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2.
Food Microbiol ; 87: 103383, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-31948624

RESUMO

Thermophilic and mesophilic lactic acid bacteria (LAB), such as Streptococcus thermophilus, Lactobacillus delbrueckii subsp. bulgaricus, Lactobacillus helveticus, and Lactococcus lactis, play a crucial role in the technological and sensory quality of Mozzarella cheese. In this study, the safety (genes encoding virulence factors and antibiotic resistance) and acidifying activity of autochthonous S. thermophilus cultures were evaluated in order to choose the most suitable strain for industrial application. The safe and good acidifying culture was tested in two buffalo Mozzarella cheese batches: Mozzarella cheeses produced with autochthonous culture (SJRP107) and commercial culture (STM5). The cultivable LAB was evaluated by culture-dependent method (plate counting) and the quantification of S. thermophilus cultures (commercial and autochthonous) were evaluated by culture-independent method RealT-qPCR (real-time quantitative polymerase chain reaction). The texture, physicochemical and proteolytic properties of the Mozzarella cheeses were similar for both batches. The nonstarter LAB count was higher during manufacture than in the storage, and the RealT-qPCR indicated the presence of S. thermophilus culture until the end of storage. S. thermophilus SJRP107 presented high potential for safety application in the production of Mozzarella cheese. Furthermore, considering the culture characteristics and their relationship with product quality, further studies could be helpful to determine their effect on the sensory characteristics of the cheese.


Assuntos
Queijo/microbiologia , Leite/microbiologia , Streptococcus thermophilus/crescimento & desenvolvimento , Streptococcus thermophilus/metabolismo , Animais , Búfalos , Queijo/análise , Qualidade de Produtos para o Consumidor , Fermentação , Microbiologia de Alimentos , Humanos , Streptococcus thermophilus/genética , Streptococcus thermophilus/isolamento & purificação , Paladar
3.
Food Microbiol ; 87: 103386, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-31948627

RESUMO

Contamination by Listeria monocytogenes is a particularly challenging problem in the food industry due to the ability of the bacterium to develop under conditions normally used for food preservation. Here, we show that the gaseous phase of Citrus limon var pompia leaf essential oil (hereafter PLEO) exerts specific anti-Listeria activity on ricotta salata cheese stored at 5 °C. The synergic effect of gaseous PLEO treatment and refrigeration was first confirmed in vitro on L. monocytogenes strains treated for 3 h with gaseous PLEO and then stored at 5 °C. Ricotta cheese was then inoculated with L. monocytogenes strains and subjected to hurdle technology with different concentrations of gaseous PLEO. Cell counts revealed gaseous PLEO to exert a bactericidal effect on L. monocytogenes 20600 DSMZ and a bacteriostatic effect on a mix of L. monocytogenes strains. Scanning and transmission electron microscopy analyses of L. monocytogenes cells suggested that gaseous PLEO targets the bacterial cell wall and plasma membrane. Chemical analyses of the liquid and vapor phases of PLEO indicated linalyl acetate to be the predominant compound, followed by limonene and the two isomers of citral, whereas EO composition analysis, although generally in line with previous findings, showed the presence of linalyl acetate for the first time. Solid-phase microextraction coupled with gas chromatography confirmed the presence of all crude oil components in the headspace of the box.


Assuntos
Antibacterianos/farmacologia , Queijo/microbiologia , Citrus/química , Listeria monocytogenes/efeitos dos fármacos , Óleos Voláteis/farmacologia , Óleos Vegetais/farmacologia , Antibacterianos/química , Membrana Celular/efeitos dos fármacos , Parede Celular/efeitos dos fármacos , Listeria monocytogenes/crescimento & desenvolvimento , Óleos Voláteis/química , Folhas de Planta/química , Óleos Vegetais/química
4.
Food Microbiol ; 87: 103392, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-31948633

RESUMO

Genetic diversity and metabolic properties of Lactococcus lactis subsp. lactis were explored using phylogenetic, pan-genomic and metatranscriptomic analysis. The genomes, used in the current study, were available and downloaded from the GenBank which were primarily related with microorganisms isolated from dairy products and secondarily from other foodstuffs. To study the genetic diversity of the microorganism, various bioinformatics tools were employed such as average nucleotide identity, digital DNA-DNA hybridization, phylogenetic analysis, clusters of orthologous groups analysis, KEGG orthology analysis and pan-genomic analysis. The results showed that Lc. lactis subsp. lactis strains cannot be sufficiently separated into phylogenetic lineages based on the 16S rRNA gene sequences and core genome-based phylogenetic analysis was more appropriate. Pan-genomic analysis of the strains indicated that the core, accessory and unique genome comprised of 1036, 3146 and 1296 genes, respectively. Considering the results of pan-genomic and KEGG orthology analyses, the metabolic network of Lc. lactis subsp. lactis was rebuild regarding its carbohydrates' metabolic capabilities. Based on the metatranscriptomic data during the ripening of the Swiss-type Maasdam cheese at 20 °C and 5 °C, it was shown that the microorganism performed mixed acid fermentation producing lactate, formate, acetate, ethanol and 2,3-butanediol. Mixed acid fermentation was more pronounced at higher ripening temperatures. At lower ripening temperatures, the genes involved in mixed acid fermentation were repressed while lactate production remained unaffected resembling to a homolactic fermentation. Comparative genomics and metatranscriptomic analysis are powerful tools to gain knowledge on the genomic diversity of the lactic acid bacteria used as starter cultures as well as on the metabolic activities occurring in fermented dairy products.


