Experimental inoculation of Salmonella Minnessota in broilers
Published: October 20, 2011
By: LB Miglino, L Pickler, LN Kuritza, MC Lourenço, E Muniz, AL Lago, E Santin - Laboratorio de Microbiología y Ornitopatología, Universidad Federal de Paraná, Curitiba, Brasil.
Summary
Bacteria of the genus Salmonella have been known for their pathogenicity. They may cause severe economic losses for poultry industry and damages to public health. The most common serotype has been S. Enteritidis, however, different serotypes have been isolated, such as S. Minnesota (SM). The poultry industry has focused efforts on reducing Salmonella incidence in live production in an effort to reduce Salmonella in the processing plant. A better understanding of the initial infection in chicks could provide approaches to control Salmonella contamination. The purpose of this study was to evaluate the pathogeny of SM in different organs at various periods after inoculation. The results showed an increased velocity in SM organ colonization in the inoculated animals; as more hours passed by, a larger number of broilers indicated the presence of SM; and healthy broiler chickens which had contact with inoculated chickens for 24 hours presented SM at the upper fraction of their gastrointestinal tract. Key Words: Epidemiology, Salmonelosis, Microbiological research.
Introduction
The world's poultry industry has grown considerably during the last years as well as the concern for the safety of the food for human consumption derived from the poultry chain. That is why the production countries create stringent legislations, mainly against the presence of agents with zoonotic potential, as for instance bacteria from the Salmonella genus.
Birds are considered the main reservoirs of these bacteria, being asymptomatic carriers that, according to Morse and Duncan (1974) are important for the infection in humans and other animals, in a direct and/or indirect form. Salmonella has the main responsibility in the cases of food intoxication in humans, associated with the consumption of poultry products (Silva, 1999). Besides, the damages related with the health of the birds are significant, due to the morbidity and mortality that this agent causes in the flocks (Ferreira, 1994).
Actually, salmonellas of the paratyphoid type stand out by the high frequency with which they have appeared (Berchieri, 2000). In this group, S. Enteritidis (SE) and S. Typhimurium (ST) are the most important, but according to Back (personal communication), the serum typification of the isolations from commercial birds since 2007 have presented a reduction in these serological varieties (serovars) while others have increased, as is the case of Salmonella Minnesota (SM), which suggests the need to carry out new studies.
Under these conditions, Brazil is a great world exporter of chicken meat who is trying to adequate itself to the measures that reduce the risk of contamination of poultry products by Salmonella. This way it is extremely important to know the pathogenesis of different serovars to establish their eradication and control programs. With this problematic in mind, the objective of this study was to evaluate the propagation of SM in the crop, the duodenum, the cecum and the liver during different periods of experimental post inoculation (PI), with groups of chicken with these bacteria, through mouth catheter, through contaminated feed, or by contact with infected birds.
Material and Methods
Sixty five broilers were used under an experimental design completely at random, with four treatments of 15 birds each, considering each bird as a repetition. In treatment T1 the birds were inoculated through an oral catheter, directly into the esophagus, with a solution of 105 colony forming units (UFC)/ml of SM. The birds in treatment T2 were fed with a ration contaminated with 105 UFC/ml of SM. The 15 animals from group T3 were identified with rings and after 24 hours of PI were transferred to the room of the T1 group to keep them in contact with the ones infected with SM. Birds from treatment T4 were not inoculated, considering this as the negative control group.
The birds were housed in disinfected rooms, with negative pressure, separate one from the other, and with controlled temperature and humidity. Water and feed were offered ad libitum, following the recommendations from the (1994) United States National Research Center (NRC). Before the inoculation, five animals were slaughtered and necropsies were performed to collect samples and to do the negativity test for the presence of Salmonella.
Table 1. Experimental design completely at random
|
Treatments
|
Challenge
|
|
T1
|
Oral inoculation with SM
|
|
T2
|
Contaminated feed with SM
|
|
T3
|
Birds in contact with T1 24 h PI
|
|
T4
|
Control not inoculated
|
At 12, 24 and 48 hours after the inoculation, five birds from each treatment were slaughtered and necropsies were performed to aseptically collect samples from the crop, the duodenum, the cecum and liver, to be tested microbiologically and trying to detect the presence or absence of SM (method adapted from Desmidt et al. 1998). The results on the presence or absence of SM were submitted to statistical analysis of chi-square (P<0.05).
Results and Discussion
Birds did not present infection with Salmonella sp. before the experimental period, which was obvious through the negativity test done when the experiment started. The results obtained through the microbiological investigation of SM in the different organs, from each group, are shown in Table 2.
