Dept. of Bacteriology,

Animal Health Research Institute,

Mansoura Provincial Laboratory.

BACTERIOLOGICAL STUDY ON SOME CAUSES OF

EARLY MORTALITY OF DUCKLINGS IN DAKAHLIA GOVERNORATE

(With 3 Tables)

By

M. El-Sayed Hatab; A.H. Moustafa

and M.M. Abd El-Latif

Received at 9/2/2005))

دراسة بکتريولوجية عن بعض مسببات النفوق المبکر فى البط

فى محافظة الدقهلية

محمد السيد محمد حطب ، عادل حسانين محمود مصطفى

 محمود محمد محمود عبد اللطيف

أجريت هذه الدراسة لمعرفة أسباب النفوق المبکر فى البط. تم جمع 100 عينة من البط عمر (1-21 يوم) وأنواع مختلفة من عدة مزارع خاصة والعينات التى تصل إلى المعمل الفرعى بالدقهلية وکانت کلها تعانى من النفوق المبکر. تم فحص هذه العينات بکتريولوجيا وذلک بزرعها على أوساط مختلفة للبکتريا وقد أمکن عزل 144 عترة بکتيرية تم تصنيفها بالطرق المورفولوجية والبيوکيميائية إلى 35 (24.30%) سيدوموناس أريجينوزا ، 25 (17.37%) إيشيريشيا کولاى، 20 (13.89%) لکل من السالمونيلاً والإستافيلوکوکس أوريس،        16 (11.11%) کليبسيلانيمونى ، 15(10.42%) رايمريلاأناتيبتفر وبروتيس فالجاريس  13 (9.02%). تم تصنيف معزولات الإيشيريشيا کولاى سيريولوجيا  إلى 7 عترات              O₇₈K₈₀(B-)) و6 (O₈₆K₆₁(B7)) و5 (O₁₂₅K₇₀(B15)) و7 (untypable). کذلک تم تصنيف عترات السالمونيلا سيريولوجيا إلى 12 عترة سالمونيلاً تيفيميورييم و8 عترات untypable. وقد أسفرت نتائج إختبارات حساسية المضادات الحيوية عن أن السيبروفلوکساسين والإنروفلوکساسين والجنتاميسين کانت أکثر المضادات الحيوية تأثيراً على معظم المعزولات.

Summary

This study was conducted toward a problem of extend early mortality of ducklings. A total of 100 freshely dead and clinically sick ducklings of different ages (1-21 days) and types obtained from private farms at Dakahlia Governorate and also cases which arrive to the Mansoura lab. The samples were dispatched to the laboratory to be examined bacteriologically for detection of the actual bacterial causes of early mortality problem in these farms. The obtained results pointed out that a total of 144 isolates were isolated. All the bacterial isolates were identified morphologically, culturally, biochemically and serologically for E. coli and Salmonella microorganisms. Pseudomonas aeruginosa was the most prevalent bacterial agent 35 (24.30%) followed by E. coli 25 (17.37) , Salmonella and Staphylococcus aureus 20 (13.89%) for each, Klebsiella pneumoniae 16 (11.11%), Riemerella anatipestifer 15 (10.42%) and Proteus vulgaris 13 (9.02%). The isolated E. coli were identified serologically into 7(O₇₈K₈₀(B-), 6(O₈₆K₆₁(B7), 5(O₁₂₅K₇₀(B15) and 7 (untypable), while the recovered Salmonella strains were identified into 12 Salmonella typhimurium and 8 (untypable). In-vitro-sensitivity pattern of isolated strains proved that ciprofloxacin, enrofloxacin, gentamycin  were the most effective drugs.

Key words: Mortality, ducklings.

Introduction

Nowadays, a great attention was payed toward ducks farming as a trial to fulfill excessive demand of the increased population from the animal protein. The ducks meat are considered to be of high protein content with high biological value.

