DNA-based identification through the use of metabarcoding has been proposed as the next step in the monitoring of biological communities, such as those assessed under the Water Framework Directive ...(WFD). Advances have been made in the field of metabarcoding, but challenges remain when using complex samples. Uneven biomass distributions, preferential amplification and reference database deficiencies can all lead to discrepancies between morphological and DNA-based taxa lists. The effects of different taxonomic groups on these issues remain understudied. By metabarcoding WFD monitoring samples, we analyzed six different taxonomic groups of freshwater organisms, both separately and combined. Identifications based on metabarcoding data were compared directly to morphological assessments performed under the WFD. The diversity of taxa for both morphological and DNA-based assessments was similar, although large differences were observed in some samples. The overlap between the two taxon lists was 56.8% on average across all taxa, and was highest for Crustacea, Heteroptera, and Coleoptera, and lowest for Annelida and Mollusca. Taxonomic sorting in six basic groups before DNA extraction and amplification improved taxon recovery by 46.5%. The impact on ecological quality ratio (EQR) scoring was considerable when replacing morphology with DNA-based identifications, but there was a high correlation when only replacing a single taxonomic group with molecular data. Different taxonomic groups provide their own challenges and benefits. Some groups might benefit from a more consistent and robust method of identification. Others present difficulties in molecular processing, due to uneven biomass distributions, large genetic diversity or shortcomings of the reference database. Sorting samples into basic taxonomic groups that require little taxonomic knowledge greatly improves the recovery of taxa with metabarcoding. Current standards for EQR monitoring may not be easily replaced completely with molecular strategies, but the effectiveness of molecular methods opens up the way for a paradigm shift in biomonitoring.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The heterogeneous nature of environmental DNA (eDNA) and its effects on species detection and community composition estimates has been highlighted in several studies in the past decades. Mostly in ...the context of spatial distribution over large areas, in fewer occasions looking at spatial distribution within a single body of water. Temporal variation of eDNA, similarly, has mostly been studied as seasonality, observing changes over large periods of time, and often only for small groups of organisms such as fish and amphibians.
We analyzed and compared small-scale spatial and temporal variation by sampling eDNA from two small, isolated dune lakes for 20 consecutive weeks. Metabarcoding was performed on the samples using generic COI primers. Molecular operational taxonomic unit (MOTUs) were used to assess dissimilarities between spatial and temporal replicates.
Our results show large differences between samples taken within one lake at one point in time, but also expose the large differences between temporal replicates, even those taken only 1 week apart. Furthermore, between-site dissimilarities showed a linear correlation with time frame, indicating that between-site differences will be inflated when samples are taken over a period of time. We also assessed the effects of PCR replicates and processing strategies on general patterns of dissimilarity between samples. While more inclusive PCR replicate strategies lead to higher richness estimations, dissimilarity patterns between samples did not significantly change.
We conclude that the dissimilarity of temporal replicates at a 1 week interval is comparable to that of spatial replicate samples. It increases, however, for larger time intervals, which suggests that population turnover effects can be stronger than community heterogeneity. Spatial replicates alone may not be enough for optimal recovery of taxonomic diversity, and cross-comparisons of different locations are susceptible to inflated dissimilarities when performed over larger time intervals. Many of the observed MOTUs could be classified as either phyto- or zooplankton, two groups that have gained traction in recent years as potential novel bio-indicator species. Our results, however, indicate that these groups might be susceptible to large community shifts in relatively short periods of time, highlighting the need to take temporal variations into consideration when assessing their usability as water quality indicators.
