Association of anurans with pathogenic fungi

In a study of 450 Amazonian anurans, we isolated yeasts and yeast-like fungi from 54 animals (Bufo granulosus, B. marinus, Dendrophrynyscus sp., Hyla geographica, H. lanciformes, Ololygon rubra, Adenomera hylaedactyla, Eleutherodactylus fenestratus, Leptodactylus fuscus, L. ocellatus, L. pentadactylus). The internal organs of these animals did not show any macroscopic anomaly nor histopathology. We recovered 105 fungal isolates from the anuran liver, lung, kidney, spleen, heart and gonad. The isolates were made up of 30 fungal species, 9 of which (48 isolates, 46%) were fungi with known pathogenic potentials, namely: Candida guilliermondii, C. parapsilosis, C. tropicalis, C. glabrata, Geotrichum candidum, Aureobasidium pullulans, Wangiella dermatitidis, Trichosporon cutaneum and Exophiala werneckii. Eleven animals harbored identical fungi in more than one of their internal organs; seven animals had more than one fungal species colonizing a single organ. Our findings indicated probable natural subclinical infections of candidiasis, geotrichosis or phaeohyphomycosis, and also symbiotic presence of non-pathogenic fungi among neotropical anurans.     

Fine structure of early human embryos frozen with 1,2 propanediol

Previous studies by a French group (Fertil Steril 44:645–651, 1985) have shown that two-to eight-cell human embryos can survive slow freeze-thawing with propanediol in a biological freezer. These embryos were assessed for morphological appearance by phase-contrast microscopy. We assessed the structure of 25 frozen-thawed one- to 12-cell embryos, obtained from our in vitro fertilization (IVF) and GIFT programmes, by phase-contrast and electron microscopy, using the same method of cryopreservation. One-fourth of the embryos examined had all cells intact, and more than one-half the embryos had over 50% of their cells well preserved. Some of these embryos had unequal blastomeres and cytoplasmic fragments. Ultrastructural assessment revealed good preservation of fine structure in the intact blastomeres of all embryos and maintenance of cell-to-cell contacts. Most cytoplasmic organelles, cell membranes, and nuclei were well preserved compared to nonfrozen controls. The cells that were cryoinjured showed varying degrees of disorganization of the cell membrane, cytosol, and cellular membranes, including swelling and disruption of the nuclear envelope. Disruption of the zona was somewhat rare. Small cytoplasmic fragments were less prone to cryoinjury than blastomeres. The use of propanediol for embryo cryopreservation seems to be feasible; frozen embryos with more than 50% cells intact have produced 10 pregnancies after embryo transfer (Fertil Steril 46:268–272, 1986). Replacement of 17 frozen embryos in seven patients has resulted in a twin pregnancy in Singapore. However, the effects of freezing on the mitotic spindles of embryonic cells need to be investigated further.     

Isolation of fungi from bats of the Amazon basin

A total of 2,886 bats captured in the Amazon Basin of Brazil were processed for the isolation of fungi. From the livers, spleens, and lungs of 155 bats (5.4%), 186 fungal isolates of the genera Candida (123 isolates), Trichosporon (26 isolates), Torulopsis (25 isolates), Kluyveromyces (11 isolates), and Geotrichum (1 isolate) were recovered. Seven known pathogenic species were present: Candida parapsilosis, C. guilliermondii, C. albicans, C. stellatoidea, C. pseudotropicalis, Trichosporon beigelii, and Torulopsis glabrata. Twenty-three culture-positive bats showed identical fungal colonization in multiple organs or mixed colonization in a single organ. The fungal isolation rates for individual bat species varied from 1 fungus per 87 bats to 3 fungi per 13 bats, and the mycoflora diversity for members of an individual fungus-bearing bat species varied from 16 fungi per 40 bats to 7 fungi per 6 bats. Of the 38 fungal species isolated, 36 had not been previously described as in vivo bat isolates. Of the 27 culture-positive bat species, 21 had not been previously described as mammalian hosts for medically or nonmedically important fungi.     

