Microbiology and Immunology
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Item Open Access A biolistic method for high-throughput production of transgenic wheat plants with single gene insertions(BioMed Central, 2018) Ismagul, A.; Yang, N.; Maltseva, E.; Iskakova, G.; Mazonka, I.; Skiba, Y.; Bi, H.; Eliby, S.; Jatayev, S.; Shavrukov, Y.; Borisjuk, N.; Langridge, P.Background: The relatively low efficiency of biolistic transformation and subsequent integration of multiple copies of the introduced gene/s significantly complicate the genetic modification of wheat (Triticum aestivum) and other plant species. One of the key factors contributing to the reproducibility of this method is the uniformity of the DNA/gold suspension, which is dependent on the coating procedure employed. It was also shown recently that the relative frequency of single copy transgene inserts could be increased through the use of nanogram quantities of the DNA during coating. Results: A simplified DNA/gold coating method was developed to produce fertile transgenic plants, via microprojectile bombardment of callus cultures induced from immature embryos. In this method, polyethyleneglycol (PEG) and magnesium salt solutions were utilized in place of the spermidine and calcium chloride of the standard coating method, to precipitate the DNA onto gold microparticles. The prepared microparticles were used to generate transgenics from callus cultures of commercial bread wheat cv. Gladius resulting in an average transformation frequency of 9.9%. To increase the occurrence of low transgene copy number events, nanogram amounts of the minimal expression cassettes containing the gene of interest and the hpt gene were used for co-transformation. A total of 1538 transgenic wheat events were generated from 15,496 embryos across 19 independent experiments. The variation of single copy insert frequencies ranged from 16.1 to 73.5% in the transgenic wheat plants, which compares favourably to published results. Conclusions: The DNA/gold coating procedure presented here allows efficient, large scale transformation of wheat. The use of nanogram amounts of vector DNA improves the frequency of single copy transgene inserts in transgenic wheat plants.Item Metadata only A comparative species listing of myxomycetes from tropical (Philippines) and temperate (United States) forests(Mushroom Research Foundation, 2014) dela Cruz, T.; Rea, M.; Tran, H.; Ko Ko, T.; Stephenson, S.In terms of their flora and fauna, tropical forests are generally reported to be characterized by higher species diversity than temperate forests. Can this also be true for microorganisms such as myxomycetes, In the present study, three lowland tropical forests in the Philippines and three mid-latitude temperate forests in north central Arkansas in the United States were surveyed for myxomycetes with the moist chamber technique as it applies to these organisms. Results indicated that more species of myxomycetes were associated with samples of aerial litter, dead but still attached plant parts, forest floor litter and woody twigs collected in Arkansas than for those collected in the Philippines. A higher value for taxonomic diversity also was noted for the temperate forests. However, species listed herein are new records for the Philippines. In the present study, a comparison of the taxonomic diversity of myxomycetes in relation to the number of moist chamber replications was carried out, and this showed that a difference of as many as eight species could occur between what was recorded from a single culture and a series of three cultures prepared from the same sample. Clearly, many of these species could be missed if only a single culture is prepared for a particular sample.Item Metadata only A divergent transcriptional landscape underpins the development and functional branching of MAIT cells(Science Immunology; American Association for the Advancement of Science, 2019) Koay, H.