Dr. Brandt is a Professor in the Departments of Ophthalmology and Visual Sciences, Medical Microbiology and Immunology, and the Clinical Cancer Center at the University of Wisconsin-Madison. His primary research interest is virology, specifically pathogenesis of herpes simplex virus; virulence genes in herpetic eye disease and herpes encephalitis; antivirals; interactions between cytokines and herpes viruses, gene delivery, and gene therapy.
Knockdown of TRIM5α or TRIM11 increases lentiviral vector transduction efficiency of human muller cells.
Exp Eye Res. 2021 Jan 10;:108436
Authors: Sauter MM, Brandt CR
The goal of this study was to determine the expression and distribution of the host restriction factors (RFs) TRIM5α and TRIM11 in non-human primate (NHP) neural retina tissue and the human Muller cell line MIO-M1. In addition, experiments were performed to determine the effect of TRIM5α and TRIM11 knockdown on FIVGFP transduction of MIO-M1 cells with the goal of devising strategies to increase the efficiency of lentiviral (LV) gene delivery vectors. Immunofluorescence (IF) studies indicated that TRIM5α and TRIM11 were localized predominantly in nuclei within the outer nuclear layer (ONL) and inner nuclear layer (INL) of NHP retina tissue. Double label IF indicated that TRIM5α and TRIM11 were localized to some of the retinal Muller cell nuclei. MIO-M1 cells expressed TRIM5α predominantly in the nucleus and TRIM11 primarily in the cytosol. FIVGFP transduction efficiency was significantly increased, at 4 and 7 days post transduction, in TRIM5α and TRIM11 knockdown clones (KD) compared to WT MIO-M1 cells. In addition, pretreatment with the proteasome inhibitor MG132 increased the transduction efficiency of FIVGFP in WT MIO-M1 cells. The nuclear translocation of NF-κB (p65), at 72 h post FIVGFP transduction, was enhanced in TRIM5α and TRIM11 KD clones. The expression of TRIM5α and TRIM11 in macaque neural retina tissue and MIO-M1 cells indicate the presence of these RFs in NHP retina and human Muller cells. Our data indicate that even partial knockdown of TRIM5α or TRIM11, or a short proteasome inhibitor pretreatment, can increase the transduction efficiency of a LV vector.
PMID: 33440192 [PubMed - as supplied by publisher]
Genomic nucleotide-based distance analysis for delimiting old world monkey derived herpes simplex virus species.
BMC Genomics. 2020 Jun 26;21(1):436
Authors: Kolb AW, Brandt CR
BACKGROUND: Herpes simplex viruses form a genus within the alphaherpesvirus subfamily, with three identified viral species isolated from Old World monkeys (OWM); Macacine alphaherpesvirus 1 (McHV-1; herpes B), Cercopithecine alphaherpesvirus 2 (SA8), and Papiine alphaherpesvirus 2 (PaHV-2; herpes papio). Herpes B is endemic to macaques, while PaHV-2 and SA8 appear endemic to baboons. All three viruses are genetically and antigenically similar, with SA8 and PaHV-2 thought to be avirulent in humans, while herpes B is a biosafety level 4 pathogen. Recently, next-generation sequencing (NGS) has resulted in an increased number of published OWM herpes simplex genomes, allowing an encompassing phylogenetic analysis.
RESULTS: In this study, phylogenetic networks, in conjunction with a genome-based genetic distance cutoff method were used to examine 27 OWM monkey herpes simplex isolates. Genome-based genetic distances were calculated, resulting in distances between lion and pig-tailed simplex viruses themselves, and versus herpes B core strains that were higher than those between PaHV-2 and SA8 (approximately 14 and 10% respectively). The species distance cutoff was determined to be 8.94%, with the method recovering separate species status for PaHV-2 and SA8 and showed that lion and pig-tailed simplex viruses (vs core herpes B strains) were well over the distance species cutoff.
CONCLUSIONS: We propose designating lion and pig-tailed simplex viruses as separate, individual viral species, and that this may be the first identification of viral cryptic species.
PMID: 32590937 [PubMed - as supplied by publisher]
Toll-like receptors 4, 5, 6 and 7 are constitutively expressed in non-human primate retinal neurons.