Assuntos
Metabolismo dos Carboidratos , Queijo/microbiologia , Lactococcus lactis/genética , Lactococcus lactis/metabolismo , Carboidratos/química , Fermentação , Microbiologia de Alimentos , Variação Genética , Genômica , Lactococcus lactis/classificação , Lactococcus lactis/isolamento & purificação , Filogenia
5.
Crit Rev Food Sci Nutr ; 60(1): 33-47, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-30285475

RESUMO

Cheese is a fermented dairy product, harboring diverse microbial communities (microbiota) that change over time and vary depending on the type of cheese and their respective starter and adjunct cultures. These microorganisms play a crucial role in determining the flavor, quality and safety of the final product. Exploring the composition of cheese microbiota and the underlying molecular mechanisms involved in cheese ripening has been the subject of many studies. Recent advances in next generation sequencing (NGS) methods and the development of sophisticated bioinformatics tools have provided deeper insights into the composition and potential functionality of cheese microbiota far beyond the information provided by culture-dependent methods. These advances, which include rRNA gene amplicon sequencing and metagenomics, have been complemented and expanded in recent years by the application of metatranscriptomics, metaproteomics and metabolomics. This paper reviews studies in which application of these meta-omics technologies has led to a better understanding of the microbial composition and functionality of cheese and highlights opportunities by which the integration of outputs from diverse multi-omics analytical platforms (cheesomics) could be used in the future to advance our knowledge of the cheese ripening process and identify biomarkers for predicting cheese flavor, quality, texture and safety, and bioactive metabolites with potential to influence human health.


Assuntos
Queijo/análise , Microbiologia de Alimentos , Microbiota , Queijo/microbiologia , Biologia Computacional , Metagenômica , Paladar
6.
Arch Virol ; 165(3): 715-718, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31873766

RESUMO

In the present study, we evaluated the degree of contamination of fresh vegetables, cheeses and jellies from disaster area in Brazil with bacteria and enteric viruses. Food samples (n = 350) were tested for Escherichia coli, Salmonella spp., Listeria monocytogenes, Staphylococcus spp., and enteric viruses (rotavirus A (RVA), human adenovirus (HAdV), hepatitis A virus (HAV), and human norovirus (HNoV). E. coli was present in 56% of the samples, Salmonella spp. was present in 14% of the samples, L. monocytogenes and Staphylococcus spp. (coagulase-positive) were present in 36% of the samples. The enteric viruses RVA and HAdV were detected in cheeses and vegetables.


Assuntos
Queijo/microbiologia , Contaminação de Alimentos/análise , Verduras/microbiologia , Adenovírus Humanos/isolamento & purificação , Brasil , Escherichia coli/isolamento & purificação , Fazendas , Vírus da Hepatite A/isolamento & purificação , Humanos , Listeria monocytogenes/isolamento & purificação , Norovirus/isolamento & purificação , Rotavirus/isolamento & purificação , Salmonella/isolamento & purificação , Staphylococcus/isolamento & purificação
7.
Food Microbiol ; 86: 103335, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31703853

RESUMO

The presence of eight common structural enterocin genes, singly or in varying combinations, in the genome of 15 antagonistic Enterococcus spp. previously isolated from artisan Greek Graviera and Galotyri retail cheeses was tested and associated with the mode of enterocin (Ent+) antilisterial activity of each isolate in three liquid culture media. The isolates were assigned to nine distinct strain genotypes of E. faecium (4 strains), E. durans (2) and E. faecalis (3). All strains were non-hemolytic, except for a cyl-positive E. faecalis genotype isolated from Galotyri cheese, which was strongly listericidal. All other strains varied from being listeriostatic to weakly listericidal in MRS and M17 broth, whereas all failed to inhibit listerial growth in skim milk. Two E. faecium strains retained strong Ent+ activity following neutralization and filter-sterilization of their MRS or M17 co-culture supernatants, whereas, all others required contact or proximity of their viable cells with L. monocytogenes cells in order to display activity. Additional studies to evaluate safety and potential synergistic effects of each strain genotype with starter LAB species in real milk environments will reveal the most active and truly harmless Enterococcus genotypes to be applied as co-starter or bioprotective adjunct cultures in traditional Greek cheese technologies.


