Genome sequences of two isolates of the yeast Candida zeylanoides: UCD849 from soil in Ireland, and AWD from an African wild dog.

We report genome sequences of two new isolates of the budding yeast Candida zeylanoides. Strain UCD849 from soil in Ireland was assembled into eight complete chromosomes. Strain AWD from an African Wild Dog in a US zoo was sequenced to draft level. The assemblies are 10.6 Mb and 99.57% identical.

Characterization of Antibiotic-Resistant Stenotrophomonas Isolates from Painted Turtles Living in the Wild.

Stenotrophomonas maltophilia is a ubiquitous multidrug-resistant opportunistic pathogen commonly associated with nosocomial infections. The purpose of this study was to isolate and characterize extended-spectrum beta-lactamase (ESBL) producing bacteria from painted turtles (Chrysemys picta) living in the wild and captured in southeastern Wisconsin. Fecal samples from ten turtles were examined for ESBL producing bacteria after incubation on HardyCHROM™ ESBL agar. Two isolates were cultivated and identified by 16S rRNA gene sequencing and whole genome sequencing (WGS) as Stenotrophomonas sp. 9A and S. maltophilia 15A. They were multidrug-resistant, as determined by antibiotic susceptibility testing. Stenotrophomonas sp. 9A was found to produce an extended spectrum beta-lactamase (ESBL) and both isolates were found to be carbapenem-resistant. EDTA-modified carbapenem inactivation method (eCIM) and the modified carbapenem inactivation method (mCIM) tests were used to examine the carbapenemase production and the test results were negative. Through WGS several antimicrobial resistance genes were identified in S. maltophilia 15A. For example a chromosomal L1 ß-lactamase gene, which is known to hydrolyze carbapenems, a L2 ß-lactamase gene, genes for the efflux systems smeABC and smeDEF and the aminoglycosides resistance genes aac(6')-lz and aph(3')-llc were found. An L2 ß-lactamase gene in Stenotrophomonas sp. 9A was identified through WGS.

Draft Genome Sequence of Paenibacillus odorifer V, Isolated from the Fecal Material of a Rabbit.

Here, we report the draft genome sequence of Paenibacillus odorifer strain V, which was isolated from the fecal material of a rabbit living in the wild. The genome size is 6,863,583 bp, with 44.35 mol% G+C content.

Pneumonia in endangered aquatic mammals and the need for developing low-coverage vaccination for their management and conservation.

Anthropogenic activities can lead to several devastating effects on the environment. The pollutants, which include the discharge of effluents, runoffs in the form of different lethal and sub-lethal concentrations of pesticides, heavy metals, and other contaminants, can harm exposed fauna and flora. The aquatic environment is the ultimate destination for many pollutants which negatively affect aquatic biodiversity and even can cause a species to become extinct. A pollutant can directly affect the behavior of an animal, disrupt cellular systems, and impair the immune system. This harm can be reduced and even mitigated by adopting proper approaches for the conservation of the target biota. Among aquatic organisms, cetaceans, such as the Yangtze finless porpoise, Irrawaddy dolphin, Ganges River dolphin, Amazon River dolphin, and Indus River dolphin, are at a higher risk of extinction because of lack of knowledge and research, and thus insufficient information with respect to their conservation status, management, and policies. Pneumonia is one of the leading causes of mass mortalities of cetaceans. This article reviews the limited research reported on stress and pneumonia induced by pollution, stress-induced pneumonia and immunosuppression, pneumonia-caused mass mortalities of aquatic mammals, and vaccination in wildlife with a specific focus on aquatic mammals, the role of genomics in vaccine development and vaccination, and the major challenges in vaccine development for biodiversity conservation.

First case of Shewanella indica isolated from a Bryde's whale (Balaenoptera edeni) stranded in the northern Beibu Gulf, China.

The purpose of this study was to culture and characterise bacteria from an intact abscess on the skin of a dead Bryde's whale (Balaenoptera edeni) which stranded in the northern Beibu Gulf, China. To grow bacteria, samples from the abscess were added to blood agar. After incubation, yellowish mucous colonies were visualized. The bacterium was firstly recognised as Shewanella algae by the VITEK® 2 System. However, by using 16S rRNA gene sequencing the bacterium was finally identified as S. indica. To characterise the bacterium, antibiotic susceptibility and virulence factors, such as hemolysis and biofilm formation were investigated. The bacterium is capable of ß-hemolysis and biofilm formation and it is also sensitive to several different classes of antibiotics, such as ß-lactams, quinolones, and aminoglycosides. To date there have been no reports of this bacterium causing infections in humans or animals. However, in this study we described the first case of S. indica isolated from an intact abscess on the back of a Bryde's whale.

