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Simple Summary: This study explores the use of alternative and non-harmful methods for monitoring endangered Mediterranean trout in small intermittent streams. Specifically, two visual survey techniques were compared: a visual survey from streambanks and an underwater visual survey using underwater cameras. The main objective was to assess the effectiveness of detecting patterns in fish occupancy in relation to a set of environmental factors. The comparison revealed that underwater camera surveys performed similarity to visual surveys from streambanks during low-flow regimes. However, visual surveys from streambanks were less effective during the highest flow regimes. The volume of pools and the percentage cover of submerged macrophytes were found to be significant environmental variables affecting fish detection probability using underwater cameras. On the other hand, the analysis of data from visual observations from streambanks indicated a clear impact of high turbulence rates on pool occupancy. In conclusion, these findings demonstrate the utility of visual methods in describing the occupancy patterns of Mediterranean trout in small streams. Monitoring the conservation status of endangered freshwater fish using less invasive methods poses challenges for ecologists and conservationists. Visual surveys have been proposed as an alternative to electrofishing, which is a standard methodology that can cause injuries, physiological stress and post-release mortality in organisms. To test the efficacy of visual methods, a study was conducted in an intermittent stream of Sardinia (Italy). Two visual methods were employed: a visual survey from streambanks (VSS) and an underwater visual survey (UVS) using cameras. The aims of this study were (1) to compare the effectiveness of these methods in detecting patch occupancy patterns and (2) to investigate the effect of environmental variables on the detection probability of Mediterranean native trout. Environmental variables characterizing pool habitats were recorded, and generalized linear models (GLMs) were employed to assess the correlation between these variables and trout presence/absence. GLM analysis revealed that UVS had higher detection probability with larger pool volume, whereas submerged macrophytes negatively affected detection probability. Detection from streambanks (VVS) was negatively affected by a high turbulence rate. In conclusion, our study suggests the utility of visual methods to describe patterns of patch occupancy of Mediterranean trout. However, methods can be differently affected by environmental variables. Therefore, monitoring programs using these methods should consider these factors to ensure a reliable description of within-stream trout distribution in intermittent streams. [ABSTRACT FROM AUTHOR]

Monitoring the conservation status of endangered freshwater fish using less invasive methods poses challenges for ecologists and conservationists. Visual surveys have been proposed as an alternative to electrofishing, which is a standard methodology that can cause injuries, physiological stress and post-release mortality in organisms. To test the efficacy of visual methods, a study was conducted in an intermittent stream of Sardinia (Italy). Two visual methods were employed: a visual survey from streambanks (VSS) and an underwater visual survey (UVS) using cameras. The aims of this study were (1) to compare the effectiveness of these methods in detecting patch occupancy patterns and (2) to investigate the effect of environmental variables on the detection probability of Mediterranean native trout. Environmental variables characterizing pool habitats were recorded, and generalized linear models (GLMs) were employed to assess the correlation between these variables and trout presence/absence. GLM analysis revealed that UVS had higher detection probability with larger pool volume, whereas submerged macrophytes negatively affected detection probability. Detection from streambanks (VVS) was negatively affected by a high turbulence rate. In conclusion, our study suggests the utility of visual methods to describe patterns of patch occupancy of Mediterranean trout. However, methods can be differently affected by environmental variables. Therefore, monitoring programs using these methods should consider these factors to ensure a reliable description of within-stream trout distribution in intermittent streams.

Brown trout (Salmo trutta), like many other freshwater species, is threated by the release in its natural environment of alien species and the restocking with allochthonous conspecific stocks. Many conservation projects are ongoing and several morphological and genetic tools have been proposed to support activities aimed to restore genetic integrity status of native populations. Nevertheless, due to the complexity of degree of introgression reached up after many generations of crossing, the use of dichotomous key and molecular markers, such as mtDNA, LDH-C1* and microsatellites, are often not sufficient to discriminate native and admixed specimens at individual level. Here we propose a reduced panel of ancestry-informative SNP markers (AIMs) to support on field activities for Mediterranean trout management and conservation purpose. Starting from the genotypes data obtained on specimens sampled in the main two Molise's rivers (Central-Southern Italy), a 47 AIMs panel was identified and validated on simulated and real hybrid population datasets, mainly through a Machine Learning approach based on Random Forest classifier. The AIMs panel proposed may represent an interesting and cost-effective tool for monitoring the level of introgression between native and allochthonous trout population for conservation purpose and this methodology could be also applied in other species. [ABSTRACT FROM AUTHOR]

