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Asset Tag: Microbial community

September 22, 2019

Molecular genetic diversity and characterization of conjugation genes in the fish parasite Ichthyophthirius multifiliis.

Ichthyophthirius multifiliis is the etiologic agent of “white spot”, a commercially important disease of freshwater fish. As a parasitic ciliate, I. multifiliis infects numerous host species across a broad geographic range. Although Ichthyophthirius outbreaks are difficult to control, recent sequencing of the I. multifiliis genome has revealed a number of potential metabolic pathways for therapeutic intervention, along with likely vaccine targets for disease prevention. Nonetheless, major gaps exist in our understanding of both the life cycle and population structure of I. multifiliis in the wild. For example, conjugation has never been described in this species, and it is unclear whether I. multifiliis undergoes sexual reproduction, despite the presence of a germline micronucleus. In addition, no good methods exist to distinguish strains, leaving phylogenetic relationships between geographic isolates completely unresolved. Here, we compared nucleotide sequences of SSUrDNA, mitochondrial NADH dehydrogenase subunit I and cox-1 genes, and 14 somatic SNP sites from nine I. multifiliis isolates obtained from four different states in the US since 1995. The mitochondrial sequences effectively distinguished the isolates from one another and divided them into at least two genetically distinct groups. Furthermore, none of the nine isolates shared the same composition of the 14 somatic SNP sites, suggesting that I. multifiliis undergoes sexual reproduction at some point in its life cycle. Finally, compared to the well-studied free-living ciliates Tetrahymena thermophila and Paramecium tetraurelia, I. multifiliis has lost 38% and 29%, respectively, of 16 experimentally confirmed conjugation-related genes, indicating that mechanistic differences in sexual reproduction are likely to exist between I. multifiliis and other ciliate species. Copyright © 2015 Elsevier Inc. All rights reserved.

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September 22, 2019

PhyloPythiaS+: a self-training method for the rapid reconstruction of low-ranking taxonomic bins from metagenomes.

Background. Metagenomics is an approach for characterizing environmental microbial communities in situ, it allows their functional and taxonomic characterization and to recover sequences from uncultured taxa. This is often achieved by a combination of sequence assembly and binning, where sequences are grouped into ‘bins’ representing taxa of the underlying microbial community. Assignment to low-ranking taxonomic bins is an important challenge for binning methods as is scalability to Gb-sized datasets generated with deep sequencing techniques. One of the best available methods for species bins recovery from deep-branching phyla is the expert-trained PhyloPythiaS package, where a human expert decides on the taxa to incorporate in the model and identifies ‘training’ sequences based on marker genes directly from the sample. Due to the manual effort involved, this approach does not scale to multiple metagenome samples and requires substantial expertise, which researchers who are new to the area do not have. Results. We have developed PhyloPythiaS+, a successor to our PhyloPythia(S) software. The new (+) component performs the work previously done by the human expert. PhyloPythiaS+ also includes a new k-mer counting algorithm, which accelerated the simultaneous counting of 4-6-mers used for taxonomic binning 100-fold and reduced the overall execution time of the software by a factor of three. Our software allows to analyze Gb-sized metagenomes with inexpensive hardware, and to recover species or genera-level bins with low error rates in a fully automated fashion. PhyloPythiaS+ was compared to MEGAN, taxator-tk, Kraken and the generic PhyloPythiaS model. The results showed that PhyloPythiaS+ performs especially well for samples originating from novel environments in comparison to the other methods. Availability. PhyloPythiaS+ in a virtual machine is available for installation under Windows, Unix systems or OS X on: https://github.com/algbioi/ppsp/wiki.

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September 22, 2019

Daily HIV pre-exposure prophylaxis (PrEP) with tenofovir disoproxil fumarate-emtricitabine reduced Streptococcus and increased Erysipelotrichaceae in rectal microbiota.

