Transcription Analysis of Streptococcus thermophilus Phages in the Lysogenic State

AbstractThe transcription of prophage genes was studied in two lysogenic Streptococcus thermophilus cells by Northern blot and primer-extension experiments. In the lysogen containing the cos-site phage Sfi21 only two gene regions of the prophage were transcribed. Within the lysogeny module an 1.6-kb-long mRNA started at the promoter of the phage repressor gene and covered also the next two genes, including a superinfection exclusion (sie) gene. A second, quantitatively more prominent 1-kb-long transcript was initiated at the promoter of the sie gene. Another prophage transcript of 1.6-kb length covered a group of genes without database matches that were located between the lysin gene and the right attachment site. The rest of the prophage genome was transcriptionally silent. A very similar transcription pattern was observed for a S. thermophilus lysogen containing the pac-site phage O1205 as a prophage. Prophages from pathogenic streptococci encode virulence genes downstream of the lysin gene. We speculate that temperate phages from lactic streptococci also encode nonessential phage genes (“lysogenic conversion genes”) in this region that increase the ecological fitness of the lysogen to further their own evolutionary success. A comparative genome analysis revealed that many temperate phages from low GC content Gram-positive bacteria encode a variable number of genes in that region and none was linked to known phage-related function. Prophages from pathogenic streptococci encode toxin genes in this region. In accordance with theoretical predictions on prophage–host genome interactions a prophage remnant was detected in S. thermophilus that had lost most of the prophage DNA while transcribed prophage genes were spared from the deletion process

Implementación del sistema de facturación electrónica en la empresa Infotec, Lima - 2020

La presente investigación buscó analizar la implementación del sistema de facturación electrónica en la empresa INFOTEC, Lima 2020; para ello se valió de una metodología de enfoque cuantitativo, de tipo descriptivo y diseño no experimental; la técnica utilizada fue la encuesta y el instrumento fue el cuestionario, el mismo que fue validado en su contenido en cuanto a los criterios de Relevancia, Coherencia y Claridad. El muestreo fue no probabilístico a conveniencia, por lo que la muestra se consideró censal con un número de 25 trabajadores pertenecientes al área contable y administrativa de la empresa en mención. Los resultados obtenidos demuestran que la implementación del sistema según las respuestas de los trabajadores, se realizó por exigencia de la Sunat (60%), en su mayoría (56%) contribuye a que los EEFF se presenten a tiempo, el personal no asiste a las capacitaciones sobre el nuevo sistema (52%), el 80% desconoce sobre la página web de publicación de comprobantes, el 84% no conoce el tipo de sistema de emisión electrónica que maneja, el 96% considera eficiente el proveedor de internet, el 84% conoce el tipo de proveedor de software, que los comprobantes se emiten según los parámetros y el procedimiento establecidos por Sunat (100%) y que cuentan con una certificación digital vigente (100%). Por lo que se concluye que la implementación del sistema de facturación electrónica es eficiente, fácil de manejo e implementación pero que requiere de una capacitación previa y posterior del personal

Microbial diversity in the human intestine and novel insights from metagenomics

Bacterial communities reside in very different ecological niches on and within the human host, such as those associated with the alimentary tract. The human gastrointestinal tract is populated with as many as 100 trillion bacterial cells, whose collective genome likely reflects the co-evolution between the microbial community and its host. Recent progress has highlighted the intriguing diversity of these bacterial populations and their important contributions to human physiology. Thus, a thorough understanding of the autochthonous component of the intestinal microbiota is expected to provide crucial information not only on how to develop therapies for various gastrointestinal diseases but also on how to choose the next generation of probiotic bacteria as part of novel functional foods. Recently, novel culture-independent approaches such as metagenomics-based techniques were shown to be crucially important for the exploration of the biodiversity of the human intestinal microbiota

Transcriptomic landscape of the kleptoplastic sea slug Elysia viridis

The longevity of kleptoplasts and sea slugs during starvation may be mediated by multiple factors, including the recognition of the plastids during feeding by multiple receptors (e.g. PRRs, CTLRs and SRs) and ROS-quenching proteins using enzymatic and nonenzymatic mechanisms. In particular, in the transcriptome of E. viridis we found that the presence of CDSs corresponds to multiple PRRs that may be involved in the plastid-recognition process; this is despite the fact that this species has a low receptor richness in comparison with other elysoids (Melo Clavijo et al., 2020). In addition, we also detected multiple enzymatic families involved in the ROS-quenching response. In contrast, the production of antioxidant compounds may contribute in only a minor way to the control of oxidative stress. A further enriched GO category in species that sequester chloroplasts corresponded to G protein-coupled receptors, which suggests that these receptors may be required for plastid recognition in Sacoglossan sea slugs, paralleling their role in other symbioses, such as the mutualism between cnidarians and dinoflagellates (Rosset et al., 2020). Sacoglossan sea slugs may also require the presence of iron ions to reduce the oxidative stress generated after plastid acquisition. All this evidence, derived from the transcriptome analysis of E. viridis, sheds interesting new light on the possible mechanisms used by sea slugs to recognize and establish kleptoplasts within their bodies.Xunta de Galicia | Ref. GPC2014/067Xunta de Galicia | Ref. ED481A-2018/30

