Biology subjects

Dixon, L. K.

Publications and source records attributed to Dixon, L. K..

4 recordsLinked to original sources

bioRxiv2024

Exploring virus-host interactions through combined proteomic approaches identifies BANF1 as a new essential factor for African Swine Fever Virus.

African swine fever virus (ASFV) causes a highly lethal disease in pigs and represents a significant threat to the global pork industry due to the lack of effective vaccines or treatments. Despite intensive research, many ASFV proteins remain uncharacterized. This study aimed to elucidate the functions of two ASFV proteins, MGF360-21R and A151R, through comprehensive analysis of their interactions with host proteins. Using affinity purification-mass spectrometry and yeast two-hybrid screening approaches, we identified the host protein barrier- to-autointegration factor 1 (BANF1) as a key interactor of both viral proteins. Biochemical and colocalization assays confirmed these interactions and demonstrated that MGF360-21R and A151R expression leads to cytoplasmic relocalization of BANF1. Functionally, BANF1 silencing significantly reduced ASFV replication, indicating its proviral role. Given BANF1s established function in regulating the cGAS/STING-dependent type I interferon (IFN-I) response, we postulated that A151R and MGF360-21R could inhibit this pathway. Using different strategies, we showed that both A151R and MGF360-21R did indeed inhibit IFN-I induction. Generation of ASFV deficient of A151R or MGF360-21R showed that both mutant viruses enhanced the host IFN response in primary porcine macrophages compared to wild-type virus. However, their capacity to inhibit this pathway could occur through mechanisms independent of BANF1. Proteomic analysis of BANF1 interactors during ASFV infection highlighted potentially roles in chromatin remodeling, nuclear transport, and innate immune response pathways. Altogether, our data provide new insights into ASFV-host interactions, identifying BANF1 as an important new host factor required for replication and uncovering novel functions for A151R and MGF360-21R. Author SummaryAfrican swine fever virus (ASFV) is a highly contagious and deadly disease affecting pigs worldwide, for which there are currently no effective vaccines or treatments. Despite extensive research, many ASFV proteins remain poorly understood. Our study investigated two ASFV proteins, MGF360-21R and A151R, to better understand their functions and interactions with host proteins. Using proteomic approaches, we found both these viral proteins interact with a host protein called barrier-to-autointegration factor 1 (BANF1). Importantly, BANF1 silencing significantly reduced ASFV replication, indicating its important role in the viral life cycle. We also showed that MGF360-21R and A151R help the virus evade the immune system by blocking the production of interferons, which are key defensive molecules against viral infections. However, this immune evasion does not seem to depend on their interaction with BANF1. Additionally, our analysis of BANF1s interactions during ASFV infection revealed potential roles in chromatin remodeling, nuclear transport, and the innate immune response. These findings provide new insights into how ASFV interacts with its host and highlight BANF1 as a critical factor in viral replication and immune evasion. Our work contributes to a better understanding of ASFV and could pave the way for developing more effective strategies to fight this virus.

microbiology↗

bioRxiv2023

Full genome sequence analysis of African swine fever virus isolates from Cameroon

African swine fever is a devastating disease of domestic pigs that has spread across the globe since its introduction into Georgia in 2007. The etiological agent is a large double-stranded DNA virus with a genome of 170 to 180 kb in length depending on the isolate. Much of the differences in genome length between isolates are due to variations in the copy number of five different multigene families that are encoded in repetitive regions that are towards the termini of the covalently closed ends of the genome. Molecular epidemiology of ASFV is primarily based on Sanger sequencing of a few conserved and variable regions, but due to the stability of the dsDNA genome changes in the variable regions occur relatively slowly. Observations in Europe and Asia have shown that changes in other genetic loci can occur and that this could be useful in molecular tracking. ASFV has been circulating in Western Africa for at least forty years. It is therefore reasonable to assume that changes may have accumulated in regions of the genome other than the standard targets over the years. At present only one full genome sequence is available for an isolate from Western Africa, that of a highly virulent isolate collected from Benin during an outbreak in 1997. In Cameroon, ASFV was first reported in 1981 and outbreaks have been reported to the present day and is considered endemic. Here we report three full genome sequences from Cameroon isolates of 1982, 1994 and 2018 outbreaks and identify novel single nucleotide polymorphisms and insertion-deletions that may prove useful for molecular epidemiology studies in Western Africa and beyond.

