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How much is enough? Effects of technical and biological replication on metabarcoding dietary analysis

ABSTRACT: DNA metabarcoding is increasingly used in dietary studies to estimate diversity, composition and frequency of occurrence of prey items. However, few studies have assessed how technical and biological replication affect the accuracy of diet estimates. This study addresses these issues using the European free?tailed bat Tadarida teniotis, involving high?throughput sequencing of a small fragment of the COI gene in 15 separate faecal pellets and a 15?pellet pool per each of 20 bats. We investigated how diet descriptors were affected by variability among (a) individuals, (b) pellets of each individual and (c) PCRs of each pellet. In addition, we investigated the impact of (d) analysing separate pellets vs. pellet pools. We found that diet diversity estimates increased steadily with the number of pellets analysed per individual, with seven pellets required to detect ~80% of prey species. Most variation in diet composition was associated with differences among individual bats, followed by pellets per individual and PCRs per pellet. The accuracy of frequency of occurrence estimates increased with the number of pellets analysed per bat, with the highest error rates recorded for prey consumed infrequently by many individuals. Pools provided poor estimates of diet diversity and frequency of occurrence, which were comparable to analysing a single pellet per individual, and consistently missed the less common prey items. Overall, our results stress that maximizing biological replication is critical in dietary metabarcoding studies and emphasize that analysing several samples per individual rather than pooled samples produce more accurate results.

SUBMITTER: Vanessa A. Mata

PROVIDER: S-BSST327 | biostudies-other |

REPOSITORIES: biostudies-other

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Project description:PurposeThe mechanisms by which depressive symptoms negatively affect clinical outcomes in patients with coronary artery disease (CAD) remain poorly understood. Previous interventions that have attempted to treat depressive symptoms in patients with CAD to improve their clinical outcomes have been disappointing. Our objectives were, among a cohort of CAD patients, to evaluate the impact of depressive symptoms over time, controlling for comorbidity, in determining both successful long-term lifestyle change (ie, increased physical activity), and cardiovascular morbidity and mortality outcomes. In addition, we examined the impact of physical activity changes over time on 2 known mediators of cardiovascular morbidity: parasympathetic tone and inflammation.MethodsClinical data were previously collected (2004-2006) from 242 elective/urgent coronary angioplasty patients who participated in a prospective randomized controlled trial evaluating the efficacy of a behavioral intervention versus an educational control to motivate physical activity over 12 months. Exclusion criteria included: (1) inability to walk; (2) enrollment in other risk-reduction trials; (3) non-English speaking; and (4) lack of cardiologist's permission to increase physical activity. Participants were assessed every 2 months for interval clinical events and physical activity. In addition, biomarkers were collected at baseline and at 12 months in a subset of 54 participants; these biomarkers included low-frequency heart rate variability (lfHRV), high-frequency heart rate variability (hfHRV), serum C-reactive protein, interleukin-6, and salivary cortisol.FindingsThe mean age of participants was 63 years and 30% were female. Overall, 37% had high depressive symptoms at baseline. Patients with high depressive symptoms who achieved an increase in physical activity of ≥336 kilocalories(kcal)/week by 12 months had significantly lower rates of cardiovascular morbidity/mortality (5.1% vs. 21.3%; odds ratio [OR], 0.20, [95% CI, 0.04-0.98]; P = 0.03). In a multivariate model examining cardiovascular morbidity/mortality in patients with high depressive symptoms, an increase in physical activity of ≥336 kcal/week reduced the risk of new cardiovascular morbidity/mortality (OR, 0.11 [95% CI, 0.02-0.81]; P < 0.03), and comorbidity increased the risk (OR, 1.58 [95% CI, 1.18-2.13]; P = 0.002). In a generalized structural equation model, increasing physical activity by ≥336 kcal/week decreased the risk of complications, and comorbidity increased the risk. Furthermore, increasing physical activity (≥336 kcal/week) predicted an increase in hfHRV, a marker of parasympathetic tone, and the increase in hfHRV predicted a reduction in the proinflammatory mediators interleukin-6 and C-reactive protein.ImplicationsThis study found a threshold in physical activity in CAD patients with depressive symptoms that is associated with a decrease in cardiovascular morbidity and mortality. Exercise maintenance at this level may improve clinical outcomes via enhanced parasympathetic tone and decreased inflammation. ClinicalTrials.gov identifier: NCT00248846.

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How much is enough? Effects of technical and biological replication on metabarcoding dietary analysis

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