Chilled Pacu (Piaractus mesopotamicus) fillets: Modeling Pseudomonas spp. and psychrotrophic bacteria growth and monitoring spoilage indicators by 1H NMR and GC-MS during storage.

This study aimed to assess the growth of Pseudomonas spp. and psychrotrophic bacteria in chilled Pacu (Piaractus mesopotamicus), a native South American fish, stored under chilling conditions (0 to 10 °C) through the use of predictive models under isothermal and non-isothermal conditions. Growth kinetic parameters, maximum growth rate (µmax, 1/h), lag time (tLag, h), and (Nmax, Log10 CFU/g) were estimated using the Baranyi and Roberts microbial growth model. Both kinetic parameters, growth rate and lag time, were significantly influenced by temperature (P < 0.05). The square root secondary model was used to describe the bacteria growth as a function of temperature. Secondary models, √µ = 0.016 (T + 10.13) and √µ =0.017 (T + 9.91) presented a linear correlation with R2 values >0.97 and were further validated under non-isothermal conditions. The model's performance was considered acceptable to predict the growth of Pseudomonas spp. and psychrotrophic bacteria in refrigerated Pacu fillets with bias and accuracy factors between 1.24 and 1.49 (fail-safe) and 1.45-1.49, respectively. Fish biomarkers and spoilage indicators were assessed during storage at 0, 4, and 10 °C. Volatile organic compounds, VOCs (1-hexanol, nonanal, octenol, and indicators 2-ethyl-1-hexanol) showed different behavior with storage time (P > 0.05). 1H NMR analysis confirmed increased enzymatic and microbial activity in Pacu fillets stored at 10 °C compared to 0 °C. The developed and validated models obtained in this study can be used as a tool for decision-making on the shelf-life and quality of refrigerated Pacu fillets stored under dynamic conditions from 0 to 10 °C.

Ultra-high irradiance (UHI) blue light: highlighting the potential of a novel LED-based device for short antifungal treatments of food contact surfaces.

Microbial food spoilage is an important cause of health and economic issues and can occur via resilient contamination of food surfaces. Novel technologies, such as the use of visible light, have seen the light of day to overcome the drawbacks associated with surface disinfection treatments. However, most studies report that photo-inactivation of microorganisms with visible light requires long time treatments. In the present study, a novel light electroluminescent diode (LED)-based device was designed to generate irradiation at an ultra-high power density (901.1 mW/cm2). The efficacy of this technology was investigated with the inactivation of the yeast S. cerevisiae. Short-time treatments (below 10 min) at 405 nm induced a ~4.5 log reduction rate of the cultivable yeast population. The rate of inactivation was positively correlated to the overall energy received by the sample and, at a similar energy, to the power density dispatched by the lamp. A successful disinfection of several food contact surfaces (stainless steel, glass, polypropylene, polyethylene) was achieved as S. cerevisiae was completely inactivated within 5 min of treatments. The disinfection of stainless steel was particularly effective with a complete inactivation of the yeast after 2 min of treatment. This ultra-high irradiance technology could represent a novel cost- and time-effective candidate for microbial inactivation of food surfaces. These treatments could see applications beyond the food industry, in segments such as healthcare or public transport. KEY POINTS : • A novel LED-based device was designed to emit ultra-high irradiance blue light • Short time treatments induced high rate of inhibition of S. cerevisiae • Multiple food contact surfaces were entirely disinfected with 5-min treatments.

High pressure sensitization of heat-resistant and pathogenic foodborne spores to nisin.

