Biological and molecular characterization of a genetically diverse avian paramyxovirus type 2 (APMV-2) isolated from pheasants

Authors: MARTINS, MATHIAS - Texas A&M Veterinary Medical Diagnostic Laboratory; Lee, Chang; MORRIS, ELLEN - Texas A&M Veterinary Medical Diagnostic Laboratory; BURNS, DEVIN - Texas A&M Veterinary Medical Diagnostic Laboratory; FICKEN, MARTIN - Texas A&M Veterinary Medical Diagnostic Laboratory; Suarez, David; DIMITROV, KIRIL - Texas A&M Veterinary Medical Diagnostic Laboratory

Submitted to: American Association of Veterinary Laboratory Diagnosticians
Publication Type: Abstract Only
Publication Acceptance Date: 10/12/2024
Publication Date: N/A
Citation: N/A

Interpretive Summary:

Technical Abstract: Avian paramyxoviruses (APMVs) have been identified in various avian species worldwide. As of 2024, twenty-two species of APMVs have been recognized and allocated to three genera within the subfamily Avulavirinae in the Paramyxoviridae family. Avian paramyxovirus type 2 (APMV-2), or Metaavulavirus yucaipaense according to ICTV, is predominantly maintained among wild birds and occasionally detected in poultry, causing mild respiratory disease in chickens and more severe infections in turkeys. This study investigates the etiology of a mild respiratory disease in pheasants submitted for necropsy at the Texas A&M Veterinary Medical Diagnostic Laboratory. Five adult pheasants with a week of weight loss and coughing history were submitted for diagnostic investigation. Tracheal swabs were tested using multiplex reverse transcription PCR (rtPCR) for Mycoplasma gallisepticum and Mycoplasma synoviae (MG and MS). Tracheas were processed, and virus isolation (VI) was attempted using SPF embryonated chicken eggs (ECE), followed by hemagglutination (HA) test and transmission electron microscopy (TEM). Whole genome sequencing was performed using sequence-independent, single-primer amplification (SISPA) reverse transcription and random amplification protocols on the MinION platform. Mean death time (MDT) determination in ECE and the intracerebral pathogenicity index (ICPI) test were performed. Antigenic characterization was conducted by hemagglutination inhibition (HI) antibody tests using a panel of reference APMVs sera and antigens. No significant alterations or relevant findings were observed during the necropsy examination. Tracheal secretions were negative for MG and MS by rtPCR. HA activity was detected in amniotic–allantoic fluid (AAF) harvested from ECE and tested negative for APMV-1 and avian influenza virus (AIV) by real-time rtPCR. Enveloped and pleomorphic viral particles of various sizes, consistent with paramyxoviruses, were observed on TEM performed on AAF. The viral isolate was classified as APMV-2 by obtaining and analyzing the complete genome (APMV-2/pheasant/Texas/G240020184/2024) (TX/24) and revealed less than 91% of the nucleotide identity with genomes available in GenBank. The genetically closest APMV-2 was APMV-2/Chicken/California/Yucaipa/56 (Yucaipa) (90.7% nt identity). BLAST analysis of the fusion protein gene yielded similar findings, with the closest available sequences less than 92% identical. The MDT could not be determined for TX/24 and Yucaipa virus, as neither killed all eggs at any tested virus dilution. Based on World Organization for Animal Health (WOAH) guidelines, the ICPI scores for TX/24 and Yucaipa fell into the lentogenic pathotype category (ICPI < 0.7). No clinical signs were observed with TX/24 isolate during the observation period, while Yucaipa resulted in one dead bird on day 2. On the HI test, the TX/24 isolate reacted with APMV-2 sera with no or minor cross-reactivity to other APMV serotypes, demonstrating an almost identical reactivity pattern observed with the Yucaipa strain. In summary, the APMV-2 isolate showed low virulence in laboratory studies, yet its potential role as the primary cause of the respiratory disease observed in pheasants cannot be ruled out. The findings from this study emphasize the necessity of expanding APMV monitoring beyond APMV-1.