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Casey Finnerty

Functional importance of deletion mut... [Appl Environ Microbiol. 2005] - PubMed - NCBI - 0 views

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    This paper shows the importance of genotypic diversity for baculovirus populations, particularly in terms of pathogenicity.
Sarah Muncy

ScienceDirect.com - Vaccine - Intranasal and intramuscular immunization with Baculoviru... - 0 views

  • An anti-malarial transmission-blocking vaccine (TBV) that prevents fertilization and/or ookinete/oocyst development within the mosquito is an attractive strategy to limit the transmission of malaria
  • The present study used this system to generate a Plasmodium vivax transmission-blocking immunogen (AcNPV-Dual-Pvs25).
  • Plasmodium vivax
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  • A variety of expression vectors (e.g., Escherichia coli, Pichia pastoris and DNA) have been used to express Pvs25 protein which has been administered alone or in combination with adjuvants
  • To date these studies suggest that the recombinant protein currently requires both not only linear, but conformation dependent epitopes, and a strong adjuvant to induce transmission-blocking antibodies.
  • Intranasal and intramuscular immunization with Baculovirus Dual Expression System-based Pvs25 vaccine substantially blocks Plasmodium vivax transmission
  • Recently, we have developed a new vaccine vector system based on the baculovirus Autographa californica nucleopolyhedrosis virus (AcNPV) termed the “Baculovirus Dual Expression System”, which drives expression of vaccine candidate antigens by a dual promoter that consists of tandemly arranged baculovirus-derived polyhedrin and mammalian-derived CMV promoters. It has been shown that AcNPV, an enveloped double-stranded DNA virus that naturally infects insects, possesses strong adjuvant properties that can activate dendritic cell-mediated innate immunity
  • Mucosal vaccines have several attractive features compared with parenteral vaccines (e.g., safety, cost-effectiveness and ease of administration), but studies on their use have been limited almost exclusively to protection against mucosally transmitted pathogens. We provide evidence that i.n. immunization is a feasible alternative for preventing malaria, which is transmitted through non-mucosal routes
  • These results are consistent with our previous work showing that intranasal immunization with the baculovirus-based vaccine induced strong systemic humoral immune responses with high titres of antigen-specific antibodies and conferred complete protection against malaria blood-stage challenge
  • which can induce immunological memory against heterologous antigens in a rodent model; however, it is precluded from clinical use due to its enterotoxicity and potential hazardous effects on olfactory nerves [22]. In contrast, a baculovirus-based delivery system may offer an attractive immunization method, as AcNPV exhibits low cytotoxicity and is incapable of replication in mammalian cells
  • The data described here adds to previously presented data showing the significant potential of the baculovirus dual expression system against the blood stages of the parasite
  • but also demonstrates clearly its ability to induce antibodies against the ookinete surface protein Pvs25, and to elicit a transmission-blocking immune response against the P. vivax isolates from endemic areas, and a transgenic rodent malaria parasite model in preliminary studies.
  • One was SMFA on peripheral blood from P. vivax infected patients.
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    Reference paper #2. Gave me information on malaria and baculoviruses.
Casey Finnerty

Using a Pan-Viral Microarray Assay (Virochip) to Screen Clinical Samples for Viral Path... - 1 views

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    A video-based publication from the peer-reviewed "Journal of Visualized Experiments". Nice introduction to the virochip method of virus screening.
Casey Finnerty

To Fight a Virus, Get a Virus: Military Bets on Mutant Pathogen - Bloomberg - 1 views

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    Describes the use of therapeutic interfering particles to treat viral disease.
Casey Finnerty

Dr. Donald A. Henderson, Who Helped End Smallpox, Dies at 87 - The New York Times - 1 views

