Differences in replication kinetics and cell tropism between neurovirulent and non-neurovirulent EHV1 strains during the acute phase of infection in horses

Abstract

Equine herpesvirus 1 (EHV1) replicates in the respiratory tract of horses, after which infected leukocytes transport virus throughout the body, resulting in abortion or nervous system disorders. Two EHV1 strains circulate in the field: neurovirulent and non-neurovirulent. To investigate differences in replication in the upper respiratory tract (URT), an experimental inoculation study in ponies was performed with both strains. Two groups of six ponies, were inoculated intranasally with 10 6.5 TCID 50 of either strain. Clinical signs, nasal shedding and viremia were evaluated. At early time points post inoculation (pi), one pony of each group was euthanized. Tissues were collected for titration and immunostainings. Number and size of EHV1-induced plaques were calculated, and individual EHV1-infected cells were quantified and characterized. Inoculation with either strain resulted in nasal shedding and replication in several tissues of the URT. Both strains replicated in a plaquewise manner in epithelium of the nasal mucosa, but replication in epithelium of the nasopharynx was largely limited to nonneurovirulent EHV1. Plaques were never able to cross the basement membrane, but individual infected cells were noticed in the connective tissue of all examined tissues for both strains. The total number of these cells however, was 3-7 times lower with non-neurovirulent EHV1 compared to neurovirulent EHV1. CD172a + cells and CD5+ lymphocytes were important target cells for both strains. Interestingly, in lymph nodes, B-lymphocytes were also important target cells for EHV1, irrespective of the strain. Viremia was detected very early pi and infected cells were mainly CD172a + for both strains. In summary, these results are valuable for understanding EHV1 pathogenesis at the port of entry, the URT.

Replication kinetics of neurovirulent versus non-neurovirulent equine herpesvirus type 1 strains in equine nasal mucosal explants

“…A big concern, however, is the heterologous nature of this animal model, which can make it difficult to make valid comparisons and extrapolations to the natural host of the virus, the horse (Walker et al, 1999). Our in vitro model is a good representation of the in vivo situation in the natural host, as the results obtained in our study were virtually identical to those obtained in a recent in vivo pathogenesis study in horses by Gryspeerdt et al (2010). Moreover, differences in invasion kinetics and infection rates between neurovirulent and non-neurovirulent EHV-1 isolates were identified, indicating that our in vitro respiratory mucosal explant system is a valuable alternative to provide novel information in addition to the currently existing EHV-1 models.…”

“…for 03P37, a neurovirulent D 752 isolate, whereas for the non-neurovirulent N 752 isolate 97P70, single infected cells were observed as early as 2 days p.i. in this in vivo experiment (Gryspeerdt et al, 2010). In our nasal explant system, we observed no differences in cell tropism between D 752 and N 752 isolates, and EHV-1-infected cells were equally identified as CD5 + T lymphocytes and CD172a + CML.…”

“…for all non-neurovirulent EHV-1 isolates carrying the N 752 genotype in their DNA polymerase when compared with neurovirulent, D 752 isolates. What the underlying mechanism is for this faster breakthrough through the BM barrier of neurovirulent isolates remains to be elucidated, but we can state that (i) the D/N 752 SNP is important for this phenomenon and (ii) the time-dependent difference in spread through the BM is not an artefact in our system, as similar observations have been made in vivo (Gryspeerdt et al, 2010). Indeed, single EHV-1-infected cells below the BM were seen starting from 1 day p.i.…”

“…The mucosa of the upper airway tract is the first line of defence against respiratory diseases (Timoney, 2004). It is also the primary replication site of EHV-1, as it is for most alphaherpesviruses (Kydd et al, 1994a;Van Maanen, 2002;Van Maanen & Cullinane, 2002;Gryspeerdt et al, 2010). Subsequently, the virus disseminates via a leukocyte-associated viraemia, which enables EHV-1 to reach end-vessel endothelia in the uterus and central nervous system (Allen & Bryans, 1986;Kydd et al, 1994b).…”

Role of gB and pUS3 in Equine Herpesvirus 1 Transfer between Peripheral Blood Mononuclear Cells and Endothelial Cells: a DynamicIn VitroModel

“…E quine herpesvirus type 1 (EHV-1) and EHV-4 are members of the Herpesviridae family and Alphaherpesvirinae subfamily (1,2). After initial replication in the upper respiratory tract, EHV-1 infects immune cells and migrates past the epithelial basement membrane to the lymph nodes and bloodstream (1)(2)(3)(4). As a result, EHV-1 is able to spread throughout the body, where it infects endothelial cells (EC), causing vascular lesions and secondary hypoxic degeneration of the affected tissues (3,5,6).…”