-F.; Su, S.; Amann-Zalcenstein, D.; Daley, S.R.; Comerford, I.; Miosge, L.; Whyte, C.E.; Konstantinov, I.E.; d'Udekem, Y.; Baldwin, T.; Hickey, P.F.; Berzins, S.P.; Mak, J.Y.W.; Sontani, Y.; Roots, C.M.; Sidwell, T.; Kallies, A.; Chen, Z.; Nüssing, S.; Kedzierska, K.; et al.MR1-restricted mucosal-associated invariant T (MAIT) cells play a unique role in the immune system. These cells develop intrathymically through a three-stage process, but the events that regulate this are largely unknown. Here, using bulk and single-cell RNA sequencing-based transcriptomic analysis in mice and humans, we studied the changing transcriptional landscape that accompanies transition through each stage. Many transcripts were sharply modulated during MAIT cell development, including SLAM (signaling lymphocytic activation molecule) family members, chemokine receptors, and transcription factors. We also demonstrate that stage 3 "mature" MAIT cells comprise distinct subpopulations including newly arrived transitional stage 3 cells, interferon-γ-producing MAIT1 cells and interleukin-17-producing MAIT17 cells. Moreover, the validity and importance of several transcripts detected in this study are directly demonstrated using specific mutant mice. For example, MAIT cell intrathymic maturation was found to be halted in SLAM-associated protein (SAP)-deficient and CXCR6-deficient mouse models, providing clear evidence for their role in modulating MAIT cell development. These data underpin a model that maps the changing transcriptional landscape and identifies key factors that regulate the process of MAIT cell differentiation, with many parallels between mice and humans.Item Metadata only A duck hepatitis B virus with a knock-out mutation in the X-ORF shows similar growth characteristics to wild-type.(Australian Centre for Hepatitis Virology, 2004) Meier, P.; Scougall, C.; Will, H.; Burrell, C.; Jilbert, A.; International Symposium on Viral Hepatitis and Liver Disease (11th : 2003 : Sydney, N.S.W.); Jilbert, A.; Grgacic, E.; Vickery, K.; Burrell, C.; Cossart, Y.Item Metadata only A dynamic relationship between mucosal T helper type 17 and regulatory T-cell populations in nasopharynx evolves with age and associates with the clearance of pneumococcal carriage in humans(ELSEVIER SCI LTD, 2016) Mubarak, A.; Ahmed, M.; Upile, N.; Vaughan, C.; Xie, C.; Sharma, R.; Acar, P.; McCormick, M.; Paton, J.; Mitchell, T.; Cunliffe, N.; Zhang, Q.Pneumococcal carriage is common in young children, which may account for the high incidence of disease in this age group. Host factors determining the clearance of carriage in humans remain unclear. We aimed to study the relationships between T helper type 17 (Th17) and Foxp3(+) regulatory T (Treg) cells in nasopharynx-associated lymphoid tissue (NALT) and carriage in children and adults. Frequencies of Th17 and Treg cells in NALT were analysed by flow cytometry in association with age and pneumococcal carriage status. Cytokine responses following pneumococcal stimulation were analysed by cytometric beads array. The frequencies of Th17 and Treg cells in NALT were inversely correlated (R -0.60). Whereas Treg cell frequency decreased with age (R -0.63), both Th17 and the Th17: Treg ratio increased with age (R 0.62 and R 0.64, respectively). Also, the Th17: Treg ratio was higher in carriage-negative than in carriage-positive children (p <0.01). Pneumococcal stimulation of tonsillar cells increased both Th17 and Treg cell numbers, but the Th17: Treg ratio and pattern of cytokine responses differed between carriage-negative and carriage-positive children. The former showed markedly higher Th17: Treg and interleukin-17A: interleukin-10 ratios than in the latter (p <0.01). Pneumococcal stimulation also induces Th17, although the capacity of this Th17 differentiation from naive T cells of young children was low, but increased with age. We demonstrated a dynamic relationship between Th17 and Treg cells in human nasopharynx that evolves with age. The balance between Th17 and Treg cells in NALT appears to be a major host factor closely associated with the clearance of Streptococcus pneumoniae from the nasopharynx.Item Open Access A genomic perspective of metal-resistant bacteria from gold particles: Possible survival mechanisms during gold biogeochemical cycling(Oxford University Press (OUP), 2020) Sanyal, S.K.; Reith, F.; Shuster, J.A bacterial consortium was enriched from gold particles that 'experienced' ca. 80 years of biotransformation within waste-rock piles (Australia). This bacterial consortium was exposed to 10 µM AuCl3 to obtain Au-tolerant bacteria. From these isolates, Serratia sp. and Stenotrophomonas sp. were the most Au-tolerant