On a Fatal Case of <Emphasis Type="Italic">Candida krusei</Emphasis> Pleural Empyema in a Pregnant Woman with Spontaneous Esophagus Perforation

Introduction  

Candida empyema thoracis can be a consequence of operation, gastropleural fistula, and esophageal perforation. Case report and review of the literature. A fatal case of a 45-year-old pregnant woman with Candida krusei empyema thoracis secondary to spontaneous esophagus perforation associated with candida esophagitis is reported. The case is contextualized among similar cases found through a PubMed search.     

Hyper response to ovarian stimulation affects the follicular fluid metabolomic profile of women undergoing IVF similarly to polycystic ovary syndrome

Introduction

During in vitro fertilization (IVF), the hyper response to controlled ovarian stimulation (COS) is a common characteristic among patients diagnosed with polycystic ovary syndrome (PCOS), although non-diagnosed patients may also demonstrate this response.

Objectives

In an effort to investigate follicular metabolic characteristics associated with hyper response to COS, the present study analyzed follicular fluid (FF) samples from patients undergoing IVF.

Methods

FF samples were obtained from patients with PCOS and hyper response during IVF (PCOS group, N?=?15), patients without PCOS but with hyper response during IVF (HR group, N?=?44), and normo-responder patients receiving IVF (control group, N?=?22). FF samples underwent Bligh and Dyer extraction, followed by metabolomic analysis by ultra-performance liquid chromatography mass spectrometry, considering two technical replicates. Clinical data was analyzed by ANOVA and chi-square tests. The metabolomic dataset was analyzed by multivariate statistics, and the significance of biomarkers was confirmed by ANOVA.

Results

Clinical data showed differences regarding follicles production, oocyte and embryo quality. From the 15 proposed biomarkers, 14 were of increased abundance in the control group and attributed as fatty acids, diacylglycerol, triacylglycerol, ceramide, ceramide-phosphate, phosphatidylcholine, and sphingomyelin. The PCOS patients showed increased abundance of a metabolite of m/z 144.0023 that was not attributed to a class.

Conclusion

The clinical and metabolic similarities observed in the FF of hyper responders with and without PCOS diagnosis indicate common biomarkers that could assist on the development of accessory tools for assessment of IVF parameters.

     

Culture-Independent Assessment of Rhizobiales-Related Alphaproteobacteria and the Diversity of <Emphasis Type="Italic">Methylobacterium</Emphasis> in the Rhizosphere and Rhizoplane of Transgenic Eucalyptus

The rhizosphere is an ecosystem exploited by a variety of organisms involved in plant health and environmental sustainability. Abiotic factors influence microorganism–plant interactions, but the microbial community is also affected by expression of heterologous genes from host plants. In the present work, we assessed the community shifts of Alphaproteobacteria phylogenetically related to the Rhizobiales order (Rhizobiales-like community) in rhizoplane and rhizosphere soils of wild-type and transgenic eucalyptus. A greenhouse experiment was performed and the bacterial communities associated with two wild-type (WT17 and WT18) and four transgenic (TR-9, TR-15, TR-22, and TR-23) eucalyptus plant lines were evaluated. The culture-independent approach consisted of the quantification, by real-time polymerase chain reaction (PCR), of a targeted subset of Alphaproteobacteria and the assessment of its diversity using PCR–denaturing gradient gel electrophoresis (DGGE) and 16S rRNA gene clone libraries. Real-time quantification revealed a lesser density of the targeted community in TR-9 and TR-15 plants and diversity analysis by principal components analysis, based on PCR–DGGE, revealed differences between bacterial communities, not only between transgenic and nontransgenic plants, but also among wild-type plants. The comparison between clone libraries obtained from the transgenic plant TR-15 and wild-type WT17 revealed distinct bacterial communities associated with these plants. In addition, a culturable approach was used to quantify the Methylobacterium spp. in the samples where the identification of isolates, based on 16S rRNA gene sequences, showed similarities to the species Methylobacterium nodulans, Methylobacterium isbiliense, Methylobacterium variable, Methylobacterium fujisawaense, and Methylobacterium radiotolerans. Colonies classified into this genus were not isolated from the rhizosphere but brought in culture from rhizoplane samples, except for one line of the transgenic plants (TR-15). In general, the data suggested that, in most cases, shifts in bacterial communities due to cultivation of transgenic plants are similar to those observed when different wild-type cultivars are compared, although shifts directly correlated to transgenic plant cultivation may be found.     

