scholarly journals Identifying optimal bioinformatics protocols for aerosol microbial community data

PeerJ ◽  
2021 ◽  
Vol 9 ◽  
pp. e12065
Author(s):  
Katie Miaow ◽  
Donnabella Lacap-Bugler ◽  
Hannah L. Buckley
Keyword(s):  
Alpha Diversity ◽  
Rrna Gene ◽  
Sequence Variant ◽  
Bioinformatics Pipeline ◽  
Taxonomic Assignment ◽  
Bioinformatic Tools ◽  
Molecular Approaches ◽  
Bacteria And Fungi ◽  
Taxonomic Assignments ◽  
Warming World

Microbes are fundamental to Earth’s ecosystems, thus understanding ecosystem connectivity through microbial dispersal is key to predicting future ecosystem changes in a warming world. However, aerial microbial dispersal remains poorly understood. Few studies have been performed on bioaerosols (microorganisms and biological fragments suspended in the atmosphere), despite them harboring pathogens and allergens. Most environmental microbes grow poorly in culture, therefore molecular approaches are required to characterize aerial diversity. Bioinformatic tools are needed for processing the next generation sequencing (NGS) data generated from these molecular approaches; however, there are numerous options and choices in the process. These choices can markedly affect key aspects of the data output including relative abundances, diversity, and taxonomy. Bioaerosol samples have relatively little DNA, and often contain novel and proportionally high levels of contaminant organisms, that are difficult to identify. Therefore, bioinformatics choices are of crucial importance. A bioaerosol dataset for bacteria and fungi based on the 16S rRNA gene (16S) and internal transcribed spacer (ITS) DNA sequencing from parks in the metropolitan area of Auckland, Aotearoa New Zealand was used to develop a process for determining the bioinformatics pipeline that would maximize the data amount and quality generated. Two popular tools (Dada2 and USEARCH) were compared for amplicon sequence variant (ASV) inference and generation of an ASV table. A scorecard was created and used to assess multiple outputs and make systematic choices about the most suitable option. The read number and ASVs were assessed, alpha diversity was calculated (Hill numbers), beta diversity (Bray–Curtis distances), differential abundance by site and consistency of ASVs were considered. USEARCH was selected, due to higher consistency in ASVs identified and greater read counts. Taxonomic assignment is highly dependent on the taxonomic database used. Two popular taxonomy databases were compared in terms of number and confidence of assignments, and a combined approach developed that uses information in both databases to maximize the number and confidence of taxonomic assignments. This approach increased the assignment rate by 12–15%, depending on amplicon and the overall assignment was 77% for bacteria and 47% for fungi. Assessment of decontamination using “decontam” and “microDecon” was performed, based on review of ASVs identified as contaminants by each and consideration of the probability of them being legitimate members of the bioaerosol community. For this example, “microDecon’s” subtraction approach for removing background contamination was selected. This study demonstrates a systematic approach to determining the optimal bioinformatics pipeline using a multi-criteria scorecard for microbial bioaerosol data. Example code in the R environment for this data processing pipeline is provided.

scholarly journals The cultivation of Pyropia haitanensis has important impacts on the seawater microbial community

2020 ◽  
Vol 32 (4) ◽  
pp. 2561-2573
Author(s):  
Wenlei Wang ◽  
Lei Wu ◽  
Kai Xu ◽  
Yan Xu ◽  
Dehua Ji ◽  
...  
Keyword(s):  
Microbial Community ◽  
Growth And Development ◽  
Alpha Diversity ◽  
18S Rrna Gene ◽  
Pyropia Haitanensis ◽  
Rrna Gene ◽  
Cultivation Period ◽  
Rrna Gene Sequencing ◽  
Bacteria And Fungi ◽  
Surrounding Seawater

