Identification of lactic acid bacteria with anti-listeria activity. Characterization and application of a bacteriocinogenic strain in the control of Listeria monocytogenes in cheese.

The purpose of the research paper was firstly to identify bacteriocin-producing lactic acid bacteria characterizing strains with anti-listeria activity and, secondly, to characterize bacteriocin evaluating its in vitro efficiency as a natural preservative and, thirdly, to evaluate the anti-listeria effect of the bacteriocinogenic strain of Lactiplantibacillus plantarum in cheeses and produce an edible film with anti-listerial effect. Of 355 lactic acid bacteria strains tested, two were able to produce bacteriocin against Listeria monocytogenes and were identified as Lactiplantibacillus plantarum and Lactiplantibacillus pentosus. A bactericidal effect of strain QS494 (Lactiplantibacillus plantarum) was observed in the first 8 h, with a reduction of 1.7 log, using cell-free supernatant with Listeria monocytogenes, where viable cells were counted on listeria selective agar. Both strains showed good technological characteristics and were without production of virulence factors. Changes in the pH of the cell-free supernatant obtained from Lactiplantibacillus plantarum did not affect its antimicrobial activity, which remained stable after heat treatments for up to 15 min at 121°C. Inhibitory activity was also observed after 12 weeks of storage at -20°C. In the evaluation of the anti-listeria effect in cheeses, a 3 log reduction in the Listeria monocytogenes count was observed in 120 h in cheeses produced with bacteriocinogenic lactic acid bacteria, while in cheeses produced with non-bacteriocinogenic culture, we observed a 2 log increase in the count. Edible films produced with the addition of precipitate from the cell free supernatant showed an antimicrobial effect against Listeria monocytogenes. Thus, the two strains studied have technological and biosafety potential.

Pasteurized milk quality in Brazil: a cross-sectional study over five years.

This research communication delineates the quality of pasteurized cow milk sold in Brazil from 2015 to 2020. A cross-sectional study was performed gathering 1749 samples, which were evaluated for microbiological and physicochemical parameters, including Salmonella spp., total and thermotolerant coliforms, freezing point, alkaline phosphatase and lactoperoxidase. The proportion of compliant and non-compliant samples was compared through the years and jurisdiction of the inspection services. Interactions between the design and response variables were assessed by log-linear analysis. Overall, a considerable non-conformity rate (12%) was found for at least one microbiological or physicochemical parameter. Post-pasteurization contamination by coliforms was the major challenge for dairy industries. Notably, the non-compliance rate for freezing point increased during the SARS-CoV-2 pandemic. In addition, the ability to comply was linked to the type of inspection service. Thus, it is suggested that the SARS-CoV-2 pandemic is affecting the dairy industries in Brazil, and we strengthen the need for more studies monitoring the quality of milk over the years, which could assist industries and regulatory agencies to ensure the compliance of pasteurized milk.

A novel approach in public health surveillance: searching the illegal dairy trade in Facebook.

This paper uses the internet-based biosurveillance to describe the Brazilian e-commerce of non-inspected dairy products and maps its traditional sale in a medium-sized city (Londrina). We searched the sales of illegal dairy products on Facebook Marketplace and conducted a census of street markets and a random sample of formal markets. Although prohibited in Brazil, consumers can easily find informal dairy products on internet and retail sale. In addition, 7% of the food products hold a false label to mimic food inspection. The e-commerce was linked with regions with higher access to technology and Human Development Index (HDI). We believe that our approach shows a step forward in the public health interventions and could be replicated stimulating debates about policies required to improve the public health surveillance.

Authenticity under threat: grated Parmesan cheese sold in Brazil.

This research communication depicts the quality and authenticity of grated Parmesan cheese sold in Brazil, with novel emphasis on ripening time. The sampling included all brands manufactured and packed at Brazilian dairy plants under federal inspection and sold in Londrina between June and September 2017. A total of 24 samples were analyzed (3 batches of 8 brands) for microbiological and compositional characteristics, including urea-PAGE electrophoresis for the quantitative determination of the milk proteins. About 80% of the samples were insufficiently ripened and 32% with low-fat content. Faced with these economically motivated food frauds, the authenticity of grated Parmesan in Brazil may be under threat. It is high recommended that future surveys and regulatory agencies go further than microbiological assessment, including and emphasizing the ripening time as the main parameter of grated Parmesan cheese.