Immune Responses of the Critically Endangered Yangtze Finless Porpoises (Neophocaena asiaeorientalis ssp. asiaeorientalis) to Escalating Anthropogenic Stressors in the Wild and Seminatural Environments.

Increasing anthropogenic stressors are potential threats to biodiversity conservation and management of Yangtze finless porpoises (YFPs). The objective of this study was to indirectly compare the habitat quality of a natural reserve, Poyang Lake and a seminatural reserve, the Tian-E-Zhou Oxbow (TZO) in terms of anthropogenic stressors by investigating different stress and immunological parameters in the blood of YFPs. Samples from a total of 74 YFPs from the TZO (n = 43) and Poyang Lake (n = 31) were collected and analyzed. The animals were divided into ontogenetic groups: male calf, female calf, juvenile female, juvenile male, and adult male, and reproductive groups: pregnant female, lactating female, and pregnant plus lactating. The blood from all the animals was analyzed for general stress (HSP14, SOD1, TXN, and FTL), metabolic stress (ACAT2 and THRA), and immunity-related genes (IL12p40, IFNγ, TNFα; IL1α, IL1ra, COX2, CRPL, IL4, and IL8) using qPCR. YFPs living in Poyang Lake showed an increased relative expression pattern for IFNγ, IL1ra, IL4, ACAT2, and CRPL across all the ontogenetic groups with significantly higher expression in adult males. In contrast, YFPs living in the TZO showed a significantly higher expression in 13 of 15 genes analyzed in the male calf group. Across the reproductive states for porpoises living in Poyang Lake, eight of the 15 genes in the pregnant female and three of the 15 genes in the pregnant plus lactating group had a significantly higher expression level. However, in YFPs living in the TZO, eight of the 15 genes showed significantly higher expression in the pregnant and lactating groups. There was significantly a higher expression of most of the genes in porpoises living in the TZO compared to the age-matched groups from porpoises living in Poyang Lake. The exception was the pregnant female group. The higher relative expression of stress and immune genes in the TZO porpoise population compared to porpoises living in Poyang Lake suggests the effects of worsening habitat quality, possibly indicating water pollution and lack of feeding resources.

Hematologic and biochemical differences between two free ranging Yangtze finless porpoise populations: The implications of habitat.

The goals of this study were to compare the serum chemistry and hematology values of wild and semi-natural free-ranging Yangtze Finless Porpoises (Neophocaena asiaeorientalis ssp. asiaeorientalis) populations and to ascertain how these values change with the different environmental condition. For this study, samples were collected from 81 YFPs, 35 living in the wild and 46 living in a semi-natural reserve. Each population was divided into 8 life history categories; Male Calf, Female Calf, Juvenile Male, Juvenile Female, Adult Male, Pregnant, Lactating and Pregnant plus Lactating. Statistically significant differences in the various parameters were observed in the same life history categories for both populations. Generally, Lipid Profile, Hepatic Enzymes, Creatine Kinase, Red Blood Cells, Hemoglobin, Hematocrit and Neutrophils were significantly higher in the Tian-E-Zhou Oxbow population while, Creatinine, Phosphate, Lactate Dehydrogenase, Bilirubin and Lymphocytes were significantly higher in the Poyang Lake YFPs. Across the groups in the Tian-E-Zhou Oxbow population, a significant decrease in serum Albumin, Alkaline Phosphatase and Calcium, while a significant increase in the Neutrophils and Platelets was observed. Similarly, in the Poyang Lake, Alkaline Phosphatase levels in the Female Calves group, High Density Lipoprotein Cholesterol in Lactating group, basophil counts in Pregnant plus Lactating group, lymphocytes counts in Juvenile Females group and Globulin and Total Protein levels in Pregnant group were significantly higher. This study in health assessments can help us to understand the effect of sex, age, reproductive status and environmental conditions on the well-being of Yangtze Finless Porpoises.

Enterococcus crotali sp. nov., isolated from faecal material of a timber rattlesnake.

