Development of a new method for extracting histamine from marine fish flesh using the salting-out technique.

A simple and practical method was developed to extract histamine from fish products using sodium chloride (NaCl). After obtaining a saline extract from fish samples, histamine was derivatized by a condensation reaction with ortho-phthalaldehyde. Fluorescence intensity was measured by a fluorimeter. The first part of this work concerned a solid-liquid extraction tested with samples from the food analysis performance assessment scheme. The best histamine extraction yield (97%) was obtained using an extraction time of 4 minutes, a temperature of 40°C, and a NaCl/water ratio of 41% (w/w). The second part focused on a liquid-liquid extraction carried out on standard solutions of histamine (45, 90, and 180 mg/kg). The use of NaCl (41%) and trichloroacetic acid [(TCA) 10%] did not show any significant difference in extraction yield. The yield obtained was 99.15-100.1% for TCA (10%) and 98.65-99.45% for NaCl (41%). The validation criteria (repeatability and reproducibility) were checked by evaluating the reliability of the method. Extraction using NaCl has proven to be an interesting alternative method for the extraction of histamine from fish, as it is reliable, inexpensive, and less hazardous.

Effect of Fishing Season and Size on the Physicochemical and Microbiological Characteristics of Salted Sardines (Sardina pilchardus).

Salting is the preferred method used by manufacturers to preserve the quality of fish. The aim of this study was to investigate the effect of size and fishing season on the physicochemical and microbiological characteristics of sardines. Two batches of sardines, caught, in winter and summer, were sorted according to size, salted, matured, and stored at 18°C. A control batch, consisting of sardines from the summer batch, was also prepared without subcutaneous fat. Various physicochemical and microbiological parameters were monitored during 12 weeks of maturation. The recorded values showed a decrease in pH, moisture content, and water activity (a measure of the water available for biological and chemical processes), while salt concentration increased. When comparing samples, the lowest values for histamine (94.3 and 48.3 mg/kg) and lipids (9.3 and 8.2%) were observed in the large and small sardines of the winter batch, respectively. In the summer batch, higher values were recorded for histamine (847.3 and 127.9 mg/kg) and lipids (14.5 and 8.6%) in the large and small sardines. Furthermore, the removal of fat in control of the summer batch resulted in lower histamine accumulation compared to the batch with fat. The microbiological parameters showed a decrease in the number of nonhalophile bacteria, while the number of halophile bacteria increased. This study showed a strong correlation between three important factors, fishing season, fish size, and histamine content, which can contribute to the successful salting of sardines and ensure the wholesomeness of the final product.

Assessment of heavy metal contamination and ecological risk in Morocco's marine and estuarine ecosystems through a combined analysis of surface sediment and bioindicator species: Donax trunculus and Scrobicularia plana.

Morocco is one of the most affected regions by heavy metal pollution worldwide. In this study, two ecosystems in Agadir Bay, southern Morocco, were studied seasonally, using surface sediment and bivalve species. The concentrations of Cu, Pb and Cd were determined using the Shimadzu AAS 7000 flame atomic absorption spectrophotometer method. Our results marked average levels corresponding to an unpolluted sediment with a low contamination degree, low ecological risk associated with metal contamination, and levels that did not exceed the thresholds set by EC, USEPA, INERIS and INRH, except for Pb in tourist beach. The principal component analysis findings revealed a positive correlation in the bioaccumulation between the two compartments, with an influence of abiotic parameters. Thus, to better manage environmental pollution in these ecosystems, authorities must implement waste treatment strategies in the surrounding harbor and tourist complexes and prohibit the use of these toxic metals in the coastal areas.

Microbiological assessment of marine and estuarine ecosystems using fecal indicator bacteria, Salmonella, Vibrio and antibiotic resistance pattern.

Marine and estuarine environments are often affected by microbiological contamination that adversely affects their use and severely impacts human health. To examine the influence of anthropogenic activities, this study used two different ecosystems in Agadir Bay, to compare fecal indicator bacteria (FIB) and bacterial pathogen profiles over two years. Vibrio target pathogens were detected at a high frequency (49.3%), while a low percentage (5.5%) was noted for Salmonella. Apart from those mentioned above, several other pathogenic bacteria were detected such as Cronobacter sakzakii, Pseudomonas fluorescens, and Aeromonas hydrophila. We also investigated the antimicrobial resistance of the pathogenic bacteria isolated. Salmonella strains were sensitive to all the antibiotics used, except ampicillin, amoxicillin + Ac clavulanic and chloramphenicol. And Vibrio strains were resistant to ampicillin, cephalothin, amikacin, and ciprofloxacin. This study highlights the limitations of FIB in assessing the microbiological quality and the importance of environmental surveys in understanding the distribution of pathogens.

