scholarly journals Airborne Bacterial and Eukaryotic Community Structure across the United Kingdom Revealed by High-Throughput Sequencing

Atmosphere ◽  
2020 ◽  
Vol 11 (8) ◽  
pp. 802
Author(s):  
Hokyung Song ◽  
Ian Crawford ◽  
Jonathan Lloyd ◽  
Clare Robinson ◽  
Christopher Boothman ◽  
...  
Keyword(s):  
16S Rrna ◽  
High Throughput ◽  
18S Rrna ◽  
High Throughput Sequencing ◽  
Gene Sequencing ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Biological Aerosols ◽  
Rrna Gene Sequencing

Primary biological aerosols often include allergenic and pathogenic microorganisms posing potential risks to human health. Moreover, there are airborne plant and animal pathogens that may have ecological and economic impact. In this study, we used high-throughput sequencing techniques (Illumina, MiSeq) targeting the 16S rRNA genes of bacteria and the 18S rRNA genes of eukaryotes, to characterize airborne primary biological aerosols. We used a filtration system on the UK Facility for Airborne Atmospheric Measurements (FAAM) research aircraft to sample a range of primary biological aerosols across southern England overflying surface measurement sites from Chilbolton to Weybourne. We identified 30 to 60 bacterial operational taxonomic units (OTUs) and 108 to 224 eukaryotic OTUs per sample. Moreover, 16S rRNA gene sequencing identified significant numbers of genera that have not been found in atmospheric samples previously or only been described in limited number of atmospheric field studies, which are rather old or published in local journals. This includes the genera Gordonia, Lautropia, and Psychroglaciecola. Some of the bacterial genera found in this study include potential human pathogens, for example, Gordonia, Sphingomonas, Chryseobacterium, Morganella, Fusobacterium, and Streptococcus. 18S rRNA gene sequencing showed Cladosporium to be the major genus in all of the samples, which is a well-known allergen and often found in the atmosphere. There were also genetic signatures of potentially allergenic taxa; for example, Pleosporales, Phoma, and Brassicales. Although there was no significant clustering of bacterial and eukaryotic communities depending on the sampling location, we found meteorological factors explaining significant variations in the community composition. The findings in this study support the application of DNA-based sequencing technologies for atmospheric science studies in combination with complementary spectroscopic and microscopic techniques for improved identification of primary biological aerosols.

scholarly journals REP-PCR Analysis to Study Prokaryotic Biodiversity from Lake Meyghan

2017 ◽  
Vol 61 ◽  
pp. 69-84 ◽  
Author(s):  
Ali Naghoni ◽  
Giti Emtiazi ◽  
Mohammad Ali Amoozegar ◽  
Zahra Etemadifar ◽  
Seyed Abolhassan Shahzadeh Fazeli
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Gene Sequencing ◽  
16S Rrna Gene Sequencing ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Pcr Analysis ◽  
Rrna Gene ◽  
White Region ◽  
Rrna Gene Sequencing

Repetitive extragenic palindromic elements-polymerase chain reaction (rep-PCR) with 16S ribosomal ribonucleic acid (16S rRNA) genes sequences successfully used for the analysis of microbial community. In this study, the prokaryotic community in Lake Meyghan described by using rep-PCR analysis along with 16S rRNA gene sequencing. The water samples were collected from Lake Meyghan in November 2013. All samples were diluted and cultured on three different media. To estimate the number of prokaryotes per milliliter of the lake we used quantitative real‑time PCR (qPCR). Rep-PCR combination with 16S rRNA gene sequencing was performed to investigate prokaryotes biodiversity in the lake. 305 strains were isolated in this work; 113 isolates for green region, 102 isolates for red region, and 90 isolates for white region. The dendrograms generated 10, 7, and 9 clusters for a 70 % similarity cut-off for green, red, and white regions, respectively. Based on rep-PCR and 16S rRNA gene sequencing, the recovered isolates were dominated by (77.5 %)Halobacteriacaeand many isolates were related to the generaHalorubrum,Haloarcula,Haloterrigena,Natrinema, andHalovivaxin the white region. In the red region more isolated strains (57.5 %) belonged toBacillaceaeand the remaining 42.5 % of isolates belonged to archaea domain,Halorubrum, andHaloarcula. In the green region members ofGammaproteobacteriawere recoverd, this region was dominant withPseudoalteromonas,Salinivibrio, andAliidiomarina.

scholarly journals High-Throughput Sequencing for Examining Salmonella Prevalence and Pathogen—Microbiota Relationships in Barn Swallows

2021 ◽  
Vol 9 ◽  
Author(s):  
Olivia N. Choi ◽  
Ammon Corl ◽  
Andrew Wolfenden ◽  
Avishai Lublin ◽  
Suzanne L. Ishaq ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
High Throughput ◽  
High Throughput Sequencing ◽  
Gene Sequencing ◽  
16S Rrna Gene Sequencing ◽  
Rrna Gene ◽  
Sequencing Data ◽  
Barn Swallows ◽  
Rrna Gene Sequencing