and reduced soluble Au as pure gold nanoparticles, indicating that passive mineralisation is a mechanism for mediating the toxic effect of soluble Au produced during particle dissolution. Genome-wide analysis demonstrated that these isolates also possessed various genes that could provide cellular defence enabling survival under heavy-metal stressed condition by mediating the toxicity of heavy metals through active efflux/reduction. Diverse metal-resistant genes or genes clusters (cop, cus, czc, znt, ars) were detected, which could confer resistance to soluble Au. Comparative genome analysis revealed that the majority of detected heavy-metal resistant genes were similar (i.e. orthologous) to those genes of Cupriavidus metallidurans CH34. The detection of heavy-metal resistance, nutrient cycling, and biofilm formation genes (pgaABCD, bsmA, hmpS) may have indirect yet important roles when dealing with soluble Au during particle dissolution. In conclusion, the physiological and genomic results suggest that bacteria living on gold particles would likely use various genes to ensure survival during Au biogeochemical cycling.Item Metadata only A large retrospective cohort study of cefazolin compared with flucloxacillin for methicillin-susceptible Staphylococcus aureus bacteraemia(Elsevier, 2018) Davis, J.; Turnidge, J.; Tong, S.Background and objectives: Antistaphylococcal penicillins (ASPs) are recommended as first-line treatment for invasive infections caused by methicillin-susceptible Staphylococcus aureus (MSSA). Cefazolin is an alternative option, but there is theoretical concern about its use as some MSSA strains produce beta-lactamases active against cefazolin. The study compared the outcomes in patients with MSSA infections treated with flucloxacillin and cefazolin. Methods: We analysed data from The Australia and New Zealand Co-operative Outcomes of Staphylococcal Sepsis (ANZCOSS) observational study, which included all consecutive unique episodes of Staphylococcus aureus bacteraemia from 27 hospital-based or independent microbiology laboratories from January 2007 to September 2013. In this retrospective analysis of prospectively collected data, we compared 30-day all-cause mortality in patients with MSSA bacteraemia treated with flucloxacillin to that in patients treated with cefazolin. Results: We included data from 7312 episodes of MSSA bacteremia and found no difference in 30-day mortality in those treated with flucloxacillin (731/6520 [11.2%, 95% CI 10.9–12.5%]) compared to cefazolin (83/792 [10.7%, 95% CI 8.4–12.8%]), OR 0.93 (95% CI 0.72–1.17). In a propensity-adjusted analysis, mortality remained non-significantly lower in the cefazolin group (aOR 0.86 [95% CI 0.65–1.14]). Conclusions: This study supports the results from previous observational studies from other regions, while extending them to Australasia and to a much larger number of patients. Although this observational study indicates cefazolin is likely to have equivalent or superior outcomes to ASPs for MSSA bacteraemia, this can only be convincingly proven by a properly designed randomised controlled trial.Item Metadata only A new locus (vsp417-4) belonging to the tsa417-like subfamily of variant-specific surface protein genes in Giardia intestinalis(Elsevier Science, 1999) Ey, P.; Darby, J.; Mayrhofer, G.Item Metadata only A novel assay for detection of hepatitis C virus-specific effector CD4⁺ T cells via co-expression of CD25 and CD134(Elsevier Science BV, 2012) Keoshkerian, E.; Helbig, K.; Beard, M.; Zaunders, J.; Seddiki, N.; Kelleher, A.; Hampartzoumian, T.; Zekry, A.; Lloyd, A.Hepatitis C virus (HCV)-specific CD4(+) effector T cell responses are likely to play a key role in the immunopathogenesis of HCV infection by promoting viral clearance and maintaining control of viraemia. As the precursor frequency of HCV-specific CD4(+) T cells in peripheral blood is low, favoured assay systems such as intracellular cytokine (ICC) or tetramer staining have limited utility for ex vivo analyses. Accordingly, the traditional lymphocyte proliferation assay (LPA) remains the gold standard, despite detecting responses in only a minority of infected subjects. Recently, we reported development and validation of a novel whole blood CD4(+) effector T cell assay based on ex vivo antigen stimulation