Isolation of Lactobacilli with probiotic properties from the human stomach

Aims: Recent evidence suggests that the human gastric microbiota is much more diverse than previously thought. The aim of this study was to assess the potential for isolating lactobacilli from the human stomach.Methods and Results: Lactobacilli were selectively cultured from gastric biopsies from 12 patients undergoing routine endoscopy. Lactobacilli were present in four of 12 biopsies. We isolated, in total 10 different strains representing five species (Lactobacillus gasseri, L. fermentum, L. vaginalis, L. reuteri and L. salivarius). The 10 isolates varied greatly in their ability to inhibit the growth of two Gram-positive bacteria and two Gram-negative bacteria. Furthermore, the acid and bile resistance profiles of the 10 isolates spanned a wide range. Conclusions: Five different Lactobacillus species were cultured from human gastric biopsies for the first time. Significance and Impact of the Study: Diverse Lactobacillus species are more prevalent in the human stomach than previously recognized, representing an untapped source of bacteria with beneficial probiotic and/or biotechnological properties

Tissue-specific transcripomes of Mytilus galloprovincialis reveal new functions

Comunicación presentada en el XII International Symposium on Genetics in Aquaculture, Santiago de Compostela, 21-27 de junio de 2015The Mediterranean mussel (Mytilus galloprovincialis) is a cosmopolitan, cultured bivalve with worldwide commercial and ecological importance. There is a need to increase our knowledge of the molecular mechanisms involved in Mytilus physiology from a qualitative and quantitative point of view. In order to start filling this gap we have used RNA-Seq to study the transcriptome of mantle, muscle and gills from naïve mussels and hemocytes exposed to distinct stimuli. After the analysis of the complete mussel transcriptome, we confirmed that we have achieved a completeness of the transcriptome of 95.16%. As mussel and oyster are model species for bivalves, we compared them and found that the shared proteins with a sequence identity over 80% represent less than 10% of their transcriptomes. In the whole mussel transcriptome it is important to highlight that the response to infectious diseases and cancer were pathways highly represented. However, only 55% of the transcripts were shared across all tissues. Hemocyte and gill transcriptomes were most different, with 60% shared transcripts, while mantle and muscle transcriptomes were most similar, with 77% shared transcripts. The transcriptomes showed characteristic expression profiles in agreement with their structures and functions: stimulated hemocytes confirmed their immune function showing a high representation of defense and immune-related expressed genes; the gills presented many transcripts assigned to both structure and recognition of non-self patterns; the mantle showed an abundance of transcripts related to reproduction and shell formation and, finally, the muscle expressed many myofibril and calcium-related proteins. However, we could find other complex and specialized functions of each tissue, not previously reported: gills and its probable osmotic and homeostatic function; muscle revealed unexpected defense functions; and in mantle additional and interesting antifungal and sensorial functions were discovered, but also hematopoiesis transcripts were exclusively expressed in mantle, confirming its possible role as the hematopoietic tissue in bivalves. This information will provide new insight to bivalves, and specially mussel, physiologyPeer reviewe

Small-scale spatial variation of meiofaunal communities in Lima estuary (NW Portugal) assessed through metabarcoding

