Progress 10/01/23 to 09/30/24
Outputs
PROGRESS REPORT Objectives (from AD-416): Objective 1. Develop CCHF virus diagnostic tests for the early detection and surveillance of Crimean-Congo Hemorrhagic Fever Virus. Develop viral detection methods for ticks. Develop direct viral detection methods for cattle, sheep, and goats. Develop antibody detection methods to determine CCHF exposure in cattle, sheep, goats, and relevant wildlife. Objective 2. Determine mechanisms of CCHF transmission. Develop CCHF tick and animal infection methods. Develop CCHF tick-animal transmission models. Objective 3. Investigate the epidemiology of CCHF and the role of ticks in maintaining reservoirs of infection in endemic settings. Determine the prevalence of CCHF in vector and animal hosts in endemic areas. Determine the competence of hard tick species in endemic areas. Determining the risk for the establishment of tick host vector in the United States considering climatic and ecological conditions. Approach (from AD-416): The research addresses the following research components in the 2022-2027 Animal Health National Program (NP 103) Action Plan: 1) Component 1: Biodefense, Problem Statement 1A, Control and eradicate foreign animal diseases. The research addresses ARS Strategic Plan Goal 4.3 and the following Performance Measure: Provide scientific information to protect animals, humans, and property from the negative effects of pests and infectious diseases, and develop and transfer tools to the agricultural community, commercial partners, and government agencies to control or eradicate domestic and exotic diseases and pests that affect animal and human health. Substantial progress was made through various collaborative projects. Ways to detect the presence of the Crimean Congo Hemorrhagic Fever virus (CCHFV) and evidence of exposure to the virus are in development or have been developed, including assays that can be used on the farm or with little equipment or infrastructure. A gap analysis of CCHFV with experts from over a dozen countries raised concerns about the availability and quality of assays to assess if an animal or person had previously been exposed to CCHFV. Working with two partners, assays were developed to detect the presence of Immunoglobulin G (IgG) as a marker for last exposure. The assays detect the presence of IgG in a suspension and use technology that is reported to be more specific and more sensitive than the technology used by current assays. These assays are also designed to detect exposure to other viruses that may be confused with CCHFV. To mitigate risk a second approach using arrays of peptides has been started. Peptides that cover different CCHFV isolates, and other closely related viruses are coated onto arrays. Samples from CCHFV-exposed animals and people, as well as samples from uninfected animals and animals infected with other viruses, are placed on the arrays to identify specific parts of the virus the immune system responds to. Once identified, the most reactive peptides are targeted for assay development. Two separate efforts are underway to aid in the identification of ticks. The first uses a matrix-assisted laser desorption/ionization time-of- flight mass spectrometry (MALDI-TOF) to identify a unique signature or fingerprint based on the protein content of the tick. This approach may also identify if a tick is carrying CCHFV. Ticks collected as part of the Centers for Disease Control Center for the Surveillance of tick-borne diseases in the Southern States have been utilized to develop tick species databases. Thirteen different species of ticks have been collected from field sites in the United States, and permits have been put in place to obtain ticks from international locations. This system will assist in efforts to monitor the changing geographic ranges or distributions of tick populations, data essential for accurate risk assessments and model development. In collaboration with partners, work was performed to establish Amblyomma variegatum and Hyalomma spp. colonies at partner locations. Efforts to investigate the impact of climate change on key ticks' host-seeking behavior using a behavioral assessment tool have been initiated. Of note, this is the first time such a system has been used in biocontainment. Tick and animal sera collections are being conducted in several locations, including Tanzania, Sierra Leone, the Democratic Republic of Congo (DRC), Uganda, Ethiopia, and Kenya. In Tanzania and Sierra Leone, a total of 54 cattle herds were sampled, 7,204 ticks were collected, and 89 milk samples were collected. Testing samples for previous exposure to Crimean Congo Hemorrhagic Fever virus (CCHFV) demonstrated a higher than anticipated seroprevalence rate with all herds having at least one positive animal and some herds at 100% positive. The seroprevalence was generally higher in the north and east of Sierra Leone. Testing of the ticks collected failed to detect the virus itself. Additional work is required but in general, results indicate widespread CCHFV exposure and infection at study sites with a trend towards age-related acquisition of anti-CCHFV antibodies. This finding reveals (at least for this limited sample set and preliminary laboratory testing) an apparent lack of association between herd-level evidence of CCHFV exposure and the presence of Hyalomma spp. ticks carrying CCHFV. In Kenya and Uganda, in-country partnerships were