J Neuroimmunol. 2018 Jun 11;:
Authors: Sauter MM, Kolb AW, Brandt CR
The purpose of this study was to characterize cell-specific expression patterns of Toll-like receptors (TLR) in non-human primate (NHP) neural retina tissue. TLR 4, 5, 6, and 7 proteins were detected by immunblotting of macaque retina tissue lysates and quantitative PCR (qPCR) demonstrated TLRs 4-7 mRNA expression. Immunofluorescence (IF) microscopy detected TLRs 4-7 in multiple cell types in macaque neural retina including Muller, retinal ganglion cells (RGC), amacrine, and bipolar cells. These results demonstrate that TLRs 4-7 are constitutively expressed by neurons in the NHP retina raising the possibility that these cells could be involved in retinal innate inflammatory responses.
PMID: 29954626 [PubMed - as supplied by publisher]
Phylogenetic and recombination analysis of the herpesvirus genus varicellovirus.
BMC Genomics. 2017 Nov 21;18(1):887
Authors: Kolb AW, Lewin AC, Moeller Trane R, McLellan GJ, Brandt CR
BACKGROUND: The varicelloviruses comprise a genus within the alphaherpesvirus subfamily, and infect both humans and other mammals. Recently, next-generation sequencing has been used to generate genomic sequences of several members of the Varicellovirus genus. Here, currently available varicellovirus genomic sequences were used for phylogenetic, recombination, and genetic distance analysis.
RESULTS: A phylogenetic network including genomic sequences of individual species, was generated and suggested a potential restriction between the ungulate and non-ungulate viruses. Intraspecies genetic distances were higher in the ungulate viruses (pseudorabies virus (SuHV-1) 1.65%, bovine herpes virus type 1 (BHV-1) 0.81%, equine herpes virus type 1 (EHV-1) 0.79%, equine herpes virus type 4 (EHV-4) 0.16%) than non-ungulate viruses (feline herpes virus type 1 (FHV-1) 0.0089%, canine herpes virus type 1 (CHV-1) 0.005%, varicella-zoster virus (VZV) 0.136%). The G + C content of the ungulate viruses was also higher (SuHV-1 73.6%, BHV-1 72.6%, EHV-1 56.6%, EHV-4 50.5%) compared to the non-ungulate viruses (FHV-1 45.8%, CHV-1 31.6%, VZV 45.8%), which suggests a possible link between G + C content and intraspecies genetic diversity. Varicellovirus clade nomenclature is variable across different species, and we propose a standardization based on genomic genetic distance. A recent study reported no recombination between sequenced FHV-1 strains, however in the present study, both splitstree, bootscan, and PHI analysis indicated recombination. We also found that the recently sequenced Brazilian CHV-1 strain BTU-1 may contain a genetic signal in the UL50 gene from an unknown varicellovirus.
CONCLUSION: Together, the data contribute to a greater understanding of varicellovirus genomics, and we also suggest a new clade nomenclature scheme based on genetic distances.
PMID: 29157201 [PubMed - in process]
Recombination Analysis of Herpes Simplex Virus Type 1 Reveals a Bias towards GC Content and the Inverted Repeat Regions.
J Virol. 2015 Apr 29;
Authors: Lee K, Kolb AW, Sverchkov Y, Cuellar JA, Craven M, Brandt CR
Herpes simplex virus type 1 (HSV-1) causes recurrent mucocutaneous ulcers, and is the leading cause of infectious blindness and sporadic encephalitis in the United States. HSV-1 has been shown to be highly recombinagenic, however to date there has been no genome wide analysis of recombination. To address this, we generated 40 recombinant HSV-1 viruses derived from two parental strains; OD4 and CJ994. The 40 OD4-CJ994 HSV-1 recombinants were sequenced using the Illumina sequencing system, and recombination breakpoints were determined for each of the recombinants using the Bootscan program. Breakpoints occurring in the terminal inverted repeats were excluded from analysis to prevent double counting, resulting in a total of 272 breakpoints in the dataset. By placing windows around the 272 breakpoints followed by Monte Carlo analysis comparing actual data to simulated data, we identified recombination bias towards both GC content and intergenic regions. A Monte Carlo analysis also suggested that recombination did not appear to be responsible for the generation of spontaneous nucleotide mutations detected following sequencing. Additionally, kernel density estimation analysis across the genome found that the large, inverted repeats comprise a recombination hotspot.