Studies in both humans and model organisms suggest that the microbiome may play a significant role in host health, including digestion and immune function. Microbiota can offer protection from exogenous pathogens through colonization resistance, but microbial dysbiosis in the gastrointestinal tract can decrease resistance and is associated with pathogenesis. Little is known about the effects of potential pathogens, such as Salmonella, on the microbiome in wildlife, which are known to play an important role in disease transmission to humans. Culturing techniques have traditionally been used to detect pathogens, but recent studies have utilized high throughput sequencing of the 16S rRNA gene to characterize host-associated microbial communities (i.e., the microbiome) and to detect specific bacteria. Building upon this work, we evaluated the utility of high throughput 16S rRNA gene sequencing for potential bacterial pathogen detection in barn swallows (Hirundo rustica) and used these data to explore relationships between potential pathogens and microbiota. To accomplish this, we first compared the detection of Salmonella spp. in swallows using 16S rRNA data with standard culture techniques. Second, we examined the prevalence of Salmonella using 16S rRNA data and examined the relationship between Salmonella-presence or -absence and individual host factors. Lastly, we evaluated host-associated bacterial diversity and community composition in Salmonella-present vs. -absent birds. Out of 108 samples, we detected Salmonella in six (5.6%) samples based on culture, 25 (23.1%) samples with unrarefied 16S rRNA gene sequencing data, and three (2.8%) samples with both techniques. We found that sex, migratory status, and weight were correlated with Salmonella presence in swallows. In addition, bacterial community composition and diversity differed between birds based on Salmonella status. This study highlights the value of 16S rRNA gene sequencing data for monitoring pathogens in wild birds and investigating the ecology of host microbe-pathogen relationships, data which are important for prediction and mitigation of disease spillover into domestic animals and humans.

scholarly journals A 16S rRNA gene sequencing and analysis protocol for the Illumina MiniSeq platform

2017 ◽  
Author(s):  
Monica Pichler ◽  
Ömer K. Coskun ◽  
Ana Sofia Ortega ◽  
Nicola Conci ◽  
Gert Wörheide ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
High Throughput ◽  
Environmental Samples ◽  
High Throughput Sequencing ◽  
Gene Sequencing ◽  
16S Rrna Gene Sequencing ◽  
Rrna Gene ◽  
Mock Community ◽  
Rrna Gene Sequencing

ABSTRACTHigh-throughput sequencing of the 16S rRNA gene is widely used in microbial ecology, with Illumina platforms being widely used in recent studies. The MiniSeq, Illumina’s latest benchtop sequencer, enables more cost-efficient DNA sequencing relative to larger sequencing platforms (e.g. MiSeq). Here we used a modified custom primer sequencing approach to test the fidelity of the MiniSeq for high-throughput sequencing of the V4 hypervariable region of 16S rRNA genes from complex communities in environmental samples. To this end, we designed an additional sequencing primer that enabled application of a dual-index barcoding method on the MiniSeq. A mock community was sequenced alongside the environmental samples as a quality control benchmark. After careful filtering procedures, we were able to recapture a realistic richness of the mock community, and identify meaningful differences in alpha and beta diversity in the environmental samples. These results show that the MiniSeq can produce similar quantities of high quality V4 reads compared to the MiSeq, yet is a cost-effective option for any laboratory interested in performing high-throughput 16S rRNA gene sequencing.IMPORTANCEWe modified a custom sequencing approach and used a mock community to test the fidelity of high-throughput sequencing on the Illumina MiniSeq platform. Our results show that the MiniSeq can produce similar quantities of high quality V4 reads compared to the MiSeq. In addition, our protocol increases feasibility for small laboratories to perform their own high-throughput sequencing of the 16S rRNA marker gene.

scholarly journals A method for high precision sequencing of near full-length 16S rRNA genes on an Illumina MiSeq

PeerJ ◽  
2016 ◽  
Vol 4 ◽  
pp. e2492 ◽  
Author(s):  
Catherine M. Burke ◽  
Aaron E. Darling
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
High Throughput ◽  
Single Molecule ◽  
Illumina Miseq ◽  
Full Length ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Bacterial Taxonomy

BackgroundThe bacterial 16S rRNA gene has historically been used in defining bacterial taxonomy and phylogeny. However, there are currently no high-throughput methods to sequence full-length 16S rRNA genes present in a sample with precision.ResultsWe describe a method for sequencing near full-length 16S rRNA gene amplicons using the high throughput Illumina MiSeq platform and test it using DNA from human skin swab samples. Proof of principle of the approach is demonstrated, with the generation of 1,604 sequences greater than 1,300 nt from a single Nano MiSeq run, with accuracy estimated to be 100-fold higher than standard Illumina reads. The reads were chimera filtered using information from a single molecule dual tagging scheme that boosts the signal available for chimera detection.ConclusionsThis method could be scaled up to generate many thousands of sequences per MiSeq run and could be applied to other sequencing platforms. This has great potential for populating databases with high quality, near full-length 16S rRNA gene sequences from under-represented taxa and environments and facilitates analyses of microbial communities at higher resolution.