established with the Uganda Virus Research Institute and the Kenya Medical Research Institute to collect ticks and detect CCHFV in and outside the cattle corridor. Approximately 180 homesteads across 6 Ugandan districts across the cattle corridor have been sampled to date. Uganda tick populations were shipped to Texas Tech University and screened for CCHFV. Two pools of Ugandan Rhipicephalus appendiculatus tick samples tested positive for CCHFV suggesting infection. Collection efforts suggest Hyalomma species are not prevalent in central Uganda, where previous outbreaks of CCHF have been reported. In Kenya, Hyalomma species were observed to be the predominant species collected in Kenyan pastoralism. Microsatellite genetic markers for Rhipicephalus appendiculatus have been optimized, and tick genotyping is in progress. Metatranscriptome and microbiome data processing of select Ugandan tick populations have been completed and data analysis is ongoing. The U.S. National Tick Collection at Georgia Southern University in Statesboro, Georgia (U.S.A.), cooperated with facilities in Salford (United Kingdom), and Makerere University, Kampala, (Uganda) to collect ticks to test for CCHFV and other pathogens. The main vector, Hyalomma ticks, is found only sporadically in the west. However, desertification from human use and global climate change may be changing the habitats and allowing for vector expansion to new areas. Ticks of different genera collected from domestic/wild animals and by dragging the vegetation among the rural communities inside and around Queen Elizabeth National Park were evaluated to survey for tick-borne pathogens and to assess the importance of the progression CCHFV, other vectors, and pathogens. In the DRC, multi-institutional partnerships were established, including with the local National Research and National Veterinary programs. Between July 2023 and June 2024, the ARS research team visited and completed sample collections in 9 provinces. 125 individual farms, slaughterhouses, and community farms were visited, and samples from over 1,000 cattle and 1,000 pigs were collected. As part of the work in the DRC, data was collected to inform agriculture husbandry practices. Approximately 60% of sites with cattle and sites with swine reported using any veterinary pharmaceuticals or dietary supplements. Only 22% of sites with cattle and 24% of sites with swine reported vaccinating their animals against any disease. Veterinary pharmaceuticals used in the DRC are commonly contributing to increased antimicrobial resistance regionally. Tetracycline antibiotics (e.g., oxytetracycline) are noted as the most frequently used therapeutic and prophylactic pharmaceuticals across the study area. The most common antibiotics used were oxytetracycline and a combination procaine penicillin G- dihydrostreptomycin sulfate drug. The most common antiparasitic agents reported were ivermectin and levamisole. Investing in proactive veterinary medicine by increasing vaccination rates has the potential for long-term national cost savings. Of note is that about half of the sites visited report notifying health officials about disease outbreaks. Approximately 50% of sites with cattle and 40% of sites with swine responded that they have reported an animal illness or death due to disease to health zones or province officials at least once. Work was initiated in Nigeria with partners at Redeemers University. Nigeria is a hub with cattle migrations from Cameroon, Niger, Burkina Faso and Chad. The border is considered porous with an estimated 1500 unofficial border crossings. In Nigeria, evidence suggests that large animals such as cattle, donkeys, and camels are mainly brought in through Borno, Sokoto, Katsina, Adamawa, and Taraba states from Eastern and central African countries. The movement and trade of animals along these borders provide a route by which numerous pathogens can come together. Coupled with the inefficient quarantine and screening of incoming animals, people and animals can be exposed to zoonotic and transboundary pathogens, where novel pathogens can quickly amplify and spread internationally, posing health risks. Samples were collected from ill or deceased animals from 3 study sites in Northern Nigeria: Kano, Sokoto, and Adamawa states. Additionally, 300 tick vectors will be collected from sampled. Partners at the Navy Medical Entomology unit initiated tick surveillance efforts in the Republic of Georgia. Next-generation metagenomic sequencing of 389 ticks was performed, and a bioinformatics pipeline was constructed to analyze the generated sequence data. Partners at the Broad Institute developed a pan-viral open reading frame (ORFs) discovery platform using Massively Parallel Ribosome Profiling (MPRP), which identified 5,381 ORFs in approximately 700 human viruses, including 4,208 noncanonical ORFs. Currently we are working on experiments to identify these ORFs and evaluate their transition in the context of viral infection. These previously unidentified ORF may shed light on how these agents cause disease and may inform vaccine development. Partners at the Broad Institute identified potential vaccine candidates. The team identified parts of CCHFV that we believe are most important to the immune system for preventing infection or disease. Expression of these targets and delivery systems were optimized. Immunogenicity studies in mice were initiated. A mini-genome