Daily PrEP is highly effective at preventing HIV-1 acquisition, but risks of long-term tenofovir disoproxil fumarate plus emtricitabine (TDF-FTC) include renal decline and bone mineral density decrease in addition to initial gastrointestinal side effects. We investigated the impact of TDF-FTC on the enteric microbiome using rectal swabs collected from healthy MSM before PrEP initiation and after 48 to 72 weeks of adherent PrEP use. The V4 region of the 16S ribosomal RNA gene sequencing showed that Streptococcus was significantly reduced from 12.0% to 1.2% (p?=?0.036) and Erysipelotrichaceae family was significantly increased from 0.79% to 3.3% (p?=?0.028) after 48-72 weeks of daily PrEP. Catenibacterium mitsuokai, Holdemanella biformis and Turicibacter sanguinis were increased within the Erysipelotrichaceae family and Streptococcus agalactiae, Streptococcus oralis, Streptococcus mitis were reduced. These changes were not associated with host factors including PrEP duration, age, race, tenofovir diphosphate blood level, any drug use and drug abuse, suggesting that the observed microbiome shifts were likely induced by daily PrEP use. Long-term PrEP resulted in increases of Catenibacterium mitsuokai and Holdemanella biformis, which have been associated with gut microbiome dysbiosis. Our observations can aid in characterizing PrEP’s side effects, which is likely to improve PrEP adherence, and thus HIV-1 prevention.

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September 22, 2019

The influence of energy harvesting strategies on performance and microbial community for sediment microbial fuel cells

Sediment microbial fuel cells (SMFCs) are being developed as potential energy sources where remote sensing and monitoring would be useful. Several energy harvesting techniques for SMFCs have emerged, but effects of these different strategies on startup, performance, and microbial community are not well understood. We investigated these effects by comparing a continuous energy harvesting (CEH) strategy with an intermittent energy harvesting (IEH) strategy. During startup, IEH systems immediately produced higher power and were cathode limited. CEH systems exhibited a gradual power increase and were anode-limited during startup. Both system types produced similar amounts of steady-state power, 1.5 mW ft-2 (16 mW m-2) when optimized. However, an IEH system using unoptimized settings could not be subsequently switched to optimal settings and produce expected power levels. The choice of energy harvester did not appear to significantly affect steady-state community structure. Anodes were dominated by ?- and d-proteobacteria while a- and ?-proteobacteria dominated cathodes. The results suggest performance and community structure are unaffected by energy harvesting strategy, but that startup conditions influence the initial amount of harvested energy and steady-state performance, suggesting future investigations into optimizing startup of these systems are critical for rapidly generating maximum power.

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September 22, 2019

Bacterial community structure in simultaneous nitrification, denitrification and organic matter removal process treating saline mustard tuber wastewater as revealed by 16S rRNA sequencing.

A simultaneous nitrification, denitrification and organic matter removal (SNDOR) process in sequencing batch biofilm reactor (SBBR) was established to treat saline mustard tuber wastewater (MTWW) in this study. An average COD removal efficiency of 86.48% and total nitrogen removal efficiency of 86.48% were achieved at 30gNaClL(-1) during 100days’ operation. The underlying mechanisms were investigated by PacBio SMRT DNA sequencing (V1-V9) to analyze the microbial community structures and its variation from low salinity at 10gNaClL(-1) to high salinity at 30gNaClL(-1). Results showed elevated salinity did not affect biological performance but reduced microbial diversity in SBBR, and halophilic bacteria gradually predominated by succession. Despite of high C/N, autotrophic ammonia-oxidizing bacteria (AOB) Nitrosomonas and ammonia-oxidizing archaea (AOA) Candidatus Nitrososphaera both contributed to ammonium oxidation. As salinity increasing, nitrite-oxidizing bacteria (NOB) were significantly inhibited, partial nitrification and denitrification (PND) process gradually contributed to nitrogen removal. Copyright © 2016 Elsevier Ltd. All rights reserved.

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September 22, 2019

Evaluating the mobility potential of antibiotic resistance genes in environmental resistomes without metagenomics.