microbiology↗

bioRxiv2023

Inefficient transmission of African swine fever virus to sentinel pigs from environmental contamination under experimental conditions

Knowledge about African swine fever virus (ASFV) transmission and its survival in the environment is mandatory to develop rational control strategies and combat this serious disease in pigs. In this study, the risk that environmental contamination poses for infection of naive pigs was investigated. Naive pigs were introduced as sentinels into contaminated pens either on the same day or up to three days after ASFV-infected pigs were removed. Three experiments were carried out in which four to six pigs per pen were inoculated with virulent ASFV isolates OURT88/1 (genotype I), Georgia 2007/1 or POL/2015/Podlaskie (genotype II), respectively. The majority of the inoculated pigs developed acute disease but with no evident haemorrhagic lesions or haemorrhagic diarrhoea and were culled at the predefined humane endpoint. The levels of ASFV DNA detected in the blood of the infected animals reached 107-9 genome copies/ml before euthanasia. Environmental swabs were taken from different surfaces in the animal rooms, as well as from faeces and urine, close to the time of introduction of the naive animals. Relatively low quantities of virus DNA were detected in the environmental samples, in the order of 103-7 genome copies. Neither clinical signs nor virus genomes were detected in the blood of any of the sentinel pigs over a period of two to three weeks after exposure, indicating that transmission from the ASFV-contaminated environment did not occur. Interestingly, viral DNA was detected in nasal and oral swabs from some of the sentinel animals at early days of exposure (ranging between 103.7-5.8 genome copies), though none of them developed ASF. The results indicate a relatively low risk of ASFV transmission from a contaminated environment in the absence of blood from infected animals.

microbiology↗

bioRxiv2023

Deletion of the gene for the African swine fever virus BCL-2 family member A179L increases virus uptake and apoptosis, but decreases virus spread in macrophages and reduces virulence in pigs

African swine fever virus encodes proteins that inhibit apoptosis including one member of the BCL-2 family, A179L. Deletion of the A179L gene from the virulent genotype I isolate Benin 97/1 compared to Benin 97/1 expressing A179L or mock-infected macrophages, resulted in increased Caspase 3 and 7 activity, annexin V binding to surface phosphatidyl serine and DNA fragmentation, measured by terminal deoxynucleotidyl transferase nick-end labelling. These results confirmed that apoptosis was induced earlier in macrophages infected with the Benin{Delta}A179L virus. Increased cell entry of the A179L gene-deleted virus was indicated at early times since up to double the numbers of cells expressed fluorescent protein from the virus genome. Yields of infectious virus were similar over a single cycle but were significantly lower for the A179L gene-deleted virus over a multi-step growth cycle. Pigs immunised and boosted with the Benin{Delta}A179L virus showed no clinical signs, although a weak cellular response to ASFV was observed showing that the virus had replicated. The immunised pigs were not protected against challenge with the virulent parental virus Benin 97/1 although viremia was lower at 3 days post-challenge compared to the control non-immune pigs. The reduced levels of virus replication in macrophages probably limited induction of a protective immune response. The results show an important role for the A179L protein in virus replication in macrophages and virulence in pigs. IMPORTANCEAfrican swine fever virus (ASFV) causes a lethal disease of pigs that has spread extensively in Africa, Europe and Asia. The virus codes for more than 150 proteins, many of which help the virus to evade the hosts defences following infection. We investigated the effect of deleting one of these genes, A179L, from the genome of an ASFV isolate that causes death of infected pigs. A179L belongs to the BCL-2 protein family, consisting of members which promote or inhibit apoptosis with A179L belonging to the latter. Deleting the A179L gene reduced ASFV replication and spread between macrophages, its main target cells. This was correlated with an increase in cell death. Pigs infected with the virus with A179L gene deleted did not show signs of disease and no virus replication was detected in blood. A low immune response was generated but the immunised pigs were not protected when challenged with the parental deadly virus. The results show that the A179L gene is important for ASFV to replicate efficiently in cells and in animals.

microbiology↗