system was established to aid in the study of different CCHFV isolates and parts of the virus. A reporter gene, firefly luciferase, was incorporated into CCHFV genotypes 2 and 3. The inclusion of a reporter gene will facilitate in vitro work. Based on these results, experiments to rescue CCHFV from full-length infectious cDNA clones have been initiated. To aid in this, we have identified and tested several reagents (anti-CCHFV glycoprotein antibodies), that can be used to confirm replicating virus following infection. Artificial Intelligence (AI)/Machine Learning (ML) A partnership was established with VecTec and ARS researchers at the National Bio and Agro-Defense Facility (NBAF) in Manhattan, Kansas, to evaluate the use of AI to identify ticks. The instrument has been deployed to the Democratic Republic of the Congo and will be sent to Liberia in August 2024. Currently, 1600 ticks have been evaluated by the instrument and local technicians. The data collected is being utilized to refine algorithms. In preparation for the use of animal studies at NBAF, a collaboration with National Institute of Allergy and Infectious Diseases (NIAID) in Frederick, Maryland, has been focused on using AI-based methods to improve whole-slide histopathologic analysis using materials from well characterized studies performed with BSL4 pathogens. In this collaboration, we have developed tools to segment glomeruli from variously stained whole-slide images on multiple animal models, including guinea pigs and ferrets. We have also used the concept of separability to show that infection with Ebola virus induces visible changes in the glomeruli of infected animals. In a similar analysis, we used the concept of separability to investigate the differences in the pathology of rhesus monkeys infected with the Marburg virus and two variants of the Ebola virus (Kikwit and Makona). We investigated many methods of using class activation mapping to highlight areas where there were histopathologic differences in animals infected with these three viruses, but class activation mapping was unable to show these histopathologic differences reliably. To have AI help identify differences in the pathology of these viruses, we developed a new methodology to show differences between two groups of histopathologic slides quickly. To do this, we trained a model to work with extremely small patches of the whole-slide image. Then we used the concept of separability and confidence to acquire the patches of the slide most strongly associated with each class. Grids of these images proved far more useful for histopathologic analysis and discovery than class activation maps. We are currently producing pipelines of this method to quickly have AI help identify subtle differences between sets of histopathologic samples. We anticipate that this will be extremely helpful in pointing out treatment-related differences in histopathology or differences in histopathologic changes induced by closely related viruses. ACCOMPLISHMENTS 01 Deployment and refinement of AI tool for identification. An AI based instrument for the identification of ticks was deployed and 1600 ticks in the Democratic Republic of the Congo have been evaluated in partnership with ARS researchers in Manhattan, Kansas. The algorithm is under refinement and the instrument will be deployed to Liberia and Turkey. 02 Collection of samples from livestock and suspect animals. In partnership with ARS researchers in Manhattan Kansas samples from livestock have been collected from the Democratic Republic of the Congo, Sierra Leone, Tanzania, Uganda and Nigeria. Samples from animals with obvious signs of disease have been collected from the Democratic Republic of the Congo and Nigeria. These samples will inform the distribution of CCHFV to all improved risk assessment. Sequencing will provide critical information to all the development of diagnostics and countermeasures with the broadest specificity. 03 Development of bioinformatics pipeline. A bioinformatics pipeline for evaluation of sequencing data for the detection of Crimean-Congo hemorrhagic fever virus (CCHFV) has been developed in partnership with ARS researchers in Manhattan, Kansas. This platform will be made publicly available and will facilitate the analysis of genetic diversity. This information will facilitate the development of diagnostics and countermeasures with the broadest specificity. 04 Initiation of tick collections in Kenya, Uganda, Sierra Leone, Liberia, Republic of Georgia, Tanzania. Ticks have been collected from Kenya, Uganda, Sierra Leone, Liberia, Republic of Georgia, Tanzania and the Democratic Republic of the Congo in partnership with ARS researchers in Manhattan, Kansas. Ticks from Uganda and Sierra Leone have been tested for Crimean-Congo hemorrhagic fever virus. Positive samples have been identified from Uganda. These samples will inform the distribution of CCHFV to all improved risk assessment. Sequencing will provide critical information to all the development of diagnostics and countermeasures with the broadest 05 Development and down selection of lead mRNA vaccine candidates for immunogenicity testing. Multiple mRNA vaccine candidates were developed and lead candidates are undergoing immunogenicity testing Congo in partnership with ARS researchers in Manhattan, Kansas. If successful the vaccine may be able to interrupt transmission and spread of Crimean Congo Hemorrhagic Fever virus.
