Probiotic administration attenuates myocardial hypertrophy and heart failure following myocardial infarction in the rat

Background—Probiotics are extensively used to promote gastrointestinal health and emerging evidence suggests that their beneficial properties can extend beyond the local environment of the gut. Here, we determined whether oral probiotic administration can alter the progression of post-infarction heart failure. Methods and Results—Rats were subjected to six weeks of sustained coronary artery occlusion and administered the probiotic Lactobacillus rhamnosus GR-1 or placebo in the drinking water ad libitum. Culture and 16s rRNA sequencing showed no evidence of GR-1 colonization or a significant shift in the composition of the cecal microbiome. However, animals administered GR-1 exhibited a significant attenuation of left ventricular hypertrophy based on tissue weight assessment as well as gene expression of atrial natriuretic peptide. Moreover, these animals demonstrated improved hemodynamic parameters reflecting both improved systolic and diastolic left ventricular function. Serial echocardiography revealed significantly improved left ventricular parameters throughout the six week follow-up period including a marked preservation of left ventricular ejection fraction as well as fractional shortening. Beneficial effects of GR-1 were still evident in those animals in which GR-1 was withdrawn at four weeks suggesting persistence of the GR-1 effects following cessation of therapy. Investigation of mechanisms showed a significant increase in the leptin to adiponectin plasma concentration ratio in rats subjected to coronary ligation which was abrogated by GR-1. Metabonomic analysis showed differences between sham control and coronary artery ligated hearts particularly with respect to preservation of myocardial taurine levels. Conclusions—The study suggests that probiotics offer promise as a potential therapy for the attenuation of heart failure.

Metaproteomics analysis reveals the adaptation process for the chicken gut microbiota

The animal gastrointestinal tract houses a large microbial community, the gut microbiota, that confers many benefits to its host, such as protection from pathogens and provision of essential metabolites. Metagenomic approaches have defined the chicken fecal microbiota in other studies, but here, we wished to assess the correlation between the metagenome and the bacterial proteome in order to better understand the healthy chicken gut microbiota. Here, we performed high-throughput sequencing of 16S rRNA gene amplicons and metaproteomics analysis of fecal samples to determine microbial gut composition and protein expression. 16 rRNA gene sequencing analysis identified Clostridiales, Bacteroidaceae, and Lactobacillaceae species as the most abundant species in the gut. For metaproteomics analysis, peptides were generated by using the Fasp method and subsequently fractionated by strong anion exchanges. Metaproteomics analysis identified 3,673 proteins. Among the most frequently identified proteins, 380 proteins belonged to Lactobacillus spp., 155 belonged to Clostridium spp., and 66 belonged to Streptococcus spp. The most frequently identified proteins were heat shock chaperones, including 349 GroEL proteins, from many bacterial species, whereas the most abundant enzymes were pyruvate kinases, as judged by the number of peptides identified per protein (spectral counting). Gene ontology and KEGG pathway analyses revealed the functions and locations of the identified proteins. The findings of both metaproteomics and 16S rRNA sequencing analyses are discussed.

Analysis of the bacterial communities present in lungs of patients with cystic fibrosis from American and British centers

The aim of this study was to determine whether geographical differences impact the composition of bacterial communities present in the airways of cystic fibrosis (CF) patients attending CF centers in the United States or United Kingdom. Thirty-eight patients were matched on the basis of clinical parameters into 19 pairs comprised of one U.S. and one United Kingdom patient. Analysis was performed to determine what, if any, bacterial correlates could be identified. Two culture-independent strategies were used: terminal restriction fragment length polymorphism (T-RFLP) profiling and 16S rRNA clone sequencing. Overall, 73 different terminal restriction fragment lengths were detected, ranging from 2 to 10 for U.S. and 2 to 15 for United Kingdom patients. The statistical analysis of T-RFLP data indicated that patient pairing was successful and revealed substantial transatlantic similarities in the bacterial communities. A small number of bands was present in the vast majority of patients in both locations, indicating that these are species common to the CF lung. Clone sequence analysis also revealed that a number of species not traditionally associated with the CF lung were present in both sample groups. The species number per sample was similar, but differences in species presence were observed between sample groups. Cluster analysis revealed geographical differences in bacterial presence and relative species abundance. Overall, the U.S. samples showed tighter clustering with each other compared to that of United Kingdom samples, which may reflect the lower diversity detected in the U.S. sample group. The impact of cross-infection and biogeography is considered, and the implications for treating CF lung infections also are discussed.