Airborne pollen monitoring is of global socio-economic importance as it provides information on presence and prevalence of allergenic pollen in ambient air. Traditionally, this task has been ...performed by microscopic investigation, but novel techniques are being developed to automate this process. Among these, DNA metabarcoding has the highest potential of increasing the taxonomic resolution, but uncertainty exists about whether the results can be used to quantify pollen abundance. In this study, it is shown that DNA metabarcoding using trnL and nrITS2 provides highly improved taxonomic resolution for pollen from aerobiological samples from the Netherlands. A total of 168 species from 143 genera and 56 plant families were detected, while using a microscope only 23 genera and 22 plant families were identified. NrITS2 produced almost double the number of OTUs and a much higher percentage of identifications to species level (80.1%) than trnL (27.6%). Furthermore, regressing relative read abundances against the relative abundances of microscopically obtained pollen concentrations showed a better correlation for nrITS2 (R2 = 0.821) than for trnL (R2 = 0.620). Using three target taxa commonly encountered in early spring and fall in the Netherlands (Alnus sp., Cupressaceae/Taxaceae and Urticaceae) the nrITS2 results showed that all three taxa were dominated by one or two species (Alnus glutinosa/incana, Taxus baccata and Urtica dioica). Highly allergenic as well as artificial hybrid species were found using nrITS2 that could not be identified using trnL or microscopic investigation (Alnus × spaethii, Cupressus arizonica, Parietaria spp.). Furthermore, perMANOVA analysis indicated spatiotemporal patterns in airborne pollen trends that could be more clearly distinguished for all taxa using nrITS2 rather than trnL. All results indicate that nrITS2 should be the preferred marker of choice for molecular airborne pollen monitoring.
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•DNA successfully extracted from microscopic pollen slides and Burkard-collected tapes.•nrITS2 and trnL DNA metabarcoding improves taxonomic resolution of airborne pollen.•Relative read abundance nrITS2 shows higher correlation with pollen counts than trnL.•Finer scale spatiotemporal patterns in pollen trends detected using nrITS2.•Detection of artificial hybrid (Alnus × spaethii) that significantly prolongs hay fever season
This prospective study aimed to study the composition and structure of the vaginal microbiota prior to
infection.
A nested case-control study was performed in 122 women, half of which acquired
within ...a year after their first visit. At the first visit, the composition and structure of vaginal microbial communities were analysed using 16S rRNA sequencing in the context of the sociodemographic and sexual risk behaviour information using logistic regression.
Five vaginal community state types (CSTs) were identified. Four CSTs were dominated by
spp., of which
(37%) and
(33%) were the most common. One CST was characterised by the absence of
spp. (25%) and the presence of an array of strict and facultative anaerobes. Multivariate logistic regression analysis revealed that women with a
dominated CST had an increased risk of
infection (p=0.04; OR: 2.6, 95% CI 1.0 to 6.6).
The distribution of CSTs dominated by
spp. agreed with previous studies. However, the frequency of dysbiosis among Caucasian women was relatively high (24%). Having vaginal microbiota dominated by
was associated with an increased risk for
infection. Therefore, we hypothesise that specific signatures of vaginal microbiota are indicative of increased host predisposition to acquiring STIs.
Corals harbor complex and diverse microbial communities that strongly impact host fitness and resistance to diseases, but these microbes themselves can be influenced by stresses, like those caused by ...the presence of macroscopic symbionts. In addition to directly influencing the host, symbionts may transmit pathogenic microbial communities. We analyzed two coral gall-forming copepod systems by using 16S rRNA gene metagenomic sequencing: (1) the sea fan Gorgonia ventalina with copepods of the genus Sphaerippe from the Caribbean and (2) the scleractinian coral Stylophora pistillata with copepods of the genus Spaniomolgus from the Saudi Arabian part of the Red Sea. We show that bacterial communities in these two systems were substantially different with Actinobacteria, Alphaproteobacteria, and Betaproteobacteria more prevalent in samples from Gorgonia ventalina, and Gammaproteobacteria in Stylophora pistillata. In Stylophora pistillata, normal coral microbiomes were enriched with the common coral symbiont Endozoicomonas and some unclassified bacteria, while copepod and gall-tissue microbiomes were highly enriched with the family ME2 (Oceanospirillales) or Rhodobacteraceae. In Gorgonia ventalina, no bacterial group had significantly different prevalence in the normal coral tissues, copepods, and injured tissues. The total microbiome composition of polyps injured by copepods was different. Contrary to our expectations, the microbial community composition of the injured gall tissues was not directly affected by the microbiome of the gall-forming symbiont copepods.
