Transboundary and Emerging Diseases

As the H9N2 subtype avian influenza virus (H9N2 AIV) evolves naturally, mutations in the hemagglutinin (HA) protein still occur, which involves some sites with glycosylations. It is widely established that glycosylation of the H9N2 AIV HA protein has a major impact on the antigenicity and pathogenicity of the virus. However, the biological implications of a particular glycosylation modification site (GMS) have not been well investigated. In this study, we generated viruses with different GMSs based on wild-type (WT) viruses. Antigenicity studies revealed that the presence of viruses with a 200G⁺/295G⁻ mutation (with glycosylation at position 200 and deletion of glycosylation at position 295 in the HA protein) combined with a single GMS, such as 87G⁺, 127G⁺, 148G⁺, 178G⁺, or 265G⁺, could significantly affect the antigenicity of the virus. Pathogenicity assays revealed that the addition of GMS, such as 127G⁺, 188G⁺, 148G⁺, 178G⁺, or 54G⁺, decreased the virulence of the virus in mice, except for 87G⁺. The removal of GMS, such as 280G⁻ or 295G⁻, increased the pathogenicity of the virus in mice. Further studies on pathogenicity revealed that 87G⁺/295G⁻ could also enhance the pathogenicity of the virus. Finally, we selected the WT, WT-87G⁺, WT-295G⁻, and WT-87G⁺/295G⁻ strains as our further research targets to investigate the detailed biological properties of the viruses. GMS, which can enhance viral pathogenicity, did not significantly affect replication or viral stability in vitro but significantly promoted the expression of proinflammatory factors to enhance inflammatory responses in mouse lungs. These findings further deepen our understanding of the influence of the glycosylation of the HA protein of H9N2 AIV on the pathogenicity and antigenicity of the virus in mice.

In 2021, a highly virulent strain of duck enteritis virus (DEV), designated as DEV XJ, was isolated from Zhejiang, China, and its complete genome, spanning 162,234 bp with 78 predicted open reading frames (ORFs), was sequenced. While showing relative homology to the DEV CV strain, DEV XJ exhibited distinctions in 38 ORFs, including various immunogenic and virulence-related genes. Amino acid variation analysis, focusing on UL6 and LORF3, indicated a high degree of homology between DEV XJ and the 2085 strain from Europe, as well as the DEV DP-AS-Km-19 strain from India. Subsequently, a full-length infectious bacterial artificial chromosome clone (BAC) of DEV XJ was successfully constructed to delve into the pathogenic mechanisms of this virulent strain. XJ BAC demonstrated substantial similarity to the parental DEV XJ in both in vitro growth properties and the induction of typical pathogenic symptoms in sheldrakes. Furthermore, the US3, LORF3, UL21, and UL36 genes were individually deleted using a two-step RED recombination approach based on the infectious BAC clone. Our findings revealed that the UL21 and UL36 genes play crucial roles in viral proliferation. Although the US3 and LORF3 genes were dispensable for viral replication and cell-to-cell transmission in vitro, they attenuated the replication and transmission efficiency of DEV compared to the WT. In summary, this study accomplished the whole-genome sequencing of a clinically virulent DEV strain and the successful construction of an infectious DEV XJ clone. Moreover, the functional roles of the above-mentioned mutant genes were preliminarily explored through the analysis of their in vitro biological characteristics.

Pigs are affected by various parvoviruses (PPVs); eight have been reported to date (PPV1–PPV8). Porcine parvovirus 1 is considered a primary agent of porcine reproductive failure (PRF), while it is unknown whether other PPVs impact porcine health. Recently, the presence of PPV2 has been confirmed in the lung, either as a single agent or in the form of coinfection with other respiratory; therefore, it has been proposed as a potential participant in the porcine respiratory disease complex (PRDC). In the present study, the presence of PPV2 alone and coinfection with other viruses (PCV2, PCV3, and PRRSV) was evaluated in lung samples obtained from pigs with respiratory signs (respiratory group: RG) (n = 146) and stillborn lungs (stillborn group: SG) (n = 19) from 82 farms in the five regions with the highest swine production in Colombia. The overall PPV2 prevalence was 37.6% (62/165), with the highest proportion mainly detected in grow-finisher pigs (62.5%), while its herd prevalence was 51.2% (42/82). The most prevalent virus was PRRSV in both groups, while PPV2 alone was found only in the RG group. The most common dual coinfection in the RG and SG was PCV2/PRRSV (17.8% and 10.5%), while the most frequent coinfections involving PPV2 in the RG were PPV2/PCV2 (7.5%) and PPV2/PRRSV (4%) and PPV2/PCV2 (5.3%) in the SG. The most common triple coinfection was PPV2/PCV2/PRRSV at 15% in the RG and 21% in the SG, while quadruple coinfection PVV2/PCV2/PCV3/PRRSV was detected only in the RG (5.5%). Histopathological evaluation of 21 PPV2-positive lungs showed variable degrees of histiocytic or lymphohistiocytic interstitial pneumonia (9%) in the RG, while no significant changes were observed in SG; in addition, neutrophilic bronchopneumonia was observed in 73.7% if cases evaluated. In situ hybridization-RNAScope® confirmed the presence of PPV2 within pulmonary lesions in 2/19 RG pigs, while no in situ detection was observed in the SG pigs. The phylogenetic evaluation of seven PPV2 sequences detected in Colombia was compared with another 102 reported sequences, indicating that the Colombian strains are located in clade 2. Our results confirm the presence of PPV2 in pigs with PRDC alone and pigs coinfected with PCV2, PCV3, and PRRSV. Likewise, its presence alone or in coinfection in stillbirths suggests that PPV2 is also involved in PRF.

