For the first time, full-length 16S rRNA sequencing method was applied to disclose the bacterial species and communities of a full-scale wastewater treatment plant using an anaerobic/anoxic/oxic (A/A/O) process in Wuhan, China. The compositions of the bacteria at phylum and class levels in the activated sludge were similar to which revealed by Illumina Miseq sequencing. At genus and species levels, third-generation sequencing showed great merits and accuracy. Typical functional taxa classified to ammonia-oxidizing bacteria (AOB), nitrite-oxidizing bacteria (NOB), denitrifying bacteria (DB), anaerobic ammonium oxidation bacteria (ANAMMOXB) and polyphosphate-accumulating organisms (PAOs) were presented, which were Nitrosomonas (1.11%), Nitrospira (3.56%), Pseudomonas (3.88%),…
Two Marinobacter sp. NP-4 and NP-6 were isolated from a deep oceanic basaltic crust at North Pond, located at the western flank of the Mid-Atlantic Ridge. These two strains are capable of using multiple carbon sources such as acetate, succinate, glucose and sucrose while take oxygen as a primary electron acceptor. The strain NP-4 is also able to grow anaerobically under 20?MPa, with nitrate as the electron acceptor, thus represents a piezotolerant. To explore the metabolic potentials of Marinobacter sp. NP-4 and NP-6, the complete genome of NP-4 and close-to-complete genome of NP-6 were sequenced. The genome of NP-4 contains…
Development of high-throughput sequencing techniques have greatly benefited our understanding about microbial ecology; yet the methods producing short reads suffer from species-level resolution and uncertainty of identification. Here we optimize PacBio-based metabarcoding protocols covering the Internal Transcribed Spacer (ITS region) and partial Small Subunit (SSU) of the rRNA gene for species-level identification of all eukaryotes, with a specific focus on Fungi (including Glomeromycota) and Stramenopila (particularly Oomycota). Based on tests on composite soil samples and mock communities, we propose best suitable degenerate primers, ITS9munngs + ITS4ngsUni for eukaryotes and selected groups therein and discuss pros and cons of long read-based…
The ruminants are one of the most successful mammalian lineages, exhibiting morphological and habitat diversity and containing several key livestock species. To better understand their evolution, we generated and analyzed de novo assembled genomes of 44 ruminant species, representing all six Ruminantia families. We used these genomes to create a time-calibrated phylogeny to resolve topological controversies, overcoming the challenges of incomplete lineage sorting. Population dynamic analyses show that population declines commenced between 100,000 and 50,000 years ago, which is concomitant with expansion in human populations. We also reveal genes and regulatory elements that possibly contribute to the evolution of the…
Members of the major candidate phylum Dependentiae (a.k.a. TM6) are widespread across diverse environments from showerheads to peat bogs; yet, with the exception of two isolates infecting amoebae, they are only known from metagenomic data. The limited knowledge of their biology indicates that they have a long evolutionary history of parasitism. Here, we present Chromulinavorax destructans (Strain SeV1) the first isolate of this phylum to infect a representative from a widespread and ecologically significant group of heterotrophic flagellates, the microzooplankter Spumella elongata (Strain CCAP 955/1). Chromulinavorax destructans has a reduced 1.2 Mb genome that is so specialized for infection that…
Variable region analysis of 16S rRNA gene sequences is the most common tool in bacterial taxonomic studies. Although used for distinguishing bacterial species, its use remains limited due to the presence of variable copy numbers with sequence variation in the genomes. In this study, 16S rRNA gene sequences, obtained from completely assembled whole genome and Sanger electrophoresis sequencing of cloned PCR products from Serratia fonticola GS2, were compared. Sanger sequencing produced a combination of sequences from multiple copies of 16S rRNA genes. To determine whether the variant copies of 16S rRNA genes affected Sanger sequencing, two ratios (5:5 and 8:2)…
Algal polysaccharides are an important bacterial nutrient source and central component of marine food webs. However, cellular and ecological aspects concerning the bacterial degradation of polysaccharide mixtures, as presumably abundant in natural habitats, are poorly understood. Here, we contextualize marine polysaccharide mixtures and their bacterial utilization in several ways using the model bacterium Alteromonas macleodii 83-1, which can degrade multiple algal polysaccharides and contributes to polysaccharide degradation in the oceans. Transcriptomic, proteomic and exometabolomic profiling revealed cellular adaptations of A. macleodii 83-1 when degrading a mix of laminarin, alginate and pectin. Strain 83-1 exhibited substrate prioritization driven by catabolite repression,…
