Detection and quantification of Bacillus cereus and its spores in raw milk by qPCR, and distinguish Bacillus cereus from other bacteria of the genus Bacillus

Introduction The raw milk is the basic raw material of dairy products, Bacillus cereus is a typical conditional pathogenic bacteria and cold-phagocytic spoilage bacteria in raw milk. This study established a qPCR method for detecting B. cereus in raw milk Materials and Methods In this study, a qPCR method for detecting B. cereus in raw milk was established. The specificity of the method was verified by using other Bacillus bacteria and pathogenic bacteria, the sensitivity of the method was evaluated by preparing recombinant plasmids and simulated contaminated samples, and the applicability of the method was verified by using pure spore DNA. The actual sample detection was completed by using the established qPCR method Results The qPCR established in this study can specifically detect B. cereus in raw milk. The LOD of the method was as low as 200 CFU/mL, and the LOQ ranged from 2 × 10 2 to 2 × 10 8 CFU/ml, the amplification efficiency of qPCR was 96.6% Conclusins The method established in this study can distinguish B. cereus from other Bacillus bacteria, and spore DNA can be used as the detection object. This method has the advantages of strong specificity, high sensitivity, wide application range and short detection time, which is expected to be applied in the dairy industry.

E-Gadget to Detect Food Freshness using IoT and ML

The food we consume plays an important role in our daily life. It provides us energy which is needed to work, grow, be active, and to learn and think. The healthy food is essential for good health and nutrition. Light, oxygen, heat, humidity, temperature and spoilage bacteria can all affect both safety and quality of perishable foods. Food kept at room temperature undergoes some chemical reactions after certain period of time, which affects the taste, texture and smell of a food. Consuming spoiled food is harmful for consumers as it can lead to foodborne diseases. This project aims at detecting spoiled food using appropriate sensors and monitoring gases released by the particular food item. Sensors will measure the different parameters of food such as pH, ammonia gas, oxygen level, moisture, etc. The microcontroller takes the readings from sensors and these readings then given as an input to a machine learning model which can decide whether the food is spoilt or not based on training data set. Also, we plan to implement a machine learning model which can calculate the lifespan of that food item. Index Terms: Arduino Uno, Food spoilage, IoT, Machine Learning, Sensors.

Mining marine metagenomes revealed a quorum-quenching lactonase with improved biochemical properties that inhibits the food spoilage bacteria Pseudomonas fluorescens

The marine environment presents great potential as a source of microorganisms that possess novel enzymes with unique activities and biochemical properties. Examples of such are the quorum-quenching (QQ) enzymes that hydrolyze bacterial quorum-sensing (QS) signaling molecules, such as N-acyl-homoserine lactones (AHLs). QS is a form of cell-to-cell communication that enables bacteria to synchronize gene expression in correlation with population density. Searching marine metagenomes for sequences homologous to an AHL lactonase from the phosphotriesterase-like lactonase (PLL) family, we identified new putative AHL lactonases (sharing 30-40% amino acid identity to a thermostable PLL member). Phylogenetic analysis indicated that these putative AHL lactonases comprise a new clade of marine enzymes in the PLL family. Following recombinant expression and purification, we verified the AHL lactonase activity for one of these proteins, named marine originated Lactonase Related Protein (moLRP). This enzyme presented greater activity and stability at a broad range of temperatures and pH, and tolerance to high salinity levels (up to 5M NaCl), as well as higher durability in bacterial culture, compared to another PLL member. The addition of purified moLRP to cultures of Pseudomonas fluorescens inhibited its extracellular protease activity, expression of the protease encoding gene, biofilm formation, and the sedimentation process in milk-based medium. These findings suggest that moLRP is adapted to the marine environment, and can potentially serve as an effective QQ enzyme, inhibiting the QS process in gram-negative bacteria involved in food spoilage. Importance Our results emphasize the potential of sequence and structure-based identification of new quorum-quenching (QQ) enzymes from environmental metagenomes, such as from the ocean, with improved stability or activity. The findings also suggest that purified QQ enzymes can present new strategies against food spoilage, in addition to their recognized involvement in inhibiting bacterial pathogen virulence factors. Future studies on the delivery and safety of enzymatic QQ strategy against bacterial food spoilage should be performed.

The Application of Bacillus subtilis for Adhesion Inhibition of Pseudomonas and Preservation of Fresh Fish

Inhibiting the growth of spoilage bacteria, such as Pseudomonas spp., is key to reducing spoilage in fish. The mucus adhesion test in vitro showed that the adhesion ability of Bacillus subtilis was positively correlated with its inhibition ability to Pseudomonas spp. In vivo experiments of tilapia showed that dietary supplementation with B. subtilis could reduce the adhesion and colonization of Pseudomonas spp. in fish intestines and flesh, as well as reduce total volatile basic nitrogen (TVB-N) production. High throughput and metabolomic analysis showed treatment with B. subtilis, especially C6, reduced the growth of Pseudomonas spp., Aeromonas spp., Fusobacterium spp., and Enterobacterium spp., as well as aromatic spoilage compounds associated with these bacteria, such as indole, 2,4-bis(1,1-dimethylethyl)-phenol, 3-methyl-1-butanol, phenol, and 1-octen-3-ol. Our work showed that B. subtilis could improve the flavor of fish by changing the intestinal flora of fish, and it shows great promise as a microecological preservative.

