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3Clinicopathological Manifestations of Pasteurella Multocida (Serotypes A: 1, 3 And 4) Infections in Commercial Chickens
in Jos, Nigeria
Dashe Yakubu1*, Raji Moshood2, Abdu Paul3, Oladele Blessing4, Okewole Philip5, Kumbish Peterside5,
Oluwadare Lola1 and Barde Israel5
1National Veterinary Research Institute, Akure, Zonal Office, Hospital Road, Akure, Ondo State, Nigeria. 2Department of Veterinary Microbiology, Ahmadu Bello University, Zaria, Nigeria. 3Department of Veterinary Medicine, Ahmadu Bello University, Zaria, Nigeria. 4Deprtment of Veterinary Pathology, Ahmadu Bello University, Zaria, Nigeria. 5Central Diagnostic Department, National Veterinary Research Institute, Vom Jos, Nigeria. *Corresponding author's Email: [email protected]
Received: 07 Sep. 2015 Accepted: 20 Oct. 2015
ABSTRACT
This pathogenicity study was conducted to document the clinicopathologic features observed in commercial chickens inoculated with Pasteurella multocida serotypes A: 1, 3 and 4. Thirty, eighteen weeks old (adult) commercial chickens were divided into five groups (A, B, C, D and E) of 6 birds each. Chickens in groups A, B and C were inoculated with 0.1 ml of Pasteurella multocida serotypes A: 1, 3 and 4 at concentrations of 106, 107 and 108 CFU/chicken respectively using intramuscular route. Group D were inoculated with 0.1 ml fowl cholera vaccine strain of P. multocida serotype A: 1 at of concentrations of 106, 107 and 108 CFU/chicken, while group E the uninfected control chickens were given normal saline. All deaths in groups A and B occurred on day 7 and mortality rates were 83.3% for group A and 50% for B. No mortality was recorded in groups C, D (vaccine strain) and E (uninfected control). Gross lesions observed were petechial and ecchymotic haemmorhages on the heart and breast muscles, congestion of the liver and lungs. Histopathological lesions observed were mononuclear cellular infiltration and pulmonary congestion. This study has shown that chickens were susceptible to both Pasteurella multocida serotypes A: 1 and 3.
Key words: Chicken, Pathogenic, Jos, Pasteurella, Serotypes
JWPR
Journal of World's Poultry Research
© 2015, Scienceline Publication
J. World's Poult. Res. 5(4): 98-103, December 25, 2015
Research Paper PII: S2322455X1500014-5
INTRODUCTION
Poultry has been declared the largest group among domesticated livestock in Nigeria with an estimated population of about 140 million (FAO, 2000). Poultry offers the quickest supply of animal protein to man, and provides comparatively greater financial profit than cattle, small ruminants or pigs (Etukudo and Adegboye, 1983; Molokwu et al., 1988). The rural poultry farming contributes 68.5% of the national meat supply; while 11.4% is from commercial chickens and 19.7% from other poultry (Wosu, 1992; Musa et al., 2009).
Despite the unquantifiable economic gains from this industry, it often suffers from disease induced losses due to sustained challenges from environmental factors, viral, bacterial, protozoan, parasitic and fungal infections among others. In Nigeria, a great loss in poultry production is attributed to these agents and it still remains as one of the major constraints hindering the success and growth of the poultry industry, where disease prevention and control measures are rare and high mortality rates are common even in vaccinated flocks in some cases. The epizootic of fowl cholera in the world poses a serious threat to poultry (Pandey,
1992; Ambali et al., 2003). The aetiologic agent of fowl cholera, Pasteurella multocida is not host specific and has demonstrated the capacity to infect and cause severe illness and death in livestock. Fowl cholera has often occurred as a sporadic disease in many parts of the world including Nigeria with grave economic consequences on livestock and humans (Ambali, 2003). Fowl cholera is widely identified as one of the major diseases of economic importance wherever intensive poultry production occurs; particularly in the United States of America (Rimler et al., 1998). However, the disease has a significant impact in less intensive systems. For example, fowl cholera was one of the main diseases seen in Mexican village chickens (Gueye, 1999). Fowl cholera was endemic in regions of Asia and Africa where it caused immense economic losses to the poultry industry due to cost of treatment, weight loss and mortality (Townsend et al., 1997). Despite the established role of P. multocida as an etiologic agent of fowl cholera in chickens in Jos, Nigeria, there is inadequate information on the clinical, gross pathologic and histopathologic features of the disease caused by P.
multocida serotypes. This study therefore seeks to document the clinicopathologic features observed in chickens inoculated with P.multocida serotypes A: 1, 3 and 4.