  • Dr. Donald A. Henderson, a leader of one of mankind’s greatest public health triumphs, the eradication of smallpox, died on Friday in Towson, Md. He was 87.
  • died in a hospice of complications of a hip fracture, including infection with antibiotic-resistant staphylococcus, a dangerous pathogen he had himself researched and raised alarms about
  • The last known case was found in a hospital cook in Somalia in 1977.
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  • The only other disease to have been banished from the earth is rinderpest, a little-known relative of measles that kills hoofed animals and once caused widespread starvation in Africa; it was eradicated in 2011.
  • Smallpox, caused by the variola virus, was long one of mankind’s most terrifying scourges.
  • it killed almost a third of its victims, often through pneumonia or brain inflammation.
  • Many others were left blind from corneal ulcerations or severely disfigured by pockmarks.
  • Because it killed 80 percent of the American Indians who caught it, it was a major factor in the European conquest of the New World.
  • In 1796, Dr. Edward Jenner, an English physician, infected a young boy with cowpox taken from a blister on a milkmaid’s hand. Cowpox, a mild disease, protected those who had it from smallpox, and the modern vaccine era began.
  • Dr. Henderson quickly realized that trying to vaccinate vast populations was futile and switched to “ring vaccination.”
  • Dr. Foege, who is considered the father of this tactic, said it was “invented by accident” during a 1967 Nigerian outbreak when he had very little vaccine on hand.
  • “The first night, we asked ourselves what we would do if we were a virus bent on immortality,”
  • In 1977, success in hand, Dr. Henderson became dean of the Johns Hopkins University School of Hygiene and Public Health.“He was an imposing guy — physically big and very confident,” said Dr. Michael J. Klag, the school’s current dean, who was a student in that era. “He did not suffer fools gladly, and you were never sure if you were a fool or not.”
  • He gloomily foresaw failure for most other disease-elimination campaigns. The “siren song of eradication,” he once wrote, had led to goals that were more “evangelical” than attainable.
  • The problem, he would explain, was that each viral foe was so different. Smallpox had many weaknesses to exploit; it had no animal host. Every case can be found because victims have pox on their faces, and one vaccination provides lifetime immunity.
  • In 2011, when Bill Gates threw the full weight of his foundation into fighting polio, he struggled to explain how he would overcome such obstacles and, at the end of an interview, turned to an aide and said aloud, “I’ve got to get my D. A. Henderson response down better.”
  • The campaign, many experts have noted, succeeded just in time. A few years later, the virus that causes AIDS spread across Africa. Because the live smallpox vaccine can grow in an immune-compromised person into a huge, rotting, ultimately fatal lesion, it would have been impossible to deploy it.
Sean Hogan

PLOS Pathogens: Different Modes of Retrovirus Restriction by Human APOBEC3A and APOBEC3... - 22 views

  • One such family of restriction factors is the apolipoprotein B editing complex 3 (A3) cellular cytidine deaminases (CDA). While A3 genes are found in all mammals, their number differs from species to species. For example, humans have 7 A3 genes (A3A to A3H) while mice have only one gene. All proteins in this family contain at least one CDA domain that deaminates carbon 4 of cytidine in single-stranded DNA, resulting in a uracil that causes G to A transitions in the opposing strand [3].
    • alexridesducati
       