Abstract Microorganisms play important roles in the growth and development of macroalgae. Still, the biodiversity of the epiphytic microbial community associated with the economically important red alga Pyropia haitanensis during the cultivation period remains uncharacterized, especially the effects of P. haitanensis cultivation on the microbial community of surrounding seawater. Here, we isolated epiphytic microbes from P. haitanensis during the thallus stage during oceanic cultivation and the conchocelis stage during industrial cultivation. The dynamic diversity patterns, as determined by 16S and 18S rRNA gene sequencing of the bacterial and fungal communities, respectively, associated with P. haitanensis and seawater in the presence and absence of algal cultivation were investigated. A notable distinction was observed between the microbial communities of seawater with and without P. haitanensis cultivation. Additionally, the alpha-diversity of seawater with P. haitanensis cultivation was significantly greater than without P. haitanensis cultivation. Cyanobacteria were the dominant species in the latter, while Rhodobacteraceae was enriched in the former. Furthermore, there were significant differences in the microbial community of P. haitanensis at the thallus and conchocelis stages. Seawater properties had significant direct effects on the microbial diversity of P. haitanensis and cultivation seawater, but not on non-cultivation seawater. The enriched microbial presence might promote thallus morphogenesis and be beneficial for the growth and development of both the thallus and conchocelis stages. These findings expand our knowledge of the bacteria and fungi that are beneficial for Pyropia nursery seeding and cultivation, as well as the effects of P. haitanensis cultivation on the seawater microbial community.

scholarly journals Surveying the sweetpotato rhizosphere, endophyte, and surrounding soil microbiomes at two North Carolina farms reveals underpinnings of sweetpotato microbiome community assembly

2019 ◽  
Author(s):  
C Pepe-Ranney ◽  
C Keyser ◽  
J Trimble ◽  
B Bissinger
Keyword(s):  
North Carolina ◽  
Community Assembly ◽  
Alpha Diversity ◽  
Rrna Gene ◽  
Sequence Variant ◽  
Skin Microbiome ◽  
Microbiome Composition ◽  
Pests And Diseases ◽  
Sub Saharan ◽  
Surrounding Soil

AbstractFarmers grow sweetpotatoes worldwide and some sub-Saharan African and Asian diets include sweetpotato as a staple, yet the sweetpotato microbiome is conspicuously less studied relative to crops such as maize, soybean, and wheat. Studying sweetpotato microbiome ecology may reveal paths to engineer the microbiome to improve sweetpotato yield, and/or combat sweetpotato pests and diseases. We sampled sweetpotatoes and surrounding soil from two North Carolina farms. We took samples from sweetpotato fields under two different land management regimes, conventional and organic, and collected two sweetpotato cultivars, ‘Beauregard’ and ‘Covington’. By comparing SSU rRNA gene amplicon sequence profiles from sweetpotato storage root skin, rhizosphere, and surrounding soil we found the skin microbiome possessed the least composition heterogeneity among samples and lowest alpha-diversity and was significantly nested by the rhizosphere in amplicon sequence variant (ASV) membership. Many ASVs were specific to a single field and/or only found in either the skin, rhizosphere, or surrounding soil. Notably, sweetpotato skin enriched for Planctomycetaceae in relative abundance at both farms. This study elucidates underpinnings of sweetpotato microbiome community assembly, quantifies microbiome composition variance within a single farm, and reveals microorganisms associated with sweetpotato skin that belong to common but uncultured soil phylotypes.

scholarly journals Responses of microbial community from tropical pristine coastal soil to crude oil contamination

2015 ◽  
Author(s):  
Daniel Morais ◽  
Victor Pylro ◽  
Ian M Clark ◽  
Penny R Hirsch ◽  
Marcos Tótola
Keyword(s):  
Microbial Community ◽  
Crude Oil ◽  
Alpha Diversity ◽  
Illumina Miseq ◽  
Oil Reservoirs ◽  
Rrna Gene ◽  
Community Shift ◽  
Campos Basin ◽  
Trindade Island ◽  
Bacteria And Fungi