followed by co-expression of CD25 and CD134 on CD4(+) T cells. Here we report adaptation of this assay to assessment of HCV-specific responses in cryopreserved peripheral blood mononuclear cells using standardised antigens, including peptide pools, viral supernatants and recombinant viral proteins. The assay allowed detection of HCV-specific CD4 responses in donors with both resolved and chronic infection. Responses were highly correlated with those revealed by LPA. Application of this assay will further define the role of CD4(+) T cells in the immunopathogenesis of HCV infection.Item Metadata only A polyphasic approach leading to the revision of the genus Planktothrix (Cyanobacteria) and its type species, P. agardhii, and proposal for integrating the emended valid botanical taxa, as well as three new species, Planktothrix paucivesiculata sp. nov.ICNP, Planktothrix tepida sp. nov.ICNP, and Planktothrix serta sp. nov.ICNP, as genus and species names with nomenclatural standing under the ICNP(Elsevier, 2015) Gaget, V.; Welker, M.; Rippka, R.; de Marsac, N.Abstract not availableItem Metadata only A putative pathway for biosynthesis of the O-antigen component, 3-deoxy-L-glycero-tetronic acid, based in the nucleotide sequence of the Vibrio cholerae O1 rfb region(Elsevier, 1995) Morona, R.; Karageorgos, L.; Manning, P.; Stroeher, U.The nucleotide sequence of a region of the rfb genes, encoding biosynthesis of the Vibrio cholerae (Vc) O1 O-antigen, was determined. Analysis of the open reading frames (ORFs) within this region has revealed similarities with a number of different classes of biosynthetic proteins and enzymes. The ORFs have been designated RfbK, RfbL, RfbM, RfbN and RfbO. RfbK is a small, acidic protein which has similarity to the family of proteins known as acyl-carrier proteins (ACP). The RfbL protein has similarity to a super-family of enzymes which adenylate their substrates as a part of their reaction mechanism. Included in these are several acetyl-CoA ligases. Alignment of RfbL with these proteins reveals a highly conserved domain containing the motif GlyXaaXaaGlyXaaPro. This resembles the ATP-binding site motif and may represent a variant of the usual motif, except that Pro replaces Gly. The VcRfbM protein has similarity with a family of long-chain, iron-containing alcohol dehydrogenases, of which the Escherichia coli K-12 fucO and adhE gene products are also members. The RfbN protein has sequence homology with LuxE and LuxC of Vibrio harveyi (Vh) and other bioluminescent bacterial species. The latter are two components of the enzyme complex which synthesizes the long-chain aldehyde used in the V. harveyi bioluminescence system. Finally, the VcRfbO protein has sequence similarity with acetyl-CoA transferases. We were able to identify a number of the gene products using a T7 expression system, confirming several of the allocated ORFs. A biosynthetic pathway for the Vc O-antigen component 3-deoxy-L-glycero-tetronic acid, based on the enzymatic functions predicted for the RfbK, RfbL, RfbM, RfbN and RfbO proteins, is presented.Item Metadata only A putative pathway for perosamine biosynthesis is the first function encoded within the rfb region of Vibrio cholerae O1(Elsevier, 1995) Stroeher, U.; Karageorgos, L.; Brown, M.; Morona, R.; Manning, P.The first four genes (rfbA,B,D,E) of the rfb region of Vibrio cholerae O1 are predicted to encode the enzymes required for the biosynthesis of perosamine, which constitutes the backbone structure of the O-antigen of the lipopolysaccharide. Based on homology to known proteins/protein families, the following functions are predicted: RfbA, phosphomannose isomerase-guanosine diphosphomannose pyrophosphorylase; RfbB, phosphomanno-mutase; RfbD, oxido reductase and RfbE, perosamine synthetase (amino-transferase). Thus, perosamine is synthesized from fructose 6-phosphate via the intermediates mannose 6-phosphate by RfbA, to mannose 1-phosphate by RfbB, to GDP-mannose by RfbA, to GDP-4-keto-6-dideoxymannose by RfbD and to GDP-perosamine by RfbE. This final product would then serve as the substrate for the addition of the tetronate, which could then be polymerized into the O-antigen for transfer to the lipid A plus core oligosaccharide and export to the cell surface. The organization of these genes are such that one would expect