Efficiency of automatic analyses of fish passages detected by an acoustic camera using Sonar5-Pro

The acoustic camera is a non-intrusive method increasingly used to monitor fish populations. Acoustic camera data are video-like, providing information on fish behaviour and morphology helpful to discriminate fish species. However, acoustic cameras used in long-term monitoring studies generate a large amount of data, making one of the technical limitations the time spent analysing data, especially for multi-species fish communities. The specific analysis software provided for DIDSON acoustic cameras is problematic to use for large datasets. Sonar5-Pro, a popular software in freshwater studies offers several advantages due to its automatic tracking tool that follows targets moving into the detection beam and distinguishes fish from other targets. This study aims to assess the effectiveness of Sonar5-Pro for detecting and describing fish passages in a high fish diversity river in low flow conditions. The tool's accuracy was assessed by comparing Sonar5-Pro outputs with a complete manual analysis using morphological and behavioural descriptors. Ninety-eight percent of the fish moving into the detection beam were successfully detected by the software. The fish swimming direction estimation was 90% efficient. Sonar5-Pro and its automatic tracking tool have great potential as a database pre-filtering process and decrease the overall time spent on data analysis but some limits were also identified. Multi-counting issues almost doubled the true fish abundance, requiring manual operator validation. Furthermore, fish length of each tracked fish needed to be manually measured with another software (SMC). In conclusion, a combination of Sonar5-Pro and SMC software can provide reliable results with a significant reduction of manpower needed for the analysis of a long-term monitoring DIDSON dataset

Impacts des souches du genre Pseudomonas protéolytiques sur la stabilité des produits laitiers transformées : maitrise et prédiction de la qualité des laits UHT

