Biology subjects

Lednicky, J. A.

Publications and source records attributed to Lednicky, J. A..

5 recordsLinked to original sources

bioRxiv2021

Niclosamide reverses SARS-CoV-2 control of lipophagy

The global effort to combat COVID-19 rapidly produced a shortlist of approved drugs with anti-viral activities for clinical repurposing. However, the jump to clinical testing was lethal in some cases as a full understanding of the mechanism of antiviral activity as opposed to pleiotropic activity/toxicity for these drugs was lacking. Through parallel lipidomic and transcriptomic analyses we observed massive reorganization of lipid profiles of infected Vero E6 cells, especially plasmalogens that correlated with increased levels of virus replication. Niclosamide (NIC), a poorly soluble anti-helminth drug identified for repurposed treatment of COVID-19, reduced the total lipid profile that would otherwise amplify during virus infection. NIC treatment reduced the abundance of plasmalogens, diacylglycerides, and ceramides, which are required for virus production. Future screens of approved drugs may identify more druggable compounds than NIC that can safely but effectively counter SARS-CoV-2 subversion of lipid metabolism thereby reducing virus replication. However, these data support the consideration of niclosamide as a potential COVID-19 therapeutic given its modulation of lipophagy leading to the reduction of virus egress and the subsequent regulation of key lipid mediators of pathological inflammation.

microbiology↗

bioRxiv2021

Transmission potential of Floridian Aedes aegypti mosquitoes for dengue virus serotype 4: Implications for estimating local dengue risk

Dengue virus serotype 4 (DENV-4) circulated in Aedes aegypti in 2016 and 2017 in Florida in the absence of human index cases, compelling a full assessment of local mosquito vector competence and DENV-4 risk. To better understand DENV-4 transmission risk in Florida, we used an expanded suite of tests to measure and compare the vector competence of both an established colony of Ae. aegypti (Orlando strain [ORL]) and a field-derived colony from Collier County, Florida in 2018 (COL) for a Haitian DENV-4 human field isolate and a DENV-4 laboratory strain (Philippines H241). We immediately noted that ORL saliva-positivity was higher for the field versus laboratory DENV-4 strains. In a subsequent comparison with the recent COL mosquito colony we also observed significantly higher midgut susceptibility of COL and ORL for the Haitian DENV-4 field strain, and significantly higher saliva-positivity rate for COL, although overall saliva virus titers were similar between the two. These data point to a potential midgut infection barrier for the DENV-4 laboratory strain for both mosquito colonies and that the marked difference in transmission potential estimates hinge on virus-vector combinations. Our study highlights the importance of leveraging an expanded suite of testing methods with emphasis on utilizing local mosquito populations and field relevant dengue serotypes and strains to accurately estimate transmission risk in a given setting. ImportanceDENV-4 was found circulating in Florida (FL) Ae. aegypti mosquitoes in the absence of human index cases in the state (2016-2017). How DENV-4 was maintained locally is unclear, presenting a major gap in our understanding of DENV-4 public health risk. We determined the baseline arbovirus transmission potential of laboratory and field colonies of Ae. aegypti for both laboratory and field isolates of DENV-4. We observed high transmission potential of field populations of Ae. aegypti and evidence of higher vertical transmission of the DENV-4 field isolate, providing clues to the possible mechanism of undetected DENV-4 maintenance in the state. Our findings also move the field forward in the development of best practices for evaluating arbovirus vector competence, with evidence that transmission potential estimates vary depending on the mosquito-virus combinations. These data emphasize the poor suitability of lab-established virus strains and the high relevance of field-derived mosquito populations in estimating transmission risk.

microbiology↗

bioRxiv2021

In vitro infection of human lung tissue with SARS-CoV-2: Heterogeneity in host defense and therapeutic response