Crude oil is still the dominant energy source in Brazil and that the consumption keeps rising since 2013, being responsible for 2.2% of the world’s energy consumption. The recent discovery of crude oil reservoirs at the Espirito Santo basin, Campos basin and Santos basin, can be considered as an excellent opportunity to supply the country’s economic and energetic demands. However, albeit the opportunity these crude oil reservoirs represent, offshore exploration offers risks to the microbiota and the whole sea life, as petroleum hydrocarbons are toxic, mutagenic, teratogenic and carcinogenic. Microbes are responsible for nutrient cycling and can degrade even very recalcitrant hydrocarbons. This work aimed to evaluate the microbial community shift (Archaea, Bacteria and Fungi) from Trindade Island coastal environment under petroleum contamination. Microcosms were assembled using Trindade Island coastal soil to create two treatments, control and contaminated (weathered crude oil at 30 g kg-1). Soils were incubated during 38 days with CO2 measurements every four hours. Total DNA was extracted, purified and submited for sequencing of 16s rRNA gene, for Bacteria and Archaea domains and Fungal ITS1 region using Illumina MiSeq platform. We compared alpha diversity, beta diversity and taxonomic shifts between controls and contaminated samples. Three days after contamination, emission rate peaked at more than 20x the control and the emissions remained higer during the whole incubation period. Microbial alpha diversity was reduced for contaminated-samples. Fungi community of contaminated samples was reduced to almost 40% of the observed species. Taxonomy comparisons showed rise of the Actinobacteria phylum and reduction of the Archaea Candidatus nitrosphaere.

scholarly journals Characterization of the mycobiome of the seagrass, Zostera marina, reveals putative associations with marine chytrids

2019 ◽  
Author(s):  
Cassandra L. Ettinger ◽  
Jonathan A. Eisen
Keyword(s):  
Zostera Marina ◽  
Marine Fungi ◽  
Its Region ◽  
Taxonomic Diversity ◽  
Its2 Region ◽  
Rrna Gene ◽  
Sequence Variant ◽  
28S Rrna ◽  
Taxonomic Assignment ◽  
28S Rrna Gene

AbstractSeagrasses are globally distributed marine flowering plants that are foundation species in coastal ecosystems. Seagrass beds play essential roles as habitats and hatcheries, in nutrient cycling and in protecting the coastline from erosion. Although many studies have focused on seagrass ecology, only a limited number have investigated their associated fungi. In terrestrial systems, fungi can have beneficial and detrimental effects on plant fitness. However, not much is known about marine fungi and even less is known about seagrass associated fungi. Here we used culture-independent sequencing of the ribosomal internal transcribed spacer (ITS) region to characterize the taxonomic diversity of fungi associated with the seagrass, Zostera marina. We sampled from two Z. marina beds in Bodega Bay over three time points to investigate fungal diversity within and between plants. Our results indicate that there are many fungal taxa for which a taxonomic assignment cannot be made living on and inside Z. marina leaves, roots and rhizomes and that these plant tissues harbor distinct fungal communities. The most prevalent ITS amplicon sequence variant (ASV) associated with Z. marina leaves was classified as fungal, but could not initially be assigned to a fungal phylum. We then used PCR with a primer targeting unique regions of the ITS2 region of this ASV and an existing primer for the fungal 28S rRNA gene to amplify part of the 28S rRNA gene region and link it to this ASV. Sequencing and phylogenetic analysis of the resulting partial 28S rRNA gene revealed that the organism that this ASV comes from is a member of Novel Clade SW-I in the order Lobulomycetales in the phylum Chytridiomycota. This clade includes known parasites of freshwater diatoms and algae and it is possible this chytrid is directly infecting Z. marina leaf tissues. This work highlights a need for further studies focusing on marine fungi and the potential importance of these understudied communities to the larger seagrass ecosystem.

scholarly journals Microbiome evaluation revealed salivary dysbiosis in addicts of betel nut preparations

2020 ◽  
Author(s):  
Faizan Saleem ◽  
Ghulam Mujtaba ◽  
Junaid Ahmed Kori ◽  
Arshad Hassan ◽  
M. Kamran Azim
Keyword(s):  
Hypervariable Region ◽  
Alpha Diversity ◽  
Oral Microbiome ◽  
P Value ◽  
Rrna Gene ◽  
Betel Nut ◽  
Taxonomic Assignment ◽  
Unifrac Distance ◽  
Genus Level ◽  
Slaked Lime