      Can these genes be exploited for antiviral therapy and if so, can it be done without harm to the host due to mutations?
  • viral cDNA accumulation
  • Packaging of A3G into virions is counteracted by HIV Vif (viral infectivity factor) protein. In virus-producer cells, Vif binds to A3G as well other A3 family members, and recruits cellular E3 ubiquitin ligase complexes, leading to ubiquitination and subsequent proteasomal degradation, thereby preventing packaging of A3G into budding virions [12]–[14]. Lentiviral Vif proteins show strong species-specificity. For example, HIV-1 Vif counteracts human A3G but only certain simian A3G homologues [15], [16]; it also does not interact with mouse A3 [17].
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  • Other members of the A3 family are believed to affect other exogenous viruses as well as endogenous retrovirus/retroelement movement within the genome. In particular, human A3A is a potent inhibitor of IAP and MusD and other retrotransposons such as LINE-1 and this inhibition is CDA-independent, at least in cultured cells [18]–[20]. A3A also inhibits adeno-associated virus replication, a nuclear-replicating parvovirus, via CDA-independent means [20]. In monocytes, A3A restricts HIV-1 infection and the decrease in A3A levels that occurs during monocyte-to-macrophage development is concomitant with increased susceptibility to HIV-1 infection [21]. A3A is not packaged into HIV virions and is thought to restrict infection by targeting incoming virus [22]–[24]. In contrast, A3A is packaged in human T-lymphotropic virus type-I virions and restricts infection, at least in transfected cells [25]. A3A preferentially deaminates cytidines that are in a TC motif [26].
  • Different A3 family members block infection by diverse retroviruses from different species, including HIV-2 [27], porcine endogenous retrovirus [28], [29], xenotropic, Friend (F-MLV) and Moloney murine leukemia virus (M-MLV) [30]–[32] and mouse mammary tumor virus (MMTV) [33]. Additionally, A3 proteins may restrict other virus families, including parvoviruses [20], [34], hepatitis B virus [35]–[37], papillomaviruses [38] and herpes simplex virus I [39]. Thus, it has been suggested that A3 proteins exist, at least in part, to prevent zoonotic transmission of viruses [40].
  • Here, we show that transgenic mice expressing the human A3A or A3G proteins restrict murine retrovirus infection in vivo in disparate ways. A3G was packaged into virions in vivo, leading to the deamination of both MLV and MMTV viral genomes. In contrast, A3A was not packaged, and appeared to restrict infection in a largely CDA-independent manner. Finally, we show that Vif/A3G interactions can be studied in this in vivo model, thus providing a potentially useful system for the analysis of small molecule inhibitors of A3 proteins and Vif.
  • To determine the level of transgene expression, we first isolated RNA from different tissues, including peripheral blood mononuclear cells (PBMCs), and performed reverse-transcribed real-time quantitative PCR (RT-qPCR). RNA from human H9 cultured cells and human and C57BL/6 mouse PBMCs served as controls. For each transgene, there was one high- (A3Ghigh, A3Ahigh) and one low- (A3Glow, A3Alow) expressing strain, defined by their relative expression in lymphoid tissues. The A3Ghigh strain expressed higher levels of the transgene than the endogenous mouse gene in spleen and thymus, but similar A3G levels in mouse and human PBMCs, while the A3Glow strain expressed approximately 10-fold lower levels in these tissues (Figure 1A). In contrast, the A3Ahigh strain expressed similar or lower levels than mouse A3; there was also about 2-fold lower expression of A3A in mouse PBMCs than in human PBMCs (Figure 1B). The A3Alow strain had very low but detectable levels of expression in several tissues. Since the β-actin regulatory region was used, transgene expression was seen in many tissues and in several at levels higher than endogenous mouse A3 (e.g. heart, brain and liver) (Figure 1A and 1B). We also performed western blots on different tissues from the 4 different mouse strains, using antiserum that detects both A3A and A3G. The relative protein expression levels were similar to that seen at the RNA level (Figure S1A and S1B).
  • We next determined if the in vivo-produced A3A and A3G proteins were functionally active. Extracts were prepared from primary splenocyte cultures and equal amounts (total protein concentration/volume) were incubated with FAM-labeled substrates containing the A3A- or A3G-preferred target sequence (S50-TTC and S50-CCC, respectively). As controls, we also performed these assays with extracts prepared from 293T cell lines transfected with A3A or A3G. Activity could be readily detected in transgenic mice expressing high levels of A3A or A3G. Further, in accord with the known specificity of the cytidine deaminases, extracts from the A3Ahigh mice deaminated the TTC- more efficiently than CCC-containing substrates, while those from A3Ghigh mice more efficiently deaminated the CCC substrate (Figure 2). For both A3Alow and A3G low, trace amounts of activity were detectable with the preferred substrates, while no activity was detectable with either endogenous mA3 or from mA3 knockout splenocytes. No deaminase activity was detected with WT mouse extracts, perhaps because the mouse protein has lower overall activity or expression. These data show that the transgenic mice expressed catalytically active human deaminases in these heterologous cells.
  • Humans have 7 APOBEC3 genes and determining how each specifically functions to inhibit retroviruses like HIV is complicated, because all 7 can be produced in a given cell type or tissue.
    • laceemarie
       
      What cell/tissue type(s) are these APOBEC3 genes naturally turned on in? 
  • To overcome this limitation, we made transgenic mice that express two of the human proteins, APOBEC3A and APOBEC3G in mice that do not express their own APOBEC3. These mice were able to effectively block infection by several mouse retroviruses
    • laceemarie
       
      What cell type(s) did they use? Does it matter which?
  • We were unable to perform similar assays with in vivo produced MMTV, because the only cell-free virus in mice is found in milk and mammary tumors and we have not yet established breeding colonies of virus-infected human A3 transgenic mice.
    • laceemarie
       
      Was this a screw up, or is it not that important to look at assays with in vivo produced MMTV. And by "not yet," does that mean they are going to? I feel like if your going to use this virus in your experimental studies, you should figure out ways to perform the assays, regardless of how you get the virus. It would appear that they knew this information before hand, so maybe an assay on MMTV is less relevant. 
    • laceemarie
       
      *you're
  • A3G and A3A inhibit retrovirus infection by different means.
    • becky214
       