Integration assay of bacteriological risks in marine environment using salmonella spp and multimarker response in the bivalve Donax trunculus: Novel biomonitoring approach.

Microbiological contamination is one of the riskiest forms of human contamination in seawater, which threaten the stability of ecosystems and human health. In this study, we study the accumulation of a pathogenic bacteria Salmonella spp; isolated from the marine environment, in the soft tissue of Donax trunculus (Mollusca, Bivalvia), a commonly used as a bioindicators species for aquatic ecosystems monitoring, under laboratory conditions during both exposure and recovery periods. These bacteria were added in seawater at three concentrations previously determined against sentinel specie at three exposure periods (24, 48 and 96 h). In a second series of experiments, exposed specimens were afterward transplanted to clean water to assess the recovery pattern. The mortality rate of bivalves was determined as biomarker of general stress. Our findings suggest that microbiological contamination by Salmonella spp was gradually incorporated into the body of D. trunculus causing a significant induction of enzymatic activity of acetylcholinesterase (AChE), Catalase (CAT), glutathione-S-transferase (GST) and malondialdehyde (MDA) levels, as a function of time and concentration. Exposure to a bacterial concentration of 5.104 bacteria/liter resulted in the mortality of more than 80% of the specimens. This study is to test the pathogenicity of Salmonella strains at concentrations close to those of the marine environment, and their effects on biomarkers, thus deducing the existence of an exponential relationship between bacterial concentrations and enzymatic response. The principal component analysis shows that the four biomarkers had similar variation with bacterial concentrations, while two groups were obtained to change following the exposure time (CAT-GST and AChE-MDA). This study provides new findings on the potential accumulation of pathogenic bacteria associated with neurotoxicity and oxidative stress in the wedge clam Donax trunculus.

Assessment of seasonal and spatial variation responses of integrated biomarkers in two marine sentinel bivalve species: Agadir Bay (Southern of Morocco).

The present study aims to assess the effects of contamination of the Agadir bay coasts using bivalves as a biomonitoring sentinel species. Seasonal variations of biochemical composition in terms of total protein content and oxidative stress biomarkers including glutathione-S-transferase, malondialdehyde, catalase and acetylcholinesterase were evaluated in the soft tissues of Scrobicularia plana and Donax trunculus specimens. The latter were collected from two sites in Agadir bay during two-year span (2018-2020). The Integrated Biomarker Response Index (IBR) was performed to classify the stress response in both species and to assess the level of exposure to xenobiotics. The data showed maximum annual values of acetylcholinesterase and malondialdehyde for Donax trunculus in Agadir beach (AG) with 6.25 nmol/mn/mg and 3 nmol/mg of protein, respectively. Those of catalase and glutathione-S-transferase for Scrobicularia plana in Oued Souss estuary (OS) were of 4.41 µmol/mn/mg and 14.43 nmol/mn/mg of protein, respectively. The studied species are considered good indicators in aquatic ecosystems.

Characteristics and dynamics of Salmonella contamination along the coast of Agadir, Morocco.

The occurrence of Salmonella enterica in the environment of tropical and desert regions has remained largely uninvestigated in many areas of the world, including Africa. In the present study, we investigated the presence of Salmonella spp. along 122 km of the coastline of Agadir (southern Morocco) in relation to environmental parameters. A total of 801 samples of seawater (243), marine sediment (279), and mussels (279) were collected from six sites between July 2004 and May 2008. The overall prevalence of Salmonella spp. was 7.1%, with the highest occurrence in mussels (10%), followed by sediment (6.8%) and seawater (4.1%). Only three serotypes were identified among the 57 Salmonella sp. strains isolated. S. enterica serotype Blockley represented 43.8% of all Salmonella strains and was identified in mussel and sediment samples. S. enterica serotype Kentucky (29.8%) was found almost exclusively in mussels, whereas S. enterica serotype Senftenberg (26.3%) was detected in sediment and seawater. Statistical analysis using generalized additive models identified seawater temperature, environmental temperature, rainfall, and solar radiation as significant factors associated with the presence of Salmonella. Rainfall was the only variable showing a linear positive effect on the presence of Salmonella in the sea, whereas the remaining variables showed more complex nonlinear effects. Twenty-eight (49.1%) Salmonella isolates displayed resistance to ampicillin (22 isolates), nalidixic acid (9 isolates), sulfonamide compounds (2 isolates), and tetracycline (1 isolate), with six of these isolates displaying multiple resistance to two of these antimicrobial agents. Pulsed-field gel electrophoresis analysis revealed homogenous restriction patterns within each serotype that were uncorrelated with the resistance pattern profiles.