Marek's disease virus morphogenesis.

Marek's disease virus (MDV) is a highly contagious virus that induces T-lymphoma in chicken. This viral infection still circulates in poultry flocks despite the use of vaccines. With the emergence of new virulent strains in the field over time, MDV remains a serious threat to the poultry industry. More than 40 yr after MDV identification as a herpesvirus, the visualization and purification of fully enveloped infectious particles remain a challenge for biologists. The various strategies used to detect such hidden particles by electron microscopy are reviewed herein. It is now generally accepted that the production of cell-free virions only occurs in the feather follicle epithelium and is associated with viral, cellular, or both molecular determinants expressed in this tissue. This tissue is considered the only source of efficient virus shedding into the environment and therefore the origin of successful transmission in birds. In other avian tissues or permissive cell cultures, MDV replication only leads to a very low number of intracellular enveloped virions. In the absence of detectable extracellular enveloped virions in cell culture, the nature of the transmitted infectious material and its mechanisms of spread from cell to cell remain to be deciphered. An attempt is made to bring together the current knowledge on MDV morphogenesis and spread, and new approaches that could help understand MDV morphogenesis are discussed.

Assembly of Marek's disease virus (MDV) capsids using recombinant baculoviruses expressing MDV capsid proteins.

The genes UL18, UL19, UL26, UL26.5, UL35 and UL38 of Marek's disease virus 1 (MDV-1) strain RB1B, encoding the homologues of herpes simplex virus type 1 (HSV-1) capsid proteins VP23, VP5, VP21-VP24, preVP22a, VP26 and VP19C, were identified and sequenced. Recombinant baculoviruses were used to express the six capsid genes in insect cells. Coexpression of the six genes or of UL18, UL19, UL26.5 and UL38 in insect cells resulted in the formation of capsids with a large core. In addition, electron microscopy of thin sections clearly revealed the presence of large numbers of small spherical particles. Experimental coinfection demonstrated that these small particles were associated with production of the preVP22a protein.

Assembly pathway of avian infectious laryngotracheitis virus.

Infectious laryngotracheitis virus (ILTV) is the causative agent of a highly contagious upper respiratory tract infection in chickens. At present, ILTV vaccines are not satisfactory because of development of a latent carrier status in vaccinated birds. Development of recombinant virus vaccines has been hampered by the limited information available on the molecular level and organization of this virus. We isolated 3 assembly intermediates, designated A, B, and C from ILTV-infected cells. Analysis of [3H]thymidine-and [35S]methionine-labeled particles, and electron microscopic studies indicated that particle A was the empty capsid, particle B was the procapsid containing scaffolding protein, and particle C was the DNA-filled capsid. The ILTV procapsids could only be found in the nucleus, which indicated that procapsids could not translocate through the nuclear membrane until they packaged the DNA. The DNA-filled capsids migrated through the nuclear membrane and obtained an envelope from the inner membrane of the nucleus. The enveloped particles then migrated through the lumen of the endoplasmic reticulum into vacuoles in the cytoplasm. Infective virions were isolated from within the infected cells, indicating that budding through the cytoplasmic membrane is not a necessary step in ILTV maturation. Abundant arrays composed of tubules about 45 to 50 nm wide were found in the cytoplasm of chicken embryonic liver cells about 30 to 38 hours after infection. Comparison of the assembly intermediates and the DNA packaging pathway of ILTV with that of bacteriophage pi 29 indicates that similarity exists.(ABSTRACT TRUNCATED AT 250 WORDS)

Nucleosomal structure of Marek's disease virus genome in transformed lymphoblastoid cell lines, MDCC-MSB1 and MDCC-RP1.

The latent MDV (Marek's disease virus) genomes are folded into nucleosomal structures in both virus-productive and -nonproductive lymphoblastoid cell lines, MDCC-MSB1 (MSB-1) and -RP1 (RP-1), respectively. There was no difference between transcriptionally active and inactive regions of MDV genome with regard to nucleosomal patterns. In order to investigate whether nucleosomal structure is correlated with the repression of the transcription of MDV genome in lymphoblastoid cells, we examined the DNaseI sensitivity of nucleosomal MDV DNA in lymphoblastoid cell lines, MSB-1 and RP-1. No difference in the DNaseI sensitivity between transcriptionally active and inactive MDV DNA regions in lymphoblastoid cells was observed.

Cohelical arrangement of the DNA strand in the core of Marek's disease virus particles.

Electron microscopic studies on Marek's disease virus demonstrated that fibrils of 20 to 30 A were arranged in a cohelical configuration within an electronopaque band of about 100 A thickness in the core of immature particles. These fibrils were presumed to be the DNA double helix itself. This configuration may represent an arrangement of the herpesvirus DNA.

Replication of turkey herpesvirus and Marek's disease virus in chick embryo skin organ culture.

Turkey herpesvirus (HVT) and an attenuated Marek's disease virus (MDV) replicated in organ cultures of chick embryo skin as assessed by immunofluorescence and/or electron microscopy. HVT-specific immunofluorescent antigen was detected in the feather follicle epithelium (FFE) and in the surface layer of the skin epidermis. Electron microscopy of infected explants revealed herpes-type cytopathology. Immature particles of both viruses appeared first in the nucleus. Oval or horseshoe-shaped non-enveloped particles of HVT and enveloped virions of MDV were seen in the cytoplasm of some transitional cells. The difference in the ability of HVT and MDV to form an envelope was believed to account for the difference in their transmissibility in chickens. The results indicated that HVT replicated in the FFE and in the epidermis of the skin. However, attempts to localise the site(s) of MDV replication by electron microscopy were unsuccessful.

Further study on the three-dimensional structure of the core of Marek's disease virus and herpesvirus of turkey.

Three-dimensional structures of the core of Marek's disease virus and herpesvirus of turkey were examined by the tilting apparatus of an electron microscope. Various types of the core found in the infected cells were considered to represent developmental stages of the viruses. The basic structure of the core consisted of a toroid surrounding a cylindrical mass, which was clearly demonstrated by tilting the core in two directions. A cylindrical mass spooled by more than two toroids, which seemed to constitute a spiral band of 10 to 20 nm, was demonstrated. The maturation process of the cores of the viruses was also discussed.