Today, there is no effective non-thermal method to inactivate unwanted bacterial spores in foods. High-Pressure (HP) process has been shown to act synergistically with moderate heating and the bacteriocin nisin to inactivate spores but the mechanisms have not been elucidated. The purpose of the present work was to investigate in depth the synergy of HP and nisin on various foodborne spore species and to bring new elements of understandings. For this purpose, spores of Bacillus pumilus, B. sporothermodurans, B. licheniformis, B. weihenstephanensis, and Clostridium sp. were suspended in MES buffer, in skim milk or in a liquid medium simulating cooked ham brine and treated by HP at 500 MPa for 10 min at 50 °C or 20 °C. Nisin (20 or 50 IU/mL) was added at three different points during treatment: during HP, during and or in the plating medium of enumeration. In the latter two cases, a high synergy was observed with the inhibition of the spores of Bacillus spp. The evaluation of the germinated fraction of Bacillus spp. spores after HP revealed that this synergy was likely due to the action of nisin on HP-sensitized spores, rather than on HP-germinated spores. Thus, the combination of nisin and HP can lead to Bacillus spp. spore inhibition at 20 °C. And Nisin can act on HP-treated spores, even if they are not germinated. This paper provides new information about the inhibition of spores by the combination of HP and nisin. The high synergy observed at low temperature has not been reported yet and could allow food preservation without the use of any thermal process.

Cellular Injuries in Cronobacter sakazakii CIP 103183T and Salmonella enterica Exposed to Drying and Subsequent Heat Treatment in Milk Powder.

Because of the ability of foodborne pathogens to survive in low-moisture foods, their decontamination is an important issue in food protection. This study aimed to clarify some of the cellular mechanisms involved in inactivation of foodborne pathogens after drying and subsequent heating. Individual strains of Salmonella Typhimurium, Salmonella Senftenberg, and Cronobacter sakazakii were mixed into whole milk powder and dried to different water activity levels (0.25 and 0.58); the number of surviving cells was determined after drying and subsequent thermal treatments in closed vessels at 90 and 100°C, for 30 and 120 s. For each condition, the percentage of unculturable cells was estimated and, in parallel, membrane permeability and respiratory activity were estimated by flow cytometry using fluorescent probes. After drying, it was clearly observable that the percentage of unculturable cells was correlated with the percentage of permeabilized cells (responsible for 20-40% of the total inactivated bacteria after drying), and to a lesser degree with the percentage of cells presenting with loss of respiratory activity. In contrast, the percentages of unculturable cells observed after heat treatment were strongly correlated with the loss of respiratory activity and weakly with membrane permeability (for 70-80% of the total inactivated bacteria after heat treatment). We conclude that cell inactivation during drying is closely linked to membrane permeabilization and that heat treatment of dried cells affects principally their respiratory activity. These results legitimize the use of time-temperature scales and allow better understanding of the cellular mechanisms of bacterial death during drying and subsequent heat treatment. These results may also allow better optimization of the decontamination process to ensure food safety by targeting the most deleterious conditions for bacterial cells without denaturing the food product.

Caco-2 Invasion by Cronobacter sakazakii and Salmonella enterica Exposed to Drying and Heat Treatments in Dried State in Milk Powder.

Due to the ability of foodborne pathogens to survive in low moisture food, the decontamination of milk powder is an important issue in food protection. The safety of food products is, however, not always insured and the different steps in the processing of food involve physiological and metabolic changes in bacteria. Among these changes, virulence properties may also be affected. In this study, the effect of drying and successive thermal treatments on the invasion capacity of Salmonella Typhimurium, Salmonella Senftenberg, and Cronobacter sakazakii was assessed. Bacteria were dried on milk powder at three different water activity levels (0.25, 0.58, and 0.80) and heated at two different temperatures (90°C and 100°C) for 30 and 120 s. After recovery, stressed bacterial populations were placed in contact with Caco-2 cells to estimate their invasion capacity. Our results show that drying increases the invasion capacity of foodborne pathogens, but that heat treatment in the dried state did not exert a selective pressure on bacterial cells depending on their invasion capacity after drying. Taken together, our findings add to the sum of knowledge on food safety in dried food products and provide insight into the effects of food processing.

Modeling the heat inactivation of foodborne pathogens in milk powder: High relevance of the substrate water activity.

