Vaccination-challenge interval markedly influences protection provided by Rispens CVI988 vaccine against very virulent Marek's disease virus challenge

Figures & data

Table 1. Summary of treatments showing treatment abbreviations and vaccination challenge interval.

Number	Treatment	Day of vaccination	Day of challenge	VCI
1	MDV d0	Unvaccinated	0	N/A
2	MDV d5	Unvaccinated	5	N/A
3	MDV d10	Unvaccinated	10	N/A
4	RIS d0	0	Unchallenged	N/A
5	RIS d5	5	Unchallenged	N/A
6	RIS d10	10	Unchallenged	N/A
7	VCI −10	10	0	−10
8	VCI −5	5	0	−5
9	VCI 0	0	0	0
10	VCI 5	0	5	5
11	VCI 10	0	10	10
12	Negative control	Unvaccinated	Unchallenged	N/A

MDV, challenged with MDV isolate 02LAR; RIS, vaccinated with Rispens CVI988; N/A, not applicable.

Table 2. Details of viruses used in the experiment.

Virus	Name	Batch number	Dose (PFU)	Source	Year of isolation
vvMDV	02LAR	P4 18/11/2009	400	University of New England, Armidale, NSW, Australia	2002
Rispens CVI988	Vaxsafe^®RIS	RIS6111	3200	Bioproperties Pty Ltd, Ringwood, Vic, Australia	1972

Table 3. Effect of vaccination with Rispens CVI988 vaccine and challenge with vvMDV isolate 02LAR in various time combinations on total mortality, mortality with MD, incidence of MD to 56 d.p.c. and PI by VCI and treatment in commercial ISA Brown chickens

VCI (days)	Treatment	Vaccination	Challenge	Total mortality (%)	Mortality with MD lesions (%)	MD incidence (%)	PI (%)
	MDV d0	Unvaccinated	02LAR	11/40 (27.5)^ab	8/40 (20)^a	21/37 (56.8)^a
	MDV d5	Unvaccinated	02LAR	9/40 (22.5)^abd	8/40 (20)^a	17/39 (43.6)^a
	MDV d10	Unvaccinated	02LAR	15/40 (37.5)^ae	11/40 (27.5)^a	22/38 (57.9)^a
	RIS d0	Rispens	Unchallenged	3/39 (7.7)^d	0/39 (0)^c	0/39 (0)^c
	RIS d5	Rispens	Unchallenged	0/50 (0)^c	0/50 (0)^c	0/50 (0)^c
	RIS d10	Rispens	Unchallenged	0/48 (0)^c	0/48 (0)^c	0/49 (0)^c
	Negative control	Unvaccinated	Unchallenged	2/20 (10)^bd	0/20 (0)^c	0/18 (0)^c
−10	VCI −10	Rispens	02LAR	6/40 (15)^bd	6/40 (15)^ab	18/40 (45)^a	20.9^c
−5	VCI −5	Rispens	02LAR	22/40 (55)^e	21/40 (52.5)^d	23/39 (59)^a	−3.6^c
0	VCI 0	Rispens	02LAR	10/40 (25)^a	8/40 (20)^a	9/40 (23.1)^b	60.4^b
5	VCI 5	Rispens	02LAR	1/30 (3.3)^d	1/30 (3.3)^bc	2/30 (6.7)^bc	84.8^ab
10	VCI 10	Rispens	02LAR	2/30 (6.7)^d	0/30 (0)^c	0/28 (0)^c	100^a
Total				81/457 (17.7)	63/457 (13.8)

Mortality is for eligible chickens from 2 d.p.c.; MD incidence from 31 d.p.c. when the first MD case occurred. ^abcdeMeans within columns not sharing a common superscript letter differ significantly. MDV, challenged with MDV isolate 02LAR; RIS, vaccinated with Rispens CVI988.

Figure 1. Survival patterns of commercial ISA Brown layer chickens vaccinated with Rispens CVI988 vaccine and/or challenged with MDV isolate 02LAR showing the effects of VCI ranging from −10 to 10 days (left; P < 0.0001), challenge with 02LAR in unvaccinated chickens (middle; P=0.38) and vaccination with Rispens CVI988 in unchallenged chickens (right; P<0.0001).

Figure 2. Association between vaccination challenge interval and (2a) MD incidence and (2b) PI. Each point represents an individual isolator.

Figure 3. Mean (least-squares means ± standard errors of the mean) serum Log₁₀ of MD antibody titre for the different VCI treatments measured at 21 and 56 d.p.c. The effect of VCI was significant (P = 0.006). ^abcMeans columns not sharing a common superscript letter differ significantly (P < 0.05).

Figure 4. Mean (least-squares means ± standard errors of the mean) serum Log₁₀ of MD antibody titre for all treatments measured at 21 and 56 d.p.c. for all treatments involving MDV challenge or 21 and 56 d.p.v. in the RIS treatments. The effect of treatment was significant (P = 0.0006).