Meiofaunal organisms play a key role in estuarine ecosystems, being responsible for significant ecological processes. However, meiofauna constitutes a particularly difficult community to be monitored through conventional morphology-based approaches. New emerging tools, such as DNA metabarcoding, facilitate the access to these communities and provide an opportunity to develop routine monitoring programs. In the present study, the small-scale spatial variation of meiofaunal communities in the Lima estuary (NW Portugal) was investigated using DNA metabarcoding. The first stage of the study aimed to establish the amount of sediment sample to be used for DNA extraction and to test six primer pairs, three of them amplifying fragments from the mitochondrial cytochrome c oxidase gene (COI) and three other the nuclear ribosomal 18S rRNA gene (18S). In a subsequent stage, sediment samples were collected in four stations along an estuarine gradient (salinity ranging between 9 and 28), in which six sampling points about 4–5 m apart were considered: three in the high intertidal and three in the mid intertidal. After the DNA extraction from sediments, COI and 18S amplicon libraries were produced and sequenced in an Illumina MiSeq platform. OTUs (operational taxonomic units) recovered by either COI or 18S displayed generally high turnover in occurrence among sampling points within a station and tidal horizon, among tidal horizons within a station, and among stations of distinct salinity (approx. 60–93%). Both markers recorded little variation among stations in OTU richness and in the taxonomic composition of the most dominant groups. However, the meiofauna detected differed qualitatively between the two markers used; Amoebozoa and Cnidaria were mostly detected with COI while Ciliophora and Platyhelminthes with 18S. In addition, the structure of the meiofauna community diverged significantly among stations and was strongly influenced by salinity and sediment features. Globally, results indicated a highly patchy distribution of meiofauna taxa in the Lima estuary, revealed by the high OTU turnover even between sampling points only a few meters apart. Hence, eDNA-based meiofauna surveys require consideration of the necessary sampling effort on relatively small spatial scales, as well as an appreciation of the tidal level-induced variation of these communities.This study was funded by the project “The NextSea: Next generation monitoring of coastal ecosystems in a scenario of global change” (operação NORTE-01-0145-FEDER-000032), supported by Norte Portugal Regional Operational Programme (NORTE 2020), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF). Maria Fais and Sofia Duarte were supported, respectively by a PhD (SFRH/BD/113547/2015) and a post-doc fellowship (SFRH/BPD/109842/2015) from FCT. The authors would like to thank Prof. Jesús Troncoso (University of Vigo) and Prof. Pedro Gomes (University of Minho) for their availability and hospitality for the lab processing, as well as Barbara Leite (University of Minho) and Manuel Mendoza (University of Vigo) for their help during the sampling and data analysis phases, respectively

Patterns of spatial and temporal variation in estuarine meiofaunal communities assessed through DNA metabarcoding: a case study in the Lima estuary (NW Portugal)

Meiobenthic organisms are key components of estuarine environments, providing invaluable functions and services. In particular, meiofaunal organisms participate actively in nutrient cycles and energy flux, supporting higher trophic levels, as well as the stabilization of sediments. Due to their small size (between 30 µm and 1mm) and high dispersal potential, meiofauna was considered to have a ubiquitous distribution. However, with the recent progress of DNA-based technologies, in particular, DNA metabarcoding, the cosmopolitan meiofauna paradox is being questioned, suggesting that meiobenthic communities can comprise also endemic and narrow-range species. Several studies on the spatial variability of meiofauna at small-scale (within meters) and at meso-scale (meters to km) have been carried out in an attempt to elucidate community structure, both using traditional approaches (such as morphological identification) and DNA-based tools. All these studies focused on the importance of environmental features (i.e., salinity, sediments’ size, percentage of organic matter) in the distribution of particular taxonomic groups, as well as of their bio-ecological characteristics, which may overall explain the spatial structure of the meiobenthic communities. However, little attention has been given to the temporal variation of meiofauna communities in estuaries, and to the best of our knowledge, no research on the spatial-temporal dynamics of Iberian estuarine meiofauna has been yet performed using DNA-based tools. Although DNA metabarcoding is assumed to i) allow a faster detection, ii) identify hard morphologically-intractable meiobenthic organisms or communities, and iii) target different taxonomic groups simultaneously, further refinement is required to better tailor this method for meiofauna profiling. A well-defined choice of marker loci and primer pairs, and the design of robust protocols for sampling effort and for the bioinformatics data processing and analysis is needed to avoid the increase of technical bias during the workflow. The main goal of the present study was to investigate spatial and temporal variations of the estuarine meiofauna communities using DNA metabarcoding. Sediments were collected in four sampling stations in the Lima estuary (North-West of Portugal), in June 2017 and June 2018, from the first 5 cm of sediment directly to 50 mL sterile falcon tubes (3 cm ø). Three samples were collected at the high and middle intertidal zones in each sampling station. For each site and yearly sampling campaign, we assessed salinity, total organic matter (TOM) and grain size. DNA was extracted from 10.0 g (± 0.50 g) of sediment from each collected sample, using the DNeasy PowerMax Isolation kit (QIAGEN®). Two different primer pairs were used for the production of amplicon libraries and their high-throughput sequencing (Illumina-MiSeq): a sub-region from the mitochondrial cytochrome c oxidase gene (COI) and the V4 hypervariable region of the nuclear ribosomal 18S rRNA gene (18S). Raw reads were quality-filtered by the removal of sequencing adapters and trimming of low-quality bases. Further de-multiplexing and read processing were carried out in mothur, using customized procedures. Reads were clustered into OTUs using a 97% similarity threshold. Then, the representative sequences of each OTU were BLASTed against the GenBank non-redundant nucleotide database and taxonomically assigned in MEGAN v.6.13. In terms of OTUs richness and turnover, our results showed that our communities globally changed along space and time. Meiobenthic structure, in general, displayed well-separated temporal and spatial clusters, with some generalist OTUs occurring along with the stations for both sampling years. Qualitatively, the two primer pairs detected different taxonomic groups attributed to meiofauna in a complementary manner. Nematoda, Platyhelminthes, Annelida, Arthropoda Crustacea, Amoebozoa and Ciliophora were the most abundant phyla, with different temporal and spatial proportions. Our results indicated that Lima estuarine meiofaunal communities are strongly influenced by the salinity, sediment grain size, and total organic matter, affecting meiofauna distribution among the sampling stations. Moreover, further differences on a time scale were found between the sampling years, probably as a consequence of greater rainfall during 2018 (https://www.pordata.pt). With a careful sampling design, laboratory and data analysis protocols, DNA metabarcoding can constitute a powerful approach for meiobenthic communities profiling, although further efforts are required for preparing dedicated meiofauna references libraryThe NextSea, under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF). MF and SF benefitted from FCT Ph.D. (SFRH/BD/113547/2015) and post-doc (SFRH/BPD/109842/2015) fellowship