Dairy industries have sometimes problems of instability of UHT milk (Ultra High Temperature) during storage. This instability usually results in gelation or phase separation. Among the multifactorial causes (Zoological enzyme, Physico-chemical ...), proteolysis of casein micelles of UHT milk by heat-resistant proteases belonging to the genus Pseudomonas during storage, is often mentioned. This work has been conducted to understand the destabilization of UHT milk with raw milk previously contaminated by proteolytic Pseudomonas. Infrared spectroscopy Fourier transform has been explored in order to detect the cause and predict the stability of milk before their technological transformations. Initially, 32 strains of the genus Pseudomonas were selected according to their proteolytic potential and the presence of protease AprX (unique extracellular protease identified from Pseudomonas isolated from milk). All strains of the species P. fluorescens and P. tolaasii and some strains of the species P. chlororaphis, P. fragi and P. aeruginosa secreted this protease. Among the species P. aeruginosa and P. fragi, some strains secreted a protease with a molecular mass different from the one of the protease aprx. Regardless of the species studied and protease secreted, there was a proteolysis variability of the 32 strains. Some strains were proteolytic, while others were not. In a second step, the monitoring of the stability of UHT milk previously and independently inoculated by nine strains of P. fluorescens, secreting AprX but with different proteolytic potential, showed that this heterogeneity of proteolysis also happened in UHT milk. Some strains destabilized UHT milk before the end of 90 days of storage, while others did not. The addition of protease AprX in raw milk before UHT treatment also resulted in a destabilization of UHT milk before the end of 90 days of storage, confirming the role of this enzyme in the phenomenon of destabilization. The study with the strain P. fragi 9.12 that secreted a protease different from AprX, showed that it was able to destabilize UHT milk during storage. This result confirmed the fact that AprX was not the only extracellular protease bacteria of the genus Pseudomonas having an effect on the destabilization of UHT milk. For each experiment performed, the destabilization resulted in a formation of aggregation, in increases of non-protein and non-casein nitrogen contents (NPN and NCN) and in decreases of hydrations and zeta potentials of caseins micelles, indicating proteolysis of casein micelles. The results obtained by HPLC analysis coupled with mass spectrometry of NCN fractions of milk destabilized by 5 proteolytic strains of P. fluorescens and the protease AprX showed that this enzyme had a large action spectrum and was able to hydrolyze β, s1, s2 and κ casein. In a third step, FTIR spectroscopy was used to detect the presence of proteolytic bacteria of the genus Pseudomonas. This exploratory phase allowed us to determine the threshold for detection of milk contaminated with bacteria of the genus Pseudomonas and the discrimination threshold of milk containing proteolytic Pseudomonas from milk containing non-proteolytic Pseudomonas (5.102 CFU / mL). However, the analysis of milk containing a smaller quantity of proteolytic Pseudomonas than non proteolytic Pseudomonas, showed us that the character proteolytic dit not appears in the IR spectrum, while the risk of proteolysis was present. Further works have to be undertaken to use FTIR spectroscopy as a tool for predicting the stability of milk.L’industrie laitière rencontre parfois des problèmes d’instabilité du lait UHT (Ultra Haute Température) au cours de son stockage. Cette instabilité se traduit généralement par une gélification ou par une séparation de phases. Parmi les causes qui sont multifactorielles (zoologique, enzymatique, physico-chimique...) la protéolyse des micelles de caséines des laits UHT au cours de leur stockage par des protéases thermorésistantes appartenant au genre Pseudomonas est souvent évoquée. Ce travail a donc été effectué dans le but de mieux comprendre la déstabilisation du lait UHT dont le lait cru a été préalablement contaminé par des bactéries protéolytiques du genre Pseudomonas. La spectroscopie infrarouge à transformée de Fourier a été explorée afin de détecter cette cause et de prédire la stabilité des laits avant leurs transformations technologiques. Dans un premier temps, 32 souches du genre Pseudomonas ont été sélectionnées en fonction de leur caractère protéolytique et de la présence de la protéase AprX (seule protéase extracellulaire identifiée à partir de Pseudomonas isolée du lait). Toutes les souches des espèces P. fluorescens et P. tolaasii ainsi que quelques souches des espèces P. chlororaphis, P. fragi et P. aeruginosa sécrétaient cette protéase. Parmi les espèces P. aeruginosa et P. fragi, certaines souches sécrétaient une protéase de masse moléculaire différente de celle de la protéase AprX. Indépendamment de l’espèce étudiée et de la protéase secrétée, il existait une variabilité de protéolyse au sein des 32 souches. Certaines souches étaient protéolytiques alors que d’autres ne l’étaient pas. Dans un second temps, le suivi de la stabilité de laits UHT préalablement et indépendamment inoculés par 9 souches de P. fluorescens, sécrétant AprX mais au caractère protéolytique différent, a montré que cette hétérogénéité de protéolyse existait aussi dans le lait UHT. Certaines souches déstabilisaient le lait UHT avant la fin des 90 jours de stockage alors que d’autres non. L’addition de la protéase AprX, dans le lait cru avant traitement UHT a également engendré une déstabilisation du lait UHT avant la fin des 90 jours de stockage confirmant le rôle de cette enzyme dans le phénomène de déstabilisation. L’étude réalisée avec la souche P. fragi 9.12 qui semblait sécréter une protéase différente d’AprX nous a montré qu’elle était capable de déstabiliser le lait UHT au cours du stockage. Ce résultat nous a conforté sur le fait qu’AprX ne serait pas la seule protéase extracellulaire des bactéries du genre Pseudomonas ayant un effet sur la déstabilisation du lait UHT. Pour chaque expérimentation effectuée, la déstabilisation se traduisait par une apparition d’agrégats, des augmentations des teneurs en azote non protéique et non caséinique (NPN et NCN) et des diminutions de l’hydratation micellaire et du potentiel zêta, nous indiquant une protéolyse des micelles de caséines. Les résultats obtenus par l’analyse HPLC couplée à la spectrométrie de masse des fractions NCN des laits déstabilisés par les 5 souches de P. fluorescens déstabilisantes et la protéase AprX, ont montré qu’AprX avait un large spectre d’action et était capable d’hydrolyser les caséines β, s1, s2 et κ. Dans un troisième temps, la spectroscopie IRTF a été utilisée dans le but de détecter la présence de bactéries du genre Pseudomonas protéolytiques. Cette phase exploratoire nous a permis de déterminer le seuil de détection d’un lait contaminé par une bactérie du genre Pseudomonas ainsi que le seuil de discrimination d’un lait contenant des Pseudomonas protéolytiques d’un lait contenant des Pseudomonas non protéolytiques (5.102 UFC/mL). Cependant, l’analyse d’un lait contenant minoritairement des Pseudomonas protéolytiques nous a montré que le caractère protéolytique ne ressortait pas sur le spectre IR alors que le risque de protéolyse était bien présent. Des travaux supplémentaires sont donc à réaliser afin que la spectroscopie IRTF soit utilisée comme un outil de prédiction de la stabilité des laits