Impacts
(N/A)
Publications
|
Progress 10/01/22 to 09/30/23
Outputs
PROGRESS REPORT Objectives (from AD-416): Objective 1. Develop CCHF virus diagnostic tests for the early detection and surveillance of Crimean-Congo Hemorrhagic Fever Virus. Develop viral detection methods for ticks. Develop direct viral detection methods for cattle, sheep, and goats. Develop antibody detection methods to determine CCHF exposure in cattle, sheep, goats, and relevant wildlife. Objective 2. Determine mechanisms of CCHF transmission. Develop CCHF tick and animal infection methods. Develop CCHF tick-animal transmission models. Objective 3. Investigate the epidemiology of CCHF and the role of ticks in maintaining reservoirs of infection in endemic settings. Determine the prevalence of CCHF in vector and animal hosts in endemic areas. Determine the competence of hard tick species in endemic areas. Determining the risk for the establishment of tick host vector in the United States considering climatic and ecological conditions. Approach (from AD-416): The research addresses the following research components in the 2022-2027 Animal Health National Program (NP 103) Action Plan: 1) Component 1: Biodefense, Problem Statement 1A, Control and eradicate foreign animal diseases. The research addresses ARS Strategic Plan Goal 4.3 and the following Performance Measure: Provide scientific information to protect animals, humans, and property from the negative effects of pests and infectious diseases, and develop and transfer tools to the agricultural community, commercial partners, and government agencies to control or eradicate domestic and exotic diseases and pests that affect animal and human health. Substantial progress was made in addressing the objectives of this research project through various collaborative projects. In diagnostics, molecular assay for the S and M segments were developed, validated, and transitioned to the field. These assays will detect the presence of the viral ribonucleic acids and allow investigators to determine if Crimean- Congo Hemorrhagic Fever (CCHF) virus is replicating. The laboratory developed and validated field deployable sequencing protocols and algorithms that will reduce errors and allow the detection of recombination events. Combinatorial arrayed reactions for multiplexed evaluation of nucleic acids (CARMEN) assays have been developed and evaluated for CCHF virus. Efforts were initiated to investigate the performance of existing serologic assays to evaluate their specificity and overall performance. Efforts to use a proteomics pipeline to identify tick species and tick- borne pathogens using matrix-assisted laser desorption/ionization time-of- flight mass spectrometry (MALDI-TOF) were initiated. If successful, the proteomics platform will identify the tick species and CCHF virus in ticks in a high throughput setting. Current efforts focus on collecting necessary materials, supplies, and equipment. This system will assist in efforts to monitor the changing geographic ranges or distributions of tick populations, data essential for accurate risk assessments and model development. In collaboration with partners, work was performed to establish Amblyomma variegatum and Hyalomma spp. tick colonies (specifically H. asiaticum, H. anatolicum, H. dendriticum, H. marginatum, H. excavatum, H. rufipes). Animal feeding will commence in September. Efforts to investigate the impact of climate change on the host-seeking behaviors of key ticks using a behavioral assessment tool have been initiated. Of note, this is the first time such a system has been used in biocontainment. Tick and animal sera samples are being conducted in several locations, including Tanzania, Sierra Leone, the Democratic Republic of Congo (DRC), Uganda, Ethiopia, and Kenya. Regional instabilities have impacted the collection of materials in some locations. In Tanzania and Sierra Leone, 54 cattle herds were sampled, 7,204 ticks and 89 milk samples were collected. Collected tick species included: Amblyomma, Hyalomma, Haemaphysalis, Boophilus and Rhipicephalus. Testing samples for antibodies to Crimean Congo Hemorrhagic Fever virus demonstrated a higher than anticipated seroprevalence rate, with all herds having at least one positive animal and some herds at 100% positive. The seroprevalence was generally higher in the North and east of Sierra Leone. Testing of the ticks collected failed to detect CCHF viral RNA. Additional work is required to confirm the serological and molecular results. Still, results generally indicate widespread CCHF virus exposure and infection at study sites with a trend towards age-related acquisition of anti-CCHFV antibodies. In Kenya and Uganda, in-country partnerships were established with the Uganda Virus Research Institute (UVRI) and the Kenya Medical Research Institute (KEMRI) to collect ticks and detect CCHFV in and outside the cattle corridor. The distribution of potential vectors for Crimean-Congo Hemorrhagic Fever Virus and CCHFV prevalence in the area will be determined. So far, around 180 homesteads across 6 Ugandan districts across the cattle corridor have been sampled. Tick species identification