Dietary iron depletion at weaning imprints low microbiome diversity and this is not recovered with oral nano Fe(III)

Alterations in the gut microbiota have been recently linked to oral iron. We conducted two feeding studies including an initial diet-induced iron-depletion period followed by supplementation with nanoparticulate tartrate-modified ferrihydrite (Nano Fe(III): considered bioavailable to host but not bacteria) or soluble ferrous sulfate (FeSO4: considered bioavailable to both host and bacteria). We applied denaturing gradient gel electrophoresis and fluorescence in situ hybridization for study-1 and 454-pyrosequencing of fecal 16S rRNA in study-2. In study-1, the within-community microbial diversity increased with FeSO4 (P = 0.0009) but not with Nano Fe(III) supplementation. This was confirmed in study-2, where we also showed that iron depletion at weaning imprinted significantly lower within- and between-community microbial diversity compared to mice weaned onto the iron-sufficient reference diet (P < 0.0001). Subsequent supplementation with FeSO4 partially restored the within-community diversity (P = 0.006 in relation to the continuously iron-depleted group) but not the between-community diversity, whereas Nano Fe(III) had no effect. We conclude that (1) dietary iron depletion at weaning imprints low diversity in the microbiota that is not, subsequently, easily recovered; (2) in the absence of gastrointestinal disease iron supplementation does not negatively impact the microbiota; and (3) Nano Fe(III) is less available to the gut microbiota.

Dynamics and diversity of the 'Atopobium cluster' in the human faecal microbiota, and phenotypic characterization of 'Atopobium cluster' isolates

This study monitored the dynamics and diversity of the human faecal 'Atopobium cluster' over a 3-month period using a polyphasic approach. Fresh faecal samples were collected fortnightly from 13 healthy donors (6 males and 7 females) aged between 26 and 61 years. Fluorescence in situ hybridization was used to enumerate total (EUB338mix) and 'Atopobium cluster' (ATO291) bacteria, with counts ranging between 1.12 × 1011 and 9.95 × 1011, and 1.03 × 109 and 1.16 × 1011 cells (g dry weight faeces)-1, respectively. The 'Atopobium cluster' population represented 0.2-22 % of the total bacteria, with proportions donor-dependent. Denaturing gradient gel electrophoresis (DGGE) using 'Atopobium cluster'-specific primers demonstrated faecal populations of these bacteria were relatively stable, with bands identified as Collinsella aerofaciens, Collinsella intestinalis/Collinsella stercoris, Collinsella tanakaei, Coriobacteriaceae sp. PEAV3-3, Eggerthella lenta, Gordonibacter pamelaeae, Olsenella profusa, Olsenella uli and Paraeggerthella hongkongensis in the DGGE profiles of individuals. Colony PCR was used to identify 'Atopobium cluster' bacteria isolated from faeces (n = 224 isolates). 16S rRNA gene sequence analysis of isolates demonstrated Collinsella aerofaciens represented the predominant (88 % of isolates) member of the 'Atopobium cluster' found in human faeces, being found in nine individuals. Eggerthella lenta was identified in three individuals (3.6 % of isolates). Isolates of Collinsella tanakaei, an 'Enorma' sp. and representatives of novel species belonging to the 'Atopobium cluster' were also identified in the study. Phenotypic characterization of the isolates demonstrated their highly saccharolytic nature and heterogeneous phenotypic profiles, and 97 % of the isolates displayed lipase activity.

Effects of orange juice formulation on prebiotic functionality using an in vitro colonic model sytem

A three-stage continuous fermentative colonic model system was used to monitor in vitro the effect of different orange juice formulations on prebiotic activity. Three different juices with and without Bimuno, a GOS mixture containing galactooligosaccharides (B-GOS) were assessed in terms of their ability to induce a bifidogenic microbiota. The recipe development was based on incorporating 2.75g B-GOS into a 250 ml serving of juice (65°Brix of concentrate juice). Alongside the production of B-GOS juice, a control juice - orange juice without any additional Bimuno and a positive control juice, containing all the components of Bimuno (glucose, galactose and lactose) in the same relative proportions with the exception of B-GOS were developed. Ion Exchange Chromotography analysis was used to test the maintenance of bimuno components after the production process. Data showed that sterilisation had no significant effect on concentration of B-GOS and simple sugars. The three juice formulations were digested under conditions resembling the gastric and small intestinal environments. Main bacterial groups of the faecal microbiota were evaluated throughout the colonic model study using 16S rRNA-based fluorescence in situ hybridization (FISH). Potential effects of supplementation of the juices on microbial metabolism were studied measuring short chain fatty acids (SCFAs) using gas chromatography. Furthermore, B-GOS juices showed positive modulations of the microbiota composition and metabolic activity. In particular, numbers of faecal bifidobacteria and lactobacilli were significantly higher when B-GOS juice was fermented compared to controls. Furthermore, fermentation of B-GOS juice resulted in an increase in Roseburia subcluster and concomitantly increased butyrate production, which is of potential benefit to the host. In conclusion, this study has shown B-GOS within orange juice can have a beneficial effect on the fecal microbiota.