Abstract Salmonids inhabiting Mediterranean rivers are of particular concern for biodiversity conservation, as they are threatened by various stressors, including habitat alterations, overfishing, climate change, and introgressive hybridization with alien species. In the Tiber River basin (Central Italy), genetic introgression phenomena of the native Salmo cettii with the non‐native Salmo trutta hinder the separate analysis of the two species, which are both included in the S. trutta complex. Little is known about the factors currently limiting the trout populations in this area, particularly with respect to climate change. With the intention of filling this gap, the aims of the current study were to (a) quantify changes in the climate and (b) analyze the distribution, status, and ecology of trout populations, in the context of changing abiotic conditions over the last decades. Fish stock assessments were carried out by electrofishing during three census periods (1998–2004, 2005–2011, and 2012–2018) at 129 sites. The trend over time of meteorological parameters provided evidence for increased air temperature and decreased rainfall. Multivariate analysis of trout densities and environmental data highlighted the close direct correlation of trout abundance with water quality, altitude, and current speed. Climate‐induced effects observed over time in the sites where trout were sampled have not yet led to local extinctions or distribution shifts, indicating a marked resilience of trout, probably due to the buffering effect of intrinsic population dynamics. Decreasing body conditions over time and unbalanced age structures support the hypothesis that variations in hydraulic regime and water temperature could overcome these compensatory effects, which may lead to a severe decline in trout populations in the near future. In a climate change context, habitat availability plays a key role in the distribution of cold‐water species, which often do not have the possibility to move upstream to reach their thermal optimum because of water scarcity in the upper river stretches.

In Italian freshwaters the introgressive hybridization with the alien Atlantic trout represents the principal threat to the native endangered Mediterranean trout. The aim of the research was to test the effectiveness of alien trout removal activities carried out by electrofishing in four streams in Central Italy. The four sites were chosen on the basis of the severe genetic introgression level of the trout populations. The removal activities were carried out from December 2014 to July 2017, for a total of 10 eradication campaigns in each site. During each field activity, five environmental parameters (flow rate, conductivity, average depth, average width, accessibility) were collected, since they could influence both the effectiveness of the electrofishing and the biological characteristics of the fish populations. A total of over 22,000 alien trout accounting for a biomass of over 700 kg were removed from the removal sites. For each site, a progressive drastic decrease over time in density and standing crop values was observed. Removal rates ranged from 94.22% to 100.00% for density, and from 96.57% to 100.00% for standing crop. The correlation analysis showed an inverse significant relationship between catchability and populations abundance, confirming that low density populations are more suitable to removal efforts. The progressive removal of specimens improved the catchability over time, indicating that even the largest populations could be eradicated, providing the necessary number of steps in a fishing season. The improved body condition and the greater growth rates observed during the eradication period, in low abundance conditions, seemed to confirm the key role that intraspecific competition and density‐dependent phenomena play in the Apennine trout population dynamics. The results obtained in the present research provided evidence that electrofishing removal could be an effective management tool for the Mediterranean trout conservation, especially in watercourses of modest dimensions in terms of flow rates, width and depth. [ABSTRACT FROM AUTHOR]

Salmonids inhabiting Mediterranean rivers are of particular concern for biodiversity conservation, as they are threatened by various stressors, including habitat alterations, overfishing, climate change, and introgressive hybridization with alien species. In the Tiber River basin (Central Italy), genetic introgression phenomena of the native Salmo cettii with the non‐native Salmo trutta hinder the separate analysis of the two species, which are both included in the S. trutta complex. Little is known about the factors currently limiting the trout populations in this area, particularly with respect to climate change. With the intention of filling this gap, the aims of the current study were to (a) quantify changes in the climate and (b) analyze the distribution, status, and ecology of trout populations, in the context of changing abiotic conditions over the last decades. Fish stock assessments were carried out by electrofishing during three census periods (1998–2004, 2005–2011, and 2012–2018) at 129 sites. The trend over time of meteorological parameters provided evidence for increased air temperature and decreased rainfall. Multivariate analysis of trout densities and environmental data highlighted the close direct correlation of trout abundance with water quality, altitude, and current speed. Climate‐induced effects observed over time in the sites where trout were sampled have not yet led to local extinctions or distribution shifts, indicating a marked resilience of trout, probably due to the buffering effect of intrinsic population dynamics. Decreasing body conditions over time and unbalanced age structures support the hypothesis that variations in hydraulic regime and water temperature could overcome these compensatory effects, which may lead to a severe decline in trout populations in the near future. In a climate change context, habitat availability plays a key role in the distribution of cold‐water species, which often do not have the possibility to move upstream to reach their thermal optimum because of water scarcity in the upper river stretches. [ABSTRACT FROM AUTHOR]