Cell lines are the mainstay in understanding the biology of COVID-19 infection, but do not recapitulate many of the complexities of human infection. The use of human lung tissue is one solution for the study of such novel respiratory pathogens. We hypothesized that a cryopreserved bank of human lung tissue allows for the in vitro study of the inter-individual heterogeneity of host response to SARS-CoV-2 infection, thus providing a bridge between studies with cell lines and studies in animal models. We generated a cryobank of tissues from 16 donors, most of whom had risk factors for severe illness from COVID-19. Cryopreserved tissues preserved 90% of cell viability and contained heterogeneous populations of metabolically active epithelial, endothelial, and immune cell subsets of the human lung. Samples were readily infectible with HCoV-OC43 and SARS-CoV-2 coronavirus strains, and demonstrated comparable susceptibility to infection. In contrast, we observed a marked donor-dependent heterogeneity in the expression of IL-6, CXCL8 and IFN{beta} in response to SARS-CoV-2 infection. Treatment of tissues with dexamethasone and the experimental drug, N-hydroxycytidine, suppressed viral growth in all samples, whereas chloroquine and remdesivir had no detectable effect. Metformin and sirolimus, molecules with predicted antiviral activity, suppressed viral replication in tissues from a subset of donors. In summary, we developed a novel system for the in vitro study of human SARS-CoV-2 infection using primary human lung tissue from a library of donor tissues. This model may be useful for drug screening and for understanding basic mechanisms of COVID-19 pathogenesis. ImportanceThe current biological systems for the study of COVID-19 are in vitro systems that differ from the human lung in many respects, and animal hosts to which the virus is not adapted. We developed another alternative for studying pathogenesis and drug susceptibility of SARS-CoV-2 in a cryopreserved bank of human lung tissues. We consider the importance of this work to relate to the practical use of this culture system as a repeatable and scalable approach that allows for the study of an important infection in relevant tissues. The tissue bank highlights the heterogeneous response to SARS-CoV-2 infection and treatment, which allows researchers to investigate why treatments work in some donors but not others.

microbiology↗

bioRxiv2020

SARS-CoV-2 viroporin triggers the NLRP3 inflammatory pathway

Cytokine storm resulting from a heightened inflammatory response is a prominent feature of severe COVID-19 disease. This inflammatory response results from assembly/activation of a cell-intrinsic defense platform known as the inflammasome. We report that the SARS-CoV-2 viroporin encoded by ORF3a activates the NLRP3 inflammasome, the most promiscuous of known inflammasomes. ORF3a triggers IL-1{beta} expression via NF{kappa}B, thus priming the inflammasome while also activating it via ASC-dependent and -independent modes. ORF3a-mediated inflammasome activation requires efflux of potassium ions and oligomerization between NEK7 and NLRP3. With the selective NLRP3 inhibitor MCC950 able to block ORF3a-mediated inflammasome activation and key ORF3a residues needed for virus release and inflammasome activation conserved in SARS-CoV-2 isolates across continents, ORF3a and NLRP3 present prime targets for intervention. SummaryDevelopment of anti-SARS-CoV-2 therapies is aimed predominantly at blocking infection or halting virus replication. Yet, the inflammatory response is a significant contributor towards disease, especially in those severely affected. In a pared-down system, we investigate the influence of ORF3a, an essential SARS-CoV-2 protein, on the inflammatory machinery and find that it activates NLRP3, the most prominent inflammasome by causing potassium loss across the cell membrane. We also define key amino acid residues on ORF3a needed to activate the inflammatory response, and likely to facilitate virus release, and find that they are conserved in virus isolates across continents. These findings reveal ORF3a and NLRP3 to be attractive targets for therapy.

microbiology↗

bioRxiv2020

Under-the-radar dengue virus infections in natural populations of Aedes aegypti mosquitoes

The incidence of locally acquired dengue infections increased during the last decade in the United States, compelling a sustained research effort on the dengue mosquito vector, Aedes aegypti, and its microbiome, which has been shown to influence virus transmission success. We examined the metavirome of four populations of Ae. aegypti mosquitoes collected in 2016-2017 from Manatee County, Florida. Unexpectedly, we discovered that dengue virus serotype 4 (DENV4) was circulating in these mosquito populations, representing the first documented case of such a phenomenon in the absence of a local DENV4 human case in this county over a two-year period. We confirmed that all of the mosquito populations carried the same DENV4 strain, assembled its full genome, validated infection orthogonally by reverse transcriptase PCR, traced the virus origin, estimated the time period of its introduction to the Caribbean region, as well as explored the viral genetic signatures and mosquito-specific virome associations that potentially mediated DENV4 persistence in mosquitoes. We discuss the significance of prolonged maintenance of these DENV4 infections in Ae. aegypti that occurred in the absence of a DENV4 human index case in Manatee County with respect to the inability of current surveillance paradigms to detect mosquito vector infections prior to a potential local outbreak. ImportanceSince 1999, dengue outbreaks in the continental United States (U.S.) involving local transmission have occurred episodically and only in Florida and Texas. In Florida, these episodes appear to be coincident with increased introductions of dengue virus into the region through human travel and migration from endemic countries. To date, the U.S. public health response to dengue outbreaks is largely reactive, and implementation of comprehensive arbovirus surveillance in advance of predictable transmission seasons, which would enable proactive preventative efforts, remains unsupported. The significance of our finding is that it is the first documented report of non-outbreak DENV4 transmission and maintenance within a local mosquito vector population in the continental U.S.in the absence of a human case during a two-year time period. Our data suggest that molecular surveillance of mosquito populations in high-risk, high tourism areas of the U.S., may allow for proactive, targeted vector control before potential arbovirus outbreaks.

ecology↗