Immunology and Microbiology Thrombomodulin Protects Against Bacterial Keratitis, Is Anti-Inflammatory, but Not Angiogenic

PURPOSE. Thrombomodulin (TM) is a multidomain, transmembrane protein with antiinflammatory properties. Thrombomodulin domain (D) 1 is lectin-like, interacting with Lewis Y antigen on lipopolysaccharide, and with HMGB1, while TMD23 is associated with angiogenic and anti-inflammatory functions. Thus, we tested if TM is protective against Pseudomonas aeruginosa keratitis and whether it enhanced corneal vascularity. METHODS. Eyes of C57BL/6 (B6) mice were injected with recombinant TM (rTM), rTMD1, or PBS subconjunctivally before and intraperitoneally after infection with P. aeruginosa. Clinical scores, photography with a slit lamp, RT-PCR, ELISA, myeloperoxidase (MPO) assay, viable bacterial plate counts, and India ink perfusion were used to assess the disease response and corneal vascularity (rTM only). RESULTS. Recombinant TM versus PBS treatment reduced clinical scores and corneal opacity. Corneal mRNA levels for HMGB1 were unchanged, but proinflammatory molecules IL-1b, CXCL2, NF-jB, TLR4, and RAGE were decreased; anti-inflammatory molecules SIGIRR and ST2 were increased. ELISA confirmed the mRNA data for HMGB1, IL-1b, and CXCL2 proteins. Both neutrophil influx and viable bacterial plate counts also were decreased after rTM treatment. Protein levels for angiogenic molecules VEGF, VEGFR-1, and VEGFR-2 were measured at 5 days post infection and were not different or reduced significantly after rTM treatment. Further, perfusion with India ink revealed similar vessel ingrowth between the two groups. Similar studies were performed with rTMD1, but disease severity, mRNA, proteins, MPO, and plate counts were not changed from controls. CONCLUSIONS. These data provide evidence that rTM treatment is protective against bacterial keratitis, does not reduce HMGB1, and is not angiogenic

Characterization of Three Ocular Clinical Isolates of P. aeruginosa: Viability, Biofilm Formation, Adherence, Infectivity, and Effects of Glycyrrhizin

We selectively characterized three isolates from Pseudomonas aeruginosa keratitis patients and how glycyrrhizin (GLY) affected them. Type III toxins were determined using polymerase chain reaction (PCR). Minimum Inhibitory Concentration (MIC) of GLY and assays for its effects on: time kill, bacterial permeability, and biofilm/adhesion were done. In vivo, C57BL/6 (B6) mice were treated topically with GLY after G81007 infection. Clinical score, photography with a slit lamp and RT-PCR were used to assess treatment effects. Isolates expressed exoS and exoT, but not exoU. MIC for all isolates was 40 mg/mL GLY and bacteriostatic effects were seen for G81007 after treatment using time kill assays. From viability testing, GLY treatment significantly increased the number of permeabilized bacteria (live/dead assay). Isolates 070490 and G81007 formed more biofilms compared with R59733 and PAO1 (control). GLY-treated bacteria had diminished biofilm compared with controls for all isolates. GLY reduced adherence of the G81007 isolate to cultured cells and affected specific biofilm associated systems tested by reverse transcription PCR (RT-PCR). In vivo, after G81007 infection, GLY treatment reduced clinical score and messenger RNA (mRNA) expression of IL-1β, TNF-α, CXCL2 and HMGB1. This study provides evidence that GLY is bacteriostatic for G81007. It also affects biofilm production, adherence to cultured cells, and an improved keratitis outcome

Refined families of dothideomycetes: Dothideomycetidae and pleosporomycetidae

The class Dothideomycetes is the largest and most ecologically diverse class of fungi, comprising endophytes, epiphytes, saprobes, human and plant pathogens, lichens, and lichenicolous, nematode trapping and rock-inhabiting taxa. Members of this class are mainly characterized by bitunicate asci with fissitunicate dehiscence, and occur on broad range of hosts in aquatic and terrestrial habitats. Since the last monograph of families of Dothideomycetes in 2013, numerous novel species, genera, families and orders have been discovered. This has expanded information which has led to the modern classification in Dothideomycetes. In this paper, we provide a refined updated document on families of Dothideomycetes with emphasis on Dothideomycetidae and Pleosporomycetidae. We accept three orders with 25 families and four orders with 94 families in Dothideomycetidae and Pleosporomycetidae, respectively. The new family Paralophiostomataceae is introduced in Pleosporales. Each family is provided with an updated description, notes, including figures to represent the morphology, list of accepted genera, and economic and ecological significances. We also provide an overall phylogenetic tree of families in Dothideomycetes based on combined analysis of LSU, SSU, rpb-2 and tef1 sequence data, and phylogenetic trees for each order in Dothideomycetidae and Pleosporomycetidae. Family-level trees are provided for the families which include several genera such as Mycosphaerellaceae and Teratosphaeriaceae. Two new genera (Ligninsphaeriopsis and Paralophiostoma) are introduced. Five new species (Biatrisopora borsei, Comoclathris galatellae, Ligninsphaeriopsis thailandica, Paralophiostoma hysterioides and Torula thailandica) are introduced based on morphology and phylogeny, together with nine new reports and seven new collections from different families

Redined families of Dothideomycetes: orders and families incertain in Dothideomycetes

peer reviewedNumerous new taxa and classifications of Dothideomycetes have been published following the last monograph of families of Dothideomycetes in 2013. A recent publication by Honsanan et al. in 2020 expanded information of families in Dothideo- mycetidae and Pleosporomycetidae with modern classifications. In this paper, we provide a refined updated document on orders and families incertae sedis of Dothideomycetes. Each family is provided with an updated description, notes, including figures to represent the morphology, a list of accepted genera, and economic and ecological significances. We also provide phylogenetic trees for each order. In this study, 31 orders which consist 50 families are assigned as orders incertae sedis in Dothideomycetes, and 41 families are treated as families incertae sedis due to lack of molecular or morphological evidence. The new order, Catinellales, and four new families, Catinellaceae, Morenoinaceae Neobuelliellaceae and Thyrinulaceae are introduced. Seven genera (Neobuelliella, Pseudomicrothyrium, Flagellostrigula, Swinscowia, Macroconstrictolumina, Pseudobogoriella, and Schummia) are introduced. Seven new species (Acrospermum urticae, Bogoriella complexoluminata, Dothiorella ostryae, Dyfrolomyces distoseptatus, Macroconstrictolumina megalateralis, Patellaria microspora, and Pseu- domicrothyrium thailandicum) are introduced base on morphology and phylogeny, together with two new records/reports and five new collections from different families. Ninety new combinations are also provided in this paper