them to be translationally coupled as part of the rfb operon. However, the absence of readily detectable promoter sequences suggests low levels of transcription, in line with other studies. The nucleotide sequence of these genes is absolutely conserved in the two isolates 569B (classical, Inaba) and O17 (El Tor, Ogawa) which were expected to show maximal sequence variation. This suggests very tight constraints on the micro-evolution within these sequences.Item Metadata only A review of the infection, genetics, and evolution of Neospora caninum: from the past to the present(Elsevier, 2013) Goodswen, S.; Kennedy, P.; Ellis, J.Abstract not availableItem Metadata only A single nucleotide polymorphism in an IgA1 protease gene determines Streptococcus pneumoniae adaptation to the middle ear during otitis media(Oxford University Press, 2021) Tikhomirova, A.; Trappetti, C.; Paton, J.C.; Watson-Haigh, N.; Wabnitz, D.; Jervis-Bardy, J.; Jardeleza, C.; Kidd, S.P.Factors facilitating the chronicity of otitis media (OM) in children are, to date, not fully understood. An understanding of molecular factors aiding bacterial persistence within the middle ear during OM could reveal pathways required for disease. This study performed a detailed analysis of Streptococcus pneumoniae populations isolated from the nasopharynx and middle ear of one OM case. Isolates were assessed for growth in vitro and infection in a mouse intranasal challenge model. Whole genome sequencing was performed to compare the nasopharyngeal and middle ear isolates. The middle ear isolate displayed a reduced rate of growth and enhanced potential to transit to the middle ear in a murine model. The middle ear population possessed a single nucleotide polymorphism (SNP) in the IgA1 protease gene igA, predicted to render its product non-functional. Allelic exchange mutagenesis of the igA alleles from the genetic variant middle ear and nasopharyngeal isolates was able to reverse the niche-adaptation phenotype in the murine model. These results indicate the potential role of a SNP in the gene encoding the IgA1 protease, in determining S. pneumoniae adaptation to the middle ear during chronic OM. In contrast, a functional IgA1 protease was associated with increased colonisation of the nasopharynx.Item Metadata only A T cell clone with three potential TCR alpha chain rearrangements expresses only one receptor combination at the cell surface(Elsevier Science, 1996) Hayball, J.; Jones, C.; Lamb, J.; Lake, R.In determining the T cell receptor (TcR) usage of various T cell clones that recognize peptide antigens derived from allergens, a particular clone (AC20) was found, that apparently expressed three different species of mRNA encoding alpha chains. The logical conclusion that the cells were not clonal was refuted by the finding of only a single beta chain rearrangement. One of the alpha chains (V alpha20), was not in frame, but two V alpha8 transcripts of different lengths were both potentially translatable. Sequence analysis suggested that the shorter transcript was generated by a secondary splice event from the longer, through the use of a splice donor sequence encoded by the J alpha38 gene segment. The efficiency of excision of the intervening sequence is such that approximately equal amounts of the long and short transcripts occur in the steady state pool of mRNA. This phenomenon has been reported previously in TcR alpha rearrangements, but it has never been made clear whether these truncated chains can form a functional TcR. Reconstitution of a TcR negative cell line with these transcripts showed that only the full length alpha chain was able to pair efficiently with the beta chain to generate a functional receptor at the cell surface.Item Metadata only Absence of high priority critically important antimicrobial resistance in Salmonella sp. isolated from Australian commercial egg layer environments(Elsevier, 2021) Veltman, T.; Jordan, D.; McDevitt, C.A.; Bell, J.; Howden, B.P.; Valcanis, M.; O'Dea, M.; Abraham, S.; Scott, P.; Kovac, J.H.; Chia, R.; Combs, B.; Chousalkar, K.; Wilson, T.; Trott, D.J.The development of antimicrobial resistance in foodborne pathogens is a growing public health concern. This study was undertaken to determine the antimicrobial susceptibility of Salmonella enterica subspecies enterica