Pleiotrophin Gene Therapy for Peripheral Ischemia: Evaluation of Full-Length and Truncated Gene Variants

Pleiotrophin (PTN) is a growth factor with both pro-angiogenic and limited pro-tumorigenic activity. We evaluated the potential for PTN to be used for safe angiogenic gene therapy using the full length gene and a truncated gene variant lacking the domain implicated in tumorigenesis. Mouse myoblasts were transduced to express full length or truncated PTN (PTN or T-PTN), along with a LacZ reporter gene, and injected into mouse limb muscle and myocardium. In cultured myoblasts, PTN was expressed and secreted via the Golgi apparatus, but T-PTN was not properly secreted. Nonetheless, no evidence of uncontrolled growth was observed in cells expressing either form of PTN. PTN gene delivery to myocardium, and non-ischemic skeletal muscle, did not result in a detectable change in vascularity or function. In ischemic hindlimb at 14 days post-implantation, intramuscular injection with PTN-expressing myoblasts led to a significant increase in skin perfusion and muscle arteriole density. We conclude that (1) delivery of the full length PTN gene to muscle can be accomplished without tumorigenesis, (2) the truncated PTN gene may be difficult to use in a gene therapy context due to inefficient secretion, (3) PTN gene delivery leads to functional benefit in the mouse acute ischemic hindlimb model.     

Reversible Control by Vitamin D of Granulocytes and Bacteria in the Lungs of Mice: An Ovalbumin-Induced Model of Allergic Airway Disease

Vitamin D may be essential for restricting the development and severity of allergic diseases and asthma, but a direct causal link between vitamin D deficiency and asthma has yet to be established. We have developed a ‘low dose’ model of allergic airway disease induced by intraperitoneal injection with ovalbumin (1 µg) and aluminium hydroxide (0.2 mg) in which characteristics of atopic asthma are recapitulated, including airway hyperresponsiveness, antigen-specific immunoglobulin type-E and lung inflammation. We assessed the effects of vitamin D deficiency throughout life (from conception until adulthood) on the severity of ovalbumin-induced allergic airway disease in vitamin D-replete and -deficient BALB/c mice using this model. Vitamin D had protective effects such that deficiency significantly enhanced eosinophil and neutrophil numbers in the bronchoalveolar lavage fluid of male but not female mice. Vitamin D also suppressed the proliferation and T helper cell type-2 cytokine-secreting capacity of airway-draining lymph node cells from both male and female mice. Supplementation of initially vitamin D-deficient mice with vitamin D for four weeks returned serum 25-hydroxyvitamin D to levels observed in initially vitamin D-replete mice, and also suppressed eosinophil and neutrophil numbers in the bronchoalveolar lavage fluid of male mice. Using generic 16 S rRNA primers, increased bacterial levels were detected in the lungs of initially vitamin D-deficient male mice, which were also reduced by vitamin D supplementation. These results indicate that vitamin D controls granulocyte levels in the bronchoalveolar lavage fluid in an allergen-sensitive manner, and may contribute towards the severity of asthma in a gender-specific fashion through regulation of respiratory bacteria.     

Engineering flavonoid glycosyltransferases for enhanced catalytic efficiency and extended sugar-donor selectivity

Flavonoids are predominantly found as glycosides in plants. The glycosylation of flavonoids is mediated by uridine diphosphate-dependent glycosyltransferases (UGT). UGTs attach various sugars, including arabinose, glucose, galactose, xylose, and glucuronic acid, to flavonoid aglycones. Two UGTs isolated from Arabidopsis thaliana, AtUGT78D2 and AtUGT78D3, showed 89 % amino acid sequence similarity (75 % amino acid sequence identity) and both attached a sugar to the 3-hydroxyl group of flavonols using a UDP-sugar. The two enzymes used UDP-glucose and UDP-arabinose, respectively, and AtUGT78D2 was approximately 90-fold more efficient than AtUGT78D3 when judged by the k _cat/K _m value. Domain exchanges between AtUGT78D2 and AtUGT78D3 were carried out to find UGTs with better catalytic efficiency for UDP-arabinose and exhibiting dual sugar selectivity. Among 19 fusion proteins examined, three showed dual sugar selectivity, and one fusion protein had better catalytic efficiency for UDP-arabinose compared with AtUGT78D3. Using molecular modeling, the changes in enzymatic properties in the chimeric proteins were elucidated. To the best of our knowledge, this is the first report on the construction of fusion proteins with expanded sugar-donor range and enhanced catalytic efficiencies for sugar donors.     

