Selective enrichment of the raw milk microbiota in cheese production: Concept of a natural adjunct milk culture.

In cheese production, microorganisms are usually added at the beginning of the process as primary starters to drive curd acidification, while secondary microorganisms, with other pro-technological features important for cheese ripening, are added as selected cultures. This research aimed to investigate the possibilities of influencing and selecting the raw milk microbiota using artisanal traditional methods, providing a simple method to produce a natural supplementary culture. We investigated the production of an enriched raw milk whey culture (eRWC), a natural adjunct microbial culture produced from mixing an enriched raw milk (eRM) with a natural whey culture (NWC). The raw milk was enriched by spontaneous fermentation for 21 d at 10°C. Three milk enrichment protocols were tested: heat treatment before incubation, heat treatment plus salt addition, and no treatment. The eRMs were then co-fermented with NWC (ratio of 1:10) at 38°C for 6 h (young eRWC) and 22 h (old eRWC). Microbial diversity during cultures' preparation was evaluated through the determination of colony forming units on selective growth media, and next-generation sequencing (16S rRNA gene amplicon sequencing). The enrichment step increased the streptococci and lactobacilli but reduced microbial richness and diversity of the eRMs. Although the lactic acid bacteria viable count was not significantly different between the eRWCs, they harbored higher microbial richness and diversity than NWC. Natural adjunct cultures were then tested in cheese making trials, following the microbial development, and assessing the chemical quality of the 120 d ripened cheeses. The use of eRWCs slowed the curd acidification in the first hours of cheese making but the pH 24 h after production settled to equal values for all the cheeses. Although the use of diverse eRWCs contributed to having a richer and more diverse microbiota in the early stages of cheese making, their effect decreased over time during ripening, showing an inferior effect to the raw milk microbiota. Even if more research is needed, the optimization of such a tool could be an alternative to the practice of isolating, geno-pheno-typing, and formulating mixed-defined-strain adjunct cultures that require knowledge and facilities not always available for artisanal cheese makers.

Evaluation of Contamination in Milk Samples Pooled From Independently Collected Quarters Within a Laboratory Setting.

The primary objective of this observational study was to evaluate the prevalence of contamination from independently collected quarter-level milk samples pooled in a laboratory and subjected to bacterial culture. To address this objective, weekly quarter-level milk samples were collected longitudinally from a cohort of 503 primiparous cows from five organic dairy farms during the first 5 weeks after calving. Individual quarter milk samples were pooled in a laboratory using aseptic technique ("lab-pooled") and subjected to bacterial culture. In the sample set of 2,006 lab-pooled milk samples, 207 (10.3%) were classified as contaminated using a standard definition (i.e., growth of three or more distinct microorganisms). Subsequent culturing of corresponding quarter-level milk samples revealed that many of the contaminated lab-pooled sample results (i.e., 46.7%) were the result of intramammary infections with different pathogens across the quarters, rather than actual contamination within any single quarter (i.e., "true contamination"). The odds of true contamination were lower when the lab-pooled sample exhibited growth of three microorganisms compared to more than 3 microorganisms. Our findings suggest that pooling of quarter samples within a laboratory setting may yield lower rates of contamination compared to those previously reported from samples composited on-farm, but that current cut-offs to define contamination may need to be evaluated for use with lab-pooled samples. Further investigation of use of lab-pooled samples may be warranted to reduce costs while still providing useful scientific insight.

Bayesian estimation of diagnostic accuracy of somatic cell counts history and on-farm milk culture using Petrifilm® to identify quarters or cows that should be treated with antimicrobials in selective treatment protocols at dry off.