is ongoing. No Hyalomma adults have been identified/collected yet but will likely be collected during the next sampling season. Most ticks from cattle identified so far belong to Rhipicephalus decoloratus. DNA and RNA isolation protocols for pooling and testing ticks for CCHF virus have been tested, optimized, and established. Molecular detection of Crimean- Congo Hemorrhagic Fever Virus in the ticks collected will start in the next few months once the ticks are shipped to Texas Tech (expected for mid-August). In the DRC, multi-institutional partnerships were established, including with the local National Research and National Veterinary programs. A preliminary landscape analysis revealed little surveillance on infectious diseases among livestock in DRC and very limited reporting structures for adverse events. Cattle and pig density were mapped, preliminary study sites were identified, and collections were initiated. New partnerships were established to facilitate monitoring tick populations in multiple regions and potential importation to the United States. Other partnerships established in 2023 include developing and using artificial intelligence for tick identification and improved low- complexity molecular diagnostics. ACCOMPLISHMENTS 01 Completion of serosurveys. ARS researchers at the National Bio Agro- Defense Facility in Manhattan, Kansas worked with partners in Sierra Leone and Tanzania to complete an expanded serosurvey. Sample collection was initiated in Uganda, Kenya, and the Democratic Republic of the Congo. This serosurvey provided the first data characterizing the presence of Crimean Congo hemorrhagic fever virus in the region. Critical is the finding that the virus may be much more prevalent than previously suspected. This data is essential for local ministries of health and agriculture. The high seroprevelance indicates that persons working closely with livestock in this region are at risk for exposure to this potentially deadly disease and Crimean Congo hemorrhagic fever should be included in the differential diagnosis in patients with unexplained illness in the region. 02 Training of in country teams. International field teams at the Sokoine University of Agriculture (Tanzania) and the University of Makeni (Sierra Leone) worked with ARS researchers from the National Bio Agro- Defense Facility in Manhattan, Kansas, and received training in tickborne disease surveillance and Crimean Congo hemorrhagic fever virus, molecular and serologic testing. The training was also completed in the Democratic Republic of the Congo using Research Electronic Data Capture (RedCap), sample collection and processing, and health assessment of animals. These capacity-building efforts are critical to ensure the rapid and accurate diagnosis of Crimean Congo hemorrhagic fever virus and other zoonotic threats in the region. Training staff to operate safely in the laboratory is another essential aspect of capacity building. Enhancing laboratory safety and capacity in the region will impact far-reaching improvements across public and animal health. The introduction of improved electronic data capture will improve the research quality in the region. The research electronic data capture is a widely used program that, once learned, can be applied to other studies. The use of this program ensures that metadata is readily linked to samples. This protects the data quality and integrity and aids in building a repository of well-characterized samples that may be of future use. 03 Validation and field testing of molecular assays. An antisense reverse transcription-polymerase chain reaction test (RTPCR) assays for the M and S segments of Crimean Congo hemorrhagic fever virus was validated in the labratory and transferred to the field for additional validation. This work was completed between ARS researchers at the National Bio and Agro-Defense Facility in Manhattan, Kansas, and its partners. These tests could improve the diagnosis of Crimean Congo hemorrhagic fever virus. Currently, there are limited assays available and of uncertain quality. The lack of diagnostics limits the ability to test for Crimean Congo hemorrhagic fever virus. If validated, these tests will have wide utility in human and animal health. 04 Development of proteomics and next-generation sequencing protocols. New protocols will reduce the error rate and allow the identification of recombinants. This data will be essential for characterizing the viral diversity and viral evolution of Crimean Congo hemorrhagic fever virus. Viral diversity is a challenge for molecular diagnostics. Even minor changes in sequences can significantly impact the performance of assays, potentially compromising their utility. Monitoring of diversity will also be essential for countermeasure development and risk assessment. Developing accurate field deployable strategies will be done between ARS researchers at the National Bio and Agro-Defense Facility in Manhattan, Kansas, and its partners, which will facilitate monitoring in even the most remote regions. The data generated from these efforts will be critical for modeling, transmission, and countermeasure efforts.