Figure 5. Association between MD incidence and pathogenic MDV load in PBL at various times post infection. Each data point represents the mean Log₁₀ MDV copy number in PBL of five chickens from each isolator and the corresponding MD incidence of that group up to 56 d.p.c. First row, unchallenged isolators excluded; second row, unchallenged and unvaccinated treatments excluded.

Table 4. Pairwise correlation of MD incidence (MD%) at day 56 with viral load in various tissues, immune organ weight and MD antibody titre of different challenge days.

Variable	Virus	Day	Correlation with MD to 56 d.p.c.	n	Lower 95%	Upper 95%	P value
PBL	MDV-1	7	0.4425	16	−0.0681	0.7694	0.0861
		14	0.8305	16	0.569	0.9394	<.0001*
		21	0.8264	16	0.5603	0.9379	<.0001*
	Rispens	7	−0.1696	10	−0.7221	0.515	0.6394
		14	−0.0668	10	−0.6683	0.5875	0.8545
		21	−0.5693	10	−0.8826	0.094	0.0858
Feather	MDV-1	7	−0.5193	16	−0.8072	−0.0318	0.0392*
		14	0.5118	16	0.0215	0.8036	0.0427*
		21	0.8386	16	0.5868	0.9425	<.0001*
	Rispens	7	−0.5559	10	−0.8782	0.1135	0.0952
		14	−0.1181	10	−0.6959	0.5527	0.7453
		21	−0.4052	10	−0.8245	0.3012	0.2453
Dust	MDV-1	7	0.0882	16	−0.4262	0.5594	0.7455
		14	−0.0915	16	−0.5617	0.4234	0.7361
		21	0.3208	16	−0.208	0.7045	0.2258
	Rispens	7	−0.0988	10	−0.6858	0.566	0.7859
		14	−0.083	10	−0.6772	0.5768	0.8197
		21	0.3316	10	−0.3767	0.7952	0.3493
Relative splenic weight (%)		14	−0.4476	16	−0.772	0.0619	0.0821
		56	0.4678	16	−0.0364	0.7821	0.0677
Relative bursal weight (%)		14	0.093	16	−0.4221	0.5628	0.7318
		56	−0.2485	16	−0.6626	0.2819	0.3534
Log10 anti-MDV titre		21	0.338	16	−0.1894	0.7141	0.2004
		56	−0.004	15	−0.5152	0.5093	0.9887

Correlation is between least-squares mean values for each isolator. Analysis includes 16 isolators excluding only treatments not challenged with pathogenic MDV. *indicates the significant correlation between subsequent pair.

Figure 6. Association between MD incidence and pathogenic MDV load in feather cells at various times post infection. Each data point represents the mean Log₁₀ MDV copy number in feather cells of five chickens from each isolator and the corresponding MD incidence of that group up to 56 d.p.c. at the end of the experiment. First row, unchallenged isolators excluded; second row, unchallenged and unvaccinated treatments excluded.

Table 5. Pairwise correlation of MD incidence (MD%) at d56 with viral load in various tissues, immune organ weight and MD antibody titre of different challenge days.

Variable	Virus	Day	Correlation with MD to 56 d.p.c.	n	Lower 95%	Upper 95%	P value
PBL	MDV-1	7	0.4871	10	−0.2056	0.8546	0.1533
		14	0.9055	10	0.6418	0.9777	0.0003*
		21	0.8739	10	0.543	0.9699	0.0009*
	Rispens	7	−0.1696	10	−0.7221	0.515	0.6394
		14	−0.0668	10	−0.6683	0.5875	0.8545
		21	−0.5693	10	−0.8826	0.094	0.0858
Feather	MDV-1	7	−0.5083	10	−0.8621	0.1784	0.1335
		14	0.7717	10	0.2763	0.9431	0.0089*
		21	0.8401	10	0.4467	0.9613	0.0023*
	Rispens	7	−0.5334	10	−0.8706	0.1449	0.1124
		14	−0.4576	10	−0.8441	0.2416	0.1835
		21	−0.5979	10	−0.8918	0.0508	0.0679
Dust	MDV-1	7	0.2616	10	−0.4406	0.7652	0.4653
		14	−0.2331	10	−0.7523	0.4647	0.5169
		21	0.4817	10	−0.2124	0.8527	0.1587
	Rispens	7	−0.0988	10	−0.6858	0.566	0.7859
		14	−0.083	10	−0.6772	0.5768	0.8197
		21	0.3316	10	−0.3767	0.7952	0.3493
Relative splenic weight (%)		14	−0.4447	10	−0.8393	0.2568	0.1978
		56	−0.0447	10	−0.6559	0.6018	0.9023
Relative bursal weight (%)		14	0.3693	10	−0.3392	0.8105	0.2937
		56	0.4575	10	−0.2417	0.844	0.1837
Log10 anti-MDV titre		21	0.358	10	−0.3507	0.8059	0.3098
		56	−0.1154	10	−0.6945	0.5546	0.751

Correlation is between least-squares mean values for each isolator. Analysis includes 10 isolators in VCI treatments involving infection with both the Rispens CVI988 vaccine virus and vvMDV isolate 02LAR. *indicates the significant correlation between subsequent pair.