Bayesian latent class models were used to estimate the test accuracy (sensitivity (Se), specificity (Sp), and predictive values (NPV and PPV)) of cow-level somatic cell counts (SCC) data, quarter-level Petrifilm® on-farm milk culture, and quarter-level standard milk bacteriology for the identification of quarters that should possibly be treated with antimicrobials at dry off in dairy cows. Data of 282 cows from 9 dairy herds in Québec, Canada, with bulk tank SCC < 250,000 cells/mL were used. Estimated median herd-prevalence of infections that should be treated was 16.2 % (95 % credibility interval (CI): 11.0-22.7). Se and Sp estimates for quarter-milk culture using Petrifilm® were 82.2 % (95 %CI: 74.0-89.5) and 62.0 % (95 %CI: 58.6-65.6), respectively. Se and Sp for quarter-milk standard bacteriology were 67.4 % (95 %CI: 55.8-81.2) and 79.6 % (95 %CI: 76.4-83.0), respectively. Se and Sp of different SCC scenarios and thresholds were estimated. For first parity cows, using only the last Dairy Herd Improvement (DHI) test SCC with a threshold of 100,000 cells/mL appeared quite accurate, with Se, Sp, PPV, NPV and reduction of antimicrobial usage of 85.6 % (95 %CI: 69.6-95.6), 86.0 % (95 %CI: 80.0-91.7), 58.0 % (95 %CI: 42.3-74.2), 96.4 % (95 %CI: 91.3-99.0), and 75.3 % (95 %CI: 70.7-79.3), respectively. For cows of ≥ 2nd parity, using only the last DHI test SCC with a threshold of 200,000 cells/mL resulted in Se, Sp, PPV, NPV and reduction of antimicrobial usage of 75.3 % (95 %CI: 55.8-87.3), 84.0 % (95 %CI: 78.8-89.3), 47.2 % (95 %CI: 32.0-63.7), 94.7 % (95 %CI: 89.0-97.6), and of 77.0 % (95 %CI: 73.3-80.3), respectively. Adding quarter-level milk culture using Petrifilm® to cows identified as unhealthy using cow-level SCC data improved the test accuracy (mainly the PPV) and further reduced the use of antimicrobials. For instance, in ≥ 2nd parity cows, using only the last DHI SCC with a threshold of 200,000 cells/mL, adding a subsequent Petrifilm® test increased the reduction from 77.0 % (95 %CI: 73.3-80.3) to 89.5 % (95 %CI: 86.7-91.8). Considering the availability of SCC data, the easiness of using just the last DHI test, and the high NPV that could be achieved, producers may consider using just the last DHI test as a potential tool to identify cows that should be treated with antimicrobials at dry off. It may be used alone or in combination with quarter-level on-farm Petrifilm® milk culture on high SCC cows to further reduce the use of antimicrobials by identifying quarters that need to be treated.

Prevalencia y características de la lactancia dolorosa. Estudio prospectivo multicéntrico en Atención Primaria / Prevalence and characteristics of breastfeeding pain. Prospective multicentric study in Primary Care

Introducción: el dolor al amamantar influye muy negativamente en el desarrollo de la lactancia y es una de las principales causas de su abandono, por lo que constituye un relevante y prioritario problema de Salud Pública. No obstante, es un aspecto infravalorado por la comunidad sanitaria y se han realizado muy pocos estudios sobre su prevalencia, sus características y la actitud de las madres que lo padecen. Pacientes y métodos: el presente trabajo forma parte del estudio Lactancia Materna y Desarrollo Infantil (LAyDI) y se centra en la investigación del dolor al amamantar durante los primeros cuatro meses en una cohorte de 203 madres lactantes adscritas a centros de salud de la Comunidad de Madrid. Resultados: se observó una alta incidencia de dolor (68,5%) entre las mujeres lactantes, que suelen padecerlo durante los primeros días, si bien en un 13% de los casos se presentó tras varias semanas. En un 44% de casos se resolvió en menos de 15 días, pero se mantuvo de forma prolongada en un 46% (incluso más de dos meses, en el 17,3%). El 30,9% de los casos se planteó en algún momento el abandono de la lactancia por dolor intenso o mantenido. Sin embargo, a pesar de la accesibilidad al apoyo profesional, mayoritariamente a cargo de las matronas, casi la mitad de las mujeres que sopesaron el destete decidió no solicitar ayuda. Conclusiones: es urgente unir esfuerzos desde diferentes campos de estudio para investigar sobre la etiopatogenia del dolor y explorar sin prejuicios alternativas seguras y efectivas para su abordaje (AU)