Assuntos
Queijo/microbiologia , Enterococcus/química , Listeria monocytogenes/efeitos dos fármacos , Leite/microbiologia , Animais , Hidrocarbonetos Aromáticos com Pontes/química , Hidrocarbonetos Aromáticos com Pontes/metabolismo , Hidrocarbonetos Aromáticos com Pontes/farmacologia , Bovinos , Meios de Cultura/química , Meios de Cultura/metabolismo , Enterococcus/genética , Enterococcus/isolamento & purificação , Enterococcus/metabolismo , Grécia , Listeria monocytogenes/crescimento & desenvolvimento
8.
Food Microbiol ; 86: 103314, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31703869

RESUMO

The aim of this study was to investigate the antibacterial effect of 460-470 nm light-emitting diode (LED460-470nm) illumination against pathogens and spoilage bacteria on the surface of agar media and packaged sliced cheese. LED460-470nm illumination highly inhibited the growth of Listeria monocytogenes and Pseudomonas fluorescens on agar media covered with oriented polypropylene (OPP) film (thickness, 0.03 mm). When sliced cheeses inoculated with L. monocytogenes or P. fluorescens and packaged with OPP film were illuminated by an LED460-470 nm at 4 or 25 °C, reduction levels of L. monocytogenes and P. fluorescens on packaged slice cheese were higher at 4 °C than at 25 °C. There were no significant differences in color between non-illuminated and illuminated sliced cheese after storage for 7 d at 4 °C. LED460-470 nm illumination at 4 °C for 4 d caused cellular injury of L. monocytogenes and P. fluorescens related to RNA, protein, and peptidoglycan metabolism, and a disruption of the cell membrane and loss of cytoplasmic components were observed from TEM results. These results suggest that LED460-470 nm illumination, in combination with refrigeration temperatures, may be applied to extend the shelf-life of packaged slice cheese and minimize the risk of foodborne disease, without causing color deterioration.


Assuntos
Queijo/microbiologia , Conservação de Alimentos/métodos , Listeria monocytogenes/efeitos da radiação , Pseudomonas fluorescens/efeitos da radiação , Contaminação de Alimentos/análise , Contaminação de Alimentos/prevenção & controle , Luz , Pseudomonas fluorescens/crescimento & desenvolvimento
9.
Food Microbiol ; 86: 103348, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31703879

RESUMO

The effects of the incorporation of the essential oils from Origanum vulgare L. (OVEO; 0.07 µL/g) and Rosmarinus officinalis L. (ROEO; 2.65 µL/g) in combination in Minas Frescal cheese on the counts of the probiotic Lactobacillus acidophilus LA-5 and Escherichia coli O157:H7 were evaluated during refrigerated storage (7 ±â€¯0.5 °C). The terpenes of OVEO and ROEO, survival of the probiotic strain during in vitro digestion, as well as the physicochemical and sensory aspects were also monitored in Minas Frescal cheese. All terpenes decreased in cheese when the storage time increased. The incorporation of OVEO and ROEO delayed the increase in L. acidophilus LA-5 counts in cheese, but did not affect its ability to survive in cheese under simulated gastrointestinal conditions. The decreases in counts of E. coli O157:H7 observed in the first 15 days of refrigerated storage were strongly correlated (r ≥ 0.82) with the terpenes detected in cheese. Scores attributed for aroma, flavor, overall impression and purchase intention of cheese with OVEO and ROEO increased with the increase of the storage time. The incorporation of OVEO and ROEO in combination could be a strategy to control E. coli O157:H7 in probiotic Minas cheese during storage; however, the amounts of these substances should be cautiously selected considering possible negative sensory impacts in this product.


Assuntos
Queijo/microbiologia , Escherichia coli O157/crescimento & desenvolvimento , Aditivos Alimentares/análise , Lactobacillus acidophilus/crescimento & desenvolvimento , Óleos Voláteis/análise , Origanum/química , Óleos Vegetais/análise , Rosmarinus/química , Queijo/análise , Escherichia coli O157/efeitos dos fármacos , Aditivos Alimentares/farmacologia , Humanos , Lactobacillus acidophilus/efeitos dos fármacos , Viabilidade Microbiana/efeitos dos fármacos , Óleos Voláteis/farmacologia , Óleos Vegetais/farmacologia , Paladar
10.
Food Microbiol ; 85: 103286, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31500709