The impact of date palm fruits and their component polyphenols, on gut microbial ecology, bacterial metabolites and colon cancer cell proliferation

The fruit of the date palm (Phoenix dactylifera L.) is a rich source of dietary fibre and polyphenols. We have investigated gut bacterial changes induced by the whole date fruit extract (digested date extract; DDE) and its polyphenol-rich extract (date polyphenol extract; DPE) using faecal, pH-controlled, mixed batch cultures mimicking the distal part of the human large intestine, and utilising an array of microbial group-specific 16S rRNA oligonucleotide probes. Fluorescence microscopic enumeration indicated that there was a significant increase in the growth of bifidobacteria in response to both treatments, whilst whole dates also increased bacteroides at 24 h and the total bacterial counts at later fermentation time points when compared with DPE alone. Bacterial metabolism of whole date fruit led to the production of SCFA, with acetate significantly increasing following bacterial incubation with DDE. In addition, the production of flavonoid aglycones (myricetin, luteolin, quercetin and apigenin) and the anthocyanidin petunidin in less than 1 h was also observed. Lastly, the potential of DDE, DPE and metabolites to inhibit Caco-2 cell growth was investigated, indicating that both were capable of potentially acting as antiproliferative agents in vitro, following a 48 h exposure. This potential to inhibit growth was reduced following fermentation. Together these data suggest that consumption of date fruits may enhance colon health by increasing beneficial bacterial growth and inhibiting the proliferation of colon cancer cells. This is an early suggestion that date intake by humans may aid in the maintenance of bowel health and even the reduction of colorectal cancer development.

Characterization of para-Nitrophenol-degrading bacterial communities in river water by using functional markers and stable isotope probing

Microbial degradation is a major determinant of the fate of pollutants in the environment. para-Nitrophenol (PNP) is an EPA listed priority pollutant with a wide environmental distribution, but little is known about the microorganisms that degrade it in the environment. We studied the diversity of active PNP-degrading bacterial populations in river water using a novel functional marker approach coupled with [13C6]PNP stable isotope probing (SIP). Culturing together with culture-independent terminal restriction fragment length polymorphism analysis of 16S rRNA gene amplicons identified Pseudomonas syringae to be the major driver of PNP degradation in river water microcosms. This was confirmed by SIP-pyrosequencing of amplified 16S rRNA. Similarly, functional gene analysis showed that degradation followed the Gram-negative bacterial pathway and involved pnpA from Pseudomonas spp. However, analysis of maleylacetate reductase (encoded by mar), an enzyme common to late stages of both Gram-negative and Gram-positive bacterial PNP degradation pathways, identified a diverse assemblage of bacteria associated with PNP degradation, suggesting that mar has limited use as a specific marker of PNP biodegradation. Both the pnpA and mar genes were detected in a PNP-degrading isolate, P. syringae AKHD2, which was isolated from river water. Our results suggest that PNP-degrading cultures of Pseudomonas spp. are representative of environmental PNP-degrading populations.

Studies on Anopheles (Kerteszia) homunculus Komp (Diptera: Culicidae)

The present findings suggest that Anopheles (Kerteszia) homunculus may comprise more than one species. The rDNA ITS2 sequence data corroborate the presence of An. homunculus l.s. in Mata Atlantica, southern Brazil, and suggest that specimens from Trinidad may belong to an unnamed morphologically similar species. There is a need for additional studies to establish the geographical distribution of An. homunculus l.s. in continental South America and in Trinidad, especially in southern Mata Atlantica, Brazil.