Pseudomonas fluorescens and Pseudomonas putida from refrigerated raw milk: genetic diversity and lipoproteolytic activity.

In this research communication the genetic diversity of Pseudomonas fluorescens (n = 67) and Pseudomonas putida (n = 44) isolated from refrigerated raw milk from bulk tank trucks were verified. The relationship between the genetic profile of the isolates and their lipoproteolytic potential was evaluated using skim milk agar and tributyrin agar (21°C/72 h). The lipoproteolytic potential (low or high), evaluated by the diameter of the halos (cm), was correlated with the number of milk producing properties that contributed to each sample (one sample = one bulk tank truck; 8-80 producers/sample) and the distance between the dairy properties and the processing plant (21-370 km). P. fluorescens was confirmed in all samples, while P. putida in 60% samples. For both species, two clusters (I and II) were observed, and the first one showed lower genotypic diversity and the presence of isolates with 100% similarity. P. fluorescens isolates presenting at least 70% similarity were 83.9% in Cluster I (n = 31) and 44.4% in Cluster II. In both clusters (I and II) observed in the P. fluorescens dendrogram, the occurrence of high proteolytic and lipolytic potential were equivalent. The higher the number of farms per milk sample, the greater the lipoproteolytic intensity of P. fluorescens isolates. In relation to P. putida isolates, 74% presented at least 50% similarity in Cluster I (n = 27) and only 35% in Cluster II (n = 17). The occurrence of high proteolysis linked to P. putida was proportional between both Clusters, but the occurrence of high lipolysis was greater in Cluster II. No significant association was detected between P. putida isolates and the variables studied. The results indicate the circulation of P. putida and P. fluorescens with 100% similarity in different milk producing regions. The level of genetic diversity was related only to the lipolytic capacity of P. putida.

Why knowledge is the best way to reduce the risks associated with raw milk and raw milk products.

In an age of flexible conditions about mandatory milk pasteurisation, this opinion-based research reflection supports the view that the knowledge and the awareness of milk-borne infections are key requirements to decrease the risks associated with raw milk. Providing an analysis of the current potential risks related to consumption of raw milk and raw milk products, we discuss the main reasons to continue to be vigilant about milk-borne pathogens and the current scenario in relation to the formal and clandestine sale of raw milk. Finally, we select some highly effective strategies to reduce the risks associated with raw milk in food services. Regardless of whether a country regulation allows or prohibits the trade of raw milk and its products, this is not the time to be negligent.

Development of a whey protein spread enriched with ß-glucan: an alternative for whey valorization.

BACKGROUND: Innovative approaches to combine whey with other ingredients and the use of new techniques in product development should be explored to meet consumers' needs and expectations. However, the question arises here of whether whey protein could be used as a suitable food matrix for supplementation with ß-glucan, an attractive glucose polymer and a physiologically functional component. The present study addresses the challenge associated with the design and characterization of whey protein spread as a substrate for ß-glucan delivery. The results are discussed on the basis of physical-chemical and microbiological characteristics, which are subsequently linked to its sensorial profile. RESULTS: A whey protein spread can be developed without the addition of NaCl, with physicochemical characteristics (pH, viscosity), microbiological counts, and sensory acceptance (color, aroma, overall impression) similar to the product with NaCl. This spread can be refrigerated for 28 days. The whey protein spread presented high whey protein content (18.67-19.17 g 100 g-1 ) and could be a good source of carbohydrates (8.30-8.68 g 100 g-1 ), with low levels of fat (0.2 g 100 g-1 ) and lactose (1.56-1.61 g 100 g-1 ). The sensorial results showed that women would prefer a product with lower salt content. CONCLUSION: This is the first study to evaluate the development of a whey protein spread enriched with ß-glucan, providing results that are of interest for the dairy sector. The combination of whey and ß-glucan can be explored industrially as a whey protein spread, with satisfactory results for physicochemical, microbiological, and sensory acceptance. © 2019 Society of Chemical Industry.

