Implications of Pulsed Electric Field Pre-Treatment on Goat Milk Pasteurization.

Goat milk is an interesting product from a nutritional and health standpoint, although its physico-chemical composition presents some technological challenges, mainly for being less stable than cow's milk at high temperatures. As pasteurization and ultra-high temperature processing are universally employed to ensure milk quality and safety, non-thermal methods, such as pulsed electric fields (PEFs), reduce the microbial load and eliminate pathogens, representing an interesting alternative for processing this product. This study demonstrates how the combined use of a PEF with short thermal processing and moderate temperature can be effective and energy-efficient in goat milk processing. A combination of thermal treatment at 63 °C after a low-intensity PEF (50 µs pulses, 3 Hz, and 10 kV·cm-1) caused the same reduction effect on the population of Listeria monocytogenes (goat's raw milk artificially spiked), as compared to a thermal treatment at 72 °C without a PEF. However, z values are significantly higher when PEF is used as a pre-treatment, suggesting that it may induce heat resistance in the survival population of L. monocytogenes. The sensitivity of L. monocytogenes to high temperatures is less pronounced in goat's milk than cow's milk, with a more pronounced impact of a PEF on lethality when combined with lower temperatures in goat's milk. The effect of a PEF on Escherichia coli viability was even more pronounced. It was also observed that thermal treatment energy needs with a PEF as a pre-treatment can be reduced by at least 50% of the total energy requirements.

The Impact of Goat Milk Pretreatment with Pulsed Electric Fields on Cheese Quality.

To reduce the microbial load in goat's milk, which is less thermally stable than cow's milk, an alternative processing method was used in this study. This involved treating the milk with pulsed electric fields (PEFs) (at 10 kV·cm-1, with 50 µs pulses for 3 Hz) and then heat-treating it at 63 °C for 6.0 s, as well as using heat treatment alone at 75 °C for 3.4 s. Cheeses were made using both types of milk treatment, and samples were collected after 5, 15, and 25 days of ripening for DNA extraction and purification, followed by high-throughput sequencing on the MiSeq Illumina sequencing platform. Analysis of the bacterial populations in the two types of cheese using various diversity indices revealed no significant differences in species richness and abundance, although there was a trend for the PEF-treated cheese to have a less diverse set of species with an uneven distribution of relative abundance. However, when examining the composition of the microbial communities in the two types of cheese using Weighted UniFrac analysis and Analysis of Similarities, there were significant differences in the presence and abundance of various species, which could have implications for the development of starter cultures. Concerning physicochemical properties (pH, aw, moisture content, total acidity and L, and a and b color parameters), the results also reveal that, generally, no significant differences were found, except for the color parameter, where cheeses treated with PEF demonstrated more whiteness (L) and yellowness (b) during ripening. Sensory scores for typicity (caprylic, goaty, and acetic) increased over time, but between treatments, only small differences were perceived by panellists in cheese with 5 days of ripening. Concerning texture firmness and cohesiveness, the PEF+HT samples presented lower values than the HT samples, even over storage time. In general, concerning quality parameters, similar behavior was observed between the treatments during the ripening period.

The Matrix Effect in the RT-PCR Detection of SARS-CoV-2 Using Saliva without RNA Extraction.

The present work focuses on the detection of SARS-CoV-2 in saliva, contributing to understanding the inhibition effect of the matrix and its influence on the results. Detection of viral genes ORF1ab, N, and E was performed by RT-PCR using saliva directly in the reaction without RNA extraction. Different amounts of saliva were spiked with increasing amounts of viral RNA from COVID-19 patients and subjected to RT-PCR detection. In parallel, 64 saliva samples from confirmed COVID-19 patients were used in two different amounts directly in the RT-PCR reaction and their results compared. The presence of saliva in the RT-PCR always causes a positive shift of the Ct values, but a very high between-person variability of its magnitude was obtained, with increases ranging from 0.93 to 11.36. Viral targets are also affected differently depending on the initial number of viral particles. Due to inhibitors present in saliva, the duplication of sample volume causes only 48 to 61% of the expected Ct value decrease depending on the viral target gene. The use of saliva has advantages, but also limitations, due to potential inhibitors present in the matrix. However, the choice of the target and the right amount of sample may significantly influence the results.