IMPORTANCE: Herpes simplex virus type 1 (HSV-1) virus is the leading cause of sporadic encephalitis and blinding keratitis in developed countries. HSV-1 has been shown to be highly recombinagenic and recombination itself appears to be a significant component of genome replication. To date there has been no genome wide analysis of recombination. Here we present the first genome wide study of recombination by generating and sequencing 40 HSV-1 recombinants derived from the OD4 and CJ994 parental strains, followed by bioinformatics analysis. Recombination breakpoints were determined, yielding 272 in the full dataset. Kernel density analysis determined that the large inverted repeats constitute a recombination hotspot. Additionally, Monte Carlo analyses found biases towards GC content, intergenic regions, and repetitive regions.
PMID: 25926637 [PubMed - as supplied by publisher]
Genomic, Phylogenetic, and Recombinational Characterization of Herpes Simplex Virus Type 2 Strains.
J Virol. 2015 Apr 8;
Authors: Kolb AW, Larsen IV, Cuellar JA, Brandt CR
Herpes simplex virus type 2 (HSV-2) is a major global pathogen, infecting 16% of people, 15-49 years old worldwide, causing recurrent, genital ulcers. Little is known about viral factors contributing to virulence, and there are currently only two genomic sequences available. In this study we determined nearly complete genomic sequences of six additional HSV-2 isolates, using the Illumina MiSeq. We report that HSV-2 has genomic overall mean distance of 0.2355%, which is lower than HSV-1. There were approximately 100 amino acid encoding SNPs/INDELs per genome. Microsatellite mapping found a bias towards intergenic regions in the non-conserved microsatellites, and a genic bias in all detected tandem repeats. Extensive recombination between the HSV-2 strains was also strongly implied. This is the first study to analyze multiple HSV-2 sequences, and will be valuable in future evolutionary, virulence, and structure-function studies.
IMPORTANCE: Herpes simplex virus type 2 (HSV-2) is an significant worldwide pathogen, causing recurrent, genital ulcers. Here we present six nearly complete HSV-2 genomic sequences, and with the addition of two previously sequenced strains, for the first time genomic, phylogenetic, and recombination analysis was performed on multiple HSV-2 genomes. Our results show that microsatellite mapping found a bias towards intergenic regions in the non-conserved microsatellites, a genic bias in all detected tandem repeats, confirm that ChHV-1 is a separate species, and that each of the HSV-2 strains is genomic mosaic.
PMID: 25855744 [PubMed - as supplied by publisher]
Peptide therapeutics for treating ocular surface infections.
J Ocul Pharmacol Ther. 2014 Nov;30(9):691-9
Authors: Brandt CR
Microbial pathogens-bacteria, viruses, fungi, and parasites-are significant causes of blindness, particularly in developing countries. For bacterial and some viral infections a number of antimicrobial drugs are available for therapy but there are fewer available for use in treating fungal and parasitic keratitis. There are also problems with current antimicrobials, such as limited efficacy and the presence of drug-resistant microbes. Thus, there is a need to develop additional drugs. Nature has given us an example of 1 potential source of new antimicrobials: antimicrobial peptides and proteins that are either present in bodily fluids and tissues constitutively or are induced upon infection. Given the nature of peptides, topical applications are the most likely use to be successful and this is ideal for treating keratitis. Such peptides would also be active against drug-resistant pathogens and might act synergistically if used in combination therapy. Hundreds of peptides with antimicrobial properties have been isolated or synthesized but only a handful have been tested against ocular pathogens and even fewer have been tested in animal models. This review summarizes the currently available information on the use of peptides to treat keratitis, outlines some of the problems that have been identified, and discusses future studies that will be needed. Most of the peptides that have been tested have shown activity at concentrations that do not warrant further development, but 1 or 2 have promising activity raising the possibility that peptides can be developed to treat keratitis.
PMID: 25250986 [PubMed - indexed for MEDLINE]
Oligonucleotides designed to inhibit TLR9 block Herpes simplex virus type 1 infection at multiple steps.