Psychrotrophs in raw milk: effect on texture, proteolysis index, and sensory evaluation of smoked provolone cheese.

BACKGROUND: Provolone can be fresh or ripened, with its taste varying from sweet to spicy. The high psychrotrophic density of raw milk is associated with thermoresistant enzymes that can change cheese characteristics such as texture, promote sensory defects and decrease industrial yield. Two batches of provolone-type smoked cheese were produced from chilled raw milk with 3 log cfu mL-1 (Treatment 1) and 7 log cfu mL-1 (Treatment 2) of psychrotrophs. The psychrotrophic (21 °C for 25 h) and physical-chemical profile of the raw milk were determined. Cheeses were evaluated by fat level, primary and secondary proteolysis index, yield, protein profile (Urea-polyacrylamide gel electrophoresis), and texture (hardness, cohesiveness, elasticity, and chewiness) at 14, 30, and 60 days of storage time. Sensorially, the cheeses were evaluated (100 tasters/period) using the triangular test. RESULTS: The treatments did not influence proteolysis index, although maturation influenced the proteolytic depth index after 60 days. The psychrotrophic population influenced αs 1- and ß-casein fractions, while maturation time influenced αs 1- and γ-casein fractions. Treatment 2 induced a 3% reduction in cheese yield. Hardness and chewiness showed a linear and positive relationship with the milk's psychrotrophic load. There was a significant difference in the fat content of the cheeses, with Treatment 2 having a lower level. The triangular test showed no difference between the cheeses. CONCLUSION: Although the larger psychrotrophic population in raw milk was associated with superior values of hardness and chewiness, as well as an increase in protein fractions indicating that proteolysis was observed, the tasters did not identify sensorial differences between the cheeses. © 2020 Society of Chemical Industry.

Validation of strip tests for the rapid screening of ethanol residues in milk.

The experiments reported in this Research Communication aimed to determine if a domestic strip test to detect ethanol residues in breast milk (Milkscreen®) is comparable to a previously established official method for detecting ethanol residues in milk. The two methods are examined in terms of overall sensitivity, robustness against storage and acidity and selectivity against formaldehyde residues. Here, Milkscreen® provided advantages, with faster results (2 min), good sensitivity (≥0.017%), no false results due formaldehyde residues and equal robustness against storage, but with lower sensitivity in acid milk samples. In summary, strip tests for the rapid detection of ethanol residues in breast milk can be used for screening purposes by dairy manufacturers, combining it with the official method to make a final diagnosis.

Lipoproteolytic capacity and potential of Pseudomonas spp. isolated from cold raw milk.

The objective of the work described in this research communication was to determine the lipoproteolytic capacity and potential of Pseudomonas spp. from the microbiota of refrigerated raw milk. The influence of temperature and bacterial population on these activities was also evaluated. Pseudomonas spp. (PS) counts (30 °C/48 h) were confirmed by PCR. Proteolytic (10% milk agar) and lipolytic capacities (PLC) (tributyrin agar) were evaluated (21 °C/72 h). Proteolytic (PP) and lipolytic potential (LP) were assessed by measuring the diameter of the halos and were categorized as low or high. A total of 91.3% PS possessed PLC. The PP of 64.16% isolates was high and was frequently observed in PS from milk samples with higher counts and lower temperatures. The LP of 70.52% isolates was low, and higher LP was associated with low microbiological counts and temperatures. Genetic studies evaluating Pseudomonas spp. strains in the milking environment and investigating the origin of these isolates could be useful to improve the quality and shelf life of dairy products.

Reduced-fat Frescal sheep milk cheese with inulin: a first report about technological aspects and sensory evaluation.