      Since they use different means of inhibition, do they work together to prevent infection? Or is an and either/or type scenario?
  • Two A3A and A3G mouse strains each were generated, expressing levels of these proteins within the range or at levels lower than that seen in human cells. This likely has relevance to what occurs in individual humans, where non-coding region polymorphisms in A3 genes alter expression levels and may influence progression to HIV-induced disease
    • laceemarie
       
      Could this have something to do with how HIV works in the HIV controllers? Where they still exhibit virus particles, but at a lower amount, don't necessarily spread the virus as much, and don't exhibit as intense of HIV symptoms?
    • ameliaobert
       
      If A3A is less clear, but does not get packaged, where must the A3A involvement with the incoming virus be located (for myeloid cells)? As well as if it is known to use retroelement retotransposition and replication inhibiton in paraoviruses by nh2 to oh independent means, how does the A3A know to be signlle to change the structure of cystine?
  • An additional limitation of previous studies done on human A3 proteins is the reliance on transfecting constructs expressing A3 proteins, which may not reflect the endogenous levels of a protein expression found in vivo
    • Sean Hogan
       
      Couldn't they mimic the in vivo environment by overexpressing necessary proteins in the host?
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    Focus paper for retrovirus presentation.
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    This will be the focus paper for 11/14.
Casey Finnerty

Fish-Killing Virus Spreading in the Great Lakes - New York Times - 0 views

  • “We anticipate that this will continue and get worse over the next few years,” said Dr. Jim Casey, associate professor of virology at Cornell University. “We fear there may be more widespread presence of the virus.”One of Dr. Casey’s colleagues researching the virus, Dr. Paul Bowser, a professor of aquatic animal medicine, added, “This is a new pathogen and for the first number of years — 4, 5 or 10 years — things are going to be pretty rough, then the animals will become more immune and resistant and the mortalities will decline.”
Sarah Muncy

ScienceDirect.com - Vaccine - Hemagglutinin Displayed Baculovirus Protects Against High... - 0 views

    • Sarah Muncy
       
      So, the baculovirus on TOP of having the H5HA on it, can also get the immune system to kick in better?
  • It is remarkable that low doses (103pfu/mouse) of BVs act as an effective adjuvant [41]. Therefore, reducing BV concentration and elongating vaccination intervals may prevent memory responses to BV administration
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  • Foreign immunogens or peptides can be displayed on the envelope of AcMNPV by fusion with the baculovirus major envelope protein gp64
  • Baculoviruses have strong adjuvant activity to promote humoral and cellular immune responses against coadministered antigens, activate dendritic cells maturation, induce the production of cytokines, chemokines, and type I IFNs
  • There are two influenza vaccine approaches licensed in the US; the inactivated, split vaccine and the live-attenuated virus vaccine. Inactivated vaccines can efficiently induce humoral immune responses but generally only poor cellular immune responses.
  • Therefore, influenza HA can be displayed on the surface of baculovirus
  • virus-like particle (VLP)
  • Even though cellular immune responses cannot confer sterilizing immunity, they are able to reduce the severity of infection and lower morbidity and mortality rates [47], and antigen-specific memory T cells are able to rapidly respond to a secondary virus infection [45]. Furthermore, cellular immune responses to the conserved epitopes contained in vaccines may provide cross-protective immunity against different subtypes of influenza virus infection
  • To confirm that each HA was incorporated on the envelope of baculoviruses, supernatants from infected Sf9 cells were used to perform hemagglutination assay
  • Most BV display strategies rely on gp64 protein which is the major envelope protein of baculovirus.
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    This paper gave me a better understanding of some aspects of my focal paper that were unclear. How to test for HA, and how baculoviruses may be adjuvants in addition to expression vectors.
Casey Finnerty

Noroviruses: The Perfect Human Pathogens? - 0 views

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    "(See the brief report by Repp and Keene on pages 1639-41.)"
Casey Finnerty

American Society for Microbiology:Norovirus Disinfection: How Much is Enough? - 0 views

  • wipe with a wet cloth, and then disinfect with chlorine.
  • Household bleach, says Duizer, is an average about 5 percent chlorine when new (concentration declines with age), or 50,000 PPM, and thus, can be diluted 50-fold to achieve the 1,000 ppm.
  • Norovirus is apparently no more resistant to cleaning and disinfection than other pathogens, says Duizer. The virus’ efficiency in causing outbreaks “is more likely due to their extremely low infectious dose
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