Comparative Genomics of Gardnerella vaginalis Strains Reveals Substantial Differences in Metabolic and Virulence Potential

Background

Gardnerella vaginalis is described as a common vaginal bacterial species whose presence correlates strongly with bacterial vaginosis (BV). Here we report the genome sequencing and comparative analyses of three strains of G. vaginalis. Strains 317 (ATCC 14019) and 594 (ATCC 14018) were isolated from the vaginal tracts of women with symptomatic BV, while Strain 409-05 was isolated from a healthy, asymptomatic individual with a Nugent score of 9.

Principal Findings

Substantial genomic rearrangement and heterogeneity were observed that appeared to have resulted from both mobile elements and substantial lateral gene transfer. These genomic differences translated to differences in metabolic potential. All strains are equipped with significant virulence potential, including genes encoding the previously described vaginolysin, pili for cytoadhesion, EPS biosynthetic genes for biofilm formation, and antimicrobial resistance systems, We also observed systems promoting multi-drug and lantibiotic extrusion. All G. vaginalis strains possess a large number of genes that may enhance their ability to compete with and exclude other vaginal colonists. These include up to six toxin-antitoxin systems and up to nine additional antitoxins lacking cognate toxins, several of which are clustered within each genome. All strains encode bacteriocidal toxins, including two lysozyme-like toxins produced uniquely by strain 409-05. Interestingly, the BV isolates encode numerous proteins not found in strain 409-05 that likely increase their pathogenic potential. These include enzymes enabling mucin degradation, a trait previously described to strongly correlate with BV, although commonly attributed to non-G. vaginalis species.

Conclusions

Collectively, our results indicate that all three strains are able to thrive in vaginal environments, and therein the BV isolates are capable of occupying a niche that is unique from 409-05. Each strain has significant virulence potential, although genomic and metabolic differences, such as the ability to degrade mucin, indicate that the detection of G. vaginalis in the vaginal tract provides only partial information on the physiological potential of the organism.     

Circulating Progenitor Cells and Vascular Dysfunction in Chronic Obstructive Pulmonary Disease

Background

In chronic obstructive pulmonary disease (COPD), decreased progenitor cells and impairment of systemic vascular function have been suggested to confer higher cardiovascular risk. The origin of these changes and their relationship with alterations in the pulmonary circulation are unknown.

Objectives

To investigate whether changes in the number of circulating hematopoietic progenitor cells are associated with pulmonary hypertension or changes in endothelial function.

Methods

62 COPD patients and 35 controls (18 non-smokers and 17 smokers) without cardiovascular risk factors other than cigarette smoking were studied. The number of circulating progenitors was measured as CD45⁺CD34⁺CD133⁺ labeled cells by flow cytometry. Endothelial function was assessed by flow-mediated dilation. Markers of inflammation and angiogenesis were also measured in all subjects.

Results

Compared with controls, the number of circulating progenitor cells was reduced in COPD patients. Progenitor cells did not differ between control smokers and non-smokers. COPD patients with pulmonary hypertension showed greater number of progenitor cells than those without pulmonary hypertension. Systemic endothelial function was worse in both control smokers and COPD patients. Interleukin-6, fibrinogen, high sensitivity C-reactive protein, vascular endothelial growth factor and tumor necrosis factor were increased in COPD. In COPD patients, the number of circulating progenitor cells was inversely related to the flow-mediated dilation of systemic arteries.

Conclusions

Pulmonary and systemic vascular impairment in COPD is associated with cigarette smoking but not with the reduced number of circulating hematopoietic progenitors. The latter appears to be a consequence of the disease itself not related to smoking habit.     

Ca(2+) shuttling in vesicles during tip growth in Neurospora crassa

Tip-growing organisms maintain an apparently essential tip-high gradient of cytoplasmic Ca(2+). In the oomycete Saprolegnia ferax, in pollen tubes and root hairs, the gradient is produced by a tip-localized Ca(2+) influx from the external medium. Such a gradient is normally dispensable for Neurospora crassa hyphae, which may maintain their Ca(2+) gradient by some form of internal recycling. We localized Ca(2+) in N. crassa hyphae at the ultrastructural level using two techniques (a) electron spectroscopic imaging of freeze-dried hyphae and (b) pyroantimoniate precipitation. The results of both methods support the presence of Ca(2+) in the wall vesicles and Golgi body equivalents, providing a plausible mechanism for the generation and maintenance of the gradient by Ca(2+) shuttling in vesicles to the apex, without exogenous Ca(2+) influx. Ca(2+) sequestration into the vesicles seems to be dependent on Ca(2+)-ATPases since cyclopiazonic acid, a specific inhibitor of Ca(2+) pumps, eliminated all Ca(2+) deposits from the vesicles of N. crassa.     

