Exploring the Individual Bacterial Microbiota of Questing Ixodes ricinus Nymphs

Ixodes ricinus is the most common hard tick species in Europe and an important vector of pathogens of human and animal health concerns. The rise of high-throughput sequencing has facilitated the identification of many tick-borne pathogens and, more globally, of various microbiota members depending on the scale of concern. In this study, we aimed to assess the bacterial diversity of individual I. ricinus questing nymphs collected in France using high-throughput 16S gene metabarcoding. From 180 dragging-collected nymphs, we identified more than 700 bacterial genera, of which about 20 are abundantly represented (>1% of total reads). Together with 136 other genera assigned, they constitute a core internal microbiota in this study. We also identified 20 individuals carrying Borreliella. The most abundant species is B. afzelii, known to be one of the bacteria responsible for Lyme disease in Europe. Co-detection of up to four Borreliella genospecies within the same individual has also been retrieved. The detection and co-detection rate of Borreliella in I. ricinus nymphs is high and raises the question of interactions between these bacteria and the communities constituting the internal microbiota.

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Comparison of sequencing data processing pipelines and application to underrepresented African human populations

BMC Bioinformatics ◽

10.1186/s12859-021-04407-x ◽

2021 ◽

Vol 22 (1) ◽

Author(s):

Gwenna Breton ◽

Anna C. V. Johansson ◽

Per Sjödin ◽

Carina M. Schlebusch ◽

Mattias Jakobsson

Keyword(s):

Best Practices ◽

High Throughput ◽

High Throughput Sequencing ◽

Variant Calling ◽

Human Populations ◽

Sequencing Data ◽

High Coverage ◽

Individual Level ◽

Bioinformatic Tools ◽

The Individual

Abstract Background Population genetic studies of humans make increasing use of high-throughput sequencing in order to capture diversity in an unbiased way. There is an abundance of sequencing technologies, bioinformatic tools and the available genomes are increasing in number. Studies have evaluated and compared some of these technologies and tools, such as the Genome Analysis Toolkit (GATK) and its “Best Practices” bioinformatic pipelines. However, studies often focus on a few genomes of Eurasian origin in order to detect technical issues. We instead surveyed the use of the GATK tools and established a pipeline for processing high coverage full genomes from a diverse set of populations, including Sub-Saharan African groups, in order to reveal challenges from human diversity and stratification. Results We surveyed 29 studies using high-throughput sequencing data, and compared their strategies for data pre-processing and variant calling. We found that processing of data is very variable across studies and that the GATK “Best Practices” are seldom followed strictly. We then compared three versions of a GATK pipeline, differing in the inclusion of an indel realignment step and with a modification of the base quality score recalibration step. We applied the pipelines on a diverse set of 28 individuals. We compared the pipelines in terms of count of called variants and overlap of the callsets. We found that the pipelines resulted in similar callsets, in particular after callset filtering. We also ran one of the pipelines on a larger dataset of 179 individuals. We noted that including more individuals at the joint genotyping step resulted in different counts of variants. At the individual level, we observed that the average genome coverage was correlated to the number of variants called. Conclusions We conclude that applying the GATK “Best Practices” pipeline, including their recommended reference datasets, to underrepresented populations does not lead to a decrease in the number of called variants compared to alternative pipelines. We recommend to aim for coverage of > 30X if identifying most variants is important, and to work with large sample sizes at the variant calling stage, also for underrepresented individuals and populations.

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Diversity Assessment of Toxic Cyanobacterial Blooms during Oxidation

Toxins ◽

10.3390/toxins12110728 ◽

2020 ◽

Vol 12 (11) ◽

pp. 728

Author(s):

Saber Moradinejad ◽

Hana Trigui ◽

Juan Francisco Guerra Maldonado ◽

Jesse Shapiro ◽

Yves Terrat ◽

...

Keyword(s):

High Throughput ◽

High Throughput Sequencing ◽

Treatment Plant ◽

Principal Component ◽

Abundant Species ◽

Chemical Oxidation ◽

Cyanobacterial Blooms ◽

Cyanobacterial Community ◽

Cell Counts ◽

Diversity Assessment

Fresh-water sources of drinking water are experiencing toxic cyanobacterial blooms more frequently. Chemical oxidation is a common approach to treat cyanobacteria and their toxins. This study systematically investigates the bacterial/cyanobacterial community following chemical oxidation (Cl2, KMnO4, O3, H2O2) using high throughput sequencing. Raw water results from high throughput sequencing show that Proteobacteria, Actinobacteria, Cyanobacteria and Bacteroidetes were the most abundant phyla. Dolichospermum, Synechococcus, Microcystis and Nostoc were the most dominant genera. In terms of species, Dolichospermum sp.90 and Microcystis aeruginosa were the most abundant species at the beginning and end of the sampling, respectively. A comparison between the results of high throughput sequencing and taxonomic cell counts highlighted the robustness of high throughput sequencing to thoroughly reveal a wide diversity of bacterial and cyanobacterial communities. Principal component analysis of the oxidation samples results showed a progressive shift in the composition of bacterial/cyanobacterial communities following soft-chlorination with increasing common exposure units (CTs) (0–3.8 mg·min/L). Close cyanobacterial community composition (Dolichospermum dominant genus) was observed following low chlorine and mid-KMnO4 (287.7 mg·min/L) exposure. Our results showed that some toxin producing species may persist after oxidation whether they were dominant species or not. Relative persistence of Dolichospermum sp.90 was observed following soft-chlorination (0.2–0.6 mg/L) and permanganate (5 mg/L) oxidation with increasing oxidant exposure. Pre-oxidation using H2O2 (10 mg/L and one day contact time) caused a clear decrease in the relative abundance of all the taxa and some species including the toxin producing taxa. These observations suggest selectivity of H2O2 to provide an efficient barrier against toxin producing cyanobacteria entering a water treatment plant.

