Biology subjects

Buczek, M.

Publications and source records attributed to Buczek, M..

7 recordsLinked to original sources

bioRxiv2024

Data of the Insect Biome Atlas: a metabarcoding survey of the terrestrial arthropods of Sweden and Madagascar

We present the data from the Insect Biome Atlas project (IBA), characterizing the terrestrial arthropod faunas of Sweden and Madagascar. Over 12 months, weekly Malaise trap samples were collected at 203 locations within 100 sites in Sweden and at 50 locations within 33 sites in Madagascar; this was complemented by soil and litter samples from each site. The field samples comprise 4,749 Malaise trap, 192 soil and 192 litter samples from Sweden and 2,566 Malaise trap and 190 litter samples from Madagascar. Samples were processed using mild lysis or homogenization, followed by DNA metabarcoding of COI (418 bp). The data comprise 698,378 non-chimeric sequence variants from Sweden and 687,866 from Madagascar, representing 33,989 (33,046 Arthropoda) and 77,599 (77,380 Arthropoda) operational taxonomic units, respectively. These are the most comprehensive data presented on these faunas so far, allowing unique analyses of the size, composition, spatial turnover and seasonal dynamics of the sampled communities. They also provide an invaluable baseline against which to gauge future changes.

ecology↗

bioRxiv2024

Cold and lonely: low-abundance microbiota and no core microbiome across a broad geographic sampling of a mosquito species in Greenland

Microbial symbionts are key players in insect ecology and evolution, but microbiota vary widely across and within host species. The microbiota of insect clades such as high-latitude mosquitoes remain understudied, despite their abundance and potential economic and medical relevance. Here, we resolve temporal and spatial variation in the abundance and diversity of bacteria and fungi associated with Ochlerotatus mosquitoes in Greenland. To this aim, we apply a comprehensive workflow combining the simultaneous characterization of host, bacterial, and fungal marker regions, contamination filtering, as well as bacterial quantification and joint species distribution modelling. We find that 573 mosquitoes collected in five regions of Greenland between 2009 and 2024 represented two species (O. nigripes and O. impiger). Their microbial communities were low-abundance, highly diverse, and dominated by environmentally versatile taxa, showing high individual-to-individual-rather than spatio-temporal variation, although taxa such as Pseudomonas and Serratia exhibited sex-specific associations. DNA from local vertebrates and potential vertebrate pathogens (e.g., Bartonella) was detected, likely reflecting blood meals. These findings suggest that Greenland Ochlerotatus mosquitoes do not rely on core microbiota, but engage in flexible associations with diverse, low-abundance microbial communities. They also point to mosquitoes as likely vectors of bacterial and fungal pathogens in the Arctic.

ecology↗

bioRxiv2024

What lurks beneath the humped back? - The patterns of bacterial abundance, diversity and distribution across communities of a dipteran family

Symbiotic microorganisms can profoundly impact insects, their life history traits, population dynamics, and evolutionary trajectories. However, microbiota remain poorly understood in natural insect communities, especially in dark taxa - i.e., hyperdiverse, yet understudied clades. Here, we implemented a novel multi-target amplicon sequencing approach to study microbiota in complex, species-rich communities. It combines four methodological innovations: (1) To establish a host taxonomic framework, we sequenced amplicons of the host marker gene (COI) and reconstructed barcodes alongside microbiota characterisation. (2) To assess microbiota abundance, we incorporated spike-in-based quantification. (3) To improve the phylogenetic resolution for the dominant endosymbiont, Wolbachia, we analysed bycatch data from the COI amplicon sequencing. (4) To investigate the primary drivers of host-microbe associations in massive multi-dimensional datasets, we performed HMSC modelling. Applying this approach to 1,842 wild-caught scuttle flies (Diptera: Phoridae) from northern Sweden, we organised them into 480 genotypes and 186 species and gained unprecedented insights into their microbiota. We found orders-of-magnitude differences in bacterial abundance and massive within-population variation in microbiota composition. Patterns and drivers differed among microbial functional categories: the distribution and abundance of facultative endosymbionts (Wolbachia, Rickettsia, Spiroplasma) were shaped by host species, genotype and sex. In contrast, many other bacterial taxa were broadly distributed across species and sites. This study highlights facultative endosymbionts as key players in insect microbiota and reveals striking variations in distributional patterns of microbial clades. It also demonstrates the power of integrative sequencing approaches in uncovering the ecological complexity and significance of symbiotic microorganisms in multi-species natural communities.

ecology↗

bioRxiv2024

A new tool in a toolbox: Addressing challenges in high-throughput microbiota surveys across diverse wild insects

With their significant effects on the biology of higher organisms, host-associated microbiota has attracted the research communitys attention. The rapid progress in sequencing techniques has greatly facilitated microbial community characterization. However, the most popular surveying technique, marker gene amplicon sequencing, has multiple caveats that are not often addressed satisfactorily, including the uncertainty about the identity of the surveyed wild-caught specimens, variable and sometimes very low abundance of microbes in some samples, or reagent- and cross-contamination. As a result, researchers often obtain incomplete, biased, and sometimes totally incorrect microbial community profiles. Here, we present a versatile, cost-effective, and high-throughput quantitative multi-target amplicon sequencing workflow for the characterization of host-associated microbial communities, combining laboratory and bioinformatic steps and addressing most of the known methodological issues. Optimized for the study of the microbiota of wild insects, it can be easily adapted for other sample types. Outputs include contamination-controlled data on the absolute abundance and identity of microbes present in insect samples, both at genotype- and OTU-level, as well as host barcodes alongside information on parasite infections. Using 1384 samples from Zackenberg Valley, NE Greenland, we demonstrate the potential of the workflow to study insect and symbiont diversity patterns across a large portion of a diverse natural community.