This research paper aimed to evaluate the role of inulin as a fat replacer on the quality of Frescal sheep milk cheese. Sheep milk and its derivatives are a promising niche in the dairy industry, mainly due to increasing interest of consumers in diversified products. Three Frescal sheep milk cheese formulations, namely whole milk cheese (WMC), semi-skimmed cheese (SSC) and semi-skimmed cheese with 5 g/100 g inulin (SSCI) were prepared. Their composition was evaluated and the feasibility of using inulin as a fat substitute was investigated. SSC and SSCI were considered 'reduced fat' or 'reduced calorie' products. The addition of inulin to SSCI cheeses yielded textural parameters (firmness, adhesiveness, cohesiveness, and gumminess) with intermediate characteristics between SSC and WMC. All the formulations presented scores higher than 7.6 in sensory analysis. In conclusion, the use of inulin in semi-skimmed sheep cheese allowed the production of cheese with texturizing properties similar to whole milk sheep cheese, enabling the development of a foodstuff with lower caloric content and beneficial characteristics valued by consumers.

X-ray diffraction has limited applicability in investigation of milk tampering.

The aim of this work was to use X-ray diffraction to identify substances used for adulteration of raw milk and to determine if crystallographic analysis can detect extraneous substances in milk. Two unknown substances were sent anonymously by employers linked to the dairy chain, who claimed that they were added directly in milk prior to water addition by truck drivers. The samples were analyzed by X-ray diffraction and submitted to physicochemical analysis. The first substance was identified by X-ray diffraction as sodium citrate, complying with its physicochemical attributes, such as the powerful ability to decrease the freezing point. The second substance was identified by X-ray diffraction as sucrose and this result was also in agreement with its ability to increase the density, decrease the freezing point and finally, to be positive for sucrose in the resorcinol qualitative test. To evaluate if X-ray diffraction can detect extraneous substances already mixed in milk, fresh raw milk samples tampered with urea, sodium hydroxide, sodium citrate and sucrose were freeze dried and analyzed by X-ray diffraction, with no detection of any extraneous substances at any percentage. This is the first report of attempted diagnosis of extraneous substances in milk by X-ray diffraction. However, this technique can be useful only when applied to identify substances used for adulteration prior to its dilution in milk, since the amorphous nature of milk seems to be a limitation for the accurate detection of extraneous substances.

Field findings about milk ethanol stability: a first report of interrelationship between α-lactalbumin and lactose.

BACKGROUND: Milk ethanol stability is not only associated with microbiological acidification, but is a phenomenon with many variables that influence the balance of soluble salts, mainly calcium ion activity. On this basis, we wanted to find out more about milk ethanol stability by studying its relationship with milk protein fractions and others major components. The influence of milk composition on ethanol stability was assessed through a predictive model comprising 180 individual raw milk samples. An additional model was used to assess the ethanol stability status as a response to the proteins fractions quantified by electrophoresis. RESULTS: Of the total samples, 68% were classified as stable and 32% as unstable to alcohol. Milk ethanol instability increased at low values of lactose content and high values of ash percentage. α-Lactalbumin (α-La) was also associated with ethanol stability, and the higher the α-La percentage the lower were the chances of ethanol instability. CONCLUSION: The lower values of α-La in unstable milk samples might be related to lower content of lactose, as α-La promotes lactose synthesis, a key component for the osmotic balance of milk and thus its ethanol stability. This is the first field report linking ethanol stability indirectly with α-La. © 2017 Society of Chemical Industry.

Milk adulteration with acidified rennet whey: a limitation for caseinomacropeptide detection by high-performance liquid chromatography.

BACKGROUND: High-performance liquid chromatography (HPLC) is widely employed to determine the caseinomacropeptide (CMP) index and to detect milk tampering with rennet whey. Prior to HPLC analysis, CMP is subject to a trichloracetic acid isolation, causing further soluble proteins in the sample to precipitate. On this basis, we aimed to determine whether rennet whey acidification could adversely affect the HPLC sensitivity with respect to detecting this peptide. RESULTS: As hypothesized, the CMP index from milk with added acidified rennet whey was, on average, half that quantified from milk with added rennet whey. Moreover, the quantum satis of acidified whey added to milk sufficient to demonstrate a HPLC CMP > 30 mg L-1 was 94% greater than that required for this threshold to be reached with rennet whey. CONCLUSION: Milk tampering with acidified rennet whey may limit the analytical sensitivity of the reversed-phase HPLC employed for the screening of CMP and, ultimately, disguise the fraudulent addition of whey to milk. © 2017 Society of Chemical Industry.