Antibiotic resistance genes are ubiquitous in the environment. However, only a fraction of them are mobile and able to spread to pathogenic bacteria. Until now, studying the mobility of antibiotic resistance genes in environmental resistomes has been challenging due to inadequate sensitivity and difficulties in contig assembly of metagenome based methods. We developed a new cost and labor efficient method based on Inverse PCR and long read sequencing for studying mobility potential of environmental resistance genes. We applied Inverse PCR on sediment samples and identified 79 different MGE clusters associated with the studied resistance genes, including novel mobile genetic elements, co-selected resistance genes and a new putative antibiotic resistance gene. The results show that the method can be used in antibiotic resistance early warning systems. In comparison to metagenomics, Inverse PCR was markedly more sensitive and provided more data on resistance gene mobility and co-selected resistances.

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September 22, 2019

Koumiss consumption alleviates symptoms of patients with chronic atrophic gastritis: A possible link To modulation of gut microbiota

Intestinal dysbiosisis closely related to a variety of medical conditions, especially gastrointestinal diseases. The present study aimed to investigate the effects of koumiss on chronic atrophic gastritis (CAG) in an out-patient clinical trial (n = 10; all female subjects aged 41-55; body mass index ranging from 19.5 to 25.8). Each patient consumed three servings of koumiss per day (i.e. 250 ml daily before each of 3 meals) for a 60-day period. The improvement of patients’ symptoms was monitored by comparing the total scores of symptoms before and after the treatment. Meanwhile, the changes in the patients’ fecal microbiota composition and specific blood parameters were determined. After the 60-day koumiss administration, significant symptom improvements were observed, as evidenced by the reduction of the total symptoms score, and changes in blood platelet and cholesterol levels. The changes in patients’ fecal microbiota composition were found. The patients’ fecal microbiota fell into two distinct enterotypes, Bacteroides dorei/ Bacteroides uniformis (BB-enterotype) and Prevotella copri (P-enterotype). Significant less Bacteroides uniformis was found in the BB-enterotype patient group, while significant more butyrate-producing bacteria (e.g. Eubacterium rectale and Faecalibacterium prausnitzii) were found in the P-enterotype patient group, following koumiss administration. After stopping koumiss consumption, the relative abundance of some biomarker taxa returned to the original level, suggesting that the gut microbiota modulatory effect was not permanent and that continuous koumiss administration was required to maintain the therapeutic effect. In conclusion, koumiss consumption could alleviate the symptoms of CAG patients. Our results may help understand the mechanism of koumiss in alleviating CAG disease symptoms, facilitating the development of such products with desired therapeutic functions.

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September 22, 2019

A comprehensive fungi-specific 18S rRNA gene sequence primer toolkit suited for diverse research issues and sequencing platforms.

Several fungi-specific primers target the 18S rRNA gene sequence, one of the prominent markers for fungal classification. The design of most primers goes back to the last decades. Since then, the number of sequences in public databases increased leading to the discovery of new fungal groups and changes in fungal taxonomy. However, no reevaluation of primers was carried out and relevant information on most primers is missing. With this study, we aimed to develop an 18S rRNA gene sequence primer toolkit allowing an easy selection of the best primer pair appropriate for different sequencing platforms, research aims (biodiversity assessment versus isolate classification) and target groups.We performed an intensive literature research, reshuffled existing primers into new pairs, designed new Illumina-primers, and annealing blocking oligonucleotides. A final number of 439 primer pairs were subjected to in silico PCRs. Best primer pairs were selected and experimentally tested. The most promising primer pair with a small amplicon size, nu-SSU-1333-5’/nu-SSU-1647-3′ (FF390/FR-1), was successful in describing fungal communities by Illumina sequencing. Results were confirmed by a simultaneous metagenomics and eukaryote-specific primer approach. Co-amplification occurred in all sample types but was effectively reduced by blocking oligonucleotides.The compiled data revealed the presence of an enormous diversity of fungal 18S rRNA gene primer pairs in terms of fungal coverage, phylum spectrum and co-amplification. Therefore, the primer pair has to be carefully selected to fulfill the requirements of the individual research projects. The presented primer toolkit offers comprehensive lists of 164 primers, 439 primer combinations, 4 blocking oligonucleotides, and top primer pairs holding all relevant information including primer’s characteristics and performance to facilitate primer pair selection.