The Bifidobacterium dentium Bd1 Genome Sequence Reflects Its Genetic Adaptation to the Human Oral Cavity

Bifidobacteria, one of the relatively dominant components of the human intestinal microbiota, are considered one of the key groups of beneficial intestinal bacteria (probiotic bacteria). However, in addition to health-promoting taxa, the genus Bifidobacterium also includes Bifidobacterium dentium, an opportunistic cariogenic pathogen. The genetic basis for the ability of B. dentium to survive in the oral cavity and contribute to caries development is not understood. The genome of B. dentium Bd1, a strain isolated from dental caries, was sequenced to completion to uncover a single circular 2,636,368 base pair chromosome with 2,143 predicted open reading frames. Annotation of the genome sequence revealed multiple ways in which B. dentium has adapted to the oral environment through specialized nutrient acquisition, defences against antimicrobials, and gene products that increase fitness and competitiveness within the oral niche. B. dentium Bd1 was shown to metabolize a wide variety of carbohydrates, consistent with genome-based predictions, while colonization and persistence factors implicated in tissue adhesion, acid tolerance, and the metabolism of human saliva-derived compounds were also identified. Global transcriptome analysis demonstrated that many of the genes encoding these predicted traits are highly expressed under relevant physiological conditions. This is the first report to identify, through various genomic approaches, specific genetic adaptations of a Bifidobacterium taxon, Bifidobacterium dentium Bd1, to a lifestyle as a cariogenic microorganism in the oral cavity. In silico analysis and comparative genomic hybridization experiments clearly reveal a high level of genome conservation among various B. dentium strains. The data indicate that the genome of this opportunistic cariogen has evolved through a very limited number of horizontal gene acquisition events, highlighting the narrow boundaries that separate commensals from opportunistic pathogens

Phages and the Evolution of Bacterial Pathogens: from Genomic Rearrangements to Lysogenic Conversion

Comparative genomics demonstrated that the chromosomes from bacteria and their viruses (bacteriophages) are coevolving. This process is most evident for bacterial pathogens where the majority contain prophages or phage remnants integrated into the bacterial DNA. Many prophages from bacterial pathogens encode virulence factors. Two situations can be distinguished: Vibrio cholerae, Shiga toxin-producing Escherichia coli, Corynebacterium diphtheriae, and Clostridium botulinum depend on a specific prophage-encoded toxin for causing a specific disease, whereas Staphylococcus aureus, Streptococcus pyogenes, and Salmonella enterica serovar Typhimurium harbor a multitude of prophages and each phage-encoded virulence or fitness factor makes an incremental contribution to the fitness of the lysogen. These prophages behave like “swarms” of related prophages. Prophage diversification seems to be fueled by the frequent transfer of phage material by recombination with superinfecting phages, resident prophages, or occasional acquisition of other mobile DNA elements or bacterial chromosomal genes. Prophages also contribute to the diversification of the bacterial genome architecture. In many cases, they actually represent a large fraction of the strain-specific DNA sequences. In addition, they can serve as anchoring points for genome inversions. The current review presents the available genomics and biological data on prophages from bacterial pathogens in an evolutionary framework