RESUMO

Hispanic style soft non-fermented cheeses, such as queso fresco (QF) have been linked to outbreaks and recalls. Salmonella is one of the main causes of these incidents. Due to lack of ripening or post-processing antimicrobial treatments, incorporating GRAS antimicrobials to production process may be a suitable approach to minimize microbial risk in QF. The aim of this study was to evaluate the efficiency of nisin (N), caprylic acid (CA) and trans-cinnamaldehyde (CN) as single or combined treatments to reduce Salmonella populations in QF during storage. Batches of QF were inoculated after curding with approx. 4 Log CFU/g of 5-strain cocktails of Salmonella and stored at 8 °C for 20 days. The final Salmonella counts in control samples ranged from 6.96 to 7.14 Log CFU/g. Application of CN at 0.6 g/kg inhibited Salmonella growth during storage, resulting in at least 3 Log CFU/g difference with the untreated controls (p < 0.05). Addition of N (0.5 g/kg) and CA (0.4 g/kg) with CN (0.3 and 0.6 g/kg) further enhanced the antimicrobial activity resulting in complete suppression of growth and even caused a 1 Log CFU/g reduction by the end of the experimental period compared to initial counts. Samples treated with the combined treatment (N, CA, CN) were evaluated in a consumer panel (n = 112). Participants preferred the control and commercial QF to the treated samples. However, treated samples with 0.3 g/kg CN were still within the acceptable range of neutral to like slightly. Results obtained, revealed that combined treatment of N, CA and CN can provide a solution to reduce the count of Salmonella in QF, whether in process or during storage.


Assuntos
Antibacterianos/farmacologia , Queijo/microbiologia , Microbiologia de Alimentos/métodos , Viabilidade Microbiana/efeitos dos fármacos , Salmonella/efeitos dos fármacos , Caprilatos/farmacologia , Contagem de Colônia Microbiana , Manipulação de Alimentos , Conservação de Alimentos , Nisina/farmacologia
11.
Food Microbiol ; 85: 103283, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31500718

RESUMO

Many countries use Escherichia coli and coliforms as indicators of sanitary quality of foods and have set limits for cheeses, including raw-milk cheeses. This paper reviewed the scientific literature for E. coli and coliform levels that are found in different types of raw milk, the fate of indicators during the manufacturing and ripening of different cheeses and the indicator levels that have been found in the finished cheeses. These studies from worldwide showed that E. coli and coliforms are found in different types of raw milk but usually at <100 CFU/ml or not found. Instances where raw milk contained indicator levels >1000 CFU/ml have mostly been attributed to unsanitary conditions/production. During cheese-making, indicators present in raw milk will often increase in numbers, but the levels decline as the acidity from lactose fermentation decreases the pH. Except for fresh cheeses that are not aged, indicator levels are further reduced by 2-3 log10 CFU/g or more, during the ripening process. As a result, indicator levels in finished cheeses are often low and within the limits of <10 or <100 CFU/g set by many countries. The cited studies also show that raw milk cheeses that are made with quality raw milk, under hygienic conditions and properly aged, should not contain high levels of indicator bacteria in the final product.


Assuntos
Bactérias/crescimento & desenvolvimento , Queijo/microbiologia , Microbiologia de Alimentos/métodos , Microbiologia de Alimentos/normas , Alimentos Crus/microbiologia , Animais , Queijo/normas , Contagem de Colônia Microbiana , Qualidade de Produtos para o Consumidor , Escherichia coli/crescimento & desenvolvimento , Contaminação de Alimentos/análise , Manipulação de Alimentos , Leite/microbiologia
12.
J Dairy Sci ; 103(1): 150-160, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31668441

RESUMO

This study aimed to evaluate the possible inhibitory effect of natural lactic acid bacteria on the growth of 2 Bacillus cereus strains. First, we evaluated the behavior of spores of B. cereus GPe2 and D43 when inoculated before cheesemaking using pasteurized or raw milk; no statistical differences were observed between cheese produced with the 2 types of milk. Then, lactic acid bacteria (LAB) were isolated from cheese at the last sampling time, identified, and tested in vitro for their antagonistic activity and organic acid production by using an HPLC method, showing antimicrobial potential. The LAB that produced larger inhibition halos (>9 mm) against B. cereus strains (LAB 3, 6, 9, 10: Lactococcus lactis ssp. lactis; LAB 7: Lactococcus lactis ssp. cremoris) were selected to produce a LAB mixture for subsequent tests. Spores of B. cereus GPe2 and D43 were inoculated in pasteurized milk before cheesemaking with or without addition of the LAB mixture at a high dosage. Bacillus cereus grew more slowly when LAB were added to the dairy matrix (with differences from 2.36 to 2.66 log cfu/g in B. cereus GPe2 and D43 growth).


Assuntos
Bacillus cereus/crescimento & desenvolvimento , Queijo/microbiologia , Microbiologia de Alimentos , Lactobacillales/fisiologia , Lactococcus lactis/fisiologia , Leite/microbiologia , Animais , Ácidos Carboxílicos
13.
J Dairy Sci ; 103(1): 128-140, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31677843