Efficiency of automatic analyses of fish passages detected by an acoustic camera using Sonar5-Pro

International audienceThe acoustic camera is a non-intrusive method increasingly used to monitor fish populations. Acoustic camera data are video-like, providing information on fish behaviour and morphology helpful to discriminate fish species. However, acoustic cameras used in long-term monitoring studies generate a large amount of data, making one of the technical limitations the time spent analysing data, especially for multi-species fish communities. The specific analysis software provided for DIDSON acoustic cameras is problematic to use for large datasets. Sonar5-Pro, a popular software in freshwater studies offers several advantages due to its automatic tracking tool that follows targets moving into the detection beam and distinguishes fish from other targets. This study aims to assess the effectiveness of Sonar5-Pro for detecting and describing fish passages in a high fish diversity river in low flow conditions. The tool's accuracy was assessed by comparing Sonar5-Pro outputs with a complete manual analysis using morphological and behavioural descriptors. Ninety-eight percent of the fish moving into the detection beam were successfully detected by the software. The fish swimming direction estimation was 90% efficient. Sonar5-Pro and its automatic tracking tool have great potential as a database pre-filtering process and decrease the overall time spent on data analysis but some limits were also identified. Multi-counting issues almost doubled the true fish abundance, requiring manual operator validation. Furthermore, fish length of each tracked fish needed to be manually measured with another software (SMC). In conclusion, a combination of Sonar5-Pro and SMC software can provide reliable results with a significant reduction of manpower needed for the analysis of a long-term monitoring DIDSON dataset

Influences of a dam on Atlantic salmon (Salmo salar) upstream migration in the Couesnon River (Mont Saint Michel Bay) using hydroacoustics

A dam has been recently modified on the Couesnon River in the vicinity of Mont Saint Michel to control and remove the volume of sediment brought into the bay by tidal waters. This paper shows that this dam has an impact on Atlantic salmon (Salmo salar) migration and that adult fish experience adaptive responses. Upstream migration activities were recorded with a 70 kHz echosounder (SIMRAD EY500 model) during two monitoring programs in 2010 and 2011, combining a total of 172 days of data collection. The echosounder was fixed at an upstream location, 8 km from the dam, and its detection beam capacity was approximately one third of the river's cross sectional area. In addition to the time of year, the upstream direction, swimming speed limit and fish size range were the specific criteria used from our database to efficiently discriminate other fish species from the Atlantic salmon. Among a total of 116,000 fishes counted during the two periods of investigations, 189 salmon were identified using those criteria. This study enabled us to identify the driving factors that influence the salmon migration in this semi-natural environment. There was a significant difference in fish activity when the dam was operating and when the river flow was under "normal" hydraulic conditions. Furthermore, during the opening and closing phases of the dam, the migration activity was scattered during the day, with peak abundance occurring mainly during the beginning of the operating phases. However, during all valve-opening phases, the salmon activity occurred mainly during the night. Therefore, the results suggest that this dam has a strong influence on fish migration activity at this particular location. (C) 2013 Elsevier Ltd. All rights reserved

