Establishing a MALDI-TOF-TOF-MS method for rapid identification of three common Gram-positive bacteria (Bacillus cereus, Listeria monocytogenes, and Micrococcus luteus) associated with foodborne diseases
Abstract:We aimed to establish a method for the rapid identification of three common Gram-positive bacteria (Bacillus cereus, Listeria monocytogenes, and Micrococcus luteus) associated with foodborne diseases. MALDI-TOF-MS was used to determine the effects of sample pretreatment, culture medium, and culture time on the identification results. Then, MALDI-TOF-TOF-MS was used to establish an optimized method and further explore the effects of culture time on secondary proteins. Among the three sample pretreatment methods… Show more
“…Similar to other authors [ 30 ], the spectra were produced by applying the Compass Satellite software, and the Microflex LT device was used for fast and accurate identification of bacterial colonies isolated from the fruit samples. A spectra score of 2.300–3.000 indicates that the identification of bacterial species has high reliability, 2.000–2.299 indicates the identification of possible species, 1.700–1.999 indicates the identification of possible genus, and 0.000–1.699 indicates that the identification result is unreliable.…”
Introduction
The ripening of fleshy fruits is a developmental process that involves changes in color, texture, aroma, nutrients, and diversity of microbiomes. Some microorganisms, specially, bacteria and molds are responsible for postharvest spoilage of fruits. Thus, this study is aimed at evaluating the alterations in microbiome and physico-chemical properties of selected fruits at different ripening stages.
Methods
Totally, 108 fresh fruit samples of Musa paradisiaca, Citrus sinensis and Carica papaya at three ripening stages were collected and processed in this study. The biochemical methods and MALD-TOF MS were used in identification. The physico-chemical properties of all samples were analyzed using standard methods.
Results
The minimum counts (6.74± 0.48–6.76± 0.42 log CFU/mL) and the maximum count (7.51± 0.43–7.96± 0.34 log CFU/mL) of AMB in all fruit samples was observed at mature green and overripe stages of the fruits, respectively. The ripening stage has significantly affected the microbial counts (P < 0.05) in all fruits, except counts of Enterobacteriaceae in banana and orange, and fungal counts in orange. The bacterial community of all fruits was predominated by B. cereus (33.7%), A. faecalis (17.3%), P. putida (15.2%), M. morganii (11.1%), S. sciuri (6.6%) and S. epidermidis (4.9%); while the fungal microbiome was constituted by Candida spp. (33.9%) followed by Saccharomyces spp. (18.1%) and Aspergillus spp. (16.3%). The ripening stages have also significantly affected the physico-chemical property in all samples. Accordingly, the lowest pH (3.53) and highest content of ascorbic acid (69.87 mg/100g) were observed in mature green oranges and overripe papaya, respectively, while the maximum concentration of total sugar (17.87%) and reducing sugar (14.20%) were recorded in overripe bananas.
Conclusion
The presence of some potential human pathogens and spoilage microorganisms in fruit samples could contribute to post-harvest product losses besides the potential health risk associated with consumption of the tainted fruits. Hence, proper safety management practices and preservation mechanisms should be developed and put in place to ensure consumers safety against pathogens besides minimizing product losses through microbial spoilage.
“…Similar to other authors [ 30 ], the spectra were produced by applying the Compass Satellite software, and the Microflex LT device was used for fast and accurate identification of bacterial colonies isolated from the fruit samples. A spectra score of 2.300–3.000 indicates that the identification of bacterial species has high reliability, 2.000–2.299 indicates the identification of possible species, 1.700–1.999 indicates the identification of possible genus, and 0.000–1.699 indicates that the identification result is unreliable.…”
Introduction
The ripening of fleshy fruits is a developmental process that involves changes in color, texture, aroma, nutrients, and diversity of microbiomes. Some microorganisms, specially, bacteria and molds are responsible for postharvest spoilage of fruits. Thus, this study is aimed at evaluating the alterations in microbiome and physico-chemical properties of selected fruits at different ripening stages.
Methods
Totally, 108 fresh fruit samples of Musa paradisiaca, Citrus sinensis and Carica papaya at three ripening stages were collected and processed in this study. The biochemical methods and MALD-TOF MS were used in identification. The physico-chemical properties of all samples were analyzed using standard methods.