RESUMO

The dairy farm environment influences the raw milk microbiota and consequently affects milk processing. Therefore, it is crucial to investigate farm management practices such as the bedding materials. The aim of this study was to evaluate the effect of recycled manure solids (RMS) as bedding material on bulk tank milk and microbiological implications for cheese quality. Bulk tank samples were collected from 84 dairy farms using RMS or straw bedding. The use of RMS did not influence thermophilic and mesophilic aerobic viable counts from spores. However, straw-milk samples gave higher values for mesophilic anaerobic spore-forming bacteria (0.44 log cfu/mL) than RMS-milk samples (0.17 log cfu/mL). The presence of thermoresistant lactic acid bacteria was not increased in milk from farms using RMS. Nevertheless, taxonomic profiles of thermoresistant bacteria isolated were different between the 2 types of milk. More Enterococcus faecalis and Streptococcus spp. were identified in RMS-milk samples. Thermoresistant enterococci and streptococci could easily end up in cheese. Therefore, milk proteolytic activities of these isolates were tested. Neither Streptococcus spp. nor Enterococcus faecium isolates exhibited proteolytic activities, whereas 53% of E. faecalis showed some. Also, only 1 vancomycin-resistant enterococcus was detected. Survival of selected RMS-milk samples isolates (3 E. faecalis and 1 Streptococcus thermophilus) was evaluated during a model Cheddar cheese manufacture. Although those strains survived well, they did not modify the acidification curve of milk. However, they might cause organoleptic defects during cheese maturing.


Assuntos
Bactérias/classificação , Roupas de Cama, Mesa e Banho/veterinária , Queijo/normas , Leite/microbiologia , Animais , Bactérias/isolamento & purificação , Roupas de Cama, Mesa e Banho/microbiologia , Queijo/microbiologia , Enterococcus/classificação , Enterococcus/isolamento & purificação , Fazendas , Microbiologia de Alimentos , Esterco/microbiologia , Reciclagem , Streptococcus/classificação , Streptococcus/isolamento & purificação , Termotolerância
14.
J Dairy Sci ; 103(2): 1238-1249, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31864732

RESUMO

Cheese is a fermented dairy product that is popular for its unique flavor and nutritional value. Recent studies have shown that microorganisms in cheese play an important role in the fermentation process and determine the quality of the cheese. We collected 12 cheese samples from different regions and studied the composition of their bacterial communities using PacBio small-molecule real-time sequencing (Pacific Biosciences, Menlo Park, CA). Our data revealed 144 bacterial genera (including Lactobacillus, Streptococcus, Lactococcus, and Staphylococcus) and 217 bacterial species (including Lactococcus lactis, Streptococcus thermophilus, Staphylococcus equorum, and Streptococcus uberis). We investigated the flavor quality of the cheese samples using an electronic nose system and we found differences in flavor-quality indices among samples from different regions. We found a clustering tendency based on flavor quality using principal component analysis. We found correlations between lactic acid bacteria and the flavor quality of the cheese samples. Biodegradation and metabolism of xenobiotics, and lipid-metabolism-related pathways, were predicted to contribute to differences in cheese flavor using Phylogenetic Investigation of Communities by Reconstruction of Unobserved States (PICRUSt). This preliminary study explored the bacterial communities in cheeses collected from different regions and their potential genome functions from the perspective of flavor quality.


Assuntos
Bactérias/isolamento & purificação , Queijo/microbiologia , Variação Genética , Bactérias/classificação , Bactérias/genética , Queijo/análise , DNA Bacteriano/análise , Microbiologia de Alimentos , Lactobacillales/genética , Lactobacillales/isolamento & purificação , Lactobacillus/genética , Lactobacillus/isolamento & purificação , Lactococcus lactis/genética , Lactococcus lactis/isolamento & purificação , Filogenia , Análise de Sequência de DNA , Streptococcus/genética , Streptococcus/isolamento & purificação , Streptococcus thermophilus/genética , Streptococcus thermophilus/isolamento & purificação
15.
J Agric Food Chem ; 67(49): 13684-13693, 2019 Dec 11.
Artigo em Inglês | MEDLINE | ID: mdl-31742396

RESUMO

A novel BL312 milk-clotting enzyme (MCE) exhibited high-level expression and remarkable milk-clotting activity (MCA) (865 ± 20 SU/mL) that was 3.3-fold higher than the control by optimizing induction conditions in recombinant Escherichia. coli harboring pET24a-proMCE. Through substrate-binding region analyses and modification, MCE-G165A was identified from nine mutants and showed a proteolytic activity of 49.4 ± 2.4 U/mL and an MCA/PA ratio of 18.2, which were respectively 1.9-fold lower and 2.0-fold higher than those of the control. The purified MCE-G165A (28 kDa) exhibited weak αs-casein, ß-casein, and strong κ-casein (κ-CN) hydrolysis levels as assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and reversed-phase high-performance liquid chromatography. The milk-clotting mechanism for MCE-G165A was the primary hydrolysis of Met106-Ala107 and Asn123-Thr124 bonds in κ-CN, as determined by mass spectrometry. MCE-G165A showed different hydrolysis sites in casein, leading to various functional peptides. Feasible methods for obtaining MCEs suitable as calf rennet substitutes are presented.