Identification and antimicrobial susceptibility of milk isolates from cows with subclinical mastitis in the northwest of Paraná State, Brazil.

Bovine mastitis is a multifactorial disease, caused mainly by bacteria, whose most traditional method of treatment is the use of antibiotics. However, incorrect administration of these drugs may lead to the emergence of resistant strains. The objective of this study was to identify the most common pathogens causing subclinical mastitis in the northwest State of Paraná, Brazil and establish their antimicrobial susceptibility profiles. A total of 507 foremilk samples were collected in 13 municipalities. Bacteria were identified and their profile of antimicrobial susceptibility were tested. The distribution of positive samples was not homogeneous among the cities. Streptococci were the most frequently genus observed (61%), followed by staphylococci (28%) and other genera (11%). Among streptococci and staphylococci, Streptococcus uberis and Staphylococcus aureus were the most frequently identified species, respectively. Overall, 40% of the isolates showed resistance to at least one of the antibiotics used in this study. A high proportion of cefoperazone resistant Staphylococcus spp. and bacitracin resistant Streptococcus were observed. The isolated microorganisms showed little resistance to enrofloxaxin. The results obtained show the importance of isolating the microorganism causing mastitis and testing its resistance to antibiotics before performing the treatment of the disease.

Etiological agents and bacterial sensitivity in subclinical mastitis in Brazil: a ten-year systematic review.

Considering the high prevalence of subclinical mastitis and its impacts on milk production, thematic studies are need to provide strategic data for its control. This study aimed at investigating the most frequent microorganisms associated with subclinical mastitis in dairy cows in Brazil through compiling the occurrence of the etiological agents and their sensitivity to antibiotics. The systematic review includes articles published between 2009 and 2019. Fiftyseven articles evaluating 22,287 milk samples were selected. The number of publications and the sample size were not homogeneous among Brazilian regions. Most of the studies and sampling were conducted in Rio Grande do Sul, whereas no studies were found in some states in the north and mid­west regions. The most frequent pathogen was Staphylococcus spp. It was isolated in all studies and had an average prevalence of 49% in the analyzed samples. Resistance to penicillin was the most frequent microbial resistance found in Brazil, with an average of 66% among the isolates evaluated. Moreover, bacterial resistance to cephalexin, cefoperazone, erythromycin, gentamicin, neomycin, penicillin, tetracycline, and trimethoprim increased over the research period. Given the territorial extension, the etiological diversity, and the lack of studies with a representative sample, the compilation of scientific data must be interpreted with caution. Regions where a greater number of studies were conducted and with numerous samples, such as the South, provided a comprehensive scenario that is closer to reality. Nevertheless, although decision making on the farm cannot be replaced by scientific studies, it can be supported by such efforts.

Listeria monocytogenes: Occurrence in beef and identification of the main contamination points in processing plants.

This study aimed to establish the occurrence of Listeria spp., especially L. monocytogenes and its main serotypes, in beef and processing plants. A total of 443 samples were obtained from equipment, installations and products from 11 meat processing establishments from Paraná state, Brazil. All samples were analyzed using USDA methodology for Listeria spp. detection, followed by species identification. The occurrence of Listeria spp. in the samples was 38.1% of which 51.4% were from equipment, 35.4% from installations and 30.2% from products. The identified species were: L. monocytogenes (12.6%), L. innocua (78.4%), L. seeligeri (1.2%), L. welshimeri (7.2%) and L. grayi (0.6%). The identified serotypes of L. monocytogenes were 1/2a and 4b. The results demonstrate the significance of equipment and installations as sources of contamination by Listeria spp. and L. monocytogenes in the processing of beef and meat products.