Freshwater habitats are under stress from agricultural land use, most notably the influx of neonicotinoid pesticides and increased nutrient pressure from fertilizer. Traditional studies investigating ...the effects of stressors on freshwater systems are often limited to a narrow range of taxa, depending heavily on morphological expertise. Additionally, disentanglement of multiple simultaneous stressors can be difficult in field studies, whereas controlled laboratory conditions do not accurately reflect natural conditions and food webs. To overcome these drawbacks, we investigated the impacts of two agricultural stressors (the neonicotinoid insecticide thiacloprid and fertilizer) in full‐factorial design in a semi‐natural research site, using environmental DNA sampling to study three different taxonomic groups representing three trophic levels: bacteria (decomposers), phytoplankton (primary producers), and chironomids (consumers). The results show considerable impact of both stressors across trophic levels, with an additive effect of fertilizer and thiacloprid on community composition at all levels. These findings suggest that agricultural stressors affect the entire food web, either directly or through cascade reactions. They are also consistent with morphological assessments that were performed in the same study site, even at a lower number of replicates. The study presented shows that the use of multimarker environmental DNA provides a more comprehensive assessment of stressor impacts across multiple trophic levels, at a higher taxonomic resolution than traditional surveys. Additionally, many putative novel bioindicators for both agricultural stressors were discovered. We encourage further investigations into stressors impacts at different trophic levels, which will lead to more effective monitoring and management of freshwater systems.
Anal and penile high-risk human papillomavirus (HPV) infection is associated with anogenital cancer, which is especially common in HIV-infected MSM. We assessed HPV prevalence and determinants in ...MSM.
Analysis of baseline data from a prospective cohort study.
MSM aged 18 years or older were recruited in Amsterdam, the Netherlands. Participants completed risk-factor questionnaires. HPV DNA was analyzed in anal and penile shaft self-swabs and genotyped using a sensitive PCR and reverse line blot assay (SPF10-PCR-DEIA-LiPA25-system). Multivariable logistic regression analyses were performed to assess determinants of high-risk HPV infection.
MSM (n = 778) were recruited in 2010-2011, of whom 317 (41%) were HIV-infected. Prevalence of anal high-risk HPV infection was 45% in HIV-negative versus 65% in HIV-infected MSM (P <0.001). HPV-16 was the most frequently detected type and was more common in HIV-infected MSM (13% in HIV-negative and 22% in HIV-infected MSM; P = 0.001). Prevalence of penile high-risk HPV infection was 16% in HIV-negative and 32% in HIV-infected MSM (P <0.001). In multivariable analyses, HIV infection remained associated with anal adjusted odds ratio (aOR) 2.2; 1.8-2.7 and penile (aOR 2.0; 1.4-2.9) high-risk HPV infection. Higher number of lifetime male sex partners was significantly associated with anal and penile high-risk HPV in HIV-negative, but not HIV-infected MSM. Receptive anal intercourse was associated with anal high-risk HPV in HIV-infected MSM.
Anal and penile high-risk HPV infections are very common in MSM. HIV infection is a strong and independent determinant for anal and penile high-risk HPV infection. Determinants for HPV infection appear to differ between HIV-negative and HIV-infected MSM.
Oral infection with human papillomavirus (HPV) is associated with a subset of head and neck cancers. We compared prevalence of, and risk factors for, oral HPV infection among HIV-negative and ...HIV-infected MSM.
Analysis of baseline data from a prospective cohort study.