Due to the ability of foodborne pathogens to survive in low moisture foods, the decontamination of these products is an important issue in food hygiene. Up to now, such decontamination has mostly been achieved through empirical methods. The intention of this work is to establish a more rational use of heat treatment cycles. The effects of thermal treatment cycles on the inactivation of dried Salmonella Typhimurium, Salmonella Senftenberg, Cronobacter sakazakii and Escherichia coli were assessed. Bacteria were mixed with whole milk powder and dried down to different water activity levels (0.11, 0.25, 0.44 and 0.58). The rate of inactivated bacteria was determined after thermal treatment at 85°C, 90°C, 95°C and 100°C, from 0s to 180s in closed vessels, in order to maintain aw during treatment. In a first step, logarithmic bacterial inactivation was fitted by means of a classical loglinear model in which temperature and aw have a significant effect (p<0.05). DT,aw values were estimated for each T, aw condition and the results clearly showed that aw is a major parameter in the thermal decontamination of dried foods, a lower aw involving greater thermal resistance. In a second step, Bigelow's law was used to determine zT, a classical parameter relative to temperature, and yaw values, a new parameter relative to aw resistance. The values obtained for zT and yaw showed that the bacterium most resistant to temperature variations is Salmonella Typhimurium, while the one most resistant to aw variations is Escherichia coli. These data will help design decontamination protocols or processes in closed batches for low moisture foods.

Listeria monocytogenes ability to survive desiccation: Influence of serotype, origin, virulence, and genotype.

Listeria monocytogenes, a bacterium that is responsible for listeriosis, is a very diverse species. Desiccation resistance has been rarely studied in L. monocytogenes, although it is a stress that is largely encountered by this microorganism in food-processing environments and that could be managed to prevent its presence. The objective of this study was to evaluate the resistance of 30 L. monocytogenes strains to moderate desiccation (75% relative humidity) and evaluate the correlation of such resistance with the strains' virulence, serotype and genotype. The results showed a great heterogeneity of strains regarding their ability to survive (loss of cultivability between 0.4 and 2.0 log). Strains were classified into three groups according to desiccation resistance (sensitive, intermediate, or resistant), and the strain repartition was analyzed relative to serotype, virulence level and environmental origin of the strains. No correlation was found between isolate origin and desiccation resistance. All serotype 1/2b strains were classified into the group of resistant strains. Virulent and hypovirulent strains were distributed among the three groups of desiccation resistance. Finally, a genomic comparison was performed based on 31 genes that were previously identified as being involved in desiccation resistance. The presence of those genes was localized among the genomes of some strains and compared regarding strain-resistance levels. High nucleotide conservation was identified between resistant and desiccation-sensitive strains. In conclusion, the findings regarding the strains of serotype 1/2b indicate potential serotype-specific resistance to desiccation, and thus, to relative humidity fluctuations potentially encountered in food-related environments. The genomic comparison of 31 genes associated to desiccation tolerance did not reveal differences among four strains which have different level of resistance to desiccation.

Drying parameters greatly affect the destruction of Cronobacter sakazakii and Salmonella Typhimurium in standard buffer and milk.

Salmonella Typhimurium and Cronobacter sakazakii are two foodborne pathogens involved in neonatal infections from milk powder and infant formula. Their ability to survive in low-moisture food and during processing from the decontamination to the dried state is a major issue in food protection. In this work, we studied the effects of the drying process on Salmonella Typhimurium and Cronobacter sakazakii, with the aim of identifying the drying parameters that could promote greater inactivation of these two foodborne pathogens. These two bacteria were dried under different atmospheric relative humidities in milk and phosphate-buffered saline, and the delays in growth recovery and cultivability were followed. We found that water activity was related to microorganism resistance. C. sakazakii was more resistant to drying than was S. Typhimurium, and milk increased the cultivability and recovery of these two species. High drying rates and low final water activity levels (0.11-0.58) had a strong negative effect on the growth recovery and cultivability of these species. In conclusion, we suggest that effective use of drying processes may provide a complementary tool for food decontamination and food safety during the production of low-moisture foods.

Recovery Estimation of Dried Foodborne Pathogens Is Directly Related to Rehydration Kinetics.

