Marek’s disease is one of the most significant infectious diseases in poultry. In birds, it can result in tumours or immunosuppression, making the birds more sensitive to other pathogens. Although MD vaccines have been widely used in China for a long time, failures of MD vaccination occur frequently on some poultry farms. The improper selection of a vaccine could be the cause of vaccine failure [5]. Birds are also more susceptible to MDV mutants that have emerged in regions with more-virulent strains than to previous MDV isolates.
The pp38 gene encodes a phosphoprotein of approximately 38 kDa that is necessary for both cell transformation and viral reactivation in the latent period [6, 7]. The pp38 gene is conserved in all strains of MDV so far isolated. Previously, it was believed that pp38 was not present in MDV strain CVI988 [8], but in recent years, the gene was shown to be expressed by CVl988, and a mutation at residue 107 (Q → R) is identified with Mab H19 [9, 10]. In this study, we found no mutation at residue 107 of the PP38 protein of the isolated strain AH1410. The pp38 gene sequence was identical to that of the vMDVs. However, at residue 109 of pp38 in the vMDVs and the very virulent MDVs (vvMDVs), such as strains GA, MD5, and RB1B, glutamate is replaced by glycine, which differs from the sequence of strain AH1410, which is similar to that of other virulent viruses isolated in China, such as GX0101 and JS0711. Therefore, the isolated virus AH1410 is not the vaccine strain.
The MEQ protein, which is considered to be the major factor responsible for the occurrence of tumours, contains 339 amino acids [4]. However, the meq gene of CVI988 has an insertion of 177 bp compared with the genes of the vMDVs, which results in a 59-amino-acid insertion relative to the MEQ proteins of the vMDVs [11–13], that inhibits the expression of the meq gene [14]. Research has also indicated that mutations at positions 71 and 77 of the MEQ protein could be distinct in the vvMDVs [15]. Compared with CVI988, position 71 of strain AH1410 is mutated (S → A), as in the vMDVs, whereas position 77 is conserved. No deletion or insertion was found at position 194 and the ORF of AH1410 meq encodes 339 amino acids. We also found mutations at positions 80, 115, 139, and 176 of MEQ in our isolate AH1410, similar to those in GX0101 [16], suggesting that this type of strain has been circulating in China for many years.
With sequence alignments of the meq, pp38, and gB genes of strain AH1410 and other MDV isolates, we found that GX0101 and AH1410 are completely identical in these three genes. A previous study demonstrated that the virulence of GX0101 was at a level between those of GA and Md5 [16]. Collectively, these data indicate that the pathogenicity of isolate AH1410 may be similar to that of isolate GX0101.
The mutations and increasing virulence of MDV are thought to be responsible for the occurrence of MD on vaccinated farms. The failure of MD vaccination may be one reason for the outbreak reported here. Notably, the onset of MD in broilers generally starts at 4–6 weeks of age. In hens, MD usually occurs at 10–12 weeks of age [17]. However, in this case, the rate of tumours reached its highest level during the peak egg-laying period (30 weeks old).
Outbreaks among egg-laying chickens have also been reported in the mining area of Karnataka, India [18], and excessive losses from MD have recently been reported in adult laying flocks over the age of 40 weeks in Japan [19]. The data indicate that these birds may have contracted these infections later in life, with subsequent clinical signs. We also found that the feathers of the chickens were positive for the virus, whereas their sera were negative for MDV antigen, indicating that the chickens’ immune responses may have been suppressed in other ways, an unusual phenomenon and a disquieting trend. Another interesting observation is that all the diseased chickens were hens, which is consistent with an observation by Heier et al., who noted that males tended to be more resistant to MD than females [20].
It is necessary to strengthen the surveillance of circulating MDVs because new variants of virulent MDVs may appear at any time, and better vaccines against MD must be developed.