Recent metagenomic analysis has revealed that our gut microbiota plays an important role in not only the maintenance of our health but also various diseases such as obesity, diabetes, inflammatory bowel disease, and allergy. However, most intestinal bacteria are considered ‘unculturable’ bacteria, and their functions remain unknown. Although culture-independent genomic approaches have enabled us to gain insight into their potential roles, culture-based approaches are still required to understand their characteristic features and phenotypes. To date, various culturing methods have been attempted to obtain these ‘unculturable’ bacteria, but most such methods require advanced techniques. Here, we have tried to isolate possible…
Bacillus sp. 7PJ-16, an endophytic bacterium isolated from a healthy mulberry stem and previously identified as Bacillus tequilensis 7PJ-16, exhibits strong antifungal activity and has the capacity to promote plant growth. This strain was studied for its effectiveness as a biocontrol agent to reduce mulberry fruit sclerotiniose in the field and as a growth-promoting agent for mulberry in the greenhouse. In field studies, the cell suspension and supernatant of strain 7PJ-16 exhibited biocontrol efficacy and the lowest disease incidence was reduced down to only 0.80%. In greenhouse experiments, the cell suspension (1.0?×?106 and 1.0?×?105 CFU/mL) and the cell-free supernatant (100-fold and…
The study of the draft genome of an Antarctic marine ciliate, Euplotes petzi, revealed foreign sequences of bacterial origin belonging to the ?-proteobacterium Francisella that includes pathogenic and environmental species. TEM and FISH analyses confirmed the presence of a Francisella endocytobiont in E. petzi. This endocytobiont was isolated and found to be a new species, named F. adeliensis sp. nov.. F. adeliensis grows well at wide ranges of temperature, salinity, and carbon dioxide concentrations implying that it may colonize new organisms living in deeply diversified habitats. The F. adeliensis genome includes the igl and pdp gene sets (pdpC and pdpE…
Pseudomonas frederiksbergensis ERDD5:01 is a psychrotrophic bacteria isolated from the glacial stream flowing from East Rathong glacier in Sikkim Himalaya. The strain showed survivability at high altitude stress conditions like freezing, frequent freeze-thaw cycles, and UV-C radiations. The complete genome of 5,746,824?bp circular chromosome and a plasmid of 371,027?bp was sequenced to understand the genetic basis of its survival strategy. Multiple copies of cold-associated genes encoding cold active chaperons, general stress response, osmotic stress, oxidative stress, membrane/cell wall alteration, carbon storage/starvation and, DNA repair mechanisms supported its survivability at extreme cold and radiations corroborating with the bacterial physiological findings. The…
Application of organic fertilisers to soil prevents erosion, improves fertility and may suppress certain soil-borne plant pathogens, but it is still unclear how different trophic groups of fungi and oomycetes respond to long-term fertilisation treatment. The objective of the study was to examine the effect of different fertilisation regimes on fungal and oomycete pathogen- and mycorrhizal symbiont diversity and community structure in both soil and roots, using PacBio SMRT sequencing. The field experiment included three fertilisation treatments that have been applied since 1989: nitrogen fertilisation (WOM), nitrogen fertilisation with manure amendment (FYM) and alternative organic fertilisation (AOF), each applied at…
Traditional fermented dairy foods have been the major components of the Mongolian diet for millennia. In this study, we used propidium monoazide (PMA; binds to DNA of nonviable cells so that only viable cells are enumerated) and single-molecule real-time sequencing (SMRT) technology to investigate the total and viable bacterial compositions of 19 traditional fermented dairy foods, including koumiss from Inner Mongolia (KIM), koumiss from Mongolia (KM), and fermented cow milk from Mongolia (CM); sample groups treated with PMA were designated PKIM, PKM, and PCM. Full-length 16S rRNA sequencing identified 195 bacterial species in 121 genera and 13 phyla in PMA-treated…
Amplicon sequencing of the 16S rRNA gene is the predominant method to quantify microbial compositions and to discover novel lineages. However, traditional short amplicons often do not contain enough information to confidently resolve their phylogeny. Here we present a cost-effective protocol that amplifies a large part of the rRNA operon and sequences the amplicons with PacBio technology. We tested our method on a mock community and developed a read-curation pipeline that reduces the overall read error rate to 0.18%. Applying our method on four environmental samples, we captured near full-length rRNA operon amplicons from a large diversity of prokaryotes. The…
Root-associated microbes play a key role in plant performance and productivity, making them important players in agroecosystems. So far, very few studies have assessed the impact of different farming systems on the root microbiota and it is still unclear whether agricultural intensification influences the structure and complexity of microbial communities. We investigated the impact of conventional, no-till, and organic farming on wheat root fungal communities using PacBio SMRT sequencing on samples collected from 60 farmlands in Switzerland. Organic farming harbored a much more complex fungal network with significantly higher connectivity than conventional and no-till farming systems. The abundance of keystone…