Assuntos
Bacillus licheniformis/enzimologia , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Caseínas/química , Quimosina/química , Quimosina/genética , Leite/química , Sequência de Aminoácidos , Animais , Proteínas de Bactérias/metabolismo , Caseínas/metabolismo , Bovinos , Queijo/análise , Queijo/microbiologia , Quimosina/metabolismo , Hidrólise , Engenharia de Proteínas , Proteólise , Alinhamento de Sequência
16.
J Dairy Sci ; 102(12): 10850-10854, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31606207

RESUMO

The aim of this study was to quantify, identify, evaluate antimicrobial resistance, and characterize the virulence factors of enteropathogenic (EPEC), Shiga-toxigenic (STEC), and enterohemorrhagic (EHEC) Escherichia coli in raw milk (RM) and legal (LMFC) and illegal (IMFC) Minas Frescal cheeses in southern and northeast Brazil. Illegal cheeses are those made without official inspection service or sanitary surveillance. We evaluated samples of RM produced in Paraná (southern) and Maranhão (northeast) States, LMFC produced using pasteurized milk in inspected industries, and IMFC potentially produced with raw milk. Mean total coliform counts were 8.4 × 104 cfu/mL for RM, 1.4 × 107 cfu/mL for LMFC, and 2.9 × 107 cfu/mL for IMFC. Mean E. coli counts were 2.4 × 103 cfu/mL for RM, 1.9 × 102 cfu/mL for LMFC, and 1.1 × 105 cfu/mL for IMFC. Among the 205 E. coli isolates from RM, 9.75% were identified as EPEC, mainly (90%) in samples from Paraná. Of the total isolates from the cheese samples, 97.4% (n = 111) came from IMFC, of which 1.8 and 2.7% were identified as EPEC and STEC, respectively; no EHEC was detected. The phylogenetic group A (60%) and typical EPEC (68%) predominated, which confirms the possible human origin of pathogenic isolates in RM and IMFC. Of these, 50% were resistant to at least one antibiotic, and streptomycin was the antimicrobial with the highest number (8) of EPEC and STEC resistant isolates. This study reports the first isolation of serogroup O28ac in Brazilian milk. We found no predominance of a specific serogroup of EPEC or STEC in milk or cheese or clonal isolates in the same sample, indicating different origins of the contamination in these products, presumably mostly related to poor hygienic handling.


Assuntos
Antibacterianos/farmacologia , Queijo/microbiologia , Escherichia coli/efeitos dos fármacos , Leite/microbiologia , Animais , Brasil , Bovinos , Farmacorresistência Bacteriana , Escherichia coli/isolamento & purificação , Humanos , Filogenia , Fatores de Virulência/análise
17.
Pesqui. vet. bras ; 39(10): 807-815, Oct. 2019. tab, ilus
Artigo em Inglês | LILACS, VETINDEX | ID: biblio-1056902

RESUMO

The most consumed cheese in Brazil, Minas Frescal cheese (MFC) is highly susceptible to microbial contamination and clandestine production and commercialization can pose a risk to consumer health. The storage of this fresh product under refrigeration, although more appropriate, may favor the growth of spoilage psychrotrophic bacteria. The objective of this study was to quantify and compare Pseudomonas spp. and other psychrotrophic bacteria in inspected and non-inspected MFC samples, evaluate their lipolytic and proteolytic activities and their metalloprotease production potentials. Twenty MFC samples were evaluated: 10 inspected and 10 non-inspected. Counts of psychrotrophic bacteria and Pseudomonas spp., evaluation of the proteolytic and lipolytic potential of the isolates, and identification of potential producers of alkaline metalloprotease (AprX) were assessed. The mean total psychrotrophic counts were 1.07 (±2.18) × 109CFU/g in the inspected samples and 4.5 (±5.86) × 108CFU/g in the non-inspected, with no significant difference (p=0.37). The average score of Pseudomonas spp. was 6.86 (±18.6) × 105 and 2.08 (±3.65) × 106 CFU/g for the inspected and non-inspected MFC samples, respectively, with no significant difference (p=0.1). Pseudomonas spp. represented 0.06% and 0.004% of psychrotrophic bacteria found in inspected and non-inspected MFC samples, respectively. Collectively, 694 psychrotrophic strains and 47Pseudomonas spp. were isolated, of which 59.9% and 68.1% were simultaneously proteolytic and lipolytic, respectively. Of the 470 psychrotrophs isolated from inspected and 224 from non-inspected cheese samples, 5.74% and 2.23% contained aprX, respectively, while 100 and 86.96% of the Pseudomonas spp. isolates in inspected and non-inspected cheese samples contained the gene. The production potential of AprX did not, however, determine the proteolytic activity on plates of these isolates under the conditions evaluated in this study. Of total, 65.63% of the psychrotrophs that contained aprX gene were confirmed as Pseudomonas spp., using genus-specific PCR. Phylogenetic analysis of the 16S rRNA gene of the other psychrotrophs that were potential producers of AprX identified them as Serratia spp. (n=7), Raoultella ornithinolytica (n=1), and Acinetobacter schindleri (n=1) in the inspected samples and Psychrobacter sanguinis (n=1) and Leuconostoc mesenteroides (n=1) in the non-inspected samples. The production conditions of Brazilian MFC of these samples, while meeting the legal determinations, are not sufficient to control Pseudomonas and other spoilage-related psychrotrophs. Thus, stricter hygienic measures are required during the formal production of this type of cheese.(AU)


