceutical  Press . NewYork , pp 248-250 .

 Skirrow , M.B. (1977) :

Campylobacter enteritis . "A new " disease . Br . Med . J. ; 2: 9-11 .

Stanley , K . N. ; Wallace , J. S. ; Currie , J. E. ; Diggle , P. J. and Jones , K . (1998):

The seasonal variation of thermophilic Campylobacter in beef  cattle , dairy cattle  and calves

J. Appl. Microbiol . , 85 (3) : 472-480 .

Terzolo , H. R. (1988) : Identification of Campylobacter from bovine and ovine and ovine feces . Rev. Argent . Microbiol . , 20 (2) : 53-68 .

Turkson , P. K. ; Lindquist , K. T. and Kapperude , G. (1988) :

Isolation of Campylobacter species and Yarsinia enterocolitica from domestic animals and human patients in Kenya . APMIS , 96(2) :141-146 .

الملخص العربى

الإسهال المعوى فى الأغنام المتسبب نتيجة العدوى ببکتريا الکامبيلوباکتر فى محافظة الأسکندرية

د/ علا عبد العزيز باشا و د/فتح الله على الشابورى

معهد بحوث صحة الحيوان

فرع الأسکندرية – قسم البکتريولوجى

إشتملت الدراسة على فحص عدد 100 مسحة شرجية من الأغنام المصابة بالإسهال المعوى لدراسة مدى الإصابة بعدوى

ميکروب الکامبيلوباکتر . و قد أسفر الفحص البکتريولوجى عن عزل ميکروب الکامبيلوباکتر جوجينى من 12 حيوان بينما

تم عزل  بکتريا الکامبيلوباکتر کولاى من 9 حيوانات مصابة فقط . ودلت دراسة تأثير المضادات الحيوية على معزولات

الکامبيلوباکتر الجوجينى و الکولاى أن أفضلها فاعلية کان الکتورامفنيکول يليه النيومايسين و الأمبيسيلين و الجنتاميسين .

بينما  کانت  معظم  المعزولات   مقاومة للتتراسيکلين و البنسلين و السلفاميزوکسازول . و تتلخص هذه  الدراسة على  أن

الکامبيلوباکتر تلعب دورا فى الإصابة بالإسهال المعوى فى الأغنام . هذا و قد تم مناقشة الأهميه الصحية العامة لميکروبات

الکامبيلوباکتر المعزولة و طرق الوقاية منهما .