A facultatively anaerobic, Gram-stain-positive bacterium, designated ETRF1T, was found in faecal material of a timber rattlesnake (Crotalus horridus). Based on a comparative 16S rRNA gene sequence analysis, the isolate was assigned to the genus Enterococcus. The 16S rRNA gene sequence of strain ETRF1T showed >97 % similarity to that of the type strains of Enterococcus rotai, E. caccae, E. silesiacus, E haemoperoxidus, E. ureasiticus, E. moraviensis, E. plantarum, E. quebecensis, E. ureilyticus, E. termitis, E. rivorum and E. faecalis. The organism could be distinguished from these 12 phylogenetically related enterococci using conventional biochemical testing, the Rapid ID32 Strep system, comparative pheS and rpoA gene sequence analysis, and comparative whole genome sequence analysis. The estimated in silico DNA-DNA hybridization values were <70 %, and average nucleotide identity values were <96 %, when compared to these 12 species, further validating that ETRF1T represents a unique species within the genus Enterococcus. On the basis of these analyses, strain ETRF1T (=CCUG 65857T=LMG 28312T) is proposed as the type strain of a novel species, Enterococcus crotali sp. nov.

Rapid identification of Robinsoniella peoriensis using specific 16S rRNA gene PCR primers.

Robinsoniella peoriensis is a Gram-positive, spore-forming anaerobic bacterium initially isolated and characterized from swine manure and feces. Since then strains of this species have been identified from a variety of mammalian and other GI tracts. More recently strains of this species have been isolated from a plethora of human infections. Therefore, it is of great interest to develop methods to rapidly identify this microorganism in the medical and other laboratories. This report describes the use of PCR primers targeting the 16S rRNA gene of R. peoriensis to identify strains of this bacterium.

Complete Genome Sequences of Enterococcus rotai LMG 26678T and Enterococcus silesiacus LMG 23085T.

The inclusion of molecular methods in the characterization of the novel species Enterococcus horridus necessitated the sequencing and assembly of the genomes of the closely related Enterococcus rotai and Enterococcus silesiacus Sequencing using Illumina technology in combination with optical mapping led to the generation of closed genomes for both isolates.

Draft Genome Sequence of Robinsoniella peoriensis Strain WTD, Isolated from the Fecal Material of a Wood Turtle.

Here, we report the draft genome of Robinsoniella peoriensis strain WTD, which was isolated from the fecal material of a wood turtle. The genome size was 7,391,415 bp with 41.1 mol% G+C.

Draft Genome Sequence of Clostridium mangenotii TR, Isolated from the Fecal Material of a Timber Rattlesnake.

Here, we report the draft genome sequence of Clostridium mangenotii strain TR, which was isolated from the fecal material of a timber rattlesnake. This bacterium is nonpathogenic but contains 68 genes involved in virulence, disease, and defense.

The effects of sodium sulfate, glycosaminoglycans, and Congo red on the structure, stability, and amyloid formation of an immunoglobulin light-chain protein.

Light-chain amyloidosis (AL) is characterized by immunoglobulin light-chain fragments aggregating into amyloid fibrils that deposit extracellularly in vital organs such as the kidney, the heart, and the liver, resulting in tissue degeneration and organ failure, leading to death. Cardiac involvement is found in 50% of AL patients and presents the most severe cases with a life expectancy of less than a year after diagnosis. In this study, we have characterized the variable domain of a cardiac AL patient light chain called AL-09. AL-09 folds as a beta-sheet and is capable of forming amyloid fibrils both in the presence of sodium sulfate and in self-seeded reactions under physiological conditions. Glycosaminoglycans such as dermatan sulfate and heparin promote amyloid formation of self-seeded AL-09 reactions, while the glycosaminoglycan chondroitin sulfate A stabilized oligomeric intermediates and did not elongate the preformed fibrils (nucleus) present in the reaction. Finally, the histological dye Congo red, known to bind to the cross beta-sheet structure of amyloid fibrils, inhibits AL-09 amyloid fibril formation in the presence of sodium sulfate and in self-seeded reactions. This paper provides insight into the impact of different reagents on light-chain stability, structure, amyloid fibril formation, and inhibition.

The cAMP-specific phosphodiesterase 4B mediates Abeta-induced microglial activation.