O mais consumido no Brasil, o queijo Minas Frescal (QMF) é altamente suscetível à contaminação microbiana e a produção e comercialização clandestina podem representar um risco para a saúde do consumidor. O armazenamento deste produto fresco sob refrigeração, embora mais apropriado, pode favorecer a multiplicação de bactérias psicrotróficas deteriorantes. O objetivo deste estudo foi quantificar e comparar Pseudomonas spp. e outras bactérias psicrotróficas em amostras de QMF inspecionadas e não inspecionadas, avaliar o potencial lipolítico, proteolítico e de produção de metaloprotease alcalina. Vinte amostras de QMF foram avaliadas: 10 inspecionadas e 10 não inspecionadas. Foram avaliadas as contagens de bactérias psicrotróficas e Pseudomonas spp., o potencial proteolítico e lipolítico dos isolados e a identificação de potenciais produtores de metaloprotease alcalina (AprX). A média total das contagens de bactérias psicrotróficas foi de 1,07 (±2,18) × 109UFC/g nas amostras inspecionadas e 4,5 (±5,86) × 108UFC/g nas não inspecionadas, sem diferença significativa (p=0,37). A média de Pseudomonasspp. foi de 6,86 (±18,6) × 105 e 2,08 (±3,65) × 106UFC/g para as amostras QMF inspecionadas e não-inspecionadas, respectivamente, sem diferença significativa (p=0,1). Pseudomonas spp. representaram 0,06% e 0,004% de bactérias psicrotróficas encontradas em amostras QMF inspecionadas e não-inspecionadas, respectivamente. Das amostras inspecionadas e não inspecionadas, foram isoladas 694 colônias psicrotróficas e 47 Pseudomonasspp., dos quais 59,9% e 68,1% foram simultaneamente proteolíticos e lipolíticos, respectivamente. Dos 470 isolados de psicrotróficos das amostras de queijo inspecionados e dos 224 isolados das não inspecionadas, 5,74% e 2,23% continham o gene aprX, respectivamente, enquanto 100 e 86,96% das Pseudomonasspp. isoladas em amostras de queijo inspecionadas e não inspecionadas continham o potencial de expressão de AprX. Esse potencial, no entanto, não determinou a atividade proteolítica em placas desses isolados nas condições avaliadas neste estudo. Do total, 65,63% dos psicrotróficos que continham o gene aprX foram confirmados como Pseudomonasspp., utilizando PCR gênero-específico. A análise filogenética do gene 16S rRNA dos outros psicrotróficos que foram produtores potenciais de AprX os identificou como Serratia spp. (n=7), Raoultella ornithinolytica (n=1) e Acinetobacter schindleri (n=1) nas amostras inspecionadas e Psychrobacter sanguinis (n=1) e Leuconostoc mesenteroides (n=1) nas amostras não inspecionadas. As condições de produção do QMF dessas amostras, atendendo às determinações legais, não são suficientes para controlar a Pseudomonas e outros psicrotróficos relacionados à deterioração. Assim, medidas higiênicas mais rígidas são necessárias durante a produção formal deste tipo de queijo.(AU)


Assuntos
Pseudomonas/isolamento & purificação , Queijo/microbiologia , Controle de Qualidade
18.
J Dairy Sci ; 102(11): 9674-9688, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31477293

RESUMO

Listeria monocytogenes can survive and grow in a variety of environments, including refrigeration, making it difficult to control and highlighting the importance of optimizing control strategies against this pathogen. Listeria phages are attractive biocontrol agents because phages bind to specific wall teichoic acids (WTA) on the bacterial cell wall, inhibiting pathogens without disrupting the normal microbiota or structure of the food. Common stresses found on dairy products can affect cell wall composition and structure and subsequently affect the efficiency of control strategies that target the cell wall. The goal of this study was to determine the effect of a range of pH and temperatures on the effectiveness of a commercial phage cocktail treatment against several strains of L. monocytogenes in a cheese matrix. We developed a laboratory-scale cheese model that was made at different pH, treated with phage, and then inoculated with L. monocytogenes. Cheeses were incubated at 6, 14, or 22°C for 14 d, and bacterial counts were determined on d 1, 7, and 14. Our data show that phage treatment has a limited ability to reduce L. monocytogenes counts at each temperature tested; however, it was more effective on specific strains of L. monocytogenes when cheese was stored at higher temperatures. More specifically, the average counts of L. monocytogenes on phage-treated cheese stored at 22°C were significantly lower than those on phage-treated cheese stored at 6 or 14°C. Similarly, phage treatment was significantly more effective at inhibiting L. monocytogenes on cheese made at higher pH (6 and 6.5) compared with counts on cheese made at pH 5.5, where L. monocytogenes did not grow. Furthermore, serotype was found to affect the susceptibility of L. monocytogenes to phage treatment; serotype 1/2 strains showed significantly higher susceptibility to phage treatment than serotype 4b strains. Overall, our results suggest the importance of considering the efficacy of phage under conditions (i.e., temperature and pH) specific to a given food matrix when applying interventions against this important foodborne pathogen.