Introduction: painful breastfeeding (BF) has a very negative impact on the establishment of BF and is one of the main causes of BF cessation, and is therefore considered an important public health problem. Unfortunately, this problem is underestimated by the health care community and few studies have been devoted to its prevalence, characteristics and the attitude of the mothers that experience it.Sample and methods: the study was conducted in the framework of the Breastfeeding and Child Development study (LAyDI) and focused on BF pain in the first 4 months post birth in a cohort of 203 lactating women managed in primary care centres of the Community of Madrid (Spain).Results: we found a high incidence of pain in BF women (68.5%), most frequently with onset in the first days post birth, although in 13% of cases it developed after several weeks. In 44% of cases, the pain resolved within 15 days, but in 46% it lasted longer (in 17%, longer than 2 months). We found that 30.9% of mothers contemplated BF cessation at some point due to severe or sustained pain. However, despite having access to professional support, mostly provided by midwives, nearly half of the women who considered weaning did not seek help.Conclusions: current efforts in different fields of study need to converge urgently to explore the aetiology and pathogenesis of pain and safe and effective approaches to its management in a nonjudgmental and unbiased environment.(AU)

Contribution of natural milk culture to microbiota, safety and hygiene of raw milk cheese produced in alpine malga.

Processing of alpine milk in malga farms is carried out under conditions that can favor contamination by coliforms, coagulase-positive staphylococci, or pathogens such as Listeria monocytogenes. With the aim to improve the hygienic characteristics and safety of cheese produced in four malga farms the use of lyophilized Natural Milk Culture prepared with selected strains was tested.. Two cheesemaking tests were carried out in the same day always starting from the same milk: in the first case following the malga recipe that uses either Natural Whey Culture or without the addition of a starter, in the second one using a Natural Milk Culture. Cheesemaking were carried out in four malga farms located in the west area of Trentino region within the same week. For hygienic and safety evaluation, aerobic colony count, coagulase-positive staphylococci, Escherichia coli, staphylococcal toxins, Listeria monocytogenes, and Salmonella spp, pH and aw were determined in raw milk from evening and morning milking, curd in vat, curd after extraction and two months-ripened cheese. Pathogens or toxins, high values of coagulase- positive staphylococci and E. coli were not found in cheese samples. However, in the curd coagulase-positive staphylococci reached values almost of 5 Log CFU/g in the two malga without starter cultures. The use of Natural Milk Culture reduced E. coli counts. In addition, DNA was extracted from cheese samples and from Natural Milk Culture and the composition of the microbial community determined by Next Generation Sequencing method. The determination of cheese microbial communities demonstrated that the use of Natural Milk Culture exerted different effects in the different malga, in any case preserving bacterial biodiversity.

Perfil microbiológico, celular e fatores de risco associados à mastite subclínica em cabras no semiárido da Paraíba / Cellular and microbiological profiles and risk factors for subclinical mastitis in goats in the semi-arid region of Paraíba

Foi realizado um estudo da mastite subclínica em nove rebanhos de cabras leiteiras no semiárido paraibano com o objetivo de determinar a ocorrência da infecção, avaliar o perfil microbiológico e celular do leite, testar a sensibilidade dos microorganismos isolados frente a antimicrobianos além de identificar os fatores de risco. Foram utilizadas 131 cabras leiteiras das quais foram colhidas 261 amostras de leite para exame microbiológico e 131 para contagem de células somáticas (CCS). Na ocasião das colheitas foi realizado o California Mastitis Test (CMT) e aplicado um questionário epidemiológico por propriedade. Houve crescimento bacteriano em 30 amostras (11,49 por cento) com 25 (83,33 por cento) dos isolados identificados como Staphylococcus coagulase negativa e cinco (16,66 por cento) Staphylococcus aureus. A média de CCS foi de 1,39x10(6) células/ml. O CMT apresentou baixa sensibilidade (46,7 por cento) e baixa especificidade (60,6 por cento) quando comparado ao exame microbiológico. A gentamicina e a associação da neomicina, bacitracina e tetraciclina foram os antimicrobianos contra os quais os microrganismos isolados apresentaram 100 por cento de sensibilidade. Penicilina e ampicilina foram os de maiores índices de resistência (66,67 por cento e 63,89 por cento, respectivamente). A caprinocultura não ser a atividade principal da propriedade e o não isolamento de animais doentes, foram identificados como fatores de risco para a mastite subclínica caprina nas propriedades estudadas. Programas de controle e profilaxia da mastite devem ser implementados enfocando as medidas de higiene na ordenha e correção dos fatores de risco identificados nesse estudo.