scholarly journals Status of the Archaeal and Bacterial Census: an Update

mBio ◽  
2016 ◽  
Vol 7 (3) ◽  
Author(s):  
Patrick D. Schloss ◽  
Rene A. Girard ◽  
Thomas Martin ◽  
Joshua Edwards ◽  
J. Cameron Thrash
Keyword(s):  
16S Rrna ◽  
Bacterial Diversity ◽  
High Throughput Sequencing ◽  
New Technologies ◽  
Full Length ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Short Read ◽  
Sequencing Technologies

ABSTRACT A census is typically carried out for people across a range of geographical levels; however, microbial ecologists have implemented a molecular census of bacteria and archaea by sequencing their 16S rRNA genes. We assessed how well the census of full-length 16S rRNA gene sequences is proceeding in the context of recent advances in high-throughput sequencing technologies because full-length sequences are typically used as references for classification of the short sequences generated by newer technologies. Among the 1,411,234 and 53,546 full-length bacterial and archaeal sequences, 94.5% and 95.1% of the bacterial and archaeal sequences, respectively, belonged to operational taxonomic units (OTUs) that have been observed more than once. Although these metrics suggest that the census is approaching completion, 29.2% of the bacterial and 38.5% of the archaeal OTUs have been observed more than once. Thus, there is still considerable diversity to be explored. Unfortunately, the rate of new full-length sequences has been declining, and new sequences are primarily being deposited by a small number of studies. Furthermore, sequences from soil and aquatic environments, which are known to be rich in bacterial diversity, represent only 7.8 and 16.5% of the census, while sequences associated with host-associated environments represent 55.0% of the census. Continued use of traditional approaches and new technologies such as single-cell genomics and short-read assembly are likely to improve our ability to sample rare OTUs if it is possible to overcome this sampling bias. The success of ongoing efforts to use short-read sequencing to characterize archaeal and bacterial communities requires that researchers strive to expand the depth and breadth of this census. IMPORTANCE The biodiversity contained within the bacterial and archaeal domains dwarfs that of the eukaryotes, and the services these organisms provide to the biosphere are critical. Surprisingly, we have done a relatively poor job of formally tracking the quality of the biodiversity as represented in full-length 16S rRNA genes. By understanding how this census is proceeding, it is possible to suggest the best allocation of resources for advancing the census. We found that the ongoing effort has done an excellent job of sampling the most abundant organisms but struggles to sample the rarer organisms. Through the use of new sequencing technologies, we should be able to obtain full-length sequences from these rare organisms. Furthermore, we suggest that by allocating more resources to sampling environments known to have the greatest biodiversity, we will be able to make significant advances in our characterization of archaeal and bacterial diversity.

scholarly journals Distribution of Hydrogen-Producing Bacteria in Tibetan Hot Springs, China

2021 ◽  
Vol 12 ◽  
Author(s):  
Li Ma ◽  
Geng Wu ◽  
Jian Yang ◽  
Liuqin Huang ◽  
Dorji Phurbu ◽  
...  
Keyword(s):  
Community Structure ◽  
Hydrogen Production ◽  
16S Rrna ◽  
High Throughput Sequencing ◽  
Hot Springs ◽  
Illumina Miseq ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
The Tibetan Plateau

Investigating the distribution of hydrogen-producing bacteria (HPB) is of great significance to understanding the source of biological hydrogen production in geothermal environments. Here, we explored the compositions of HPB populations in the sediments of hot springs from the Daggyai, Quzhuomu, Quseyongba, and Moluojiang geothermal zones on the Tibetan Plateau, with the use of Illumina MiSeq high-throughput sequencing of 16S rRNA genes and hydA genes. In the present study, the hydA genes were successfully amplified from the hot springs with a temperature of 46–87°C. The hydA gene phylogenetic analysis showed that the top three phyla of the HPB populations were Bacteroidetes (14.48%), Spirochaetes (14.12%), and Thermotogae (10.45%), while Proteobacteria were absent in the top 10 of the HPB populations, although Proteobacteria were dominant in the 16S rRNA gene sequences. Canonical correspondence analysis results indicate that the HPB community structure in the studied Tibetan hot springs was correlated with various environmental factors, such as temperature, pH, and elevation. The HPB community structure also showed a spatial distribution pattern; samples from the same area showed similar community structures. Furthermore, one HPB isolate affiliated with Firmicutes was obtained and demonstrated the capacity of hydrogen production. These results are important for us to understand the distribution and function of HPB in hot springs.

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