EFEKTIVITAS LARUTAN DAN PARUTAN KUNYIT DALAM MENGAWETKAN IKAN BANDENG (CHANOS CHANOS)

ABSTRACT In the content contained in milkfish such as protein and fat is a suitable medium for the growth of spoilage bacteria or other microorganisms. One way to maintain its quality by providing preservatives to inhibit bacterial growth is to provide natural preservatives such as turmeric.The purpose of this study was to determine the effectiveness of turmeric as a preservative for milkfish (chanos chanos). This research is a quasi-experimental type of research, namely the treatment of the use of turmeric grater and solution as a preservative in milkfish (chanos chanos) to determine the physical durability of milkfish.The results of the physical condition examination were seen from the appearance of the eyes, surface mucus, meat, body texture in the preservation of milkfish using 65 grams of turmeric solution, showing contact time 1 hour, 8 hours, 16 hours, 21 hours organoleptic value did not exceed the minimum limit of 7, but contact time 22 hours shelf life has been damaged. The results of physical condition examination on milkfish preservation samples using 65 gr turmeric grated effectively preserved at the contact time of 23 hours while the contact time of 24 hours has been damaged. While the milkfish without treatment have suffered damage in the span of 8 hours.The conclusion obtained is that grated turmeric is better able to preserve milkfish in a long time compared to using turmeric solution. Suggestion We recommend using turmeric natural preservatives with grated turmeric to preserve milkfish because the durability of using turmeric grated is longer compared to the turmeric solution seen from physical conditions.

Changes in microbial composition and quality characteristics of yellowfin tuna under different storage temperature

Yellowfin tuna is one of the commercially important fish varieties, and inappropriate storing may deteriorate its safety and quality. This study aimed to investigate the microbial composition and quality characteristics of yellowfin tuna stored at different temperatures for varying amounts of time. With an increase in the storage temperature and storage time, the biogenic amines, the total volatile basic nitrogen TVB-N, and the total viable cell count steadily increased, which influenced the quality of tuna. The most significant histamine concerning food safety reached levels of 21.25, 235.05, 1166.18, and 3799.29 mg/kg, respectively. The values of total viable cell counts were increased to 7.04, 7.97, 8.24, and 8.91 log CFU/g after storage at 0, 4, 10, and 20 °C for 12 days, 7 days, 7 days, 3 days, respectively. Additionally, changes in microbial composition were evaluated by high-throughput sequencing, and the results showed that Pseudomonas was the dominant spoilage bacteria in yellowfin tuna. The bacterial dynamics and their correlation with biogenic amines and TVB-N in yellowfin tuna were analyzed. A positive correlation between Pseudomonas, Shewanella, Morganella, Acinetobacter, and biogenic amines was found. Pseudomonas showed significant correlation with histamine, cadaverine, and putrescine. This study provides insights into yellowfin tuna quality and microbial composition, which provide theoretical guidance for maintaining seafood safety and quality during distribution and storage.

Assessment of the Spoilage Microbiota during Refrigerated (4 °C) Vacuum-Packed Storage of Fresh Greek Anthotyros Whey Cheese without or with a Crude Enterocin A-B-P-Containing Extract

Although fresh whey cheeses are prone to rapid deterioration, mainly by psychrotrophic Gram-negative bacteria and lactic acid bacteria (LAB), data on the specific spoilage species in traditional Greek whey cheeses are scarce. Therefore, this study quantified growth and characterized the primary spoilage bacteria in fresh Anthotyros whey cheeses stored at 4 °C in a vacuum for 40 days, without or with an added 5% (v/w) of an enterocin A-B-P crude extract (CEntE). Psychrotrophic Pseudomonas spp., Aeromonas spp., Hafnia spp. and Serratia spp. grew faster than LAB during early storage. However, LAB outgrew the Gram-negative bacteria and prevailed by mid to late storage in all cheese batches, causing a strong or milder batch-dependent natural acidification. Two major non-slime-producing and two minor biotypes of Leuconostoc-like bacteria, all identified as Leuconostoc mesenteroides by 16S rRNA sequencing, dominated the LAB association (76.7%), which also included four subdominant Carnobacterium maltaromaticum biotypes (10.9%), one Leuconostoc lactis biotype (3.3%) and few Lactococcus (1.6%), mesophilic Lactobacillus (0.8%) and Enterococcus (0.8%). Growth and distribution of LAB and Gram-negative species were strongly batch-dependent and plant-dependent. The CEntE neither retarded growth nor altered the whey cheese spoilage association but enhanced LAB growth and the declines of Gram-negative bacteria by late storage.

Antimicrobial Properties of Lyophilized Extracts of Olive Fruit, Pomegranate and Orange Peel Extracts against Foodborne Pathogenic and Spoilage Bacteria and Fungi In Vitro and in Food Matrices

Several novel antimicrobials with different concentrations of olive, pomegranate, and orange fruit pulp extracts were produced from agricultural byproducts and, after lyophilization, their antimicrobial activity and potential synergistic effects were evaluated in vitro and in food samples against foodborne pathogenic and spoilage bacteria and fungi. The Minimum Inhibitory of the tested bacteria was 7.5% or 10%, while fungi were inhibited at a concentration of 10% or above. The optical density of bacterial and yeast cultures was reduced to a different extent with all tested antimicrobial powders, compared to a control without antimicrobials, and mycelium growth of fungi was also restricted with extracts containing at least 90% olive extract. In food samples with inoculated pathogens and spoilage bacteria and fungi, the 100% olive extract was most inhibitory against E. coli, S. typhimurium, and L. monocytogenes in fresh burger and cheese spread samples (by 0.6 to 1.8 log cfu/g), except that S. typhimurium was better inhibited by a 90% olive and 10% pomegranate extract in burgers. The latter extract was also the most effective in controlling the growth of inoculated fungi (Αspergillus niger, Penicillium italicum, Rhodotorula mucilaginosa) in both yogurt and tomato juice samples, where it reduced fungal growth by 1–2.2 log cfu/g at the end of storage period. The results demonstrate that these novel encapsulated extracts could serve as natural antimicrobials of wide spectrum, in order to replace synthetic preservatives in foods and cosmetics.