AbstractBetel nut addiction is recognized as the causative agent of oral microbiome dysbiosis and other systematic disorders. A number of betel nut preparations containing ingredients such as slaked lime, catechu extract and tobacco are being commonly used particularly in South Asia. The underlying variations in the oral microbiome due to usage of betel nut preparations are poorly understood. We evaluated salivary microbiome in response to chewing of betel nut preparation(s). In order to assess the microbiome dynamics, metagenomic analysis of 16S rRNA gene (V3-V4 hypervariable region) from salivary bacteria in chewers of betel nut preparation (n = 16) and non-chewers (n = 55) was carried out by Greengenes and SILVA ribosomal sequence databases. It was observed that Gutka chewers demonstrated lower alpha diversity and number of bacterial genera than the non-chewers. Taxonomic assignment on phylum level revealed Firmicutes (p-value = 0.042 at 95% confidence interval) to be significantly more abundant in Gutka chewers in comparison with non-chewers. Beta diversity analysis at genus level by weighted unifrac distance matrices unveiled both groups to be divergent from each other. On the genus level, Veillonella (p-value = 0.015), Streptococcus (p-value = 0.026), Leptotrichia (p-value = 0.022) and Serratia (p-value = 0.022) species appeared to be significantly more abundant in Gutka chewers in comparison to non-chewers. The present study suggests salivary dysbiosis in response to gutka chewing and concludes that gutka chewers possess higher abundance of acidogenic and aciduric bacteria. This study contributes additional information regarding oral microbiome variations with response to gutka consumption.

scholarly journals Molecular phylogeny of heritable symbionts and microbiota diversity analysis in phlebotominae sand flies and Culex nigripalpus from Colombia

2021 ◽  
Vol 15 (12) ◽  
pp. e0009942
Author(s):  
Rafael J. Vivero-Gomez ◽  
Víctor A. Castañeda-Monsalve ◽  
María Claudia Atencia ◽  
Richard Hoyos-Lopez ◽  
Gregory D. Hurst ◽  
...  
Keyword(s):  
Molecular Phylogeny ◽  
Natural Infection ◽  
Alpha Diversity ◽  
Illumina Miseq ◽  
P Value ◽  
Rrna Gene ◽  
Sand Flies ◽  
Culex Nigripalpus ◽  
Bacteria And Fungi ◽  
Stress Susceptibility

Background Secondary symbionts of insects include a range of bacteria and fungi that perform various functional roles on their hosts, such as fitness, tolerance to heat stress, susceptibility to insecticides, reproduction. These endosymbionts could have the potential to shape microbial communites and high potential to develop strategies for mosquito-borne disease control. Methodology/Principal findings The relative frequency and molecular phylogeny of Wolbachia, Microsporidia and Cardinium were determined of phlebotomine sand flies and mosquitoes in two regions from Colombia. Illumina Miseq using the 16S rRNA gene as a biomarker was conducted to examine the microbiota. Different percentages of natural infection by Wolbachia, Cardinium, and Microsporidia in phlebotomines and mosquitoes were detected. Phylogenetic analysis of Wolbachia shows putative new strains of Lutzomyia gomezi (wLgom), Brumptomyia hamata (wBrham), and a putative new group associated with Culex nigripalpus (Cnig) from the Andean region, located in Supergroup A and Supergroup B, respectively. The sequences of Microsporidia were obtained of Pi. pia and Cx. nigripalpus, which are located on phylogeny in the IV clade (terrestrial origin). The Cardinium of Tr. triramula and Ps. shannoni were located in group C next to Culicoides sequences while Cardinium of Mi. cayennensis formed two putative new subgroups of Cardinium in group A. In total were obtained 550 bacterial amplicon sequence variants (ASVs) and 189 taxa to the genus level. The microbiota profiles of Sand flies and mosquitoes showed mainly at the phylum level to Proteobacteria (67.6%), Firmicutes (17.9%) and Actinobacteria (7.4%). High percentages of relative abundance for Wolbachia (30%-83%) in Lu. gomezi, Ev. dubitans, Mi. micropyga, Br. hamata, and Cx. nigripalpus were found. ASVs assigned as Microsporidia were found in greater abundance in Pi. pia (23%) and Cx. nigripalpus (11%). An important finding is the detection of Rickettsia in Pi. pia (58,8%) and Bartonella sp. in Cx. nigripalpus. Conclusions/Significance We found that Wolbachia infection significantly decreased the alpha diversity and negatively impacts the number of taxa on sand flies and Culex nigripalpus. The Principal Coordinate Analysis (PCoA) is consistent, which showed statistically significant differences (PERMANOVA, F = 2.4744; R2 = 0.18363; p-value = 0.007) between the microbiota of sand flies and mosquitoes depending on its origin, host and possibly for the abundance of some endosymbionts (Wolbachia, Rickettsia).