ecology↗

bioRxiv2024

Implementing high-throughput insect barcoding in microbiome studies: impact of non-destructive DNA extraction on microbiome reconstruction

BackgroundSymbiotic relationships with diverse microorganisms are crucial for many aspects of insect biology. However, while our understanding of insect taxonomic diversity and the distribution of insect species in natural communities is limited, we know much less about their microbiota. In the era of rapid biodiversity declines, as researchers increasingly turn towards DNA-based monitoring, developing and broadly implementing approaches for high-throughput and cost-effective characterization of both insect and insect-associated microbial diversity is essential. We need to verify whether approaches such as high-throughput barcoding, a powerful tool for identifying wild insects, would permit subsequent microbiota reconstruction in these specimens. MethodsHigh-throughput barcoding ("megabarcoding") methods often rely on non-destructive approaches for obtaining template DNA for PCR amplification by leaching DNA out of insect specimens using alkaline buffers such as HotSHOT. This study investigated the impact of HotSHOT on microbial abundance estimates and the reconstructed bacterial community profiles. We addressed this question by comparing quantitative 16S rRNA amplicon sequencing data for HotSHOT-treated or untreated specimens of 16 insect species representing six orders and selected based on the expectation of limited variation among individuals. ResultsWe find that in 13 species, the treatment significantly reduced microbial abundance estimates, corresponding to an estimated 15-fold decrease in amplifiable 16S rRNA template on average. On the other hand, HotSHOT pre-treatment had a limited effect on microbial community composition. The reconstructed presence of abundant bacteria with known significant effects was not affected. On the other hand, we observed changes in the presence of low-abundance microbes, those close to the reliable detection threshold. Alpha and beta diversity analyses showed compositional differences in only a few species. ConclusionOur results indicate that HotSHOT pre-treated specimens remain suitable for microbial community composition reconstruction, even if abundance may be hard to estimate. These results indicate that we can cost-effectively combine barcoding with the study of microbiota across wild insect communities. Thus, the voucher specimens obtained using megabarcoding studies targeted at characterizing insect communities can be used for microbiome characterizations. This can substantially aid in speeding up the accumulation of knowledge on the microbiomes of abundant and hyperdiverse insect species.

microbiology↗

bioRxiv2024

Pinpointing the microbiota of tardigrades: what is really there?

Microbiota have been proposed as an important aspect of tardigrade biology, but little is known about their diversity and distribution. Here, we attempted to characterize the microbiota of 44 cultured species of tardigrades using 16S rRNA amplicon sequencing, using different specimen pooling strategies, various DNA extraction kits, and multiple types of controls. We also estimated the number of microbes in samples using synthetic DNA spike-ins. Additionally, we reanalyzed data from previous studies. Our results suggest that the microbial community profiles of cultured tardigrades are dominated by bacterial OTUs and genotypes originating from food, medium, or laboratory reagents. We found microbial strains consistently enriched in certain tardigrades (relative to the culture media and controls), which indicates likely symbiotic associations, but the reads representing putative true tardigrade-associated microbes rarely exceeded 20% of the datasets. Some of the identified tardigrade-associated microbes matched symbionts identified by other studies. However, we also identified serious contamination issues with previous studies of tardigrade microbiome, making some of their conclusions questionable. We conclude that tardigrades are not universally dependent on specialized microbes and highlight the necessary safeguards in future studies of the microbiota of microscopic organisms.

microbiology↗

bioRxiv2022

Optimizing insect metabarcoding using replicated mock communities

Metabarcoding (high-throughput sequencing of marker gene amplicons) has emerged as a promising and cost-effective method for characterizing insect community samples. Yet, the methodology varies greatly among studies and its performance has not been systematically evaluated to date. In particular, it is unclear how accurately metabarcoding can resolve species communities in terms of presence-absence, abundances, and biomass. Here we use mock community experiments and a simple probabilistic model to evaluate the performance of different metabarcoding protocols. Specifically, we ask four questions: (Q1) How consistent are the recovered community profiles across replicate mock communities?; (Q2) How does the choice of lysis buffer affect the recovery of the original community?; (Q3) How are community estimates affected by differing lysis times and homogenization?; and (Q4) Is it possible to obtain adequate species abundance estimates through the use of biological spike-ins? We show that estimates are quite variable across community replicates. In general, a mild lysis protocol is better at reconstructing species lists and approximate counts, while homogenization is better at retrieving biomass composition. Tiny insects are more likely to be detected in lysates, while some tough species require homogenization to be detected. Results are less consistent across biological replicates for lysates than for homogenates. Some species are associated with strong PCR amplification bias, which complicates the reconstruction of species counts. Yet, with adequate spike-in data, species abundance can be determined with roughly 40% standard error for homogenates, and with roughly 50% standard error for lysates, under ideal conditions. In the latter case, however, this often requires species-specific reference data, while spike-in data generalizes better across species for homogenates. We conclude that a non-destructive, mild lysis approach shows the highest promise for presence/absence description of the community, while also allowing future morphological or molecular work on the material. However, homogenization protocols perform better for characterizing community composition, in particular in terms of biomass.

ecology↗