Expression of Xylella fastidiosa Fimbrial and Afimbrial Proteins during Biofilm Formation

Complete sequencing of the Xylella fastidiosa genome revealed characteristics that have not been described previously for a phytopathogen. One characteristic of this genome was the abundance of genes encoding proteins with adhesion functions related to biofilm formation, an essential step for colonization of a plant host or an insect vector. We examined four of the proteins belonging to this class encoded by genes in the genome of X. fastidiosa: the PilA2 and PilC fimbrial proteins, which are components of the type IV pili, and XadA1 and XadA2, which are afimbrial adhesins. Polyclonal antibodies were raised against these four proteins, and their behavior during biofilm development was assessed by Western blotting and immunofluorescence assays. In addition, immunogold electron microscopy was used to detect these proteins in bacteria present in xylem vessels of three different hosts (citrus, periwinkle, and hibiscus). We verified that these proteins are present in X. fastidiosa biofilms but have differential regulation since the amounts varied temporally during biofilm formation, as well as spatially within the biofilms. The proteins were also detected in bacteria colonizing the xylem vessels of infected plants.Aggregative growth is a common feature in the microbial world, and its discovery radically changed our concept of microbial growth dynamics. A cellular aggregate adhering to a surface is known as a biofilm. It has important characteristics, such as greater resistance to antimicrobial compounds (34, 54), increased capacity of the cells to take up nutrients from the environment (59), and higher detoxification efficiency resulting from an increase in expression of genes encoding efflux pumps (43). These characteristics give the biofilm cells a great adaptive advantage.Biofilm growth also confers advantages to plant pathogens by promoting virulence and protection against plant defense responses. Bacteria can colonize different niches in the plant, from aerial surfaces to roots and the vascular system, and biofilm formation can play a role at all of these sites of colonization. In the vessels, biofilms are very important since the cells need to survive in a competitive habitat where plant defense compounds are produced in response to infection (7).Biofilm development is divided into at least the following five phases: (i) reversible attachment, (ii) irreversible attachment, (iii) beginning of maturation, (iv) mature biofilm, and (v) dispersion (13, 50). In Xylella fastidiosa strain 9a5c, the maturation phase occurs between days 15 and 20 in vitro, while dispersion occurs between days 25 and 30, as observed by our analysis of biofilm formation using different methods, including scanning electron microscopy and quantification of exopolysaccharides, biomass, and the total protein (unpublished data). The establishment and development of biofilms of plant-colonizing bacteria share several features with the establishment and development of biofilms of human bacterial pathogens, such as regulation by quorum sensing, nutrient starvation regulation, and phase variation. Motility is also an important factor not only for the initiation and development of the biofilm but also for dispersion (50). Attachment is mediated by surface-associated structures, which include both polysaccharides and proteins classified as fimbrial and afimbrial adhesins, depending on the structure to which they contribute. Fimbrial adhesins form filamentous structures, while afimbrial adhesins produce projections on the outer membrane (23).X. fastidiosa, a Gram-negative phytopathogen that grows as a biofilms in both plant xylem vessels and the cybarium of insect vectors, is a major threat to plant production around the world. In Brazil, it has a major economic impact on citriculture since it causes citrus variegated chlorosis disease (CVC) (35, 39, 42). The biofilm formed by X. fastidiosa blocks the xylem vessels of susceptible citrus plants, impairing water flow. This blockage leads to a drastic reduction in fruit size (32) and, consequently, severe economic losses resulting from reduced plant productivity (4).Due to the economic damage caused by CVC, there has been a major effort to generate more information about its biology. This led to sequencing of the genome of the pathogen. The X. fastidiosa genome harbors a wide variety of genes encoding adhesins (53). Bacterial cell surface adhesins are important in the initial phases of adherence to surfaces, as well as in bacterium-bacterium interactions and microcolony development (15). Insight into X. fastidiosa has also come from genome analysis of a strain of X. fastidiosa which causes Pierce''s disease of grape (58). Studies of this strain showed that the cellular aggregation process involves type I and type IV fimbrial adhesins. The two types of fimbriae present different adhesion forces that help bacteria adhere to a substrate (10, 30). Adhesion proteins have also been demonstrated to mediate adherence to carbohydrates of leafhopper foregut surfaces (27). In addition, both fimbrial and afimbrial adhesins are important for plant pathogenicity (38, 41). However, the expression of these proteins during X. fastidiosa biofilm formation either in vitro or in planta is still poorly understood. For X. fastidiosa strains causing CVC, nothing is known about the role of these proteins in pathogenicity or biofilm formation, although some adhesion-encoding genes, such as pilA2, pilC, xadA1, and xadA2, were found to be upregulated either in virulent strains of X. fastidiosa or during biofilm formation (12, 14). These results suggest that the biofilm mode of growth is important for successful colonization of the citrus host by X. fastidiosa strains that cause CVC. In this work we focused on the temporal expression of the PilA2 and PilC fimbrial proteins and XadA1 and XadA2, which are afimbrial adhesins, during in vitro development of X. fastidiosa CVC biofilms. We demonstrated that the temporal and spatial patterns of expression of the fimbrial and afimbrial adhesins are very different during biofilm development in vitro. Moreover, we also verified that these adhesins are present in X. fastidiosa cells in symptomatic plants of three different hosts (citrus, periwinkle, and hibiscus).     