Myxobolus cordeiroi n. sp., a parasite of Zungaro jahu (Siluriformes: Pimelodiade) from Brazilian Pantanal: Morphology, phylogeny and histopathology

This work is part of an ongoing investigation into the characteristics of Myxozoan parasites of freshwater fish in Brazil and was carried out using morphology, histopathology and molecular analysis. A new Myxosporea species (Myxobolus cordeiroi) is described infecting the jau catfish (Zungaro jahu). Fifty jau specimens were examined and 78% exhibited plasmodia of the parasite. The plasmodia were white and round, measuring 0.3-2.0 mm in diameter and the development occurred in the gill arch, skin, serosa of the body cavity, urinary bladder and eye. The spores had an oval body and the spore wall was smooth. Partial sequencing of the 18S rDNA gene resulted in a total of 505 bp and the alignment of the sequences obtained from samples in different organs revealed 100% identity. In the phylogenetic analysis, the Myxobolus species clustered into two clades-one primarily parasites of freshwater fish and the other primarily parasites of marine fish. M. cordeiroi n. sp. was clustered in a basal position in the freshwater fish species clade. The histological analysis revealed the parasite in the connective tissue of the different infected sites, thereby exhibiting affinity to this tissue. The plasmodium was surrounded by an outer collagen capsule of fibers with distinct orientation from the adjacent connective tissue and an inner layer composed of delicate collagen fibrils-more precisely reticular fibers. The development of the parasite in the cornea and urinary bladder caused considerable stretching of the epithelium. (C) 2009 Elsevier B.V. All rights reserved.

Host-parasite-environment relationship, morphology and molecular analyses of Henneguya eirasi n. sp parasite of two wild Pseudoplatystoma spp. in Pantanal Wetland, Brazil

A new myxosporean species, Henneguya eirasi n. sp., is described parasitizing the gill filaments of Pseudoplatystoma corruscans and Pseudoplatystoma fasciatum (Siluriformes: Pimelodidae) caught in the Patanal Wetland of the state of Mato Grosso, Brazil. The parasite formed white, elongated plasmodia measuring up to 3 mm. Mature spores were ellipsoidal in the frontal view, measuring 37.1 +/- 1.8 mu m in total length, 12.9 +/- 0.8 mu m in body length, 3.4 +/- 0.3 mu m in width, 3.1 +/- 0.1 mu m in thickness and 24.6 +/- 2.2 mu m in the caudal process. Polar capsules were elongated and equal in size, measuring 5.4 +/- 0.5 mu m in length and 0.7 +/- 0.1 mu m in width. Polar filaments had 12-13 coils. Histopathological analysis revealed that the parasite developed in the sub-epithelial connective tissue of the gill filaments and the plasmodia were surrounded by a capsule of host connective tissue. The plasmodia caused slight compression of the adjacent tissues, but no inflammatory response was observed in the infection site. Ultrastructure analysis revealed a single plasmodial wall connected to the ectoplasmic zone through numerous pinocytotic canals. The plasmodial wall exhibited numerous projections and slightly electron-dense material was found in the ectoplasm next to the plasmodial wall, forming a line just below the wall. Partial sequencing of the 18S rDNA gene of H. eirasi n. sp. obtained from P. fasciatum resulted in a total of 1066 bp and this sequence did not match any of the Myxozoa available in the GenBank. Phylogenetic analysis revealed the Henneguya species clustering into clades following the order and family of the host fishes. H. eirasi n. sp. clustered alone in one clade, which was the basal unit for the clade composed of Henneguya species parasites of siluriform ictalurids. The prevalence of the parasite was 17.1% in both fish species examined. Parasite prevalence was not influenced by season, host sex or host size. (C) 2011 Elsevier B.V. All rights reserved.

PHYLOGENETIC AFFINITIES OF "CHANTRANSIA" STAGES IN MEMBERS OF THE BATRACHOSPERMALES AND THOREALES (RHODOPHYTA)

This study evaluated the phylogenetic relationship among samples of ""Chantransia"" stage of the Batrachospermales and Thoreales from several regions of the world based on sequences of two genes-the plastid-encoded RUBISCO LSU gene (rbcL) and the nuclear SSU ribosomal DNA gene (SSU rDNA). All sequences of ""Chantransia macrospora"" were shown to belong to Batrachospermum macrosporum based on both molecular markers, confirming evidence from previous studies. In contrast, nine species are now associated with ""Chantransia pygmaea,"" including seven species of the Batrachospermales and two of the Thoreales. Therefore, the presence of ""C. macrospora"" in a stream can be considered reliable evidence that it belongs to B. macrosporum, whereas the occurrence of ""C. pygmaea"" does not allow the recognition of any particular species, since it is associated with at least nine species. Affinities of ""Chantransia"" stages to particular taxa were congruent for 70.5% of the samples comparing the rbcL and SSU analyses, which were associated with the same or closely related species for both markers. Sequence divergences have been reported in the ""Chantransia"" stage in comparison to the respective gametophyte, and this matter deserves further attention.