Hagfish phylogeny and taxonomy,with description of the new genus Rubicundus (Craniata,Myxinidae)

A recent phylogenetic analysis of the Myxinidae based on the 16S rRNA gene resulted in synonymization of Paramyxine with Eptatretus. This created homonymy of Paramyxine fernholmi with Eptatretus fernholmi and Paramyxine wisneri with Eptatretus wisneri. In order to resolve this nomenclatural dilemma, we made a more extensive phylogenetic assessment of the Myxinidae and examined the nomenclature of the family. We used 75 sequences (37 of which new for this study) of a 561 bp fragment of the 16S rRNA gene, representing 33 species, and 72 sequences (37 of which new for this study) of a 687 bp fragment of the cytochrome c oxidase subunit I (COI) gene, representing 23 species, to reconstruct the phylogeny of Myxinidae. The monophyly of the subfamily Myxininae, traditionally characterized by having a single pair of external gill openings, was rejected (0.50 Bayesian posterior probability) by the 16S analysis, but supported by the COI and combined COI+16S analyses (0.99 and 0.81 Bpp, respectively). The monophyly of the subfamily Eptatretinae, characterized by having several pairs of external gill openings, was not supported by the 16S analysis and rejected by the COI and combined COI+16S analysis due to the placement of Eptatretus lopheliae as the earliest branch of Myxinidae (0.71 and 0.57 Bpp, respectively). Eptatretus lopheliae and Eptatretus rubicundus formed a monophyletic group and were allocated to a new genus, Rubicundus, characterized by the presence of an elongated tubular nostril and reddish coloration. A new monotypic subfamily, Rubicundinae, was proposed for Rubicundus. The synonymy of the genera Paramyxine and Quadratus with Eptatretus was confirmed. E. fernholmi is renamed Eptatretus luzonicus. Eptatretus wisneri was renamed Eptatretus bobwisneri. Petromyzon cirrhatus Forster, 1801, Homea banksii Fleming, 1822, and Bdellostoma forsteri Müller, 1836 are synonyms, but no type specimens are known to exist. Petromyzon cirrhatus was designated as type species of Eptatretus, conserving present usage. Gastrobranchus dombeyi Shaw, 1804 has priority over other names for Chilean myxinids. Bdellostoma stoutii was designated as type species of Polistotrema Gill. The validity of the Western Atlantic Myxine limosa as distinct from the Eastern Atlantic Myxine glutinosa was confirmed.     

Abnormal Organization of White Matter Network in Patients with No Dementia after Ischemic Stroke

Structural changes after ischemic stroke could affect information communication extensively in the brain network. It is likely that the defects in the white matter (WM) network play a key role in information interchange. In this study, we used graph theoretical analysis to examine potential organization alteration in the WM network architecture derived from diffusion tensor images from subjects with no dementia and experienced stroke in the past 5.4–14.8 months (N = 47, Mini-Mental Screening Examination, MMSE range 18–30), compared with a normal control group with 44 age and gender-matched healthy volunteers (MMSE range 26–30). Region-wise connectivity was derived from fiber connection density of 90 different cortical and subcortical parcellations across the whole brain. Both normal controls and patients with chronic stroke exhibited efficient small-world properties in their WM structural networks. Compared with normal controls, topological efficiency was basically unaltered in the patients with chronic stroke, as reflected by unchanged local and global clustering coefficient, characteristic path length, and regional efficiency. No significant difference in hub distribution was found between normal control and patient groups. Patients with chronic stroke, however, were found to have reduced betweenness centrality and predominantly located in the orbitofrontal cortex, whereas increased betweenness centrality and vulnerability were observed in parietal-occipital cortex. The National Institutes of Health Stroke Scale (NIHSS) score of patient is correlated with the betweenness centrality of right pallidum and local clustering coefficient of left superior occipital gyrus. Our findings suggest that patients with chronic stroke still exhibit efficient small-world organization and unaltered topological efficiency, with altered topology at orbitofrontal cortex and parietal-occipital cortex in the overall structural network. Findings from this study could help in understanding the mechanism of cognitive impairment and functional compensation occurred in patients with chronic stroke.     

Early Diagnosis of Neurodegenerative Diseases – The Long Awaited Holy Grail and Bottleneck of Modern Brain Research – 19th HUPO BPP Workshop

The HUPO Brain Proteome Project (HUPO BPP) held its 19th workshop in Dortmund, Germany, from May 22 to 24, 2013. The focus of the spring workshop was on strategies and developments concerning early diagnosis of neurodegenerative diseases