Environmental DNA (eDNA) metabarcoding approaches to surveillance have great potential for advancing biodiversity monitoring and fisheries management. For eDNA metabarcoding, having a genetic reference sequence identified to fish species is vital to reduce detection errors. Detection errors will increase when there is no reference sequence for a species or when the reference sequence is the same between different species at the same sequenced region of DNA. These errors will be acute in high biodiversity systems like the Mekong River Basin, where many fish species have no reference sequences and many congeners have the same or very similar sequences. Recently developed tools allow for inspection of reference database coverage and the sequence similarity between species. These evaluation tools provide a useful pre-deployment approach to evaluate the breadth of fish species richness potentially detectable using eDNA metabarcoding. Here we combined established species lists for the Mekong River Basin, resulting in a list of 1345 fish species, evaluated the genetic library coverage across 23 peer-reviewed primer pairs, and measured the species specificity for one primer pair across four genera to demonstrate that coverage of genetic reference libraries is but one consideration before deploying an eDNA metabarcoding surveillance program. This analysis identifies many of the eDNA metabarcoding knowledge gaps with the aim of improving the reliability of eDNA metabarcoding applications in the Mekong River Basin. Genetic reference libraries perform best for common and commercially valuable Mekong fishes, while sequence coverage does not exist for many regional endemics, IUCN data deficient, and threatened fishes.

Microbes play a critical role in plant litter decomposition and influence the fate of carbon in rivers and riparian zones. When decomposing low‐nutrient plant litter, microbes acquire nitrogen (N) and phosphorus (P) from the environment (i.e., nutrient immobilization), and this process is potentially sensitive to nutrient loading and changing climate. Nonetheless, environmental controls on immobilization are poorly understood because rates are also influenced by plant litter chemistry, which is coupled to the same environmental factors. Here we used a standardized, low‐nutrient organic matter substrate (cotton strips) to quantify nutrient immobilization at 100 paired stream and riparian sites representing 11 biomes worldwide. Immobilization rates varied by three orders of magnitude, were greater in rivers than riparian zones, and were strongly correlated to decomposition rates. In rivers, P immobilization rates were controlled by surface water phosphate concentrations, but N immobilization rates were not related to inorganic N. The N:P of immobilized nutrients was tightly constrained to a molar ratio of 10:1 despite wide variation in surface water N:P. Immobilization rates were temperature‐dependent in riparian zones but not related to temperature in rivers. However, in rivers nutrient supply ultimately controlled whether microbes could achieve the maximum expected decomposition rate at a given temperature. Collectively, we demonstrated that exogenous nutrient supply and immobilization are critical control points for decomposition of organic matter.

Intensive fishing is altering the functioning of aquatic ecosystems worldwide, threatening both biodiversity and food security. No‐fishing reserves have proven effective at restoring food‐web structure and enhancing fisheries in marine ecosystems, but remain virtually untested in freshwater systems. Using experiments inside and outside of community‐created riverine reserves in Thailand, we describe a trophic cascade across six trophic levels, from humans to algal responses to nutrient availability. Protection from fishing profoundly reconfigures fish communities – greatly increasing biodiversity, biomass, and body size – yet mean trophic position was unaffected. Cascade dynamics from fish to algae were observed regardless of protection status, although fishing intensified trophic interactions through strong effects on grazing insect behavior. The marked effectiveness of these small, grassroots reserves offers an important conservation‐planning model for protecting food webs and augmenting fishery yields in biodiverse tropical rivers.