Several microbial infections are responsible for the early mortality of ducklings and losses of duck industry. Many isolated microorganisms were associated with duck mortality as: Salmonella, E.coli, Pseudomonas, Pasteurella, Proteus, Klebsiella and Staphylococci (Bhowmik & Ray, 1987; EL-Gharib et al., 1993 and Istania, 1993). Antibiotics are used therapeutically and for prophylaxis in intensive domestic birds farming. However of bacteria resistant to antibiotics emerge even under control use of antibiotics (Helm et al. 1999). This study was done to investigate the possible bacterial causes of the  problem.

Materials and Methods

     A- Samples

A total of 100 freshly dead and clinical sick ducklings of different ages (1-21 days) and breeds were obtained from private farms and also cases which arrive Mansoura Laboratory. The Samples were dispatched to the laboratory without delay to be examined bacteriologically for isolation and identification of microorganisms agents. 

      B- Clinical and PM examination:

Ailing ducklings were examined clinically, then sacrified and immersed in a disinfectant before being autopsied. Gross pathological changes were recorded, summarized and presented with results for both freshely dead & clinically sick ducklings.

     C- Media:

- Nutrient agar media (Oxoid CM3).

- Blood agar media (Nutrient agar base Oxoide CM3 + 5-10%     defibrinated sheep blood).

- Eosin methylene blue (Oxoid CM69).

- MacConkey agar (Oxoid CM7).

- Rappaport vassiliadis broth (Oxoid CM 669).

- Xylose Lysine desoxycholate agar (Oxoid CM 469).

   D- Serological identification of Salmonella and E. coli:

Samonella and E. coli isolates were identifed according to (Edward and E wing, 1972) 

   E- Bacteriological examination:

Bacteriological samples were collected aseptically from the deep tissue of liver, lung, bone marrow, heart blood and brain. Each specimens was divided into 3 portions under aseptic condition. The first part was streaked onto predried surface of Blood agar, Nutrient agar, MacConkey agar (Oxoid, CM7) and Eosin methylene blue (EMB) (Oxoid, CM69), incubated aerobic at 37^oC for 24 hours. The second part was  inoculated into Rappaport Vassiliadis broth (RV) ( Oxoid, CM 669) incubated at 42^oc, after 24 hours incubation, loopfules from R.V. enrichement were streaked onto Xylose lysine desoxycholate  agar plate ( XLD) (Oxoid, CM 469) with incubation at 37^oc for 24 hours. The third part was streaked over blood agar plate containing 0.05% yeast extract, 5% newborn calf serum and 5 mg/100ml gentamycine, incubated in a candle Jar at 37^oc for 24-48 hours.

The growing colonies on various plates were examined morphologically and biochemically tests as described by Baily & Scott, (1974); Cruickshank et al. (1975) and Carter (1984). 

            The identified E. coli strains were tested for enterotoxin production through grown the E. coli isolate in trypticase soya broth at 37^oc in stationary culture overnight. Culture was centrifuged at 4000 rev/min. for 20 minutes. The supernatant was tested using commercially VET-RPLA kits (reversed passive latex agglutination) from Oxoid (TD 0920 A) following the manufacturer's direction.

The biochemically identified E. coli and Salmonella isolates were subjected for serological identification using avaliabe E. coli test agglutinating sera (BioMerieux, 1986) and diagnostic Salmonella agglutinating antisera (Denka Selken Co. LTD, Tokyo, Japan) according to manufacturer's  instruction.

In vitro antibiotic sensitivity test:

The disc diffusion technique was performed using Muller-Hinton medium (BioMerieux, France) on isolated bacteria from examined samples according to National Committee for Clinical Laboratory Standards (1984) and Quinn et al (1994).

Results

The clinical signs and PM lesions: The recorded clinical signs were diarrhea, ataxia, tremor of head and neck, coma, affected ducklings lie on their back, paddling their legs, septicemia. The P.M. examination of duckling revealed the presence of congestion in the internal organs         (liver, spleen, intestine) and enlarged gall bladder.  

The results of bacteriological examination were recorded in Table 1, 2 and 3.