Knowledge of the composition of the gut microbiota in freshwater fish living in their natural habitat has taxonomic and ecological importance. Few reports have been produced on the composition of the gut microbiota and on the presence of LAB in the intestines of freshwater fish that inhabit river environments. In this study, we investigated the LAB community that was present in the gastrointestinal tract (GIT) of Mediterranean trout (Salmo macrostigma) that colonized the Biferno and Volturno rivers of the Molise region (Italy). The partial 16S rRNA gene sequences of these strains were determined for the species-level taxonomic placement. The phylogenetic analysis revealed that the isolated LABs belonged to seven genera (Carnobacterium, Enterococcus, Lactobacillus, Lactiplantibacillus, Vagococcus, Lactococcus, and Weissella). The study of the enzymatic activities showed that these LABs could contribute to the breakdown of polysaccharides, proteins, and lipids. In future studies, a greater understanding of how the LABs act against pathogens and trigger the fish immune response may provide practical means to engineer the indigenous fish microbiome and enhance disease control and fish health.

Mediterranean trout is a freshwater fish of particular interest with economic significance for fishery management, aquaculture and conservation biology. Unfortunately, native trout populations’ abundance is significantly threatened by anthropogenic disturbance. The introduction of commercial hatchery strains for recreation activities has compromised the genetic integrity status of native populations. This work assessed the fine-scale genetic structure of Mediterranean trout in the two main rivers of Molise region (Italy) to support conservation actions. In total, 288 specimens were caught in 28 different sites (14 per basins) and genotyped using the Affymetrix 57 K rainbow-trout-derived SNP array. Population differentiation was analyzed using pairwise weighted FST and overall F-statistic estimated by locus-by-locus analysis of molecular variance. Furthermore, an SNP data set was processed through principal coordinates analysis, discriminant analysis of principal components and admixture Bayesian clustering analysis. Firstly, our results demonstrated that rainbow trout SNP array can be successfully used for Mediterranean trout genotyping. In fact, despite an overwhelming number of loci that resulted as monomorphic in our populations, it must be emphasized that the resulted number of polymorphic loci (i.e., ~900 SNPs) has been sufficient to reveal a fine-scale genetic structure in the investigated populations, which is useful in supporting conservation and management actions. In particular, our findings allowed us to select candidate sites for the collection of adults, needed for the production of genetically pure juvenile trout, and sites to carry out the eradication of alien trout and successive re-introduction of native trout.

Environmental, climate and historical factors are important to explain patterns of freshwater biodiversity and population dynamics in the Mediterranean area. This region is one of the most important areas for the maintenance of native lineages for brown trout. The aim of this study was the identification of the main drivers for the spread and the distribution of genetic introgression between alien brown trout and two native Mediterranean salmonids (brown and marble trout). Estimates of mitochondrial and nuclear introgression were from both the literature and original data and were used as dependent variables in a multivariate framework, correlating them to a suite of environmental and climate parameters. The last glacial maximum appeared as an important factor explaining the geographic pattern of alien brown trout genes throughout the Alps. Here, native populations of Mediterranean salmonids persisted in former refugia. Throughout the Italian Peninsula and major islands, geological setting of catchment and current climate conditions are key factors for securing the persistence of native trout populations. The reevaluation of genetic data regarding the spread of alien brown trout lineage into Mediterranean salmonids populations with a landscape approach allowed us to reveal the role of important factors implicated with the current pattern of distribution of remnant native populations of salmonids. This information provides new insights for improving conservation strategies and management of taxa threatened by the incipient global climate changes. [ABSTRACT FROM AUTHOR]

Overexploitation and consequent depletion of the breeding stock is one of multiple stressors affecting Mediterranean trout Salmo cettii Rafinesque, 1810 in central Italy. The practice of total Catch and Release (C&R), combined with gear restrictions and seasonal closures, reduces catch-related impacts. To test for the effects of C&R management, the goals of the study were to: (i) analyse the status of trout inhabiting 8 C&R areas of the Nera River Basin, and (ii) compare the results with those from 11 exploited areas and 15 no-fishing areas. Fish data were collected by electrofishing from 1997 to 2020. Abundance, growth, age structure, and relative weight were estimated. Trout populations in the C&R sectors exhibited high abundances, well-balanced age structures, and a congruous number of age classes according to the longevity of the species. However, the abundance values never reached those of the no-fishing areas. The results provided evidence for an unbalanced population structure in fishing areas, where intensive size-selective harvest caused negative demographic effects, especially on older fish, and the legal-limit length was not adequate for maximal yield length. C&R angling appeared to maintain the fishery value of trout stocks favouring the conservation of native trout, although a certain impact on fish population status remains and thus, it cannot completely replace the adoption of more restrictive criteria. [ABSTRACT FROM AUTHOR]

