Impact of heterophil granulocyte depletion caused by 5-fluorouracil on infectious bursal disease virus infection in specific pathogen free chickens

Figures & data

Table 1.  Heterophile granulocytes in blood and bursa lesion score

		Heterophile granulocyte count^a			Bursa lesion score^b
Days after 5-FU treatment	Days after IBDV inoculation	Group 1	Group 2	Group 3	Group 1	Group 2	Group 3
5	0	29, 31, 41	26, 18, 15	35, 15, 20	0, 1, 1	0, 0, 0	2, 1, 1
6	1	49, 44, –	16, 19, 18	37, 25, 38	1, 3, 1	0, 0, 0	3, 2, 2
7	2	20, 10, 10	50, 41, 12	0, 0, 10	1, 1, 1	3, 4, 4	4, 4, 3
8	3	4, 1, 0	24, 23, 50	8, 1, 1	2, 2, 1	4, 4, 4	4, 4, 4
15	10	3, 2, 0	ND	40, 6, 14	0, 0, 0	ND	4, 4, 4
Three chickens were examined on each sampling day. ND, not done; –, sample missing. ^aThe heterophile granulocyte counts are indicated as percentage of blood heterophils. ^bThe bursa lesion score values for groups 1 to 3 indicate the percentage of bursa tissue involved in changes as follows: 0 = 0%, 1 to 25% = 1, 25 to 50% = 2, 51 to 75% = 3, 76 to 100% = 4.

Figure 1.  Reactions to IBDV infection in 5-FU-pretreated and non-pretreated chickens. 1a: 5-FU and IBDV inoculation (group 3): 3 days after IBDV inoculation (p.i.), folded surface epithelium was observed (arrows); the medullary epithelial reticular cells form “empty holes”. Anticytokeratin immunostaining. Magnification, 40×. 1b: IBDV inoculation (group 2): 2 days p.i., B-cell depletion. Depletion started in the centre of the medulla and inner part of the cortex. Bu-1b staining. Magnification, 35×. 1c: IBDV inoculation (group 2): 2 days p.i., the follicular cortex flooded by heterophil granulocytes (arrows). Semithin section stained with toluidin blue. Magnification, 500×. 1d: 5-FU and IBDV inoculation (group 3): 2 days p.i., reduction in the number of heterophil granulocytes (arrows). Compare with 1c. Semithin section stained with toluidin blue. Magnification, 500×. 1e: IBDV inoculation (group 2): 2 days p.i., huge numbers of vimentin-positive cells in the medulla and surface epithelium of the follicles. Magnification, 140×. 1f: 5-FU and IBDV inoculation (group 3): 2 days p.i., the number of vimentin-positive cells in the medulla reduced as compared with group 2 (see 1e). Magnification, 140×.

Table 2.  Histological development in the bursa of Fabricius 5 to 8 days after 5-FU treatment (0 to 3 days after IBDV inoculation)

Cell type	Group 1, 5-FU	Group 2, IBDV	Group 3, 5-FU + IBDV
B cells	Mild depletion	Increasing depletion starting from the medulla and inner part of the cortex on day 2 p.i.	Mild depletion in the medulla from day 0 p.i., increasing in the medulla and cortex
BSDCs	Not done	Present in mature form in the medulla and along the cortico-medullary border 2 days p.i. Most of the cells had disappeared 3 days p.i.	Elevated number of immature forms, aggregated at the cortico-medullary border from 2 days p.i. Few mature BSDCs in the medulla
Epithelial cells	Normal surface epithelium	Folded surface epithelium 3 days p.i.	Folded surface epithelium 3 days p.i.
Heterophile granulocytes	Mild depletion	Increased number, mostly in cortex, 2 days p.i.	Depletion from 2 days p.i.
Multinucleated giant cells	Not done	Virus containing cells in cortex 3 days p.i.	B cells and virus-containing cells in the medulla on day 2 p.i. and virus containing cells in the cortex 3 days p.i.
T cells	Not done	Increase and aggregation, mainly in cortex 2 days p.i.	Increase and aggregation, mainly in cortex 2 days p.i.

Figure 2.  BSDCs and T cells during IBDV infection. 2a: 5-FU and IBDV inoculation (group 3): 2 days after IBDV infection. The cell contains apoptotic lymphocytes (A) and two demarcated bodies, in which the virus particles are associated with an electron-dense substance (arrow). Part of one of the demarcated bodies is outlined and shown in 2b. Lipid droplets (L) in the cytoplasm. Magnification, 10 000×. 2b: Detail from 2a. In the demarcated body, the virus particles are intermingled with electron dense substance. Magnification, 40 000×. 2c: IBDV inoculation (group 2): 3 days p.i., CD3-positive T cells aggregate in the follicular cortex. Medulla contains only scattered T cells. Magnification, 140×. 2d: IBDV inoculation (group 2): 2 days p.i., the 74.3-positive BSDCs aggregate along the cortico-medullary border in several follicles. Magnification, 160?×. 2e: IBDV inoculation (group 2): 3 days p.i., the 74.3-positive BSDCs seem to be completely aggregated in the medulla. Magnification, 140×. 2f: IBDV inoculation (group 2): 2 days p.i., the anti-IgG staining confirm observations obtained by 74.3 monoclonal antibodies. The IgG-positive BSDCs aggregate along the cortico-medullary border. Compare with 2d. Magnification, 70×.