Discussion

Various clinical signs were recorded, diarrhea, ataxia, tremor of head & neck, coma, affected ducklings lie on their backs, paddling their legs, septicemia. Bacterial infection of the examined ducklings is one of the main causes of early mortality and the present study deals with the pathogenic bacteria responsible for early mortality of ducklings. Pseudomonas aeruginosa, E.coli, salmonella, Riemerella anatipestifer, Kilebsiella pneumoniae, Proteus vulgaris and Staphylococcus aureus were isolated in percentage of 24.3, 17.37, 13.89, 10.42, 11.11, 9.02 and 13.89 respectively. Similar findings were investigated by (El-Gharib et al. 1993 and  Istania, 1993).

Pseudomonas aeruginosa was the most prevalent bacterial agent 35 (24.3%). Pseudomonas infection of birds are of great important  because epidemics may spread rapidly through flocks and is considered as one of the most dangerous disease, it causes morbidity and mortality and the clinical signs including septicemia, diarrhea and respiratory signs (Soumaya, 1992 and Tanois & Samia, 1999).

Collibacillosis is a common systemic disease of world wide economic importance in birds, the major clinical signs of E.coli infection in young birds, acute septicemia that can cause sudden death (Leithnes and Heller, 1992).

 In this study, all the isolated 25 (17.37%) E. coli strains from examined duckling samples were enterotoxigenic produce heat labile enterotoxin when tested by VET-RPLA kits, and serologically identified as 28% E. coli O₇₈ K₈₀ (B-), 24% E. coli O₈₆ K₆₁ (B₇), 20% E. coli O₁₂₅ K₇₀ (B₁₅) and 28% untypable strains (Table 2).

The results in Table (1) pointed out that salmonella species were isolated from 20 (13.89%). This finding as nearly agree with the observations of El-Gharib et al. (1993). and Scott et al. (1984). The isolated Salmonella strains were serotyped as 12 (60%) Salmonella typhimurium and 8 (40%) untypable strains (Table2). Salmonella typhimurium had been reported as the most common species isolated from examined ducks (50%) Scott et al. (1984).

Generally, Riemerella anatipestifer is a Gram-negative, non-motile non sporforming rod, oxidase and catalase positive, not grow on MacConkey, negative indol and not hemolysis blood agar. R. anatipestifer is a contagious disease of domestic duck, it is known as duck septicemia, characterized by fibrinous pericarditis, perihepatitis, air saculitis, caseous salpingitis and meningitis (Sandhu, 1986). Results in Table (1) showed that Riemerella anatipestifer were isolated in 15 (10.42%), our results nearly similar with Ziedler et al. (1984), reported high mortality in the fattening flocks of ducks occurred due to pasteurella anatipestifer outbreaks. Regarding the isolation of Klebsiella pneumoniae Proteus vulgaris and Staphylococcus aureus,they were isolated from examined duckling samples at the incidence rate 11.11, 9.02 and 13.89% respectively. This finding similar with (El-Gharib et al. 1993 and Istania, 1993). The high mortality rate in duckling might be attributed to septicemic shock to the toxic effect associated with lipopolysaccharide fraction of the Proteus organisms (Wilison & Miles, 1975).

In vitro, the susceptibility distribution of each isolated pathogen to different antibiotics is presented in Table (3). The typical pattern of highly effective compounds were observed for zones of ciprofloxacin, enrofloxacin and gentamycin. These findings corresponded with those reported by (Khodary & El-Sayed, 1997; Turbahn et al. 1997 and Rahman et al., 1999).

In conclusion, the information given by the achieved results revealed that several microorganisms were inciriminated in the early mortality of the ducks and therefore strict hygienic measurements should be applied on the egg laying, hatcheries, and good management during the production.

References

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Table 1: The incidence of different bacterial isolates recovered from the examined samples.

* The percentage was calculated according to the total isolates (144)

Table 2: Serological identification of isolated E. coli and Salmonella  strains .

[*] All strains are LT toxin