MATERIAL AND METHODS
Management and housing of experimental chicken
Thirty, 18 weeks old (adult) commercial chicken were provided by the National Veterinary Research institute, Vom, Nigeria for this experiment. The chickens were divided into five groups (A, B, C, D and E) of 6 chickens each. Groups A, B, C and D were further sub-divided into three sub-groups, with each group consisting of two chickens. One week before the onset of the study, chickens were screened for the presence of P. multocida by taking oro-pharyngeal swabs. All chickens were fed with commercial pelleted layer feed and water ad libitum before and during the experiment. The house with floor space of the pen was 4 x 5 m2 measurement in size was cleaned disinfected prior to the commencement of experiment. Chickens were allowed to acclimatize for two weeks prior to the commencement of the pathogenicity study.
Source of Pasteurella multocida
The bacterium P. multocida serotypes A: 1, 3 and 4 were recovered by the authors from chickens with clinical cases of fowl cholera in Jos, Nigeria in 2012. The isolates were confirmed by biochemical test, Microbact, Polymerase Chain Reaction and Multiplex PCR. They were sorotyped at United State Department of Agriculture in USA.
Inoculation of Chickens
All groups (A, B, C, D and E) were used for the pathogenicity test. Pasteurella multocida serotypes were reconstituted into three different concentrations for groups A, B and C; every chicken in first sub-group of A was administered 0.1 ml intramuscularly of 106 CFU (serotype A:1- field strain) contained in Typtose Soya broth, the second sub-group received 107 CFU of the same serotypes and the last sub-group received 108 CFU of P. multocida serotype A: 1, the administration of serotype A: 3 and A: 4 of P. multocida at the same doses were replicated for sub-groups B and C respectively. The chickens in group D were inoculated with P. multocida serotype A: 1 (fowl cholera vaccine strain) at the same concentrations of 106, 107 and 108 CFU. Chickens in group E (uninfected control) received normal saline at 0.1 ml intramuscularly. Death occurred within the period of twenty one days of the experiment was recorded. Findings from clinical, postmortem examination of carcasses and histopathological features were recorded.
Clinical and pathological examinations
Chickens in all the groups were observed for clinical signs and mortality post inoculation. Postmortem examinations were conducted on chickens that died and tissue section of the spleen, liver, heart and lungs were prepared for histology. The section of
grossly affected organs were fixed in 10% buffered formalin; paraffin embedded tissues were sectioned at 5 microns, stained with haematoxylin and eosin (H & E) and mounted on glass slides and examined under light microscope at X200 and X400 respectively.
Statistical Analysis
The entry and sorting of the primary data was performed with Microsoft excel, 2010. Descriptive statistical analysis was conducted using statistical package for social sciences IBM SPSS (version 21.0) (2012) and the results were summarized as percentages in tables.
RESULTS
Ninety two percent of the experimental chickens in groups A and B became sick within 24 h post inoculation. Clinical signs manifested were depression, inappetence, ruffled feathers, dyspnoea, sitting on hock and these signs continued for four days. All deaths in groups A and B occurred on day 7 and mortality rates were 83.3% for group A and 50% for B. No clinical sign or mortality was recorded in groups D (vaccine strain), C and E (uninfected control) (Table 1). Lesions observed at postmortem examinations in chickens inoculated with P. multocida serotypes A: 1 and A: 3 were prominent keel with congested heart, liver, kidneys and lungs (Table 2 and Figure 1). Frothy exudates from the lungs, petechial and ecchymotic haemorrhages were seen on the heart, subserosal haemorrhages (Figure 2). Lymphocytic, heterophilic and macrophagic cellular infiltration in the lungs and heart were observed (Figure 3).
DISCUSSION
Pasteurella multocida is one of the bacterial agents responsible for significant economic losses in the poultry industry worldwide. The report of this study indicated that only serotypes A: 1 and A: 3 caused clinical disease in chickens with high morbidity and mortality. Interestingly, these serotypes also demonstrated various degrees of gross and histopathological lesions in the affected chickens. In a similar study conducted in India by Kumar et al. (2004), opined that majority of P. multocida belonging to serotypes A: 1, 3 and 4 were associated with fowl cholera in chickens. The findings in this study differ from that of Kumar and others in that serotype A: 4 in this study did not cause mortality in chickens. Similarly, Snipes et al. (1988) also asserted that not all P. multocida isolates are pathogenic to chickens. Previous study conducted on fowl cholera by Glisson et al. (2003) indicated that P. multocida serotypes vary from one country to the other. Another report of a study from Central Saudi Arabia, by Elfak et al. (2002) reported the involvement of P. multocida s erotypes 1, 3, 4 and 5 in an outbreak of fowl cholera in ostriches. The serious implication of these reports is that the production of universal fowl cholera vaccine would be difficult considering the fact that P. multocida serotypes vary from one country to the other and immunity is
known to be serotype specific. The congested liver, spleen, lungs as well as petechial and ecchymotic haemorrhages that were observed at postmortem in the experimentally challenged chickens, also confirmed the findings of Visut et al. (2010). The result of this pathogenicity study indicated that all the experimental chickens inoculated with serotypes A: 1 and A: 3 came down with the clinical signs of fowl cholera similar to those reported by Rimler and Glisson (1997). This has buttressed the fact that adult chickens above 16 weeks of age were susceptible to fowl cholera. Host variation in the susceptibility to P. multocida serotypes was observed as different clinical signs and gross lesions manifested in the chickens inoculated by the same pathogenic agent. These findings collaborated the findings of Botzler (1991) who also observed that clinical signs of fowl cholera in birds vary depending
on various factors such as age, species, dose of strain, route of entry of the bacterium and form of the disease.