MSM aged 18 years or older were recruited from three study sites in Amsterdam, the Netherlands. Participants completed a self-administered risk-factor questionnaire. Oral-rinse and gargle specimens were analyzed for HPV DNA and genotyped using a highly sensitive PCR and reverse line blot assay short PCR fragment (SPF)10-PCR-DNA Enzyme Immuno Assay (DEIA)/LiPA25 system.
In 2010-2011, 794 MSM were included, of whom 767 participants had sufficient data for analysis. Median age was 40.1 years interquartile range (IQR) 34.8-47.5 and 314 men were HIV-infected (40.9%). Any of 25 typable HPV types was present in 24.4% of all oral samples. Oncogenic HPV types were detected in 24.8 and 8.8% of oral samples from HIV-infected and HIV-negative MSM, respectively (P < 0.001). Of these high-risk types, HPV-16 was the most common (overall 3.4%). Oral infection with high-risk HPV was associated with HIV infection in multivariable analysis (P < 0.001). Increasing age was significantly associated with oral HPV infection in HIV-negative, but not in HIV-infected MSM.
Oral HPV infection is very common among MSM. HIV infection was independently associated with high-risk oral HPV infection, suggesting an important role of HIV in oral HPV infection.
Classical ecological theory posits that species partition resources such that each species occupies a unique resource niche. In general, the availability of more resources allows more species to ...co-occur. Thus, a strong relationship between communities of consumers and their resources is expected. However, correlations may be influenced by other layers in the food web, or by the environment. Here we show, by studying the relationship between communities of consumers (land snails) and individual diets (from seed plants), that there is in fact no direct, or at most a weak but negative, relationship. However, we found that the diversity of the individual microbiome positively correlates with both consumer community diversity and individual diet diversity in three target species. Moreover, these correlations were affected by various environmental variables, such as anthropogenic activity, habitat island size, and a possibly important nutrient source, guano runoff from nearby caves. Our results suggest that the microbiome and the environment explain the absence of correlations between diet and consumer community diversity. Hence, we advocate that microbiome inventories are routinely added to any community dietary analysis, which our study shows can be done with relatively little extra effort. Our approach presents the tools to quickly obtain an overview of the relationships between consumers and their resources. We anticipate our approach to be useful for ecologists and environmentalists studying different communities in a local food web.
The effects of single or multiple concordant HPV infections at various anatomical sites on type-specific HPV seropositivity are currently unknown. In this cross-sectional study we assessed whether ...high-risk HPV infections at various anatomical sites (i.e., anal canal, penile shaft, and oral cavity), as well as concordant infections at multiple anatomical sites, were associated with type-specific seropositivity in HIV-positive and HIV-negative MSM. MSM aged ≥ 18 years were recruited in Amsterdam, the Netherlands (2010-2011). Baseline anal, penile, and oral samples were analyzed for HPV DNA and genotyped using a highly sensitive PCR and reverse line blot assay. Virus-like particle (VLP) based multiplex immunoassay was used to asses HPV-specific serum antibodies against L1 VLPs. The associations between HPV infections and type-specific seropositivity of seven high-risk HPV types (7-hrHPV: types 16, 18, 31, 33, 45, 52, 58) were estimated using logistic regression analyses with generalized estimating equations. We found that 86% of 306 HIV-positive MSM and 62% of 441 HIV-negative MSM were seropositive for at least one 7-hrHPV type. 69% of HIV-positive and 41% of HIV-negative MSM were infected with at least one 7-hrHPV type at the anus, penis, or oral cavity. In multivariable analyses, 7-hrHPV seropositivity was associated with type-specific anal (and not penile) 7-hrHPV infection, and did not significantly increase with a higher number of infected anatomical sites. Oral 7-hrHPV infection showed a positive, albeit non-significant, association with seropositivity. In conclusion, seropositivity among MSM appears to be largely associated with anal HPV infection, irrespective of additionally infected anatomical sites.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
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