Comparison of local Salmonella pullorum antigen with imported product in whole blood agglutination test

Background: Salmonellosis is considered as one of the most important diseases in poultry as it causes devastating losses in chicken industry. Proper identification of the infected and carrier birds is required to control the disease among chickens. In field situation whole blood agglutination test is performed in order to identify carriers of Pullorum and Fowl typhoid particularly, in breeder operations. In this test, serum antibodies are detected by using a specially made antigen for this purpose. In Sri Lanka, three antigen products are used commonly in whole blood agglutination test. Aim: This study was carried out to compare these two locally available S. Pullorum antigen products and to determine any difference in the efficacy. Materials and Methods:“Shaver Brown” commercial layer birds (70 in number) were used in the experiment. Birds were 9 inoculated orally with 1.8X10 cfu/ml of S. Pullorum at 16 weeks age. After Three weeks post inoculation, blood was collected from each bird and Whole blood agglutination test was performed using both antigen products. Fifteen (15) inoculated hens were selected randomly and cloacal swabs were cultured on cultured Agar on same day of serum collected. Results: In this study, there was no significant difference observed between two antigens to detect carrier birds by whole blood agglutination test. Salmonella was not isolated from cloacal swabs since no observed excretion of Salmonella Pullorum through faces. All cloacal swabs gave negative results, when cultured on artificial Agar. Conclusion: Both antigen can be used effectively to detect carrier birds under the control program in country.


Introduction
infections of the ovarian follicles [3].The disease can be transmitted through eggs to the chicks (6,7).The genus Salmonella, of the family Enteroba-However, chronic carriers are the most important cteriaceae contains over 2400 serotypes which may reservoirs of the bacterium that transmit to rest of the infect wide range of animals [1].Since bacteria often birds [3,4].excreted in feces, Organisms may be present in water, The Pullorum disease can be controlled by soil, animal feeds, raw meat, and offal and in vegetable continuous eliminating of carrier birds by bloodmaterials [1].In global contest, Isolations of Salmonellae testing of potential breeding bird followed by culling are reported more often from poultry and its products [7].Breeders that test negative produce non-infected than rest of the livestock and other animal species in eggs later non-infected chicks (7,8).Since S. Pullorum world [2,3].and S. Gallinarum possess similar somatic antigens, Chicken is the natural hosts for S. Pullorum and both have been shown positive reaction during whole S.Gallinarum[1], infects young chicks at two to three blood agglutination test [9].However, antibodies from weeks of age [1].Characteristic lesions include whitish live or killed vaccine can be interfered with the testing nodules throughout the lungs and focal necrosis of and prohibited to use in breeder flock, Sri Lanka [7].liver and spleen [1,4,5].It was reported in previous The whole blood agglutination test is a simple and less studies that a greater percentage of female than male costly test, greatest value when used on a flock basis.stay as reactors, due to the sequestered nature of local [1,7,9].Serological prevalence of Salmonella Pullorum Statistical analysis: MINITAB statistical software is monitored routinely in every breeder farm, Sri was used and binary logistic regression was the Lanka from 16 weeks onwards [7].Retesting is carried methodology adopted in data analysis.out at intervals of 60-90 days if the flock shows more

Results
than 1% positive reactors that allow in rapid whole blood agglutination test [7].However, there was believed In this study, there was no significant difference among farmers that different antigens gathered observed between two antigens to detect carrier birds different results and local antigens are not met up to the by whole blood agglutination test.Salmonella was not standards of imported products.The objective of this isolated from cloacal swabs since no observed study is to compare the reading of WBAT with reputed excretion of Salmonella Pullorum through faces.All imported products in order to maintain unique reading cloacal swabs gave negative results, when cultured on through the control program.
artificial Agar.same hatch were used for the study.They were reared Logistic Regression there were no statistically significant different observed in both antigens.
Antigen products: Two antigens namely, one from Veterinary Research Institute (A) and one of the Imported brand antigen (B)were used in the study, the A was produced by the given standard in the OIE (8). of each serum sample were prepared (1, 1:2, 1:4, 1:8, All samples gave negative results and there was 1:16,1:32).Whole blood agglutination test was no any pink colony, when cultured on brilliant green performed as described by Quinn et al. (2003).
agar.No Salmonella organsim was isolated from the cloacal swabs taken from serologically positive birds.Fecal swabs: Fifteen serologically positive birds were selected randomly and cloacal swabs were taken Discussion from them on the same day of blood collection.The In this study both antigen were detected swabs were pre-enriched in peptone water (incubated antibodies in chicken infected with S. Pullorum.There at 37°C for 48 hours) and then inoculated into 10 ml of was no significant difference between "A" and "B." tetrathionate broth and incubated at 37°C for 24 hours.
The results were the same in diluted serum tested up to The loopful of tetrathionate broth was inoculated on 1:32, since there is unique reading using both antigens, artificial media (Brilliant Green).accuracy of the national control program would not at Bacteriology division who helped to complete this affected by local antigen used.The testing was done in research activity, also the research committee at the room temperature to minimize the variables such Faculty of Veterinary Medicine and Animal science, as environment temperature, dust particles and quality Director/VRI and Director General of DAP&H for of the blood used [1].However, those factors were not granting permission to carry out the research, finally negligible in the field level at mass screening.In this the staff at both Karandhagolla and Marawilla NLDB experiment relatively large doses were administered farm.on the same day though this condition is not common (A-Local antigen, B -Imported branded antigen) at the Veterinary Research Institute, Gannoruwa under Since P value was 0.092 (P > 0.05) of the test by Binary deep liter management system.

SalmonellaFigure- 1 .
Figure-1.Number of positive samples against serum hour before testing.. Then five two fold serial dilutions dilution levels.ofeach serum sample were prepared (1, 1:2, 1:4, 1:8, All samples gave negative results and there was 1:16,1:32).Whole blood agglutination test was no any pink colony, when cultured on brilliant green performed as described byQuinn et al. (2003).agar.No Salmonella organsim was isolated from the cloacal swabs taken from serologically positive birds.Fecal swabs: Fifteen serologically positive birds were selected randomly and cloacal swabs were taken Discussion from them on the same day of blood collection.The In this study both antigen were detected swabs were pre-enriched in peptone water (incubated antibodies in chicken infected with S. Pullorum.There at 37°C for 48 hours) and then inoculated into 10 ml of was no significant difference between "A" and "B." tetrathionate broth and incubated at 37°C for 24 hours.The results were the same in diluted serum tested up to The loopful of tetrathionate broth was inoculated on 1:32, since there is unique reading using both antigens, artificial media (Brilliant Green).

Table - 1
. Number of positive samples for each dilution level.Ethical clearance: the ethical clearance for the study was given by the Animal Ethical Review Committee at of serum Veterinary Research Institute, Sri Lanka.