eDNA metabarcoding reveals the role of habitat specialization and spatial and environmental variability in shaping diversity patterns of fish metacommunities.

Information is scarce on how environmental and dispersal processes interact with biological features of the organisms, such as their habitat affinity, to influence patterns in biodiversity. We examined the role of habitat specialist vs. generalist species, and the spatial configuration, connectivity, and different environmental characteristics of river-floodplain habitats to get a more mechanistic understanding of alpha and beta diversity of fish metacommunities. We used environmental DNA metabarcoding to characterize species (taxa) richness and composition in two separate floodplains of the river Danube (Austria and Hungary) during two different hydrological conditions. Results showed that differences in the number of generalist and specialist species and their responses to connectivity and environmental gradients influenced patterns in alpha and beta diversity. Of the components of beta diversity, richness difference (nestedness) showed consistently higher values than replacement (turnover), mainly due to the decrease of specialist species along the connectivity gradient (i.e., from the mainstem to the most isolated oxbows). Variance in both alpha and beta diversity could be well predicted by a set of local and regional variables, despite high environmental variability, which characterizes river-floodplain ecosystems. Of these, the joint or shared variance fractions proved to be the most important, which indicates that the effects of local and regional processes cannot be unambiguously separated in these river-floodplain systems. Local scale environmental variables were more important determinants of both alpha and beta diversity in the low water period than in the high water period. These results indicate the differential role of local and regional processes in community organization during different hydrological conditions. Maintenance of both local and regional scale processes are thus important in the preservation of alpha and beta diversity of floodplain fish metacommunities, which should be considered by environmental management.

Hydrology is a major influence on amphibian abundance in a large European floodplain.

River-floodplain ecosystems play a crucial role in connecting landscape patches through hydrological connectivity, but they are among the most threatened ecosystems. Floodplains provide important habitat for amphibians by connecting aquatic and terrestrial habitats. Modifications to floodplain hydrology can impact amphibian communities, yet few studies have examined amphibian metacommunities in floodplain wetlands.In this study, we assessed patterns in amphibian breeding abundance in one of the largest floodplains of the Danube River, Hungary, relative to hydrological connectivity and multi-scale variables at 30 waterbody sites. Our aim was to determine whether these patterns aligned with the pond-permanence gradient hypothesis, where breeding amphibian abundance is predicted to be highest in ephemeral ponds without large predatory fish. We used Bayesian hierarchical modelling to estimate multi-species abundance from repeated survey (count) data collected over one breeding season.We detected the eggs and larvae of four amphibian species. The best model of abundance included covariates describing two principal component axes associated with waterbody hydrology and landscape composition within a 500-m radius of a site. There was a positive relationship between mean community abundance at a site and hydrological disconnection from the main river channel; however, the common toad (Bufo bufo) was associated with hydrologically connected waterbodies. There was a positive relationship between mean community abundance and a high proportion of forest cover and low cover of agricultural land within a 500-m radius around a site, although this relationship was clear for only two species. There was no support for models containing the number of large predatory fish species detected at a site.Although our results showed that amphibian abundance declined with hydrological connectivity, based on model selection we could not ascribe this relationship to an increased number of large predatory fish species detected in waterbodies close to the main river channel. Differences in life history and habitat requirements are likely to have explained interspecific responses to hydrological connectivity. Our results underscore the importance of addressing amphibian abundance at multiple spatial scales in floodplain wetlands, as landscape composition partly explained patterns in abundance.Application of multi-species abundance modelling allowed us to investigate environmental relationships for common and infrequently detected species. Habitat restoration programmes in floodplains should provide waterbodies disconnected from main river channels as potential amphibian breeding sites and protect or restore forest as terrestrial habitat.

Effect of 80% ethanol or 10% formalin fixation, freezing at -20 °C and staining on Myxobolus (Myxosporea) spores to be deposited in parasitological collections.