Currently, PRRSV-1 causes a large number of clinical infections in Chinese swine herds, and the prevalence of new strains has presented great challenges. In this study, the novel PRRSV-1 strain SL-01 was isolated, with a genome length of 14,978 bp, and genetic evolution analysis revealed that it belonged to a new subtype branch. Sequence homology analysis showed that the strain was only 82.2%–86.7% identical to the current classical PRRSV-1 strains. In particular, the novel strain exhibited a unique deletion pattern in Nsp2. In addition, GP3 and GP4 of the SL-01 strain showed four consecutive amino acid deletions in the highly variable regions at amino acids 243–248 and 63–68, respectively. Further challenges in piglet and pregnant sow demonstrated that the SL-01 strain could cause the piglet fever and death but less pathogenic to pregnant sow. Overall, the characterization and pathogenicity of a novel PRRSV-1 strain were first explored and provide a prevention for pig farms.

Whereas bovine has been demonstrated as the main reservoir of influenza D virus (IDV), this virus was first isolated in a pig and is regularly detected in some swine populations. However, the role of swine in IDV ecology, as well as the outcomes of IDV infection in pigs, is still unclear. This study aimed to provide additional information on pathogenesis, transmission, and adaptation of a bovine-origin IDV in swine. An infection and transmission study, using an IDV strain isolated following a first passage on pig of a bovine IDV, was conducted on specific pathogen-free pigs, including inoculated and direct contact pigs. Two routes of inoculation were tested, i.e., nasal and tracheal. None of the inoculated or their contact pigs showed clinical signs, but all of them shed the virus in nasal secretions and seroconverted. Virus shedding started earlier in pigs inoculated intranasally as well as in their contact pigs, compared to pigs inoculated intratracheally and associated contacts, suggesting that the viral replication occurred preferentially in the upper respiratory tract. Sequencing data brought to light a mutation on hemagglutinin-esterase-fusion protein (L118F) in the bovine IDV-derived isolate obtained after the first passage on pig. This mutation was fixed in all viral strains obtained in this study, either from inoculated or contact pigs, and was maintained over the second and third passages on swine. The L118F mutation could be linked to the adaptation of the parental bovine IDV to the swine host and might have contributed to an efficient viral multiplication and subsequent pig-to-pig transmission.

H5N8 HPAI is a highly infectious avian disease that now poses a serious threat and potential risk to poultry farming, wild birds, and public health. In this study, to investigate the seasonality and transmission directionality of global H5N8 HPAI, the spatial and temporal analysis of H5N8 HPAI was conducted using time series decomposition and directional distribution analysis. An ecological niche model was developed for H5N8 HPAI in poultry to identify areas at high risk of H5N8 HPAI in poultry and associated risk factors. The results indicated that three global pandemics of H5N8 HPAI emerged from 2014 to 2022, all showing a southeast–northwest distribution direction. H5N8 HPAI occurred more frequently in winter and less frequently in summer. The southwestern border region and the southeastern region of North America, the southern region of South America, most of Europe, the southern border region and the northern border region of Africa, and the southwestern region and the southeastern region of Asia provide the suitable environment for the occurrence of H5N8 HPAI in poultry. Chicken density, duck density, population density, bio1 (annual mean temperature), and land cover were considered important variables for the occurrence of H5N8 HPAI in poultry. This study can help optimize the use of resources and provide new information for policymakers to carry out prevention and control efforts.