Results
The minimum counts (6.74± 0.48–6.76± 0.42 log CFU/mL) and the maximum count (7.51± 0.43–7.96± 0.34 log CFU/mL) of AMB in all fruit samples was observed at mature green and overripe stages of the fruits, respectively. The ripening stage has significantly affected the microbial counts (P < 0.05) in all fruits, except counts of Enterobacteriaceae in banana and orange, and fungal counts in orange. The bacterial community of all fruits was predominated by B. cereus (33.7%), A. faecalis (17.3%), P. putida (15.2%), M. morganii (11.1%), S. sciuri (6.6%) and S. epidermidis (4.9%); while the fungal microbiome was constituted by Candida spp. (33.9%) followed by Saccharomyces spp. (18.1%) and Aspergillus spp. (16.3%). The ripening stages have also significantly affected the physico-chemical property in all samples. Accordingly, the lowest pH (3.53) and highest content of ascorbic acid (69.87 mg/100g) were observed in mature green oranges and overripe papaya, respectively, while the maximum concentration of total sugar (17.87%) and reducing sugar (14.20%) were recorded in overripe bananas.
Conclusion
The presence of some potential human pathogens and spoilage microorganisms in fruit samples could contribute to post-harvest product losses besides the potential health risk associated with consumption of the tainted fruits. Hence, proper safety management practices and preservation mechanisms should be developed and put in place to ensure consumers safety against pathogens besides minimizing product losses through microbial spoilage.
“…LC-ESI-MS/MS is the main technology in microbiological research, unlike MALDI, which occupies the clinical microbiology laboratory market despite its low possibility to target specific protein biomarkers or to handle complex microbial samples [ 136 ]. MALDI-ToF/ToF MS, complementary to MALDI-ToF MS and LC-ESI-MS/MS techniques, is useful for characterization of protein patterns, protein biomarkers, and whole proteomes for pathogenic bacteria [ 19 ] as well as for identification of bacteria based on secondary protein peaks, improving the identification of foodborne pathogens, such as Bacillus cereus , Listeria monocytogenes , and Micrococcus luteus [ 137 ]. Furthermore, MALDI-ToF/ToF coupled with MALDI-MSI can assure the discrimination between isomeric compound in ion images [ 138 ].…”
Section: Applications Of Maldi-ms/ms-based Proteomicsmentioning
Matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS) is one of the most widely used techniques in proteomics to achieve structural identification and characterization of proteins and peptides, including their variety of proteoforms due to post-translational modifications (PTMs) or protein–protein interactions (PPIs). MALDI-MS and MALDI tandem mass spectrometry (MS/MS) have been developed as analytical techniques to study small and large molecules, offering picomole to femtomole sensitivity and enabling the direct analysis of biological samples, such as biofluids, solid tissues, tissue/cell homogenates, and cell culture lysates, with a minimized procedure of sample preparation. In the last decades, structural identification of peptides and proteins achieved by MALDI-MS/MS helped researchers and clinicians to decipher molecular function, biological process, cellular component, and related pathways of the gene products as well as their involvement in pathogenesis of diseases. In this review, we highlight the applications of MALDI ionization source and tandem approaches for MS for analyzing biomedical relevant peptides and proteins. Furthermore, one of the most relevant applications of MALDI-MS/MS is to provide “molecular pictures”, which offer in situ information about molecular weight proteins without labeling of potential targets. Histology-directed MALDI-mass spectrometry imaging (MSI) uses MALDI-ToF/ToF or other MALDI tandem mass spectrometers for accurate sequence analysis of peptide biomarkers and biological active compounds directly in tissues, to assure complementary and essential spatial data compared with those obtained by LC-ESI-MS/MS technique.
“…These bacterial strains were chosen in order to have a good representativity of bacterial morphologies, cell grouping and cell wall structures, which are important factors influencing resistance to antibacterial molecules. Moreover, the species studied are described as human pathogens regarding E. coli [18] and S. saprophyticus [19], or in food poisoning with B. subtilis [20] and M. luteus [21]. The first group (Table 1, Cn.1, entries 1-8) was chosen for its various functions led by the amino acids (R 1 and R 2 ), as well as the presence of 2 or no chlorides (R 3 and R 5 ).…”
Amino acid Schiff base Cu(II) complexes were synthesized under microwave irradiation using methanol as a solvent, to maximize the best conditions to obtain the attained compounds, containing aromatics possessing no, one or two chlorine atoms. The compounds’ antibacterial activities were tested against Gram-positive and Gram-negative bacteria, and the most active were tested for their antioxidant activities, and as E. coli, in particular, was found to be sensitive to these compounds, their interaction with this bacterium was investigated. It was found that, depending on the amino acid used for the formation of the Schiff base ligand, its LogPo/w mono-chlorinated or bis-chlorinated compounds are the most efficient against the tested bacteria.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.