Antiviral Res. 2014 Sep;109:83-96
Authors: Sauter MM, Gauger JJ, Brandt CR
Herpes simplex virus type 1 (HSV-1) is an important human pathogen which requires activation of nuclear factor-kappa B (NFκB) during its replication cycle. The persistent nature of HSV-1 infection, and the emergence of drug-resistant strains, highlights the importance of research to develop new antiviral agents. Toll-like receptors (TLRs) play a prominent role during the early antiviral response by recognizing viral nucleic acid and gene products, activating NFκB, and stimulating the production of inflammatory cytokines. We demonstrate a significant effect on HSV-1 replication in ARPE-19 and Vero cells when oligonucleotides designed to inhibit TLR9 are added 2h prior to infection. A greater than 90% reduction in the yield of infectious virus was achieved at oligonucleotide concentrations of 10-20 μM. TLR9 inhibitory oligonucleotides prevented expression of essential immediate early herpes gene products as determined by immunofluorescence microscopy and Western blotting. TLR9 oligonucleotides also interfered with viral attachment and entry. A TLR9 inhibitory oligonucleotide containing five adjacent guanosine residues (G-ODN) exhibited virucidal activity and inhibited HSV-1 replication when added post-infection. The antiviral effect of the TLR9 inhibitory oligonucleotides did not depend on the presence of TLR9 protein, suggesting a mechanism of inhibition that is not TLR9 specific. TLR9 inhibitory oligonucleotides also reduced NFκB activity in nuclear extracts. Studies using these TLR inhibitors in the context of viral infection should be interpreted with caution.
PMID: 24995383 [PubMed - indexed for MEDLINE]
A cationic peptide, TAT-Cd°, inhibits herpes simplex virus type 1 ocular infection in vivo.
Invest Ophthalmol Vis Sci. 2013 Feb 05;54(2):1070-9
Authors: Jose GG, Larsen IV, Gauger J, Carballo E, Stern R, Brummel R, Brandt CR
PURPOSE: To test the in vivo activity of a peptide derived from the protein transducing domain of the human immunodeficiency virus (HIV) Tat protein, TAT-Cd°, in a murine herpes simplex type 1 (HSV-1) keratitis model.
METHODS: the efficacy of TAT-CD° was assessed in a postinfection treatment model with different concentrations (1 mg/mL, 0.1 mg/mL, 0.01 mg/mL) of the peptide in one of four delivery vehicles: artificial tears, PBS, methylcellulose, and aquaphor cream. Treatment began within 4 or 24 hours postinfection. Viral titers in the tear film were determined by plaque assay.
RESULTS: TAT-Cd° reduced the severity of keratitis in all of the delivery vehicles tested when treatment started, 4 hours postinfection. Peptide in the tears or PBS delivery vehicle had the most significant reduction in disease severity and delayed the onset of vascularization and stromal keratitis. The percentage of mice presenting with disease was also significantly reduced and viral titers were reduced by 1 log at 24 hours postinfection in mice treated with 1 mg/mL TAT-Cd°, suggesting that inhibiting replication early is sufficient to achieve clinical effects. Lower concentrations were not effective and delaying treatment by 24 hours was also not effective.
CONCLUSIONS: This study shows that TAT-Cd° is an effective antiviral against HSV-1 strain KOS when applied shortly postinfection and that aqueous-based formulations are more suitable.
PMID: 23341013 [PubMed - indexed for MEDLINE]
Using HSV-1 genome phylogenetics to track past human migrations.
PLoS One. 2013;8(10):e76267
Authors: Kolb AW, Ané C, Brandt CR
We compared 31 complete and nearly complete globally derived HSV-1 genomic sequences using HSV-2 HG52 as an outgroup to investigate their phylogenetic relationships and look for evidence of recombination. The sequences were retrieved from NCBI and were then aligned using Clustal W. The generation of a maximum likelihood tree resulted in a six clade structure that corresponded with the timing and routes of past human migration. The East African derived viruses contained the greatest amount of genetic diversity and formed four of the six clades. The East Asian and European/North American derived viruses formed separate clades. HSV-1 strains E07, E22 and E03 were highly divergent and may each represent an individual clade. Possible recombination was analyzed by partitioning the alignment into 5 kb segments, performing individual phylogenetic analysis on each partition and generating a.phylogenetic network from the results. However most evidence for recombination spread at the base of the tree suggesting that recombination did not significantly disrupt the clade structure. Examination of previous estimates of HSV-1 mutation rates in conjunction with the phylogenetic data presented here, suggests that the substitution rate for HSV-1 is approximately 1.38 × 10(-7) subs/site/year. In conclusion, this study expands the previously described HSV-1 three clade phylogenetic structures to a minimum of six and shows that the clade structure also mirrors global human migrations. Given that HSV-1 has co-evolved with its host, sequencing HSV-1 isolated from various populations could serve as a surrogate biomarker to study human population structure and migration patterns.