Assuntos
Bacteriófagos , Queijo/microbiologia , Microbiologia de Alimentos , Listeria monocytogenes/virologia , Animais , Carga Bacteriana , Humanos , Concentração de Íons de Hidrogênio , Análise dos Mínimos Quadrados , Listeria monocytogenes/classificação , Listeria monocytogenes/efeitos dos fármacos , Listeria monocytogenes/crescimento & desenvolvimento , Sorogrupo , Temperatura , Fatores de Tempo
19.
Photochem Photobiol Sci ; 18(11): 2707-2716, 2019 Nov 06.
Artigo em Inglês | MEDLINE | ID: mdl-31556891

RESUMO

In this study, the optimal parameters for the photodynamic inactivation (PDI) of Staphylococcus aureus in bacterial suspensions and in cheese were assessed using a water-soluble curcumin salt as the photosensitizer (PS). The in vitro study aimed at finding the optimal concentration and light dose to promote S. aureus photokilling. Four main groups were proposed: CONTROL (L-C-), LIGHT (L+C-), CUR (L-C+) and PDI (L+C+). A fixed light dose (LED, 450 ± 10 nm, 10 J cm-2) was applied using four different PS concentrations (0.75, 1.0, 1.5 and 3.0 mg mL-1). The dose also varied from 10-100 J cm-2 for a fixed concentration. High inactivation rates were observed for all light doses, with a maximum reduction of 7.58 log10 at 100 J cm-2 (p ≪ 0.05). Saturation of the PDI effect was observed after a 10 minute illumination time, as well as a slight decrease in the S. aureus population for increasing illumination times in the L+C- group. As an application, the concentration showing the best decontamination performance in vitro (0.75 mg mL-1) was applied to decontaminate cheese in loco. PDI in two types of coalho cheese, a rennet-coagulated cheese commonly consumed in Brazil, was investigated. The results showed no significant inactivation in unpasteurized cheese, but a 4.34 log10 reduction for t > 5 min in pasteurized specimens. In conclusion, the present PDI-catalyzed curcumin photosensitizer inactivated S. aureus at statistically significant levels in vitro, in pasteurized cheese, but not in unpasteurized specimens.


Assuntos
Queijo/microbiologia , Curcumina/farmacologia , Contaminação de Alimentos/prevenção & controle , Fármacos Fotossensibilizantes/farmacologia , Staphylococcus aureus/efeitos dos fármacos , Luz
20.
J Dairy Sci ; 102(12): 10790-10798, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31563304

RESUMO

Considering the growing consumption of artisanal foods worldwide, we aimed to evaluate the microbial safety of Serro artisanal cheese (SAC), produced in Minas Gerais State, Brazil. This cheese is produced with raw milk using 1 of 2 natural starter cultures: "pingo" and "rala." A total of 53 SAC samples (pingo = 8; rala = 45) were obtained from different farmers and subjected to conventional and molecular assays to detect and enumerate Listeria monocytogenes, Salmonella spp., coagulase-positive staphylococci (CPS), diarrheagenic Escherichia coli, Mycobacterium tuberculosis, and Brucella abortus. The SAC samples were also subjected to an ELISA to detect classical staphylococcal enterotoxins (CSE: SEA, SEB, SEC, SED, SEE) and to PCR assays to detect staphylococcal enterotoxin-related genes (sea, seb, sec, sed, see). Coagulase-positive staphylococci isolates were obtained and tested by the same assays to detect their potential in CSE production and presence of CSE-related genes. None of the SAC samples showed any of the screened food-borne pathogens and zoonotic agents, and none showed the presence of CSE by phenotypic and genotypic approaches. Despite the absence of microbial hazards, mean counts of CPS in SAC samples were 5.2 log cfu/g (pingo starter) and 4.6 log cfu/g (rala starter), indicating poor hygiene practices during production. None of the tested CPS isolates (n = 116) produced CSE or presented CSE-related genes. Despite the relative microbial safety, hygienic conditions during SAC production must be improved to meet official guidelines established in Brazil.


Assuntos
Queijo/microbiologia , Microbiologia de Alimentos , Inocuidade dos Alimentos , Animais , Brasil , Bovinos , Enterotoxinas/isolamento & purificação , Listeria monocytogenes/isolamento & purificação , Leite/microbiologia , Reação em Cadeia da Polimerase , Salmonella/isolamento & purificação , Staphylococcus/isolamento & purificação
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