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September 22, 2019

A response to Lindsey et al. “Wolbachia pipientis should not be split into multiple species: A response to Ramírez-Puebla et al.”.

In Ramírez-Puebla et al. [18] we compared 34 Wolbachia genomes and constructed phylogenetic trees using genomic data. In general, our results were congruent with previously reported phy- logenetic trees [5,9]. Our datasets were carefully selected, checked and analyzed avoiding horizontally transferred genes. In the case of the wAna genome we did not use the raw data, but the assem- bled genome [22] and 31 genes were used to compare in a dataset of conserved proteins. To confirm our conclusions a new phyloge- nomic analysis was performed excluding the wAna strain in the dataset (Fig. 1). The same topology was obtained, therefore indi- cating that the results were not affected by the presence of this particular strain.

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September 22, 2019

Candidatus Dactylopiibacterium carminicum, a nitrogen-fixing symbiont of Dactylopius cochineal insects (Hemiptera: Coccoidea: Dactylopiidae)

The domesticated carmine cochineal Dactylopius coccus (scale insect) has commercial value and has been used for more than 500?years for natural red pigment production. Besides the domesticated cochineal, other wild Dactylopius species such as Dactylopius opuntiae are found in the Americas, all feeding on nutrient poor sap from native cacti. To compensate nutritional deficiencies, many insects harbor symbiotic bacteria which provide essential amino acids or vitamins to their hosts. Here, we characterized a symbiont from the carmine cochineal insects, Candidatus Dactylopiibacterium carminicum (betaproteobacterium, Rhodocyclaceae family) and found it in D. coccus and in D. opuntiae ovaries by fluorescent in situ hybridization, suggesting maternal inheritance. Bacterial genomes recovered from metagenomic data derived from whole insects or tissues both from D. coccus and from D. opuntiae were around 3.6?Mb in size. Phylogenomics showed that dactylopiibacteria constituted a closely related clade neighbor to nitrogen fixing bacteria from soil or from various plants including rice and other grass endophytes. Metabolic capabilities were inferred from genomic analyses, showing a complete operon for nitrogen fixation, biosynthesis of amino acids and vitamins and putative traits of anaerobic or microoxic metabolism as well as genes for plant interaction. Dactylopiibacterium nif gene expression and acetylene reduction activity detecting nitrogen fixation were evidenced in D. coccus hemolymph and ovaries, in congruence with the endosymbiont fluorescent in situ hybridization location. Dactylopiibacterium symbionts may compensate for the nitrogen deficiency in the cochineal diet. In addition, this symbiont may provide essential amino acids, recycle uric acid, and increase the cochineal life span.

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September 22, 2019

Analysis of gut microbiota – An ever changing landscape.

In the last two decades, the field of metagenomics has greatly expanded due to improvement in sequencing technologies allowing for a more comprehensive characterization of microbial communities. The use of these technologies has led to an unprecedented understanding of human, animal, and environmental microbiomes and have shown that the gut microbiota are comparable to an organ that is intrinsically linked with a variety of diseases. Characterization of microbial communities using next-generation sequencing-by-synthesis approaches have revealed important shifts in microbiota associated with debilitating diseases such as Clostridium difficile infection. But due to limitations in sequence read length, primer biases, and the quality of databases, genus- and species-level classification have been difficult. Third-generation technologies, such as Pacific Biosciences’ single molecule, real-time (SMRT) approach, allow for unbiased, more specific identification of species that are likely clinically relevant. Comparison of Illumina next-generation characterization and SMRT sequencing of samples from patients treated for C. difficile infection revealed similarities in community composition at the phylum and family levels, but SMRT sequencing further allowed for species-level characterization – permitting a better understanding of the microbial ecology of this disease. Thus, as sequencing technologies continue to advance, new species-level insights can be gained in the study of complex and clinically-relevant microbial communities.