PMID: 24146849 [PubMed - indexed for MEDLINE]
Ocular distribution, spectrum of activity, and in vivo viral neutralization of a fully humanized anti-herpes simplex virus IgG Fab fragment following topical application.
Antimicrob Agents Chemother. 2012 Mar;56(3):1390-402
Authors: Berdugo M, Larsen IV, Abadie C, Deloche C, Kowalczuk L, Touchard E, Dubielzig R, Brandt CR, Behar-Cohen F, Combette JM
Herpes simplex ocular infection is a major cause of corneal blindness. Local antiviral treatments exist but are associated with corneal toxicity, and resistance has become an issue. We evaluated the biodistribution and efficacy of a humanized anti-herpes simplex virus (anti-HSV) IgG FAb fragment (AC-8; 53 kDa) following repeated topical administration. AC-8 was found in the corneal epithelium, anterior stroma, subepithelial stromal cells, and retinal glial cells, with preferential entry through the ocular limbus. AC-8 was active against 13 different strains of HSV-1, with 50% and 90% mean effective concentrations (MEC(50) and MEC(90), respectively) ranging from 0.03 to 0.13 μg/ml, indicating broad-spectrum activity. The in vivo efficacy of AC-8 was evaluated in a mouse model of herpes-induced ocular disease. Treatment with low-dose AC-8 (1 mg/ml) slightly reduced the ocular disease scores. A greater reduction of the disease scores was observed in the 10-mg/ml AC-8-treated group, but not as much as with trifluridine (TFT). AC-8 treatment reduced viral titers but less than trifluridine. AC-8 did not display any toxicity to the cornea or other structures in the eye. In summary, topical instillation of an anti-HSV FAb can be used on both intact and ulcerated corneas. It is well tolerated and does not alter reepithelialization. Further studies to improve the antiviral effect are needed for AC-8 to be considered for therapeutic use.
PMID: 22203590 [PubMed - indexed for MEDLINE]
Antiviral activity of the EB peptide against zoonotic poxviruses.
Virol J. 2012 Jan 06;9:6
Authors: Altmann SE, Brandt CR, Jahrling PB, Blaney JE
BACKGROUND: The EB peptide is a 20-mer that was previously shown to have broad spectrum in vitro activity against several unrelated viruses, including highly pathogenic avian influenza, herpes simplex virus type I, and vaccinia, the prototypic orthopoxvirus. To expand on this work, we evaluated EB for in vitro activity against the zoonotic orthopoxviruses cowpox and monkeypox and for in vivo activity in mice against vaccinia and cowpox.
FINDINGS: In yield reduction assays, EB had an EC50 of 26.7 μM against cowpox and 4.4 μM against monkeypox. The EC50 for plaque reduction was 26.3 μM against cowpox and 48.6 μM against monkeypox. A scrambled peptide had no inhibitory activity against either virus. EB inhibited cowpox in vitro by disrupting virus entry, as evidenced by a reduction of the release of virus cores into the cytoplasm. Monkeypox was also inhibited in vitro by EB, but at the attachment stage of infection. EB showed protective activity in mice infected intranasally with vaccinia when co-administered with the virus, but had no effect when administered prophylactically one day prior to infection or therapeutically one day post-infection. EB had no in vivo activity against cowpox in mice.
CONCLUSIONS: While EB did demonstrate some in vivo efficacy against vaccinia in mice, the limited conditions under which it was effective against vaccinia and lack of activity against cowpox suggest EB may be more useful for studying orthopoxvirus entry and attachment in vitro than as a therapeutic against orthopoxviruses in vivo.
PMID: 22225618 [PubMed - indexed for MEDLINE]
Multiplex sequencing of seven ocular herpes simplex virus type-1 genomes: phylogeny, sequence variability, and SNP distribution.
Invest Ophthalmol Vis Sci. 2011 Nov 25;52(12):9061-73
Authors: Kolb AW, Adams M, Cabot EL, Craven M, Brandt CR
PURPOSE: Little is known about the role of sequence variation in the pathology of HSV-1 keratitis virus. The goal was to show that a multiplex, high-throughput genome-sequencing approach is feasible for simultaneously sequencing seven HSV-1 ocular strains.