isolated from the Australian commercial egg layer industry. S. enterica subspecies enterica (n=307) isolated from Australian commercial layer flock environments (2015-2018) were obtained from reference, research and State Government laboratories from six Australian states. All Salmonella isolates were serotyped. Antimicrobial susceptibility testing (AST) for 16 antimicrobial agents was performed by broth microdilution. Antimicrobial resistance genes and sequence types (STs) were identified in significant isolates by whole genome sequencing (WGS). Three main serotypes were detected, S. Typhimurium (n=61, 19.9%), S. Senftenburg (n=45, 14.7%) and S. Agona (n=37, 12.1%). AST showed 293/307 (95.4%) isolates were susceptible to all tested antimicrobial agents and all isolates were susceptible to amoxicillin-clavulanate, azithromycin, ceftiofur, ceftriaxone, ciprofloxacin, colistin, florfenicol, gentamicin, kanamycin and trimethoprim-sulfamethoxazole. Low levels of non-susceptibility were observed to streptomycin (2.3%, n=7), sulfisoxazole (2.0%, n=6), chloramphenicol (1.3%, n=4) and tetracycline (1.0%, n=3). Very low levels of non-susceptibility were observed to ampicillin (2/307; 0.7%) and cefoxitin (2/307; 0.7%). Two isolates (S. Havana and S. Montevideo), exhibited multidrug-resistant phenotypes to streptomycin, sulfisoxazole and tetracycline and possessed corresponding antimicrobial resistance genes (aadA4, aac(6')-Iaa, sul1, tetB). One S. Typhimurium isolate was resistant to ampicillin and tetracycline, and possessed both tetA and blaTEM-1B. WGS also identified these isolates as belonging to ST4 (S. Montevideo), ST578 (S. Havana) and ST19 (S. Typhimurium). The absence of resistance to highest priority critically important antimicrobials as well as the extremely low level of AMR generally among Australian commercial egg layer Salmonella isolates likely reflect Australia's conservative antimicrobial registration policy in food-producing animals and low rates of antimicrobial use within the industry.Item Metadata only Absorption and presentation of antigens by epithelial cells of the small intestine: Hyptheses and predictions relating to the pathogenesis of coeliac disease(University of Adelaide, 1995) Mayrhofer, G.SummaryEffects of the route of antigen absorption and presentation in the small intestine are discussed in relation to outcomes from antigen exposure. It is proposed that antigens presented by epithelial cells to intrinsic T cell populations in the mucosa induce anti‐inflammatory immune responses, while antigens presented to conventional T cells by lamina propria APC predisposes to inflammation. The pathogenesis of coeliac disease is attributed to three factors: a defect in antigen processing by epithelial cells; the intrinsic properties of the gliadins; and the HLA‐D haplotype of the individual. It is suggested that MHC class II molecules play a role in antigen uptake by enterocytes, in addition to their function in antigen presentation to T cells.Item Open Access Activity of bacteriophages in removing biofilms of pseudomonas aeruginosa isolates from chronic rhinosinusitis patients(Frontiers, 2017) Fong, S.; Drilling, A.; Morales, S.; Cornet, M.; Woodworth, B.; Fokkens, W.; Psaltis, A.; Vreugde, S.; Wormald, P.Introduction: Pseudomonas aeruginosa infections are prevalent amongst chronic rhinosinusitis (CRS) sufferers. Many P. aeruginosa strains form biofilms, leading to treatment failure. Lytic bacteriophages (phages) are viruses that infect, replicate within, and lyse bacteria, causing bacterial death. Aim: To assess the activity of a phage cocktail in eradicating biofilms of ex vivo P.aeruginosa isolates from CRS patients. Methods: P. aeruginosa isolates from CRS patients with and without cystic fibrosis (CF) across three continents were multi-locus sequence typed and tested for antibiotic resistance. Biofilms grown in vitro were treated with a cocktail of four phages (CT-PA). Biofilm biomass was measured after 24 and 48 h, using a crystal violet assay. Phage titrations were performed to confirm replication of the phages. A linear mixed effects model was applied to assess the effects of treatment, time, CF status, and multidrug resistance on the biomass of the biofilm. Results: The isolates included 44 strain types. CT-PA treatment significantly