Gracilariopsis mclachlanii sp nov and Gracilariopsis persica sp nov of the Gracilariaceae (Gracilariales, Rhodophyceae) from the Indian Ocean

Two new species of Gracilariopsis from the Indian Ocean are proposed-Gracilariopsis (Gp.) mclachlanii Buriyo, Bellorin et M. C. Oliveira sp. nov. from Tanzania and Gracilariopsis persica Bellorin, Sohrabipour et E. C. Oliveira sp. nov. from Iran-based on morphology and DNA sequence data (rbcL gene and SSU rDNA). Both species fit the typical features of Gracilariopsis: axes cylindrical throughout, freely and loosely ramified up to four orders, with an abrupt transition in cell size from medulla to cortex, cystocarps lacking tubular nutritive cells and superficial spermatangia. Nucleotide sequence comparisons of rbcL and SSU rDNA placed both species into the Gracilariopsis clade as distinct species from all the accepted species for this genus, forming a deeply divergent lineage together with some species from the Pacific. The new species are very difficult to distinguish on morphological grounds from other species of Gracilariopsis, stressing the importance of homologous molecular marker comparisons for the species recognition in this character-poor genus.

Phylogeny, species limits, and biogeography of the Brazilian lizards of the genus Eurolophosaurus (Squamata : Tropiduridae) as inferred from mitochondrial DNA sequences

Phylogenetic relationships and divergence times for 10 populations of the three recognized ""species"" of Brazilian lizards of genus Eurolophosaurus were estimated from 1229 bp of cyt b, COI, 12S, and 16S rRNA mitochondrial gene segments. Eurolophosaurus is monophyletic and the basal split within the genus separates E divaricatus from a clade comprising E amathites and E nanuzae. Three populations of E divaricatus, which occurs along the western bank of Rio S (a) over tildeo Francisco, were consistently grouped together. Oil the east bank of the river, E amathites and E nanuzae from state of Bahia were recovered as the sister group of E nanuzae populations from state of Minas Gerais. The paraphyly of E nanuzae and the high divergence levels among populations of E divaricatus strongly suggest that species limits in Eurolophosaurus should be revised. Even considering an extreme evolutionary rate of 2.8% sequence divergence per million years for the four gene segments analyzed together, E. divaricatus would have separated from the two other species by at least 5.5 my ago, and E. amathites from E nanuzae populations from Bahia and Minas Gerais, respectively, by 1.5 and 3.5 my. The paleolacustrine hypothesis and changes in the course of the river potentially explain faunal divergence in the area, but divergences are much older than previously admitted. (C) 2007 Elsevier Inc. All rights reserved.

Comparative Cytogenetics and Molecular Phylogeography in the Group Astyanax altiparanae - Astyanax aff. bimaculatus (Teleostei, Characidae)

The genus Astyanax comprises small characin fish of the neotropical region. The so-called `yellow-tailed characins` compose one of the most widely distributed Astyanax groups. A. altiparanae and A. aff. bimaculatus, are evolutionarily closely related and commonly found in several Brazilian hydrographic basins. In the present work, chromosomal data of specimens of A. altiparanae and A. aff. bimaculatus from 4 hydrographic basins in the states of Sao Paulo (Upper Tiete, Paranapanema, Ribeira de Iguape) and Rio de Janeiro (Guapimirim) are shown. All the populations showed 50 chromosomes, with different karyotypic formula. Although only a single Ag-NOR bearing chromosome pair was observed, all populations possess multiple cistrons of 18S rDNA. FISH with the 5S rDNA probe showed single signals at the interstitial position of one metacentric chromosome pair. C-bands are distributed in the terminal and interstitial regions of several chromosomes. However, the As-51 satDNA are frugally located in a few chromosomes of fishes from Upper Tiete, Paranapanema and Guapimirim Rivers, being absent in individuals of A. aff. bimaculatus from Ribeira de Iguape River basin. Beside these 4 populations, molecular phylogeography studies were also performed in individuals from Middle and Lower Tiete River basin and from 2 additional collection sites in the Paranapanema and Ribeira de Iguape River basins. The phylogeographic analysis using 2 mtDNA regions (totalizing 1.314 bp of ND2 and ATPase6/8 genes) of 8 populations of the group of `yellow-tailed characins` from 3 major hydrographic basins showed structuring of populations, suggesting a correlation between chromosomal (nuclear) and molecular (mitochondrial) data. Copyright (C) 2011 S. Karger AG, Basel