Microglial activation is a key player in the degenerative process that accompanies the deposition of amyloid-beta (Abeta) peptide into senile plaques in Alzheimer's disease (AD) patients. The goal of this study is to identify novel genes involved in microglial activation in response to Abeta peptide. Prompted by the fact that soluble Abeta(1-42) (sAbeta(1-42))-stimulated primary rat microglia produce more tumor necrosis factor-alpha (TNF-alpha) than fibrillar Abeta(1-42) (fAbeta(1-42))-stimulated microglia, we examined gene expression in these cells following stimulation using cDNA arrays. This analysis confirms the upregulation caused by both sAbeta(1-42) and fAbeta(1-42) of pro-inflammatory molecules such as TNF-alpha, interleukin-1beta and macrophage inflammatory protein-1alpha. In addition, other transcripts not previously described in the context of Abeta-induced microglial activation were identified. The modulation of some of these genes within microglial cells seems to be specific to sAbeta(1-42) as compared to fAbeta(1-42) suggesting that different forms of Abeta may activate distinct pathways during the progression of AD. Importantly, we demonstrate that Pde4B, a cAMP-specific phosphodiesterase, is upregulated by Abeta and results in an increased production of TNF-alpha. Inhibition of Pde4B reduces by up to 70% the release of TNF-alpha from sAbeta-stimulated microglial cells, implicating cAMP as an important mediator of Abeta-induced microglial activation.

Human thiopurine S-methyltransferase pharmacogenetics: variant allozyme misfolding and aggresome formation.

Thiopurine S-methyltransferase (TPMT) catalyzes the S-methylation of thiopurine drugs. TPMT genetic polymorphisms represent a striking example of the potential clinical value of pharmacogenetics. Subjects homozygous for TPMT*3A, the most common variant allele for low activity, an allele that encodes a protein with two changes in amino acid sequence, are at greatly increased risk for life-threatening toxicity when treated with standard doses of thiopurines. These subjects have virtually undetectable levels of TPMT protein. In this study, we tested the hypothesis that TPMT*3A might result in protein misfolding and aggregation. We observed that TPMT*3A forms aggresomes in cultured cells and that it aggregates in vitro, functional mechanisms not previously described in pharmacogenetics. Furthermore, there was a correlation among TPMT half-life values in rabbit reticulocyte lysate, aggresome formation in COS-1 cells, and protein aggregation in vitro for the three variant allozymes encoded by alleles that include the two TPMT*3A single-nucleotide polymorphisms. These observations were compatible with a common structural explanation for all of these effects, a conclusion supported by size-exclusion chromatography and CD spectroscopy. The results of these experiments provide insight into a unique pharmacogenetic mechanism by which common polymorphisms affect TPMT protein function and, as a result, therapeutic response to thiopurine drugs.

Stereoselective interactions of peptide inhibitors with the beta-amyloid peptide.

Residues 16-20 of the beta-amyloid peptide (A beta) function as a self-recognition element during A beta assembly into fibers. Peptides containing this motif retain the ability to interact with A beta and, in some cases, potently inhibit its assembly. Replacing L- with D-amino acids could stabilize such peptides and permit their evaluation as therapeutic agents for Alzheimer's disease. Here we have assessed the effect that such a chiral reversal has on inhibitory potency. D-enantiomers of five peptides, KLVFFA, KKLVFFA, KFVFFA, KIVFFA, and KVVFFA, were unexpectedly more active as inhibitors in an in vitro fibrillogenesis assay. Circular dichroism showed that D-KLVFFA more effectively prevented A beta adopting the beta-sheet secondary structure correlated with fibrillogenesis. Electron microscopy showed that fiber formation was also more strongly inhibited by D-KLVFFA. Heterochiral inhibition was confirmed using D-A beta, on the principle that enantiomeric proteins exhibit reciprocal chiral biochemical interactions. With D-Abeta, L-KLVFFA was the more potent inhibitor, rather than d-KLVFFA. Most significantly, D-peptides were more potent at reducing the toxicity of both A beta1-40 and A beta 1-42 toward neuronal cells in culture. This unforeseen heterochiral stereoselectivity of A beta for D-peptide inhibitors should be considered during future design of peptide-based inhibitors of A beta neurotoxicity and fibrillogenesis.

Are there naturally occurring pleomorphic bacteria in the blood of healthy humans?

Dark-field microscopy of blood from healthy individuals revealed the existence of pleomorphic microorganisms. These bacteria exhibited limited growth and susceptibility to antibiotics and could be detected by fluorescent in situ hybridization and flow cytometry. They were further characterized by analysis of their 16S rRNA and gyrB genes.

Isolation of Campylobacter fetus subsp. fetus from a patient with cellulitis.

Campylobacter fetus subsp. fetus is a gram-negative, slender, spirally curved bacterial pathogen. It has been isolated from human blood, spinal fluid, and abscesses, but cellulitis associated with bacteremia is rare. We report its isolation from a blood culture of a human patient with cellulitis as well as difficulties encountered in determining the identity of the subspecies of C. fetus.