scholarly journals Surveying the Sweetpotato Rhizosphere, Endophyte, and Surrounding Soil Microbiomes at Two North Carolina Farms Reveals Underpinnings of Sweetpotato Microbiome Community Assembly

2020 ◽  
Vol 4 (1) ◽  
pp. 75-89
Author(s):  
C. Pepe-Ranney ◽  
C. Keyser ◽  
J. K. Trimble ◽  
B. Bissinger
Keyword(s):  
North Carolina ◽  
Community Assembly ◽  
Alpha Diversity ◽  
Small Subunit ◽  
Rrna Gene ◽  
Sequence Variant ◽  
Skin Microbiome ◽  
Small Subunit Rrna ◽  
Pests And Diseases ◽  
Surrounding Soil

Farmers grow sweetpotatoes worldwide and some sub-Saharan African and Asian diets include sweetpotato as a staple, yet the sweetpotato microbiome is conspicuously less studied relative to crops such as maize, soybean, and wheat. Studying sweetpotato microbiome ecology may reveal paths to engineer the microbiome to improve sweetpotato yield, and/or combat sweetpotato pests and diseases. We sampled sweetpotatoes and surrounding soil from two North Carolina farms. We took samples from sweetpotato fields under two different land management regimes, conventional and organic, and collected two sweetpotato cultivars, ‘Beauregard’ and ‘Covington’. By comparing small subunit rRNA gene amplicon sequence profiles from sweetpotato storage root skin, rhizosphere, and surrounding soil, we found the skin microbiome possessed the least composition heterogeneity among samples, lowest alpha-diversity, and was significantly nested by the rhizosphere in amplicon sequence variant (ASV) membership. Many ASVs were specific to a single field and/or only found in either the skin, rhizosphere, or surrounding soil. Notably, sweetpotato skin enriched for Planctomycetaceae in relative abundance at both farms. This study elucidates underpinnings of sweetpotato microbiome community assembly, quantifies microbiome composition variance within a single farm, and reveals microorganisms associated with sweetpotato skin that belong to common but uncultured soil phylotypes. [Formula: see text] Copyright © 2020 The Author(s). This is an open access article distributed under the CC BY 4.0 International license .

scholarly journals Genomes from Bacteria Associated with the Canine Oral Cavity: a Test Case for Automated Genome-Based Taxonomic Assignment

2019 ◽  
Author(s):  
David A. Coil ◽  
Guillaume Jospin ◽  
Aaron E. Darling ◽  
Corrin Wallis ◽  
Ian J. Davis ◽  
...  
Keyword(s):  
16S Rrna ◽  
Oral Cavity ◽  
16S Rrna Gene ◽  
Marker Gene ◽  
Pcr Amplification ◽  
Rrna Gene ◽  
Test Case ◽  
Bacterial Genomes ◽  
Taxonomic Assignment ◽  
Taxonomic Assignments

AbstractTaxonomy for bacterial isolates is commonly assigned via sequence analysis. However, the most common sequence-based approaches (e.g. 16S rRNA gene-based phylogeny or whole genome comparisons) are still labor intensive and subjective to varying degrees. Here we present a set of 33 bacterial genomes, isolated from the canine oral cavity. Taxonomy of these isolates was first assigned by PCR amplification of the 16S rRNA gene, Sanger sequencing, and taxonomy assignment using BLAST. After genome sequencing, taxonomy was revisited through a manual process using a combination of average nucleotide identity (ANI), concatenated marker gene phylogenies, and 16S rRNA gene phylogenies. This taxonomy was then compared to the automated taxonomic assignment given by the recently proposed Genome Taxonomy Database (GTDB). We found the results of all three methods to be similar (25 out of the 33 had matching genera), but the GTDB approach was less subjective, and required far less labor. The primary differences in the remaining taxonomic assignments related to proposed taxonomy changes by the GTDB team.