Impacts
(N/A)
Publications
|
Progress 10/01/21 to 09/30/22
Outputs
PROGRESS REPORT Objectives (from AD-416): Objective 1. Develop CCHF virus diagnostic tests for the early detection and surveillance of Crimean-Congo Hemorrhagic Fever Virus. Develop viral detection methods for ticks. Develop direct viral detection methods for cattle, sheep, and goats. Develop antibody detection methods to determine CCHF exposure in cattle, sheep, goats, and relevant wildlife. Objective 2. Determine mechanisms of CCHF transmission. Develop CCHF tick and animal infection methods. Develop CCHF tick-animal transmission models. Objective 3. Investigate the epidemiology of CCHF and the role of ticks in maintaining reservoirs of infection in endemic settings. Determine the prevalence of CCHF in vector and animal hosts in endemic areas. Determine the competence of hard tick species in endemic areas. Determining the risk for the establishment of tick host vector in the United States considering climatic and ecological conditions. Approach (from AD-416): The research addresses the following research components in the 2022-2027 Animal Health National Program (NP 103) Action Plan: 1) Component 1: Biodefense, Problem Statement 1A, Control and eradicate foreign animal diseases. The research addresses ARS Strategic Plan Goal 4.3 and the following Performance Measure: Provide scientific information to protect animals, humans, and property from the negative effects of pests and infectious diseases, and develop and transfer tools to the agricultural community, commercial partners, and government agencies to control or eradicate domestic and exotic diseases and pests that affect animal and human health. To address the objectives of this research project, collaborative projects were initiated with the University of Texas Medical Branch (UTMB) and the University of California Davis (UCD). To maximize funds at ARS in Manhattan, Kansas, UCD and UTMB teamed to capitalize on each teams unique expertise. UCD is leading the field collection of tick and sera samples from cattle, the local training, and capacity building for collection and serologic and molecular testing of samples. UTMB brings unique experience in the development of assays, next generation sequencing, as well as experience working in biosafety level-4. The partnership with another maximum containment laboratory is critical while location facilities are still coming online. The joint project directly addresses Objective 3: Investigate the epidemiology of Crimean Congo Hemorrhagic Fever virus (CCHFV) and the role of ticks in maintaining reservoirs of infection in endemic settings and Objective 2. Determine mechanisms of CCHFV transmission and proteomics will be performed on vertebrate and tick cells infected with CCHFV, to compare the expression profiles and identify potential pathogenesis and maintenance pathways. In addition, the project will begin to address Objective 1: develop CCHFV diagnostic tests for the early detection and surveillance of CCHFV. Development of molecular assays including protocols for next generation sequencing will be performed as part of this effort. In addition, a commercially available assay will be compared to a gold standard in house research assay. Efforts to develop and validate assays to be used for research and diagnostic purposes are well advanced. An antisense reverse transcription polymerase chain reaction (RTPCR) assay for the S segment of CCHF has been developed and validated. A second assay for the M segment equivalent has been developed and is currently undergoing validation. The development of the anti-sense assay will be critical to demonstrate CCHF infection and replication. Protocols for next generation sequencing (NGS) using a field deployable Min-Ion have been optimized and are undergoing validation. We expect these protocols to be field tested in early 2022. The necessary equipment, reagents and controls have been acquired for the matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry (MS) platform, and protocol optimization has been completed. Lastly, efforts focused on tick host identification have down selected polymerase chain reaction (PCR) amplification combined with probe blotting as the most promising method. All reagents, equipment, and primers and probes needed were ordered. Despite travel restrictions and prioritization of COVID-19 related activities, substantial progress has been accomplished. Critical staff members were hired and trained at UCD and UTMB. Remote trainings with partner