Table 1. Experimental design.

Group	N°animals (n)	Inoculum 1 Day 0	Inoculum 2 Day 15	N°animals (n)	Inoculum 3 Day 30
G1	10	H7N2 LPAIVa	H7N1 HPAIVb	6	H5N1 HPAIVc
G2	10	Saline solution	H7N1 HPAIV	–	–
G3	10	Saline solution	Saline Solution	6	H5N1 HPAIV

Thirty 2-week old SPF-chickens were randomly distributed into three groups. Animals received the first inoculum (day 0) and 2 weeks later (day 15), birds were challenged with the respective inoculum 2. Six birds from G1 and G2 were consecutively infected 2 weeks later (day 30) with the third inoculum.

Abbreviations: LPAIV = low pathogenic avian influenza virus; HPAIV = highly pathogenic avian influenza virus.

^aChickens from G1 were inoculated intranasally with LPAIV A/Anas plathyrhynchos/Spain/1877/2009 (H7N2) (10^5.5 ELD₅₀). Birds from G2 and G3 received saline solution.

^bChickens from G1 and G2 were intranasally challenged with HPAIV A/FPV/Rostock/34 (H7N1) (10^5.5 ELD₅₀) 15 days after the pre-exposure to H7N2 LPAIV. Birds from G3 received saline solution.

^cChickens from G1 and G3 were inoculated intranasally with 10^4.5 ELD₅₀ of A/Great crested grebe/Basque Country/06.03249/2006 (H5N1) 15 days after the challenge with H7N1 HPAIV.