Sequence distribution and intercooperativity detection for two ligands simultaneously binding to DNA

A method for detecting and quantifying the cooperativity in the simultaneous binding of two ligands, A and B, to DNA (intercooperativity; omega(AB)) is proposed. This involves the determination of an apparent affinity constant K(app) for one of the ligands (A) in the limit of its null saturation (nu(Alpha) --> 0), in the presence of the second one (B). A definition for this constant is given and an expression is derived corresponding to a simple model of competitive binding to an unbranched three-state homogeneous polar lattice with nearest-neighbor interactions (Markov chain). The ratio between the apparent and intrinsic affinity constants of one ligand in the maximum saturation limit of the other one becomes omega(2)(AB), and thus can be graphically obtained from K(app)(A) vs nu(B) plots. All the frequencies that define the sequence distribution of ligands can be easily calculated by introducing some generalized statistical weights for the free lattice monomer in a standard sequence generating function procedure. A model of fluorescence quenching emission is obtained from such frequencies under the hypothesis of a short-range electron transfer mechanism of the deactivation; it confirms, as suggested by the binding model, an outstanding influence of the intercooperativity on the distribution.     

Analysis of three separate probes suggests the absence of endocytosis in Neurospora crassa hyphae

Reports of the existence of endocytosis in filamentous fungi have been conflicting and inconclusive. For this reason, we have tested three independent markers in Neurospora crassa: the electron opaque marker lanthanum (La) and the fluorescent probes Lucifer yellow (LY) and FM4-64. Both La and LY were endocytosed by Saccharomyces cerevisiae cells, which were used as positive controls for endocytosis, but the probes did not accumulate in N. crassa hyphae. Only FM4-64 became internalized into N. crassa hyphae, but it induced abnormal changes in membrane systems and its internalization could be explained by mechanisms other than endocytosis. Together, our results suggest that endocytosis does not occur in N. crassa hyphae and question whether the styryl dyes do in fact reliably report normal endocytosis in filamentous fungi.     

Calcium-supplemented University of Wisconsin solution in long-term myocardial preservation

The purpose of this study was to assess the effects of the addition of calcium to University of Wisconsin solution in long-term myocardial perfusion. In a heterotopic heart transplantation model, performed in pigs, the donor heart was preserved for 24 hours by means of continuous perfusion in this solution, without (24hUW group) or with calcium, 2.4 mmol/L (24hUW+Ca). During this period, the oxygenation and pH of the solution were measured, as were the calcium and lactate concentrations and enzyme release. After two hours of reperfusion, samples were collected from both ventricles for the morphological study. In the control group, there were no signs that reperfusion had triggered the calcium paradox. The addition of this cation to the preservation solution improved the intercellular junction integrity but, at the same time, favored intracellular calcium overload. This is manifested by increased enzyme release during preservation (LDH: 242+/-95 vs 140+/-25; CK: 668+/-371 vs 299+/-83 (U/L). p<0.01 in both cases) and signs of ventricular contracture: hardness and stiffness were significantly more prominent than in the group without calcium supplementation. Moreover, in comparison with the control group, the structural morphology of 24hUW+Ca is characterized by the more prominent and extensive presence of contraction bands and disorganized actin structure. Thus, under the experimental conditions employed in this study, we consider the addition of calcium to Wisconsin solution to be unadvisable.     

Enhancement of aryl hydrocarbon hydroxylase activity in cultured rodent cells by X-irradiation.

X-irradiation (500 rads) was found to enhance the aryl hydrocarbon hydroxylase (AHH) activity of three cell lines. Radiation followed by induction with benz (a) anthracene (5–15 μg/ml) produced a synergistic effect on AHH. These effects were highly significant and were observed most dramatically with a hamster tumor cell line, A(T_l)Cl-3,a nd to a lesser extent in secondary hamsters embryo cells and mouse $\text{C}\text{3}\text{H}\text{/10}\text{T}\text{1}\text{2}$ CL8 cells.     

Draft genome sequence of Bacteroides vulgatus PC510, a strain isolated from human feces

Although Bacteroides vulgatus is one of the most prevalent microorganisms in the human gastrointestinal tract, little is known about the genetic potential of this species. Here, we describe the annotated draft genome sequence of B. vulgatus PC510 isolated from human feces.     

Gut microbiome composition and metabolomic profiles of wild western lowland gorillas (Gorilla gorilla gorilla) reflect host ecology

The metabolic activities of gut microbes significantly influence host physiology; thus, characterizing the forces that modulate this micro‐ecosystem is key to understanding mammalian biology and fitness. To investigate the gut microbiome of wild primates and determine how these microbial communities respond to the host's external environment, we characterized faecal bacterial communities and, for the first time, gut metabolomes of four wild lowland gorilla groups in the Dzanga‐Sangha Protected Areas, Central African Republic. Results show that geographical range may be an important modulator of the gut microbiomes and metabolomes of these gorilla groups. Distinctions seemed to relate to feeding behaviour, implying energy harvest through increased fruit consumption or fermentation of highly fibrous foods. These observations were supported by differential abundance of metabolites and bacterial taxa associated with the metabolism of cellulose, phenolics, organic acids, simple sugars, lipids and sterols between gorillas occupying different geographical ranges. Additionally, the gut microbiomes of a gorilla group under increased anthropogenic pressure could always be distinguished from that of all other groups. By characterizing the interplay between environment, behaviour, diet and symbiotic gut microbes, we present an alternative perspective on primate ecology and on the forces that shape the gut microbiomes of wild primates from an evolutionary context.