Proteolytic destabilization of casein micelle in UHT-milk during storage: impact of different strains of Pseudomonas fluorescens

International audienceUHT sterilization combined with an aseptic packaging provides drinking milk that is microbiologically safe and can be stored at ambient temperature for several months. In some cases, commercial UHT milk can be destabilized by formation of a gel or sediment. Different studies have shown that residual activities of heat-resistant proteases of Pseudomonas fluorescens modify physico-chemical properties of casein micelles and lead to destabilization of UHT-milk. In our previous study, destabilization happened due to one strain of Pseudomonas fluorescens. In this present work, a wider range of strains were tested to explore further their impact on the quality of UHT-milk and to highlight the phenomena that cause proteolysis of casein micelles. Nine strains of Pseudomonas fluorescens secreting the heat-resistant protease aprX were selected as a function of their proteolytic potential. Each strain was inoculated in microfiltered milk, heat-treated at 140°C for 4 seconds and stored 90 days at 20°C. The physico-chemical characteristics of casein micelles were appreciated by evaluating their aggregation and measuring their size, zeta potential and hydration. In parallel, global proteolysis was determined by evaluating non casein nitrogen (NCN) and non protein nitrogen (NPN) contents. After 90 days of storage, destabilization of UHT-milks contaminated with the strains of Pseudomonas fluorescens was visual with presence of aggregates. Moreover, physico-chemical characteristics of casein micelles showed roughly the same evolution. Zeta potential, size, NCN and NPN contents increased whereas hydration decreased. The higher proteolytic activity was, the faster destabilization of milk happened. These results confirm that destabilization of UHT-milk occurs due to proteolysis of casein micelles. It suggests also that the heat-resistant protease aprX causes this proteolysis. To check this assumption, an experiment with inoculation of the purified enzyme in microfiltered milk is planned. Identification of cleavage sites is also a future project

Proteolytic destabilization of casein micelle in UHT-milk during storage: impact of different strains of Pseudomonas fluorescens

International audienceUHT sterilization combined with an aseptic packaging provides drinking milk that is microbiologically safe and can be stored at ambient temperature for several months. In some cases, commercial UHT milk can be destabilized by formation of a gel or sediment. Different studies have shown that residual activities of heat-resistant proteases of Pseudomonas fluorescens modify physico-chemical properties of casein micelles and lead to destabilization of UHT-milk. In our previous study, destabilization happened due to one strain of Pseudomonas fluorescens. In this present work, a wider range of strains were tested to explore further their impact on the quality of UHT-milk and to highlight the phenomena that cause proteolysis of casein micelles. Nine strains of Pseudomonas fluorescens secreting the heat-resistant protease aprX were selected as a function of their proteolytic potential. Each strain was inoculated in microfiltered milk, heat-treated at 140°C for 4 seconds and stored 90 days at 20°C. The physico-chemical characteristics of casein micelles were appreciated by evaluating their aggregation and measuring their size, zeta potential and hydration. In parallel, global proteolysis was determined by evaluating non casein nitrogen (NCN) and non protein nitrogen (NPN) contents. After 90 days of storage, destabilization of UHT-milks contaminated with the strains of Pseudomonas fluorescens was visual with presence of aggregates. Moreover, physico-chemical characteristics of casein micelles showed roughly the same evolution. Zeta potential, size, NCN and NPN contents increased whereas hydration decreased. The higher proteolytic activity was, the faster destabilization of milk happened. These results confirm that destabilization of UHT-milk occurs due to proteolysis of casein micelles. It suggests also that the heat-resistant protease aprX causes this proteolysis. To check this assumption, an experiment with inoculation of the purified enzyme in microfiltered milk is planned. Identification of cleavage sites is also a future project

Déstabilisation des micelles de caséine dans les laits UHT au cours de leur stockage. Implication des protéases extracellulaires appartenant au genre Pseudomonas