The preparation of myxosporeans for the description of myxospores and their preservation as type material in parasitological collections show great variations. Most frequently, formalin and ethanol are used for fixation and Giemsa solution for staining spores. In this work, authors studied the effect of 80% ethanol and 10% formalin fixation, freezing at -20 °C and staining on the size and transparency of two Myxobolus species of cyprinid fishes, M. bramae and M. bliccae spore, and recommended a new method for the deposition of type material to parasitological collections in museums. The studies have commended that fresh spores from mature plasmodia are the best material for measuring the size and studying the inner structures, the number of polar tubules in polar capsules and the morphological characters of the intercapsular appendix. The obtained quantitative data suggest that cryo- and chemical preservation do not have a notable negative effect on spores compared to fresh samples but they decrease the transparency of spores. Staining the spores with Ziehl-Neelsen has proved to be a useful method for studying the fine structure without size reduction, while Giemsa staining induced a shrinkage of spores so it seems to be not ideal for description of a new species. When treating spores of Myxobolus spp. with Lugol's solution, iodinophilous vacuoles in the sporoplasm were not recognised but visualisation of the coils of polar tubules was enhanced. As a type material for newly described species, authors suggest phototypes and spores fixed in 80% ethanol to be deposited into collections, as this preservation method is suitable for subsequent research, such as re-measurements and molecular analysis.

The mixed phylogenetic origin of northern pike (Esox lucius Linnaeus 1758) populations in the Middle Danubian drainage.

BACKGROUND: Pikes, members of genus Esox, are widespread freshwater predators of the northern hemisphere, and important sport fish also. From the Carpathian basin only one species, the northern pike (E. lucius) is noted. At the same time the pike stocks living in this area show high level of phenotypic variance (e.g. various body pattern) and its growth varies highly both among and within populations. These features usually explained by the environmental diversity of the area. Whereas we think that genetic reasons -e.g. the appearance of other/new pike species in the area- may also be responsible for these observed features. Since as no detailed information have been published from the pike populations of this area, so far; we conducted phylogenetic and morphological assay on 88 pike specimens, collected from 49 Middle Danubian sampling sites. RESULTS: Our phylogenetic surveys showed that the northern pike appear in the study area solely, but all the three of its major lineages (Northern, Circumpolar, Southern) were indicated. Only six specimens represent the Northern lineage, collected from the western part of the study area. The Circumpolar and Southern lineages were common in the Carpathian basin, but the Southern lineage showed higher levels of haplotype diversity than the Circumpolar clade. Which indicates that only the Southern lineage is native in the area, while the other two groups could have appeared in the Middle Danubian system either spontaneously or by human introduction. Moreover, the different clades appeared in the same populations, suggesting the opportunity of inter-lineage hybridisation. From the studied morphologicalal features, the number of scales on the lateral line and the head length showed significant differences among the lineages. At the same time the body pattern of the studied individuals seems to be rather influenced by the ontogenic changes than phylogeny. CONCLUSIONS: The high phenotypic variability of Middle Danubian northern pike populations may be due that all of its three major clades appeared and came in secondary contact in the area. In the within watershed spread of the non-native lineages the human stocking/transfer may play a considerable role.

Scavenging behaviour and size-dependent carcass consumption of the black bullhead (Ameiurus melas).

This study examined the size-dependent scavenging behaviour of black bullheads Ameiurus melas under laboratory conditions, using common bleak Alburnus alburnus and pumpkinseed Lepomis gibbosus carcasses. Video camera observations showed that the activity of A. melas was higher at night, but substantial daytime activity was also recorded. Larger A. melas were more active than their smaller conspecifics, especially at night. All size classes exhibited a well-defined sequence of consuming different parts of the carcasses independent of size, but larger individuals tended to consume carcasses more efficiently. Carcasses of the softer-bodied A. alburnus were consumed more readily than those of the bonier L. gibbosus, independent of size. This scavenging behaviour of A. melas might facilitate the invasion success of the species.

New record of metacercariae of the North American Posthodiplostomum centrarchi (Digenea, Diplostomidae) in pumpkinseed (Lepomis gibbosus) in Hungary.

Two species of the genus Posthodiplostomum (Digenea: Diplostomatidae) (Posthodiplostomum brevicaudatum Nordmann, 1832 and Posthodiplostomum cuticola Nordmann, 1832) are known as parasites of Hungarian native fishes. Metacercariae of P. cuticola are widespread in Europe and cause black spot disease. Several species of Posthodiplostomum were described also from North America but none of them has been isolated in Hungary up to now. Posthodiplostomum centrarchi Hoffman, 1958 has been detected recently in pumpkinseeds (Lepomis gibbosus L., 1758) in several European countries. Posthodiplostomum centrarchi was isolated for the first time in Hungary from pumpkinseeds caught in the Maconka water reservoir in 2015. Thereafter, several natural waters (e.g. the River Danube, Lake Balaton and the Sió channel) were sampled in order to determine its presence and distribution. Only the native species P. cuticola was detected in Lake Balaton on cyprinids but a relatively high infection rate of P. centrarchi was observed in the Sió channel close to the lake. Pathological changes were absent, and metacercariae were mostly attached to the surface of the liver, kidney and heart. The phylogenetic analysis of the ITS and COI sequences of P. centrarchi and P. cuticola clustered into two distinct branches, which was in agreement with the morphological results.