Impact of Thermal Pretreatment of Saliva on the RT-PCR Detection of SARS-CoV-2.

The use of saliva directly as a specimen to detect viral RNA by RT-PCR has been tested for a long time as its advantages are relevant in terms of convenience and costs. However, as other body fluids, its proven inhibition effect on the amplification reaction can be troublesome and compromise its use in the detection of viral particles. The aim of the present work is to demonstrate that saliva pretreatment may influence the RT-PCR amplification of three gene targets of SARS-CoV-2 significantly. A pool of RNA from confirmed COVID-19 patients was used to test the influence of heat pretreatment of saliva samples at 95°C for 5, 10, 15 and 20 min on the amplification performance of ORF1ab, E, and N SARS-CoV-2 genes. Prolonged heating at 95°C significantly improves the Ct value shift, usually observed in the presence of saliva, increasing the limit of detection of viral genes ORF1ab, E, and N. When tested using a cohort of COVID-19 patients' saliva, the increased time of heat pretreatment resulted in a significant increase in the detection sensitivity.

Performance of saliva as a specimen to detect SARS-CoV-2.

Massive testing to detect SARS-CoV-2 is an imperious need in times of epidemic but also presents challenges in terms of its concretization. The use of saliva as an alternative to nasopharyngeal swabs (NPS) has advantages, being more friendly to the patient and not requiring trained health workers, so much needed in other functions. This study used a total of 452 dual samples (saliva and NPS) of patients suspected of having COVID-19 to compare results obtained for the different specimens when using RT-PCR of RNA extracted from NPS and saliva, as well as saliva directly without RNA extraction. SARS-CoV-2 was not detected in 13 saliva (direct) of the 80 positive NPS samples and in 16 saliva (RNA) of a total of 76 NPS positive samples. Sensitivity of detection of viral genes ORF1ab, E and N in saliva is affected differently and detection of these genes in saliva samples presents great variability when NPS samples present Ct-values above approximately 20, with sensitivities ranging from 76.3% to 86.3%. On average an increase in 7.3 Ct-values (average standard deviation of 4.78) is observed in saliva samples when compared to NPS. The use of this specimen should be carefully considered due to the false negative rate and the system used for detection may be also very relevant since the different viral genes are affected differently in terms of detection sensitivity using saliva.

Fresh-Cut Bell Peppers in Modified Atmosphere Packaging: Improving Shelf Life to Answer Food Security Concerns.

The influence of modified atmosphere packaging (MAP, 10% O2 and 45% CO2) on the quality characteristics of fresh-cut green, red and yellow bell peppers (Capsicum annuum L. var annuum) was investigated. Packaging film bags (Krehalon MLF40-PA/PE) with fresh-cut bell peppers were stored for up to 17 days at 5 °C. The in-package O2 level ranged between 10 and 15%, respecting the current recommendations for fresh-cut vegetable products. Initial CO2 levels were higher than commonly used (from 5 to 10%), decreasing progressively over time due to the permeability of the selected polyethylene film. At the end of the storage period, they stabilized between 2 and 5%. A small variation in texture, moisture, titratable acidity, pH and microbial growth was observed during the storage period, as well as a good color retention and sensory properties maintenance. Negligible losses in the antioxidant activity and bioactive compounds (total phenol, flavonoid, anthocyanin and carotenoid content) were noted at the end of the study. Sensory analysis showed that panelists could not detect significant differences among sampling periods. A PCA with predictive biplots confirmed the existence of significant correlations. The products retain their initial characteristics without severe loss of quality until at least the 17th storage day. Given the current commercial shelf life of fresh-cut bell peppers, ranging from 9 to 14 days, the described treatment enabled an increase of at least 3 days (20%) of the products shelf life, reducing food waste and contributing to food security.