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A simulation framework for correlated count data of features subsets in high-throughput sequencing or proteomics experiments

Statistical Applications in Genetics and Molecular Biology ◽

10.1515/sagmb-2015-0082 ◽

2016 ◽

Vol 15 (5) ◽

Cited By ~ 1

Author(s):

Jochen Kruppa ◽

Frank Kramer ◽

Tim Beißbarth ◽

Klaus Jung

Keyword(s):

High Throughput ◽

Protein Interactions ◽

Count Data ◽

Dna Microarrays ◽

High Throughput Sequencing ◽

Correlated Data ◽

Correlation Structure ◽

Shrinkage Estimators ◽

Distribution Parameters ◽

The Individual

AbstractAs part of the data processing of high-throughput-sequencing experiments count data are produced representing the amount of reads that map to specific genomic regions. Count data also arise in mass spectrometric experiments for the detection of protein-protein interactions. For evaluating new computational methods for the analysis of sequencing count data or spectral count data from proteomics experiments artificial count data is thus required. Although, some methods for the generation of artificial sequencing count data have been proposed, all of them simulate single sequencing runs, omitting thus the correlation structure between the individual genomic features, or they are limited to specific structures. We propose to draw correlated data from the multivariate normal distribution and round these continuous data in order to obtain discrete counts. In our approach, the required distribution parameters can either be constructed in different ways or estimated from real count data. Because rounding affects the correlation structure we evaluate the use of shrinkage estimators that have already been used in the context of artificial expression data from DNA microarrays. Our approach turned out to be useful for the simulation of counts for defined subsets of features such as individual pathways or GO categories.

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Spatiotemporal Dynamics of the Bacterial Microbiota on Lacustrine Cladophora Glomerata (Chlorophyta)

10.1101/096347 ◽

2016 ◽

Author(s):

Michael J. Braus ◽

Linda E. Graham ◽

Thea L. Whitman

Keyword(s):

High Throughput ◽

High Throughput Sequencing ◽

Open Water ◽

Spatial And Temporal Variation ◽

Cladophora Glomerata ◽

Growth Season ◽

Organic C ◽

Lake Mendota ◽

Bacterial Microbiota ◽

Sequencing Studies

ABSTRACTThe branched periphytic green alga Cladophora glomerata, often abundant in nearshore waters of lakes and rivers worldwide, plays important ecosystem roles, some mediated by epibiotic microbiota that benefit from host-provided surface, organic C, and O2. Previous microscopy and high throughput sequencing studies have indicated surprising epibiont taxonomic and functional diversity, but have not included adequate consideration of sample replication or the potential for spatial and temporal variation. Here we report the results of 16S rRNA amplicon-based phylum-to-genus taxonomic analysis of Cladophora-associated bacterial epibiota sampled in replicate from three microsites and at six times during the open-water season of 2014, from the same lake locale (Picnic Point, Lake Mendota, Dane Co., WI, USA) explored by high throughput sequencing studies in two previous years. Statistical methods were used to test null hypotheses that the bacterial community: 1) is homogeneous across microsites tested, and 2) does not change over the course of a growth season or among successive years. Results indicated a dynamic microbial community that is more strongly influenced by sampling day during the growth season than by microsite variation. A surprising diversity of bacterial genera known to be associated with the key function of methane-oxidation (methanotrophy)-including relatively high-abundance of Crenothrix, Methylomonas, and Methylocaldum–showed intra-seasonal and inter-annual variability possibly related to temperature differences, and microsite preferences possibly related to variation in methane abundance. By contrast, a core assemblage of bacterial genera seems to persist over a growth season and from year-to-year, possibly transmitted by a persistent attached host resting stage.

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Bacterial Microbiota Analysis Demonstrates That Tick Can Acquire Bacteria From Habitat and Host Blood Meal

10.21203/rs.3.rs-882619/v1 ◽

2021 ◽

Author(s):

Si-Si Li ◽

Xiao-Yu Zhang ◽

Xue-Jiao Zhou ◽

Kai-Li Chen ◽

Abolfazl Masoudi ◽

...