Impact of urbanization on abundance and phenology of caterpillars and consequences for breeding in an insectivorous bird.

Urbanization can have marked effects on plant and animal populations' phenology, population size, predator-prey, interactions and reproductive success. These aspects are rarely studied simultaneously in a single system, and some are rarely investigated, e.g., how insect phenology responds to urban development. Here, we study a tri-trophic system of trees, phytophagous insects (caterpillars), and insectivorous birds (Great Tits) to assess how urbanization influences (1) the phenology of each component of this system, (2) insect abundance, and (3) avian reproductive success. We use data from two urban and two forest sites in Hungary, central Europe, collected over four consecutive years. Despite a trend of earlier leaf emergence in urban sites, there is no evidence for an earlier peak in caterpillar abundance. Thus, contrary to the frequently stated prediction in the literature, the earlier breeding of urban bird populations is not associated with an earlier peak in caterpillar availability. Despite this the seasonal dynamics of caterpillar biomass exhibited striking differences between habitat types with a single clear peak in forests, and several much smaller peaks in urban sites. Caterpillar biomass was higher in forests than urban areas across the entire sampling period, and between 8.5 and 24 times higher during the first brood's chick-rearing period. This higher biomass was not associated with taller trees in forest sites, or with tree species identity, and occurred despite most of our focal trees being native to the study area. Urban Great Tits laid smaller clutches, experienced more frequent nestling mortality from starvation, reared fewer offspring to fledging age, and their fledglings had lower body mass. Our study strongly indicates that food limitation is responsible for lower avian reproductive success in cities, which is driven by reduced availability of the preferred nestling diet, i.e., caterpillars, rather than phenological shifts in the timing of peak food availability.

Problem-solving performance and reproductive success of great tits in urban and forest habitats.

Success in problem solving, a form of innovativeness, can help animals exploit their environments, and recent research suggests that it may correlate with reproductive success. Innovativeness has been proposed to be especially beneficial in urbanized habitats, as suggested by superior problem-solving performance of urban individuals in some species. If there is stronger selection for innovativeness in cities than in natural habitats, we expect problem-solving performance to have a greater positive effect on fitness in more urbanized habitats. We tested this idea in great tits (Parus major) breeding at two urban sites and two forests by measuring their problem-solving performance in an obstacle-removal task and a food-acquisition task. Urban pairs were significantly faster problem-solvers in both tasks. Solving speed in the obstacle-removal task was positively correlated with hatching success and the number of fledglings, whereas performance in the food-acquisition task did not correlate with reproductive success. These relationships did not differ between urban and forest habitats. Neophobia, sensitivity to human disturbance, and risk taking in the presence of a predator did not explain the relationships of problem-solving performance either with habitat type or with reproductive success. Our results suggest that the benefit of innovativeness in terms of reproductive success is similar in urban and natural habitats, implying that problem-solving skills may be enhanced in urban populations by some other benefits (e.g. increased survival) or reduced costs (e.g. more opportunities to gain practice with challenging tasks).

Does urbanization facilitate individual recognition of humans by house sparrows?

Wild animals living in proximity to humans may benefit from recognizing people individually and adjusting their behaviour to the potential risk or gain expected from each person. Although several urban-dwelling species exhibit such skills, it is unclear whether this is due to pre-existing advanced cognitive abilities of taxa predisposed for city life or arises specifically in urban populations either by selection or through ontogenetic changes facilitated by exposure to humans. To test these alternatives, we studied populations of house sparrows (Passer domesticus) along the urbanization gradient. We manipulated the birds' experience (hostile or not) associated with humans with different faces (masks) and measured their behavioural responses to the proximity of each person. Contrary to our expectations, we found that while rural birds showed less fear of the non-hostile than of the hostile or an unfamiliar person, urban birds made no distinction. These results indicate that house sparrows are less able to recognize individual humans or less willing to behaviourally respond to them in more urbanized habitats with high human population density. We propose several mechanisms that may explain this difference, including reduced pay-off of discrimination due to a low chance of repeated interactions with city people, or a higher likelihood that city people will ignore them.