METHODS: A genome sequencer was used to sequence the HSV-1 ocular isolates TFT401, 134, CJ311, CJ360, CJ394, CJ970, and OD4, in a single lane. Reads were mapped to the HSV-1 strain 17 reference genome by high-speed sequencing. ClustalW was used for alignment, and the Mega 4 package was used for phylogenetic analysis (www.megasoftware.net). Simplot was used to compare genetic variability and high-speed sequencing was used to identify SNPs (developed by Stuart Ray, Johns Hopkins University School of Medicine, Baltimore, MD, http://sray.med.som.jhml.edu/SCRoftware/simplot).
RESULTS: Approximately 95% to 99% of the seven genomes were sequenced in a single lane with average coverage ranging from 224 to 1345. Phylogenetic analysis of the sequenced genome regions revealed at least three clades. Each strain had approximately 200 coding SNPs compared to strain 17, and these were evenly spaced along the genomes. Four genes were highly conserved, and six were more variable. Reduced coverage was obtained in the highly GC-rich terminal repeat regions.
CONCLUSIONS: Multiplex sequencing is a cost-effective way to obtain the genomic sequences of ocular HSV-1 isolates with sufficient coverage of the unique regions for genomic analysis. The number of SNPs and their distribution will be useful for analyzing the genetics of virulence, and the sequence data will be useful for studying HSV-1 evolution and for the design of structure-function studies.
PMID: 22016062 [PubMed - indexed for MEDLINE]
Virus aggregating peptide enhances the cell-mediated response to influenza virus vaccine.
Vaccine. 2011 Oct 13;29(44):7696-703
Authors: Jones JC, Settles EW, Brandt CR, Schultz-Cherry S
Given the poor immunogenicity of current H5N1 influenza vaccines, additives and adjuvants remain a viable solution for increasing efficacy. Here, we demonstrate that a 20-amino acid peptide (EB) possessing influenza antiviral activity also enhances the immune response to H5N1 vaccination in mice. The addition of EB to formalin-inactivated whole-virus vaccine induced virion aggregation and these aggregates were readily engulfed by phagocytic cells in vitro. In vivo, mice vaccinated with a suboptimal dose of inactivated vaccine containing EB peptide had reduced morbidity, improved viral clearance, and faster recovery than mice receiving vaccine alone. This phenomenon was not accompanied by an increase in virus-specific antibodies. Instead, cell-mediated immunity was enhanced as demonstrated by increased interferon-γ production from splenocytes. This data demonstrates that the EB peptide may a useful adjuvant for boosting the efficacy of poorly immunogenic influenza vaccines.
PMID: 21839131 [PubMed - indexed for MEDLINE]
Sequence variation in the herpes simplex virus U(S)1 ocular virulence determinant.
Invest Ophthalmol Vis Sci. 2011 Jun 28;52(7):4630-8
Authors: Kolb AW, Schmidt TR, Dyer DW, Brandt CR
PURPOSE: The herpes simplex virus type 1 (HSV-1) U(S)1 gene encodes host-range and ocular virulence determinants. Mutations in U(S)1 affecting virulence are known in strain OD4, but the genomic variation across several strains is not known. The goal was to determine the degree of sequence variation in the gene from several ocular HSV isolates.
METHODS: The U(S)1 gene from six ocular HSV-1 isolates, as well as strains KOS and F, were sequenced, and bioinformatics analyses were applied to the data.
RESULTS: Strains 17, F, CJ394, and CJ311 had identical amino acid sequences. With the other strains, most of the variability was concentrated in the amino-terminal third of the protein. MEME analysis identified a 63-residue core sequence (motif 1) present in all α-herpesvirus U(S)1 homologs that were located in a region identified as structured. Ten amino acids were absolutely conserved in all the α-herpesvirus U(S)1 homologs and were all located in the central core. Consensus-binding motifs for cyclin-dependent kinases and pocket proteins were also identified.
CONCLUSIONS: These results suggest that significant sequence variation exists in the U(S)1 gene, that the α22 protein contains a conserved central core region with structurally variable regions at the amino- and carboxyl termini, that 10 amino acids are conserved in α-herpes U(S)1 homologs, and that additional host proteins may interact with the HSV-1 U(S)1 and U(S)1.5 proteins. This information will be valuable in designing further studies on structure-function relationships and on the role these play in host-range determination and keratitis.