countries were performed. International field teams at the Sokoine University of Agriculture (Tanzania) and University of Makeni (Sierra Leone) received remote training in tickborne disease surveillance, CCHFV biology and safe collection and processing of samples. In Sierra Leone, a total of 4 study staff members and 8 government district livestock officers were also trained. In Tanzania, 4 staff members and 2 government livestock officers were trained. This training was essential for future efforts. A pilot seroprevalence survey of CCHFV in cattle was performed. Due to COVID-19 travel restrictions the work was primarily performed by in- country partners after receipt of remote virtual training. In total 754 cattle from 37 cattle herds were sampled (20 in Sierra Leone; 17 in Tanzania) including milk specimens from 89 animals.Teams have also collected 7,204 ticks from cattle. Tanzania cattle serology results indicate widespread CCHFV exposure and infection at study sites with a trend towards age-related acquisition of anti-CCHFV antibodies. Further testing and evaluation of a commercial enzyme linked immunosorbent assay (c-ELISA) (ID-VET) with current gold- standard ELISAs will be conducted at both Sierra Leone and Tanzania partner sites during the latter part of 2022. Other progress includes UCD obtaining the necessary USDA import permits. UCD is in in the process of establishing a Amblyomma variegatum as well as a Hyalomma marginatum and Hyalomma anatolicum colony. COVID-19 related travel delays and interruptions greatly impacted the UCD and UTMB team�s ability to travel to Sierra Leone and Tanzania to accomplish laboratory testing and training. The UCD team plans to resume travel and conduct PCR testing of these tick specimens in September to October of 2022. Despite these limitations significant progress was achieved. ACCOMPLISHMENTS 01 Completion of initial serosurvey. Collection of ticks and a serosurvey study was initiated. Samples were analyzed to determine the seroprevelance of Crimean Congo Hemorrhagic Fever virus (CCHFV) in Sierra Leone and Tanzania. This study was essential for down selection of future study sites and provides some of the first insights into CCHFV prevalence in these regions. Work was performed by in-country partners with remote support from researchers from ARS in Manhattan, Kansas, University of California Davis, and University of Texas Medical Branch. 02 Training of in-country teams. International field teams at the Sokoine University of Agriculture (Tanzania) and University of Makeni (Sierra Leone) received remote training in tickborne disease surveillance and Crimean Congo Hemorrhagic Fever virus (CCHFV), establishing their competency with the necessary skills and knowledge accomplish project goals. A total of 4 staff members and 8 District Livestock officers in Sierra Leone, and 4 staff members and 2 Livestock Officers in Tanzania were trained. Trainings were performed by University of California Davis and University of Texas Medical Branch staff in partnership with ARS scientists in Manhattan, Kansas. 03 Validation of anti-sense assay. An antisense reverse transcription polymerase chain reaction (RTPCR) essay for the S segment of Crimean Congo Hemorrhagic Fever virus (CCHFV) has been developed and validated at University of Texas Medical Branch (UTMB). A second assay for the M segment equivalent has been developed and is currently undergoing validation at UTMB. The development of the anti-sense assay will be critical to distinguish CCHFV infection and replication. Work was performed by University of California Davis scientists in partnership with ARS researchers in Manhattan, Kansas. 04 Development of proteomics and next generation sequencing protocols. Protocols for next generation sequencing (NGS) using a field deployable Min-Ion have been optimized and are undergoing validation. The necessary equipment, reagents and controls have been acquired for the Matrix-assisted laser desorption/ionization-time of flight (MALDI-Tof) platform and protocol optimization has been performed. Work was performed by University of California Davis scientists in partnership with ARS researchers in Manhattan, Kansas.
Impacts
(N/A)
Publications
- Rodriguez, S.E., Hawman, D.W., Sorvillo, T.E., O�Neal, T.J., Bird, B.H., Rodriguez, L.L., Feldmann, H., Bergeron, �., Nichol, S.T., Montgomery, J.M. , Spiropoulou, C.F., Spengler, J.R. 2022. Immunobiology of Crimean-Congo hemorrhagic fever. Antiviral Research. https://doi.org/10.1016/j.antiviral. 2022.105244.
|