Table 2. Results in the number of birds positive to Salmonella in the different organs, whose samples were obtained after the inoculation
|
Trat.
|
12h PI
|
24h PI
|
48h PI
|
|||||||||
|
Crop
|
Duod.
|
Cecum
|
Liver
|
Crop
|
Duod.
|
Cecum
|
Liver
|
Crop
|
Duod.
|
Cecum
|
Liver
|
|
|
T1
|
5/5ª
|
2/5ab
|
4/5ª
|
1/5
|
4/5ª
|
3/5ª
|
5/5ª
|
1/5
|
5/5ª
|
4/5ª
|
5/5ª
|
2/5ª
|
|
T2
|
4/5ª
|
4/5ª
|
3/5ª
|
0/5
|
5/5ª
|
4/5ª
|
2/5ab
|
1/5
|
4/5ª
|
3/5ª
|
5/5ª
|
3/5ª
|
|
T3
|
0/5b
|
0/5b
|
0/5b
|
0/5
|
0/5b
|
0/5b
|
0/5b
|
0/5
|
2/5ab
|
1/5ab
|
0/5b
|
0/5b
|
|
T4
|
0/5b
|
0/5b
|
0/5b
|
0/5
|
0/5b
|
0/5b
|
0/5b
|
0/5
|
0/5b
|
0/5b
|
0/5b
|
0/5b
|
a,b Different letters in the same column show differences statistically significant (P>0.05), according to the chi-square test..
Birds in the group not inoculated continued negative to the presence of SM during all the experimental period. The animals from the inoculated group through the mouth (VO) and those of the group infected with contaminated feed, presented after 12 hours PI, the presence of the bacteria in the crop, duodenum and cecum, and one bird in group T1 also presented SM in the liver. Desmidt et al (1996), studying the pathogenesis of SE (phagotype 4), inoculated through VO a solution containing 2 x 104 UFC/ml, in chickens one day old and only 3 hours PI he was able to isolate the bacterium from the crop and the gastrointestinal tract, and 12 hours PI the same was isolated from visceral organs, for example the liver. Nevertheless, Gorham et al (1991), when inoculating 7 day old birds with a solution containing 107 UFC/ml of SE (phagotype 13a), he was only able to isolate the bacterium from the liver and the gastrointestinal tract four days PI.
In this work, at 48 hours PI, 100% of the birds in groups T1 and T2 had the presence of Salmonella in the cecum. In the liver, the same was found in two or three birds from these groups, respectively. In the birds from group T3, kept in contact with those from group T1, during 24 hours PI, the presence of SM was observed in the crop and the duodenum 24 hours after the transference, showing that the chickens from group T1 were eliminating the bacterium to the environment, but its propagation by contact to the liver of the contaminated birds did not happen in this same period, which it was observed in the birds inoculated or that consumed the contaminated feed, this may suggest that the bacterial contamination is faster when the animals receive the inoculums VO ( orally) than when this happens by contact from bird to bird.
During all the experiment it was not possible to find neither clinical alterations nor macroscopic lesions in the birds. The Salmonella investigation did not allow the counting of UFC, this suggests that SM suffered interferences coming perhaps from the immune system of the birds or from other bacteria.
Conclusion
It is concluded that the groups of orally infected birds with SM, the propagation of this bacterium into the different organs tested was faster when this animals were compared with the healthy birds that were kept in contact with them during 24 hours.
Bibliography
Berchieri JA. 2000. Salmoneloses aviárias. pp. 185-195. In: Berchieri Júnior A & Macari M (Eds.). Doenças das aves. Facta, Campinas.
Gorham SL, Kadavil K, Lambert H, Vaughan E, Pert B, Abel J. 1991. Persistence of Salmonella enteritidis in young chickens. Avian Pathol. 20:433-437.
Desmidt M, Ducatelle R, Haesebrouck F. 1996. Pathogenesis of Salmonella enteritidis phage type four after experimental infection of young chickens. Vet Microbiol. 56:99-109.
Desmidt M, Ducatelle R, Haesebrouck F. 1998. Serological and Bacteriological observations on experimental infection with Salmonella Hadar in chickens. Vet Microbiol. 60:259-269.
Ferreira AJP. 1994. Salmonelose Aviária. Relatório do Departamento de Patologia, Faculdade de Medicina Veterinária e Zootecnia, USP. SP.
Morse EV & Duncan MA. 1974. Salmonellosis an environmental health problem. J Am Vet Med Assoc. 165:1015-1019.
National Research Council (NRC). 1994. Nutrient requirements of poultry. 9th rev.ed. National Academy Press: Washington, D.C.
Silva EAJ. 1999. Manual de Controle Higiênico - Sanitário em Alimentos. 3ª ed. Varela, São Paulo.
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