Keyword(s):

Life Cycle ◽

High Throughput Sequencing ◽

Developmental Stages ◽

Tick Control ◽

Hard Tick ◽

Haemaphysalis Longicornis ◽

Bacterial Microbiota ◽

Bacterial Richness ◽

Host Blood ◽

Blood Meals

Abstract Exploring the bacterial microbiota is imperative to tick control since it has an important role in tick physiology and vector capacity. The life cycle of ticks consists of parasitic and non-parasitic stages, with a diversity of habitats and host blood meals. Whether and how these factors, such as tick developmental stages, tick organs, habitats and host blood meals affect tick bacterial microbiota is poorly elucidated. In the present study, we investigated the bacterial microbiotas of hard tick Haemaphysalis longicornis, their blood meals and habitats using 16S rRNA high-throughput sequencing. The bacterial richness and diversity in ticks varied depending on the tick developmental stage, feeding status and the tick organs. Results showed that fed ticks present a higher bacterial richness suggesting that ticks may acquire bacteria from blood meals. The significant overlap of the bacteriota of fed ticks and the host blood also support this possibility. Another possibility is that blood meals can stimulate the proliferation of certain bacteria. However, most shared bacteria cannot transmit throughout the tick life cycle, as they were not present in tick eggs. The most shared bacteria between ticks and habitats are genus of Staphylococcus, Pseudomonus, Enterobacter, Acinetobacer and Stenotrophomonas, some of them are also present in tick organ, suggesting that these environmental bacteria cannot be completely washed away and can be acquired by ticks. As tick reproductive organ, ovary showed the lowest bacterial richness and diversity compared to other organs. The predominant proportion of Coxiella in fed females and ovary further demonstrated that this genus is required for H. longicornis reproduction system. These findings further reveal that the bacterial composition of ticks is influenced by a variety of factors and will help in subsequent studies of the function of these bacteria.

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Cpipe: a shared variant detection pipeline designed for diagnostic settings

10.1101/020388 ◽

2015 ◽

Author(s):

Simon Sadedin ◽

Harriet Dashnow ◽

Paul James ◽

Melanie Bahlo ◽

Denis C Bauer ◽

...

Keyword(s):

Open Source ◽

High Throughput ◽

Genetic Disease ◽

High Throughput Sequencing ◽

Clinical Settings ◽

Clinical Genetic ◽

Disease Diagnostics ◽

Variant Detection ◽

Reproducible Analysis ◽

The Individual

The benefits of implementing high throughput sequencing in the clinic are quickly becoming apparent. However, few freely available bioinformatics pipelines have been built from the ground up with clinical genomics in mind. Here we present Cpipe, a pipeline designed specifically for clinical genetic disease diagnostics. Cpipe was developed by the Melbourne Genomics Health Alliance, an Australian initiative to promote common approaches to genomics across healthcare institutions. As such, Cpipe has been designed to provide fast, effective and reproducible analysis, while also being highly flexible and customisable to meet the individual needs of diverse clinical settings. Cpipe is being shared with the clinical sequencing community as an open source project and is available at http://cpipeline.org.

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Extensive Profiling of a Complex Microbial Community by High-Throughput Sequencing

Applied and Environmental Microbiology ◽

10.1128/aem.68.6.3055-3066.2002 ◽

2002 ◽

Vol 68 (6) ◽

pp. 3055-3066 ◽

Cited By ~ 90

Author(s):

Janet E. Hill ◽

Robyn P. Seipp ◽

Martin Betts ◽

Lindsay Hawkins ◽

Andrew G. Van Kessel ◽

...

Keyword(s):

Microbial Community ◽

Microbial Communities ◽

High Throughput ◽

High Throughput Sequencing ◽

Animal Health ◽

Nucleotide Sequences ◽

Molecular Diagnostic ◽

Reference Database ◽

16S Rrna Sequence ◽

Microbiological Methods

ABSTRACT Complex microbial communities remain poorly characterized despite their ubiquity and importance to human and animal health, agriculture, and industry. Attempts to describe microbial communities by either traditional microbiological methods or molecular methods have been limited in both scale and precision. The availability of genomics technologies offers an unprecedented opportunity to conduct more comprehensive characterizations of microbial communities. Here we describe the application of an established molecular diagnostic method based on the chaperonin-60 sequence, in combination with high-throughput sequencing, to the profiling of a microbial community: the pig intestinal microbial community. Four libraries of cloned cpn60 sequences were generated by two genomic DNA extraction procedures in combination with two PCR protocols. A total of 1,125 cloned cpn60 sequences from the four libraries were sequenced. Among the 1,125 cloned cpn60 sequences, we identified 398 different nucleotide sequences encoding 280 unique peptide sequences. Pairwise comparisons of the 398 unique nucleotide sequences revealed a high degree of sequence diversity within the library. Identification of the likely taxonomic origins of cloned sequences ranged from imprecise, with clones assigned to a taxonomic subclass, to precise, for cloned sequences with 100% DNA sequence identity with a species in our reference database. The compositions of the four libraries were compared and differences related to library construction parameters were observed. Our results indicate that this method is an alternative to 16S rRNA sequence-based studies which can be scaled up for the purpose of performing a potentially comprehensive assessment of a given microbial community or for comparative studies.

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