Drying is a common process which is used to preserve food products and technological microorganisms, but which is deleterious for the cells. The aim of this study is to differentiate the effects of drying alone from the effects of the successive and necessary rehydration. Rehydration of dried bacteria is a critical step already studied in starter culture but not for different kinetics and not for pathogens. In the present study, the influence of rehydration kinetics was investigated for three foodborne pathogens involved in neonatal diseases caused by the consumption of rehydrated milk powder: Salmonella enterica subsp. enterica serovar Typhimurium, Salmonella enterica subsp. enterica serovar Senftenberg and Cronobacter sakazakii. Bacteria were dried in controlled relative humidity atmospheres and then rehydrated using different methods. Our results showed that the survival of the three pathogens was strongly related to rehydration kinetics. Consequently, rehydration is an important step to consider during food safety assessment or during studies of dried foodborne pathogens. Also, it has to be considered with more attention in consumers' homes during the preparation of food, like powdered infant formula, to avoid pathogens recovery.

Control of Relative Air Humidity as a Potential Means to Improve Hygiene on Surfaces: A Preliminary Approach with Listeria monocytogenes.

Relative air humidity fluctuations could potentially affect the development and persistence of pathogenic microorganisms in their environments. This study aimed to characterize the impact of relative air humidity (RH) variations on the survival of Listeria monocytogenes, a bacterium persisting on food processing plant surfaces. To assess conditions leading to the lowest survival rate, four strains of L. monocytogenes (EGDe, CCL500, CCL128, and LO28) were exposed to different RH conditions (75%, 68%, 43% and 11%) with different drying kinetics and then rehydrated either progressively or instantaneously. The main factors that affected the survival of L. monocytogenes were RH level and rehydration kinetics. Lowest survival rates between 1% and 0.001% were obtained after 3 hours of treatment under optimal conditions (68% RH and instantaneous rehydration). The survival rate was decreased under 0.001% after prolonged exposure (16h) of cells under optimal conditions. Application of two successive dehydration and rehydration cycles led to an additional decrease in survival rate. This preliminary study, performed in model conditions with L. monocytogenes, showed that controlled ambient RH fluctuations could offer new possibilities to control foodborne pathogens in food processing environments and improve food safety.

Taxonomic composition and trophic structure of the continental bony fish assemblage from the early late cretaceous of Southeastern Morocco.

The mid-Cretaceous vertebrate assemblage from south-eastern Morocco is one of the most diversified continental vertebrate assemblages of this time worldwide. The bony fish component (coelacanths, lungfishes and ray-finned fishes) is represented by relatively complete specimens and, mostly, by fragmentary elements scattered along 250 kilometres of outcrops. Here we revisit the bony fish assemblage by studying both isolated remains collected during several fieldtrips and more complete material kept in public collections. The assemblage comprises several lungfish taxa, with the first mention of the occurrence of Arganodus tiguidiensis, and possibly two mawsoniid coelacanths. A large bichir cf. Bawitius, is recorded and corresponds to cranial elements initially referred to 'Stromerichthys' from coeval deposits in Egypt. The ginglymodians were diversified with a large 'Lepidotes' plus two obaichthyids and a gar. We confirm here that this gar belongs to a genus distinctive from Recent gars, contrary to what was suggested recently. Teleosteans comprise a poorly known ichthyodectiform, a notopterid, a probable osteoglossomorph and a large tselfatiiform, whose cranial anatomy is detailed. The body size and trophic level for each taxon are estimated on the basis of comparison with extant closely related taxa. We plotted the average body size versus average trophic level for the Kem Kem assemblage, together with extant marine and freshwater assemblages. The Kem Kem assemblage is characterized by taxa of proportionally large body size, and by a higher average trophic level than the trophic level of the extant compared freshwater ecosystems, but lower than for the extant marine ecosystems. These results should be regarded with caution because they rest on a reconstructed assemblage known mostly by fragmentary remains. They reinforce, however, the ecological oddities already noticed for this mid-Cretaceous vertebrate ecosystem in North Africa.