The super-species Salmo trutta is taxonomically divided into a number of semi-species, local varieties, morphs, and possible hybrids due to restocking. Their genetic status is nevertheless not always clear, although relevant for management and conservation practices. We used mitochondrial haplotypes, nuclear microsatellites, and LDH- C1 genotyping to study the trout population of Lake Posta Fibreno, one of the few extant populations of the semi-species Salmo ( trutta) macrostigma ( S.t.m.). Levels of introgression from Northern European stocks were estimated and the possible genetic separation and/or differential introgression were evaluated in the three generally recognized morphological variants fibreno, leopardo, and rossa. We show that trout from Posta Fibreno belong to the Adriatic group. The three morphotypes are genetically uniform, regardless of their apparently stable morphological differences. Low levels of introgression from non-autochthonous stocks of Northern European origin were detected, but these figures are not statistically different among the three morphs, ruling out the possibility that one or more morphs are in fact hybrid phenotypes. The relative 'purity' of the S.t.m. population form Posta Fibreno was compared with available accounts of introgression in other Italian populations, showing that it is in fact one of the least affected, and of significant interest for conservation. [ABSTRACT FROM AUTHOR]

Knowledge of the composition of the gut microbiota in freshwater fish living in their natural habitat has taxonomic and ecological importance. Few reports have been produced on the composition of the gut microbiota and on the presence of LAB in the intestines of freshwater fish that inhabit river environments. In this study, we investigated the LAB community that was present in the gastrointestinal tract (GIT) of Mediterranean trout (Salmo macrostigma) that colonized the Biferno and Volturno rivers of the Molise region (Italy). The partial 16S rRNA gene sequences of these strains were determined for the species-level taxonomic placement. The phylogenetic analysis revealed that the isolated LABs belonged to seven genera (Carnobacterium, Enterococcus, Lactobacillus, Lactiplantibacillus, Vagococcus, Lactococcus, and Weissella). The study of the enzymatic activities showed that these LABs could contribute to the breakdown of polysaccharides, proteins, and lipids. In future studies, a greater understanding of how the LABs act against pathogens and trigger the fish immune response may provide practical means to engineer the indigenous fish microbiome and enhance disease control and fish health. [ABSTRACT FROM AUTHOR]

Simple Summary: Mediterranean trout is one of the most threatened freshwater fish at risk of extinction. A complex pattern of climatic and anthropogenic pressures has dramatically compromised its biodiversity. In particular, the introduction of non-native trout represents one of the most important threats with negative impact on intraspecific diversity of native populations. The introgressive hybridization between native and alien trout reduces the fitness of native trout and creates hybrid swarms, resulting in native genome extinction. Recently, several conservation projects have been proposed to restore the genetic integrity status of native Mediterranean trout. In this study, we report the first use of the Affymetrix 57 K rainbow-trout-derived SNP array in research on Italian Mediterranean trout populations. The results provide insight into the genetic relationships and spatial distribution of two trout populations inhabiting the Volturno and Biferno rivers (Central-Southern Italy) and provide useful information for the identification of a fine-scale genetic structure, as well as the determination of subpopulations and their related habitats. These data are crucial to undertake effective conservation and management strategies with the aim to preserve native trout and recover autochthonous genetic heritage in such rivers. Overall, our outcomes support the use of the rainbow-trout-derived SNP array to identify SNPs that are informative in relation to the Mediterranean trout genome. Mediterranean trout is a freshwater fish of particular interest with economic significance for fishery management, aquaculture and conservation biology. Unfortunately, native trout populations' abundance is significantly threatened by anthropogenic disturbance. The introduction of commercial hatchery strains for recreation activities has compromised the genetic integrity status of native populations. This work assessed the fine-scale genetic structure of Mediterranean trout in the two main rivers of Molise region (Italy) to support conservation actions. In total, 288 specimens were caught in 28 different sites (14 per basins) and genotyped using the Affymetrix 57 K rainbow-trout-derived SNP array. Population differentiation was analyzed using pairwise weighted FST and overall F-statistic estimated by locus-by-locus analysis of molecular variance. Furthermore, an SNP data set was processed through principal coordinates analysis, discriminant analysis of principal components and admixture Bayesian clustering analysis. Firstly, our results demonstrated that rainbow trout SNP array can be successfully used for Mediterranean trout genotyping. In fact, despite an overwhelming number of loci that resulted as monomorphic in our populations, it must be emphasized that the resulted number of polymorphic loci (i.e., ~900 SNPs) has been sufficient to reveal a fine-scale genetic structure in the investigated populations, which is useful in supporting conservation and management actions. In particular, our findings allowed us to select candidate sites for the collection of adults, needed for the production of genetically pure juvenile trout, and sites to carry out the eradication of alien trout and successive re-introduction of native trout. [ABSTRACT FROM AUTHOR]