Notable histopathological findings in this study indicated a moderate to severe lymphocytic, heterophilic and macrophagic cellular infiltration in lungs and heart. These findings confirmed report of Shilpa and Verma (2006). The findings could possibly signify that the chickens were responding to the inciting bacterial agent, which is a common characteristic in acute phase of fowl cholera. The severe involvement of visceral organs such as congestion noticed in lungs, heart and spleen of chickens clearly demonstrate the septicaemic nature of fowl cholera; this could possibly explain the profound debilitation, massive flock morbidity and mortality observed in chickens suffering from the acute form during outbreaks of fowl cholera.
_Table 1. Mortality of chickens after inoculated with P. multocida serotypes A: 1 A: 3 and A: 4
Groups P. multocida serotypes Mortality
2 at 108 CFU
A A: 1 (Field Strain) 2 at 107 CFU
1 at 106 CFU
2 at 108 CFU
B A: 3 1 at 107 CFU
0 at 106 CFU 0 at 108 CFU
C A: 4 0 at 107 CFU
0 at 106 CFU 0 at 108 CFU
D A: 1 (Fowl Cholera Vaccine Strain) 0 at 107 CFU
0 at 106 CFU 0 at 106 CFU
E Control (Normal Saline) 0 at 106 CFU
0 at 106 CFU
Table 2. Gross lesions in 18 weeks old chickens inoculated with Pasteurella multocida serotypes A: 1, A: 3 and A: 4
Gross lesions Group A Group B Group C Group D Group E
Congested liver 6 (100%) 6 (100%) - - -
Congested heart 6 (100%) 6 (100%) - - -
Congested kidneys 6 (100%) 6 (100%) - - -
Congested lungs 6 (100%) 6 (100%) - - -
Frothy exudates from lung 3 (50%) 0 (0%) - - -
Pet. and Ecc. hae on heart 6 (100%) 6 (100%) - - -
Fatty degeneration 1 (16.7%) 0 (0%) - - -
Egg yolk peritonitis 3 (50%) 3 (50%) - - -
Prominent keel 6 (100%) 6 (100%) - - -
Pet: Petechial, Ecc: Ecchymotic, Hae: Haemorrhages
To cite this paper: Yakubu D, Moshood R, Paul A, Blessing O, Philip O, Peterside K, Lola O and Israel B. 2015. Clinicopathological Manifestations of Pasteurella
Figure 1. A - Congested liver in a chicken aged 18 weeks inoculated with Pasteurella multocida serotypes A: 3, in Jos,
Nigeria
Figure 2. A - Haemorrhages in subserosal layer of sternal bone of a chicken aged 18 weeks inoculated with Pasteurella
multocida serotypes A: 1 in Jos, Nigeria.
To cite this paper:
Figure 3. A: The lung of a chicken aged 18 weeks infected with Pasteurella multocida A:3=108. Congested blood vessels (small arrow), areas with obliterated alveoli and infiltrated with inflammatory cells (big arrow) H and E Stain. X
200; B: normal lung of a chicken aged 18 weeks at X 400.
CONCLUSSION
The findings of present study have revealed that chickens were highly susceptible to infections with P. multocida serotypes A: 1 and 3. These two serotypes of P. multocida were found to cause high morbidity, mortality, clinical, gross and histopathologic features in the experimental chickens.
Recommendations
It is therefore recommended that chickens should be vaccinated against fowl cholera with a polyvalent fowl cholera vaccine consisting of P. multocida serotypes A: 1 and 3 in order to protect them against P. multocida infections.
Acknowledgement
The authors wish to express their profound gratitude to the staff of Central Diagnostic Department, National Veterinary Research Institute, Vom, Nigeria
for assisting in carrying out post mortem examination on the carcasses.
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