Blood Pressure, Serum Glucose, Cholesterol, and Triglycerides in Dogs with Different Body Scores.

The objective of this research was to determine the frequency for the occurrence of MS in dogs, using the criteria determined, and to correlate the criteria of dogs that would characterize the MS with different body condition score (BCS). 271 dogs with different body scores were studied, with 101 dogs with BCS 4-5; 101 dogs with BCS 6-7; and 69 dogs with BCS 8-9. Among the dogs studied, 62 (22,87%) had two or more inclusion criteria for MS. Of these, 28 had BCS 6-7, while 34 dogs had BCS 8-9. Therefore, 27,72% of overweight dogs had inclusion criteria for MS and 49,27% of obese ones had two or more inclusion criteria for MS. When only overweight and obese dogs were considered as a total population, it was observed that 36,47% got inclusion criteria for the MS. No dog with BCS 4-5 showed two or more inclusion criteria for MS. The metabolic syndrome, according to the parameters for inclusion defined in the literature, was observed in 22,87% of the animals studied and in 36% of dogs overweight or obese. Furthermore, MS was most common in obese (49%) compared to overweight dogs (27%).

Natural infection by endoparasites among free-living wild animals.

The objective of this study was to investigate the frequency of occurrence and variety of intestinal parasites among free-living wild animals. Fecal samples from wild mammals and birds at rehabilitation centers in the states of Mato Grosso do Sul and São Paulo were analyzed by sedimentation and flotation-centrifugation methods. Parasite eggs, oocysts, cysts and/or trophozoites were found in 71% of the samples. Cryptosporidium sp. oocysts were detected in fecal samples from oncillas (Leopardus tigrinus) and scaly-headed parrots (Pionus maximiliani). Giardia cysts were identified in the feces of a gray brocket (Mazama gouazoubira). Among the most common parasites found, there were eggs from Toxocara cati, Toxascaris leonina and Ancylostoma tubaeforme, and from Cestoda. Several Enterobius sp. eggs were found in the feces of red howler monkeys (Alouatta seniculus). It can be concluded from this study that despite the small number of samples, the diversity of parasites found was noteworthy. Additional information about parasite endofauna in wild animals is needed, since their presence might suggest that there could be proximity to and interactions with domestic animals and/or humans. In addition, further studies on parasites from free-living wild animals are of prime importance for understanding the intensity of anthropic changes in wild environments.

Hipocalcemia no deslocamento de abomaso de bovinos: estudo de 39 casos / Hypocalcemia in displacement the abomasum in cattle: study of 39 cases

Este estudo avaliou o status do cálcio sérico em 39 vacas com deslocamento de abomaso (DA), provenientes de 30 propriedades leiteiras selecionadas, na região de Campos Gerais no Paraná. O diagnóstico do deslocamento de abomaso foi realizado por percussão auscultatória, além dos sinais clínicos e informações da anamnese. Previamente ao procedimento cirúrgico, amostras de sangue foram coletadas para a mensuração dos níveis sanguíneos de cálcio, albumina, proteína e glicose. Como grupo controle, amostras de sangue foram coletadas de vacas hígidas que se encontravam em semelhante período de lactação. Das 39 vacas com deslocamento de abomaso, 35 apresentaram hipocalcemia e no grupo controle, apenas um animal. Concentrações sanguíneas de glicose e proteína foram inferiores nos animais com DA, quando comparados com animais do grupo controle.(AU)

This study evaluated the status of serum calcium in 39 Holstein cows with displacement of the abomasum (DA), from 30 dairy farms selected in the region of Campos Gerais, Paraná state, Brazil.. The diagnosis of abomasal displacement was performed by auscultation and percussion, besides the clinical signs and history information. Before surgery, blood samples were collected to measure blood levels of calcium, albumin, protein and glucose. As a control group, blood samples were collected from healthy cows that were in similar period of lactation. Of the 39 cows with abomasal displacement, 35 had hypocalcemia and in the control group only one cow. Blood glucose and protein concentrations were lower in the animals with DA when compared with the control group.(AU)