reduced biofilm biomass at both 24 and 48 h post-treatment (p < 0.0001), regardless of CF status or antibiotic resistance. Biomass was decreased by a median of 76% at 48 h. Decrease in biofilm was accompanied by a rise in phage titres for all except one strain. Conclusion: A single dose of phages is able to significantly reduce biofilms formed in vitro by a range of P.aeruginosa isolates from CRS patients. This represents an exciting potential and novel targeted treatment for P. aeruginosa biofilm infections and multidrug resistant bacteria.Item Open Access Adjuvant selection for influenza and RSV prefusion subunit vaccines(MDPI, 2021) Isaacs, A.; Li, Z.; Cheung, S.T.M.; Wijesundara, D.K.; McMillan, C.L.D.; Modhiran, N.; Young, P.R.; Ranasinghe, C.; Watterson, D.; Chappell, K.J.Subunit vaccines exhibit favorable safety and immunogenicity profiles and can be designed to mimic native antigen structures. However, pairing with an appropriate adjuvant is imperative in order to elicit effective humoral and cellular immune responses. In this study, we aimed to determine an optimal adjuvant pairing with the prefusion form of influenza haemagglutinin (HA) or respiratory syncytial virus (RSV) fusion (F) subunit vaccines in BALB/c mice in order to inform future subunit vaccine adjuvant selection. We tested a panel of adjuvants, including aluminum hydroxide (alhydrogel), QS21, Addavax, Addavax with QS21 (AdQS21), and Army Liposome Formulation 55 with monophosphoryl lipid A and QS21 (ALF55). We found that all adjuvants elicited robust humoral responses in comparison to placebo, with the induction of potent neutralizing antibodies observed in all adjuvanted groups against influenza and in AdQS21, alhydrogel, and ALF55 against RSV. Upon HA vaccination, we observed that none of the adjuvants were able to significantly increase the frequency of CD4+ and CD8+ IFN-γ+ cells when compared to unadjuvanted antigen. The varying responses to antigens with each adjuvant highlights that those adjuvants most suited for pairing purposes can vary depending on the antigen used and/or the desired immune response. We therefore suggest that an adjuvant trial for different subunit vaccines in development would likely be necessary in preclinical studies.Item Metadata only Aire-Deficient C57BL/6 Mice Mimicking the Common Human 13-Base Pair Deletion Mutation Present with Only a Mild Autoimmune Phenotype(Amer Assoc Immunologists, 2009) Hubert, F.; Kinkel, S.; Crewther, P.; Cannon, P.; Webster, K.; Link, M.; Uibo, R.; O'Bryan, M.; Meager, A.; Forehan, S.; Smyth, G.; Mittaz, L.; Antonarakis, S.; Peterson, P.; Heath, W.; Scott, H.Autoimmune regulator (AIRE) is an important transcription regulator that mediates a role in central tolerance via promoting the "promiscuous" expression of tissue-specific Ags in the thymus. Although several mouse models of Aire deficiency have been described, none has analyzed the phenotype induced by a mutation that emulates the common 13-bp deletion in human APECED (autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy) by disrupting the first plant homeodomain in exon 8. Aire-deficient mice with a corresponding mutation showed some disturbance of the medullary epithelial compartment, but at the phenotypic level their T cell compartment appeared relatively normal in the thymus and periphery. An increase in the number of activated T cells was evident, and autoantibodies against several organs were detected. At the histological level, lymphocytic infiltration of several organs indicated the development of autoimmunity, although symptoms were mild and the quality of life for Aire-deficient mice appeared equivalent to wild-type littermates, with the exception of male infertility. Vbeta and CDR3 length analysis suggested that each Aire-deficient mouse developed its own polyclonal autoimmune repertoire. Finally, given the prevalence of candidiasis in APECED patients, we examined the control of infection with Candida albicans in Aire-deficient mice. No increase in disease susceptibility was found for either oral or systemic infection. These observations support the view that additional genetic and/or environmental factors contribute substantially to the overt nature of autoimmunity associated with Aire mutations, even for mutations identical to those found in humans with APECED.