Bacterial community in the rhizosphere and rhizoplane of wild type and transgenic eucalyptus

The rhizosphere is a niche exploited by a wide variety of bacteria. The expression of heterologous genes by plants might become a factor affecting the structure of bacterial communities in the rhizosphere. In a greenhouse experiment, the bacterial community associated to transgenic eucalyptus, carrying the Lhcb1-2 genes from pea (responsible for a higher photosynthetic capacity), was evaluated. The culturable bacterial community associated to transgenic and wild type plants were not different in density, and the Amplified Ribosomal DNA Restriction Analysis (ARDRA) typing of 124 strains revealed dominant ribotypes representing the bacterial orders Burkholderiales, Rhizobiales, and Actinomycetales, the families Xanthomonadaceae, and Bacillaceae, and the genus Mycobacterium. Principal Component Analysis based on the fingerprints obtained by culture-independent Denaturing Gradient Gel Electrophoresis analysis revealed that Alphaproteobacteria, Betaproteobacteria and Actinobacteria communities responded differently to plant genotypes. Similar effects for the cultivation of transgenic eucalyptus to those observed when two genotype-distinct wild type plants are compared.     

The pentose phosphate pathway constitutes a major metabolic hub in pathogenic Francisella

Metabolic pathways are now considered as intrinsic virulence attributes of pathogenic bacteria and thus represent potential targets for antibacterial strategies. Here we focused on the role of the pentose phosphate pathway (PPP) and its connections with other metabolic pathways in the pathophysiology of Francisella novicida. The involvement of the PPP in the intracellular life cycle of Francisella was first demonstrated by studying PPP inactivating mutants. Indeed, we observed that inactivation of the tktA, rpiA or rpe genes severely impaired intramacrophage multiplication during the first 24 hours. However, time-lapse video microscopy demonstrated that rpiA and rpe mutants were able to resume late intracellular multiplication. To better understand the links between PPP and other metabolic networks in the bacterium, we also performed an extensive proteo-metabolomic analysis of these mutants. We show that the PPP constitutes a major bacterial metabolic hub with multiple connections to glycolysis, the tricarboxylic acid cycle and other pathways, such as fatty acid degradation and sulfur metabolism. Altogether our study highlights how PPP plays a key role in the pathogenesis and growth of Francisella in its intracellular niche.     