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September 22, 2019

Saliva and tooth biofilm bacterial microbiota in adolescents in a low caries community.

The oral cavity harbours a complex microbiome that is linked to dental diseases and serves as a route to other parts of the body. Here, the aims were to characterize the oral microbiota by deep sequencing in a low-caries population with regular dental care since childhood and search for association with caries prevalence and incidence. Saliva and tooth biofilm from 17-year-olds and mock bacteria communities were analysed using 16S rDNA Illumina MiSeq (v3-v4) and PacBio SMRT (v1-v8) sequencing including validity and reliability estimates. Caries was scored at 17 and 19 years of age. Both sequencing platforms revealed that Firmicutes dominated in the saliva, whereas Firmicutes and Actinobacteria abundances were similar in tooth biofilm. Saliva microbiota discriminated caries-affected from caries-free adolescents, with enumeration of Scardovia wiggsiae, Streptococcus mutans, Bifidobacterium longum, Leptotrichia sp. HOT498, and Selenomonas spp. in caries-affected participants. Adolescents with B. longum in saliva had significantly higher 2-year caries increment. PacBio SMRT revealed Corynebacterium matruchotii as the most prevalent species in tooth biofilm. In conclusion, both sequencing methods were reliable and valid for oral samples, and saliva microbiota was associated with cross-sectional caries prevalence, especially S. wiggsiae, S. mutans, and B. longum; the latter also with the 2-year caries incidence.

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September 22, 2019

Effects of metal and metalloid pollutants on the microbiota composition of feces obtained from twelve commercial pig farms across China.

Understanding the metal and metalloid contamination and microbiota composition of pig feces is an important step required to support the design and implementation of effective pollution control and prevention strategies. A survey was implemented in 12 locations across China to investigate the content of metals and metalloids, and the main composition of the microbial communities of commercially reared pigs during two growth periods, defined as the early (Q group) and the later fattening growth phases (H group). These data showed widespread Al, Mn, Cu, Zn, and Fe pollution in pig feces. The concentration of Zn in the Q group feces was nearly two times higher than the levels measured in the H group. The microbial composition of the Q group exhibited greater richness of operational taxonomic units (OTUs) and fewer bacteria associated with zoonotic diseases compared with the microbial composition of the H group. Spearman rank correlation analysis showed that Cu and northern latitudes had a significant positive effect on the richness of bacterial communities in pig feces. Zn and Cd exhibited the biggest impact on microbial community composition based on canonical correspondence analysis. Functional metagenomic prediction indicated that about 0.8% genes present in the pig feces bacteria community are related to human diseases, and significantly more predicted pathogenic genes were detected in the H group than in the Q group. These results support the need to monitor heavy metal contamination and to control for zoonotic pathogens disseminated from pig feces in Chinese pig farms. Copyright © 2018. Published by Elsevier B.V.

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September 22, 2019

An intact gut microbiota may be required for lactoferrin-driven immunomodulation in rats

Lactoferrin can modulate both the host immunity and gut microbiota. However, whether the immune modulation requires the gut microbiota has not been directly shown. Thus, our study compared (1) lactoferrin-driven immunomodulation profiles and (2) changes in fecal phylogenic metagenome with and without antibiotics-induced dysbiosis in rats. Rats receiving only lactoferrin but not both lactoferrin and antibiotics had a Th-1 type cytokine serum profile. Significant differences were detected between the fecal microbiota of the lactoferrin and control groups at day 19 and/or day 33 but not initially, with a shift in the major contributors for community dissimilarity to Clostridium, Lactobacillus, and Oscillibacter valericigenes. The antibiotics-induced dysbiosis enriched the proinflammatory phyla, Proteobacteria and Deferribacteres, together with the anti-inflammatory species, Akkermansia muciniphila, while suppressed some butyrate-producers from the Firmicutes phylum. Our study shows that an intact microbiota is necessary for lactoferrin-driven immunomodulation.

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