PMID: 21519032 [PubMed - indexed for MEDLINE]
Identification of the minimal active sequence of an anti-influenza virus peptide.
Antimicrob Agents Chemother. 2011 Apr;55(4):1810-3
Authors: Jones JC, Settles EW, Brandt CR, Schultz-Cherry S
The antiviral peptide, entry blocker (EB), inhibits influenza virus replication by preventing attachment to cells. Here, we identified the minimal and optimal EB sequence that retained antiviral activity with a 50% inhibitory concentration (IC(50)) and 50% effective concentration (EC(50)) similar to those of the full-length EB peptide and several truncated variants that possessed up to 10-fold lower IC(50)s. These data have implications for improving the antiviral efficacy of EB-derived peptides while decreasing production costs and easing synthesis.
PMID: 21220525 [PubMed - indexed for MEDLINE]
A cationic TAT peptide inhibits Herpes simplex virus type 1 infection of human corneal epithelial cells.
J Ocul Pharmacol Ther. 2010 Dec;26(6):541-7
Authors: Larsen IV, Brandt CR
UNLABELLED: Abstract Purpose: To determine if a peptide, TAT-Cd(0), inhibits Herpes simplex virus type 1 infection of human corneal epithelial cells.
METHODS: TAT-Cd(0) and a control peptide, E(50,51)TAT-Cd(0), were added at various times throughout infection with the lacz-expressing hrR3 virus, and viral replication was measured by β-galactosidase activity. Toxicity was assessed using a dye reduction assay.
RESULTS: The CC(50) value for TAT-Cd(0) was ∼100 μM. In assays with peptide present at all times, TAT-Cd(0) was 150-fold more active than E(50,51)TAT-Cd(0) (EC(50) 0.2 vs. 30.0 μM). The EC(50) values of TAT-Cd(0) for entry inhibition, cell protection, virus inactivation, and inhibition of attachment were 0.1, 0.4, 9.5, and 3.0 μM, respectively. TAT-Cd(0) was less effective when added 1 h postinfection (EC(50) = 30.0 μM).
CONCLUSIONS: TAT-Cd(0) is an effective inhibitor of Herpes simplex virus type 1 infection in human corneal epithelial cells and affects multiple steps before, or very early, in infection. The peptide has potential as an antiviral and further studies are warranted.
PMID: 21029018 [PubMed - indexed for MEDLINE]
The virucidal EB peptide protects host cells from herpes simplex virus type 1 infection in the presence of serum albumin and aggregates proteins in a detergent-like manner.
Antimicrob Agents Chemother. 2010 Oct;54(10):4275-89
Authors: Bultmann H, Girdaukas G, Kwon GS, Brandt CR
The linear cationic amphiphilic EB peptide, derived from the FGF4 signal sequence, was previously shown to be virucidal and to block herpes simplex type I (HSV-1) entry (H. Bultmann, J. S. Busse, and C. R. Brandt, J. Virol. 75:2634-2645, 2001). Here we show that cells treated with EB (RRKKAAVALLPAVLLALLAP) for less than 5 min are also protected from infection with HSV-1. Though protection was lost over a period of 5 to 8 h, it was reinduced as rapidly as during the initial treatment. Below a 20 μM concentration of EB, cells gained protection in a serum-dependent manner, requiring bovine serum albumin (BSA) as a cofactor. Above 40 μM, EB coprecipitated with BSA under hypotonic conditions. Coprecipitates retained antiviral activity and released active peptide. NaCl (≥0.3 M) blocked coprecipitation without interfering with antiviral activity. As shown for β-galactosidase, EB below 20 μM acted as an enzyme inhibitor, whereas above 40 to 100 μM EB, β-galactosidase was precipitated as was BSA or other unrelated proteins. Pyrene fluorescence spectroscopy revealed that in the course of protein aggregation, EB acted like a cationic surfactant and self associated in a process resembling micelle formation. Both antiviral activity and protein aggregation did not depend on stereospecific EB interactions but depended strongly on the sequence of the peptide's hydrophobic tail. EB resembles natural antimicrobial peptides, such as melittin, but when acting in a nonspecific detergent-like manner, it primarily seems to target proteins.
PMID: 20643896 [PubMed - indexed for MEDLINE]