A subclinical mastitis study was conducted in nine dairy goat herds in the semi-arid region of Paraíba state, Northeastern Brazil, to determine the occurrence of infection, to evaluate microbiological and cellular profiles of the milk, to test the sensitivity of isolated microorganisms to antimicrobials, and to identify risk factors. One hundred thirty-one dairy goats were used, 261 samples were collected for microbiological culture and 131 samples for somatic cells count (SCC). During collection, the California Mastitis Test (CMT) was conducted and an epidemiological questionnaire was applied for each herd. There was bacterial growth in 30 samples (11.49 percent), with 25 (83.33 percent) coagulase-negative Staphylococcus and five (16.66 percent) Staphylococcus aureus isolated. The SCC mean was 1.39x10(6) cells/ml. CMT presented low sensitivity (46.7 percent) and low specificity (60.6 percent) compared with microbiological culture. Gentamicin and the association of neomycin, bacitracin and tetracyclin were the antimicrobials against which the microorganisms isolated showed 100 percent sensitivity. Penicillin and ampicillin had the greatest resistance rates (66.67 percent and 63.89 percent, respectively). Goat breeding is not the main activity on the farms and do not isolate diseased animals were identified as risk factors for caprine subclinical mastitis. Program for the control and prophylaxis of mastitis must be implemented focusing primarily on hygiene cares at milking and correction of the risk factors identified in this study.

Perfil microbiológico, celular e fatores de risco associados à mastite subclínica em cabras no semiárido da Paraíba / Cellular and microbiological profiles and risk factors for subclinical mastitis in goats in the semi-arid region of Paraíba

Foi realizado um estudo da mastite subclínica em nove rebanhos de cabras leiteiras no semiárido paraibano com o objetivo de determinar a ocorrência da infecção, avaliar o perfil microbiológico e celular do leite, testar a sensibilidade dos microorganismos isolados frente a antimicrobianos além de identificar os fatores de risco. Foram utilizadas 131 cabras leiteiras das quais foram colhidas 261 amostras de leite para exame microbiológico e 131 para contagem de células somáticas (CCS). Na ocasião das colheitas foi realizado o California Mastitis Test (CMT) e aplicado um questionário epidemiológico por propriedade. Houve crescimento bacteriano em 30 amostras (11,49 por cento) com 25 (83,33 por cento) dos isolados identificados como Staphylococcus coagulase negativa e cinco (16,66 por cento) Staphylococcus aureus. A média de CCS foi de 1,39x10(6) células/ml. O CMT apresentou baixa sensibilidade (46,7 por cento) e baixa especificidade (60,6 por cento) quando comparado ao exame microbiológico. A gentamicina e a associação da neomicina, bacitracina e tetraciclina foram os antimicrobianos contra os quais os microrganismos isolados apresentaram 100 por cento de sensibilidade. Penicilina e ampicilina foram os de maiores índices de resistência (66,67 por cento e 63,89 por cento, respectivamente). A caprinocultura não ser a atividade principal da propriedade e o não isolamento de animais doentes, foram identificados como fatores de risco para a mastite subclínica caprina nas propriedades estudadas. Programas de controle e profilaxia da mastite devem ser implementados enfocando as medidas de higiene na ordenha e correção dos fatores de risco identificados nesse estudo.

A subclinical mastitis study was conducted in nine dairy goat herds in the semi-arid region of Paraíba state, Northeastern Brazil, to determine the occurrence of infection, to evaluate microbiological and cellular profiles of the milk, to test the sensitivity of isolated microorganisms to antimicrobials, and to identify risk factors. One hundred thirty-one dairy goats were used, 261 samples were collected for microbiological culture and 131 samples for somatic cells count (SCC). During collection, the California Mastitis Test (CMT) was conducted and an epidemiological questionnaire was applied for each herd. There was bacterial growth in 30 samples (11.49 percent), with 25 (83.33 percent) coagulase-negative Staphylococcus and five (16.66 percent) Staphylococcus aureus isolated. The SCC mean was 1.39x10(6) cells/ml. CMT presented low sensitivity (46.7 percent) and low specificity (60.6 percent) compared with microbiological culture. Gentamicin and the association of neomycin, bacitracin and tetracyclin were the antimicrobials against which the microorganisms isolated showed 100 percent sensitivity. Penicillin and ampicillin had the greatest resistance rates (66.67 percent and 63.89 percent, respectively). Goat breeding is not the main activity on the farms and do not isolate diseased animals were identified as risk factors for caprine subclinical mastitis. Program for the control and prophylaxis of mastitis must be implemented focusing primarily on hygiene cares at milking and correction of the risk factors identified in this study.