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- Hensley, Lisa
- 2023
- 2022
- Animal Health (103)

Research Project: Viral Ecology of Henipaviruses in Endemic Settings and Intervention Strategies to Prevent their Spread to Domestic Animals **Location:
                            Zoonotic and Emerging Disease Research** 2023 Annual Report **Objectives**
                                

Objective 1.  Characterize the ecology of henipahviruses with a focus on the One-Health concept.  
- Conduct the molecular characterization of new and emerging henipahviruses, including phylogenetics, and network analysis.
- Examine known or emerging henipahviruses that may have an impact on animal agriculture, including their potential impact on public health.
Objective 2. Elucidate the host-pathogen interactions of henipahviruses infections. 
- Investigate virus-specific factors and viral molecular markers associated with infectivity, pathogenicity and transmissibility of henipahviruses in susceptible animal species including virus tissue tropism and replication. 
- Investigate host-specific factors associated with the infectivity, pathogenicity and transmissibility in different animal species.
- Characterize the innate and adaptive immune response to henipahviruses infections in animal models that are either susceptible, tolerant, or resistant to infection.
Objective 3.  Develop surveillance strategies and early warning systems for henipahviruses.
- Improve surveillance strategies to detect henipaviruses in high-risk countries.
- Establish a formal laboratory network for henipavirus surveillance that includes standardized specimen collection, laboratory testing scheme, quality control, specimen referral and accreditation. **Approach**
                                

Henipahviruses are members of the family Paramyxoviridae, order Mononegavirales. The name henipavirus was recommended for the genus of both Hendra virus and Nipah virus.  Henipaviruses have a large host range, unlike other members of the Paramyxoviridae, which generally have a very narrow host range. The cell attachment protein, unlike many other members for the paramyxovirus subfamily, does not have haemagglutinating activity and as a consequence does not bind sialic acid on the surface of cells.  The natural reservoir of the henipaviruses are fruit bats mainly from the genus Pteropus (flying foxes).  Nucleic acid and antibody signatures of exposure to Nipah virus or Nipah-like viruses has been documented in a diversity of bat species across the globe. The threat for a natural introduction of henipaviruses in the United States is low, but there is significant concern that henipaviruses could be used for nefarious purposes to harm agriculture and people. Both Hendra virus and Nipah virus are on the HHS and USDA list of overlap Select Agents and Toxins. Henipaviruses are listed as APHIS Tier 3 high-consequence foreign animal diseases and pests. Henipaviruses are promiscuous in their ability to cause severe morbidity in several animal species, including people, and human infections result in a very high mortality rate. The mortality rate associated with Nipah virus infections in pigs has been reported to be approximately 2.5% in adult pigs – high morbidity, but low mortality. Mortality rates in humans however are significantly higher and range from 40% (Malaysia) to 75% (up to 100%) in Bangladesh. The animal reservoir includes several species of bats, and henipaviruses may thus be readily available in these wildlife reservoirs. **Progress Report**
                                

Substantial progress was made in addressing the objectives of this project. To address Objective 1. Characterize the ecology of henipaviruses with a focus on the One-Health concept, including the characterization of new and emerging henipahviruses and their impact on animal agriculture, including their potential impact on public health partnerships, were established in regions where the bats that have been demonstrated to carry the virus are found. Samples will be collected in West and Central Africa, Puerto Rico, Bangladesh, and Indonesia. New collaborative agreements were established to facilitate sampling in Bangladesh, Indonesia, and Puerto Rico. Sample transport is expected to be difficult from Indonesia and Bangladesh and will require either working closely with established capacity or establishing capacity in the region. 
To begin to address Objective 2, the transmissibility of henipaviruses in susceptible animal species, including virus tissue tropism and replication, including the host-specific factors associated with the infectivity, pathogenicity, and transmissibility in different animal species, the innate and adaptive immune response to henipaviruses infections in animal models that are either susceptible, tolerant, or resistant to infection strategies to perform single-cell sequencing in maximum containment have been developed. Single-cell sequencing will allow scientists to map changes in the host responses and changes in the virus and work around the challenges of limited reagents for different species. This approach will be implemented with partners to characterize host and viral changes in available models and samples. As part of this work, samples will be collected to allow detailed mapping of immune responses. This information will inform vaccine development and identification of critical epitopes. To better understand Nipah virus pathogenesis, a partnership was developed to use artificial intelligence to facilitate pathologic analysis. The algorithms developed will hopefully identify commonalities and differences in susceptible models.
Researchers at the National Bio and Agro-Defense Facility (NBAF), in Manhattan, Kansas, established partnerships will also facilitate addressing Objective 3. Develop surveillance strategies and early warning systems for henipaviruses, including improving surveillance strategies to detect henipaviruses in high-risk countries and establishing a formal laboratory network for henipavirus surveillance that includes standardized specimen collection, laboratory testing scheme, quality control, specimen referral and accreditation. Sensitive, specific, and easy-to-use diagnostics are essential for surveillance activities. First-generation combinatorial arrayed reactions for multiplexed evaluation of nucleic acids (CARMEN) molecular assays have been developed and will be further tested this fall in partner laboratories.
Lastly, in anticipation of work with swine at NBAF, experimental work with swine  has been initiated with partners. Initial studies were performed with pigs and Monkeypox (MPOX). MPOX was selected from available viruses as risk from accidental exposure to the virus could be minimized with an approved vaccine. In addition, the experiment allowed USDA scientists to address critical questions regarding the risk to U.S. livestock from a rapidly spreading zoonotic virus. The studies demonstrated that pigs are susceptible to mpox virus and could transmit the virus even without significant clinical disease. The work also allowed staff to gain experience in containment and challenges they may encounter. **Accomplishments**
                                

1. Development of single cell sequencing techniques. ARS researchers in Manhattan, Kansas, worked with partners to develop single-cell sequencing techniques suitable for use in laboratories. The technique is essential for characterizing the host response to viral infection and the potential genetic changes that may occur in pathogens in various host species, and will address a challenge that all investigators have with limited reagents. This protocol will be essential for evaluating vaccine responses and understanding host susceptibility, immunity, and transmissibility. This technique has the potential to be incorporated into all future studies and provide critical data for risk assessments and countermeasure development. 3. Development of first generation combinatorial arrayed reactions for multiplexed evaluation of nucleic acids (CARMEN). ARS researchers in Manhattan, Kansas, worked with partners to develop the first-generation combinatorial arrayed reactions for multiplexed evaluation of nucleic acids (CARMEN). A rapid, sensitive assay was developed using combinatorial arrayed reactions for multiplexed evaluation of nucleic acids (CRISPR)-based technology and the combinatorial arrayed reactions for multiplexed evaluation of nucleic acids (CARMEN) platform. The assay will be further tested this fall in partner laboratories. This technique will open the door for new diagnostics that are more field deployable, lower cost, and have the same or lower level of detection compared to current molecular assay using polymerase chain reaction. This technology also has the advantage that it can be rapidly developed, manufactured, and adjusted to address genetic variations or shifts.


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