Larvae of the invasive species <Emphasis Type="Italic">Limnoperna fortunei</Emphasis> (Bivalvia) in the diet of fish larvae in the Paraná River,Argentina

To assess the trophic impact of the planktonic larvae of the invasive Asian bivalve Limnoperna fortunei, introduced in South America around 1990, we investigated the gut contents of fish larvae collected at monthly intervals between October 2000 and March 2001 at three locations along a 600 km stretch of the Paraná River, and during November 2004 in two areas of marginal lagoons connected to the river near the city of San Nicolás, Argentina. Zooplankton was also collected in the lagoons in 2004. In total, 11,956 fish larvae were retrieved, of which 1,511 were used for detailed analyses. Of the 15 fish taxa collected, 11 had veligers of L. fortunei in their gut. Fish larvae with empty guts represented 60% (San Nicolás) to 72% (Paraná River) of the total number of fish. Proportions of feeding fish larvae with L. fortunei veligers in their guts varied between 20% (San Nicolás) and 56% (Paraná River); in 15% of the guts analyzed, L. fortunei was the only food item recorded. For those specimens that had consumed L. fortunei larvae and any other food item, L. fortunei was the most important item in 55% (Paraná River) to 71% (San Nicolás) of the animals in terms of biomass. No major temporal or spatial changes in the diet were observed along the Paraná River, but the relative biomass contribution of L. fortunei larvae differed strongly in fishes of different developmental stage. In protolarvae and mesolarvae, veligers accounted for 30–35% of the gut contents. In metalarvae, veligers accounted for only 3%, indicating enhanced food supply for the earliest fish life stages. Comparison of the relative proportions of the three main zooplankton types (L. fortunei veligers, cladocerans, and copepods) in the water and in larval fish guts indicates that L. fortunei is always selected positively over the other two prey types. While our results strongly suggest that the expansion of L. fortunei results in an enhanced food supply for local fish populations, they do not necessarily imply that the overall effect on the ecosystem in general, and on the fish fauna in particular is beneficial.     

Proline Exogenously Supplied or Endogenously Overproduced Induces Different Nutritional,Metabolic, and Antioxidative Responses in Transgenic Tobacco Exposed to Cadmium

Journal of Plant Growth Regulation - Proline plays adaptive roles in plant tolerance to cadmium (Cd)-induced stress, but many gaps remain to be elucidated as the responses triggered by exogenously...     

Automated Detection of Soma Location and Morphology in Neuronal Network Cultures

Automated identification of the primary components of a neuron and extraction of its sub-cellular features are essential steps in many quantitative studies of neuronal networks. The focus of this paper is the development of an algorithm for the automated detection of the location and morphology of somas in confocal images of neuronal network cultures. This problem is motivated by applications in high-content screenings (HCS), where the extraction of multiple morphological features of neurons on large data sets is required. Existing algorithms are not very efficient when applied to the analysis of confocal image stacks of neuronal cultures. In addition to the usual difficulties associated with the processing of fluorescent images, these types of stacks contain a small number of images so that only a small number of pixels are available along the z-direction and it is challenging to apply conventional 3D filters. The algorithm we present in this paper applies a number of innovative ideas from the theory of directional multiscale representations and involves the following steps: (i) image segmentation based on support vector machines with specially designed multiscale filters; (ii) soma extraction and separation of contiguous somas, using a combination of level set method and directional multiscale filters. We also present an approach to extract the soma’s surface morphology using the 3D shearlet transform. Extensive numerical experiments show that our algorithms are computationally efficient and highly accurate in segmenting the somas and separating contiguous ones. The algorithms presented in this paper will facilitate the development of a high-throughput quantitative platform for the study of neuronal networks for HCS applications.