L’industrie laitière rencontre parfois des problèmes d’instabilité du lait UHT au cours de son stockage. L’instabilité des laits se traduit généralement par une gélification ou par une séparation de phases. Les causes sont multifactorielles (zoologique, enzymatique, physico chimique...). Cette instabilité pourrait venir de la protéolyse des micelles de caséines des laits UHT au cours de leurs stockages. Récemment, il a été démontré que les protéases des cellules somatiques et la plasmine n’avaient pas d’effet sur le phénomène de déstabilisation alors que la protéase AprX d’une souche de P. fluorescens (espèce psychrotrophe la plus retrouvée dans le lait) avait un effet marqué sur la déstabilisation (I.Gaucher, 2008). Ces résultats n’ayant été obtenus qu’avec une seule souche, il était donc essentiel de reproduire ce phénomène de déstabilisation et de le caractériser sur une plus grande gamme de souche du genre Pseudomonas afin de mieux comprendre les mécanismes mis en jeu.Pour cela, six souches du genre Pseudomonas ont été inoculées dans du lait microfiltré (lait épuré au niveau des cellules somatiques et de la flore bactérienne) puis traité thermiquement (140°C/4sec). Après 1 mois de stockage, la déstabilisation était visible. Des agrégats étaient également détectés. En parallèle, nous avons trouvé une augmentation de la taille et de la charge des micelles de caséines, une baisse de l’hydratation micellaire ainsi qu’une augmentation de l’azote non caséinique et de l’azote non protéique pour les laits ensemencés par rapport aux laits témoins. Les résultats obtenus traduisaient bien la protéolyse des micelles de caséines. L’outil spectrométrie de masse va être par la suite utilisé pour permettre de déterminer les peptides issus de ces protéolyses. Nous espérons de cette manière mieux comprendre le mode d’action de ces protéases. Sur les 6 souches, 5 secrètent la protéase aprX. La déstabilisation de ces laits serait donc due à cette protéase thermorésistante au traitement UHT. Le mécanisme proposé est résumé dans la figure 1. Afin de confirmer cette hypothèse, il a été prévu d’injecter une quantité définie de l’enzyme directement dans du lait UHT et d’en apprécier la déstabilisation

The biodiversity of the microbiota producing heat-resistant enzymes responsible for spoilage in processed bovine Milk and dairy products

Raw bovine milk is highly nutritious as well as pH-neutral, providing the ideal conditions for microbial growth. The microbiota of raw milk is diverse and originates from several sources of contamination including the external udder surface, milking equipment, air, water, feed, grass, feces, and soil. Many bacterial and fungal species can be found in raw milk. The autochthonous microbiota of raw milk immediately after milking generally comprises lactic acid bacteria such as Lactococcus, Lactobacillus, Streptococcus, and Leuconostoc species, which are technologically important for the dairy industry, although they do occasionally cause spoilage of dairy products. Differences in milking practices and storage conditions on each continent, country and region result in variable microbial population structures in raw milk. Raw milk is usually stored at cold temperatures, e.g., about 4°C before processing to reduce the growth of most bacteria. However, psychrotrophic bacteria can proliferate and contribute to spoilage of ultra-high temperature (UHT) treated and sterilized milk and other dairy products with a long shelf life due to their ability to produce extracellular heat resistant enzymes such as peptidases and lipases. Worldwide, species of Pseudomonas, with the ability to produce these spoilage enzymes, are the most common contaminants isolated from cold raw milk although other genera such as Serratia are also reported as important milk spoilers, while for others more research is needed on the heat resistance of the spoilage enzymes produced. The residual activity of extracellular enzymes after high heat treatment may lead to technological problems (off flavors, physico-chemical instability) during the shelf life of milk and dairy products. This review covers the contamination patterns of cold raw milk in several parts of the world, the growth potential of psychrotrophic bacteria, their ability to produce extracellular heat-resistant enzymes and the consequences for dairy products with a long shelf life. This problem is of increasing importance because of the large worldwide trade in fluid milk and milk powder