scholarly journals Gut microbiome composition in the Hispanic Community Health Study/Study of Latinos is shaped by geographic relocation, environmental factors, and obesity

2019 ◽  
Vol 20 (1) ◽  
Author(s):  
Robert C. Kaplan ◽  
Zheng Wang ◽  
Mykhaylo Usyk ◽  
Daniela Sotres-Alvarez ◽  
Martha L. Daviglus ◽  
...  
Keyword(s):  
Community Health ◽  
Gut Microbiome ◽  
Alpha Diversity ◽  
Amplicon Sequencing ◽  
Shannon Index ◽  
Health Study ◽  
Rrna Gene ◽  
Cross Sectional ◽  
Hispanic Community ◽  
The Usa

Abstract Background Hispanics living in the USA may have unrecognized potential birthplace and lifestyle influences on the gut microbiome. We report a cross-sectional analysis of 1674 participants from four centers of the Hispanic Community Health Study/Study of Latinos (HCHS/SOL), aged 18 to 74 years old at recruitment. Results Amplicon sequencing of 16S rRNA gene V4 and fungal ITS1 fragments from self-collected stool samples indicate that the host microbiome is determined by sociodemographic and migration-related variables. Those who relocate from Latin America to the USA at an early age have reductions in Prevotella to Bacteroides ratios that persist across the life course. Shannon index of alpha diversity in fungi and bacteria is low in those who relocate to the USA in early life. In contrast, those who relocate to the USA during adulthood, over 45 years old, have high bacterial and fungal diversity and high Prevotella to Bacteroides ratios, compared to USA-born and childhood arrivals. Low bacterial diversity is associated in turn with obesity. Contrasting with prior studies, our study of the Latino population shows increasing Prevotella to Bacteroides ratio with greater obesity. Taxa within Acidaminococcus, Megasphaera, Ruminococcaceae, Coriobacteriaceae, Clostridiales, Christensenellaceae, YS2 (Cyanobacteria), and Victivallaceae are significantly associated with both obesity and earlier exposure to the USA, while Oscillospira and Anaerotruncus show paradoxical associations with both obesity and late-life introduction to the USA. Conclusions Our analysis of the gut microbiome of Latinos demonstrates unique features that might be responsible for health disparities affecting Hispanics living in the USA.

scholarly journals Comparative analysis of bacterioplankton assemblages from two subtropical karst reservoirs of southwestern China with contrasting trophic status

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Qiang Li ◽  
Yadan Huang ◽  
Shenglin Xin ◽  
Zhongyi Li
Keyword(s):  
Trophic Status ◽  
Alpha Diversity ◽  
Decisive Role ◽  
Sensu Stricto ◽  
Karst Area ◽  
Southwestern China ◽  
Rrna Gene ◽  
Karst Water ◽  
Bacterioplankton Community ◽  
Generation Sequencing

AbstractAlthough bacterioplankton play an important role in aquatic ecosystems, less is known about bacterioplankton assemblages from subtropical karst reservoirs of southwestern China with contrasting trophic status. Here, 16S rRNA gene next-generation sequencing coupled with water chemistry analysis was applied to compare the bacterioplankton communities from a light eutrophic reservoir, DL Reservoir, and a mesotrophic reservoir, WL Reservoir, in subtropical karst area of southwestern China. Our findings indicated that Proteobacteria, Firmicutes, Actinobacteria, Bacteroidetes, Cyanobacteria and Verrucomicrobia dominated bacterioplankton community with contrasting relative frequency in the two subtropical karst reservoirs. Proteobacteria and Bacteroidetes were the core communities, which played important roles in karst biogeochemical cycles. Though WT, TN and DOC play the decisive role in assembling karst aquatic bacterioplankton, trophic status exerted significantly negative direct effects on bacterioplankton community composition and alpha diversity. Due to contrasting trophic status in the two reservoirs, the dominant taxa such as Enterobacter, Clostridium sensu stricto, Candidatus Methylacidiphilum and Flavobacteriia, that harbor potential functions as valuable and natural indicators of karst water health status, differed in DL Reservoir and WL Reservoir.

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