Metataxonomic and immunological analysis of milk from ewes with or without a history of mastitis.

Mastitis is a highly prevalent condition that has a great impact on milk production and animal welfare, and often requires substantial management efforts. For this reason, it is generally considered an important threat to the dairy industry. Many microbial, host, and environmental factors can protect against, predispose to, or influence the development of mastitis. The objective of this work was to characterize the milk microbiota of Manchega ewes, and to compare samples from animals with and without a history of mastitis. We analyzed milk samples from 36 ewes belonging to 2 different farms (18 ewes from each farm) using culture-dependent and culture-independent techniques. We also analyzed several immune compounds to investigate associations of mastitis with 3 main variables: farm; history of mastitis or no mastitis; and parity number. Both culture-dependent and culture-independent techniques showed that ewe milk harbored a site-specific complex microbiota and microbiome. Staphylococcus epidermidis was the main species driving the difference between farm A (where it was the dominant species) and B (where it was not). In contrast, samples from farm B were characterized by the presence of a wide spectrum of other coagulase-negative staphylococci. Some of these species have already been associated with subclinical intramammary infections in ruminants. Of the 10 immune compounds assayed in this study, 3 were related to a history of mastitis [IL-8, IFN-γ, and IFN-gamma-induced protein 10 (IP-10)]. Increases in IL-8 concentrations in milk seemed to be a feature of subclinical mastitis in sheep, and in this study, this immune factor was detected only in samples from ewes with some episodes of mastitis and from the group with the highest somatic cell count. We also observed a positive correlation between the samples with the highest somatic cell count and IFN-γ and IP-10 levels. Our results suggest that these 3 compounds could be used as biomarkers for the negative selection of mastitis-prone animals, particularly when somatic cell count is very high.

Novel mixed-linkage ß-glucan activated by c-di-GMP in Sinorhizobium meliloti.

An artificial increase of cyclic diguanylate (c-di-GMP) levels in Sinorhizobium meliloti 8530, a bacterium that does not carry known cellulose synthesis genes, leads to overproduction of a substance that binds the dyes Congo red and calcofluor. Sugar composition and methylation analyses and NMR studies identified this compound as a linear mixed-linkage (1 → 3)(1 → 4)-ß-D-glucan (ML ß-glucan), not previously described in bacteria but resembling ML ß-glucans found in plants and lichens. This unique polymer is hydrolyzed by the specific endoglucanase lichenase, but, unlike lichenan and barley glucan, it generates a disaccharidic → 4)-ß-D-Glcp-(1 → 3)-ß-D-Glcp-(1 → repeating unit. A two-gene operon bgsBA required for production of this ML ß-glucan is conserved among several genera within the order Rhizobiales, where bgsA encodes a glycosyl transferase with domain resemblance and phylogenetic relationship to curdlan synthases and to bacterial cellulose synthases. ML ß-glucan synthesis is subjected to both transcriptional and posttranslational regulation. bgsBA transcription is dependent on the exopolysaccharide/quorum sensing ExpR/SinI regulatory system, and posttranslational regulation seems to involve allosteric activation of the ML ß-glucan synthase BgsA by c-di-GMP binding to its C-terminal domain. To our knowledge, this is the first report on a linear mixed-linkage (1 → 3)(1 → 4)-ß-glucan produced by a bacterium. The S. meliloti ML ß-glucan participates in bacterial aggregation and biofilm formation and is required for efficient attachment to the roots of a host plant, resembling the biological role of cellulose in other bacteria.

Mini-Tn7 vectors for stable expression of diguanylate cyclase PleD* in Gram-negative bacteria.

BACKGROUND: The cyclic diguanylate (c-di-GMP) is currently considered an ubiquitous second messenger in bacteria that influences a wide range of cellular processes. One of the methodological approaches to unravel c-di-GMP regulatory networks involves raising the c-di-GMP intracellular levels, e.g. by expressing a diguanylate cyclase (DGC), to provoke phenotypic changes. RESULTS: We have constructed mini-Tn7 delivery vectors for the integration and stable expression of the pleD* gene encoding a highly active DGC, which can be used to artificially increase the intracellular levels of c-di-GMP in Gram negative bacteria. The functionality of these new vectors has been validated in several plant-interacting α- and γ-proteobacteria. Similarly to vector plasmid-borne pleD*, the genome-borne mini-Tn7pleD* constructs provide significant increases in intracellular c-di-GMP, provoking expected phenotypic changes such as enhanced polysaccharide production, biofilm formation and reduced motility. However, the mini-Tn7pleD* constructs resulted far more stable in the absence of antibiotics than the plasmid-based pleD* constructs. Furthermore, we have also implemented an inducible system to modulate pleD* expression and intracellular c-di-GMP rises "on demand". CONCLUSIONS: mini-Tn7pleD* constructs are very stable and are maintained during bacterial free-living growth as well as during interaction with eukaryotic hosts, in the absence of selective pressure. This high stability ensures experimental homogeneity in time and space with regard to enhancing c-di-GMP intracellular levels in bacteria of interest.

A Stochastic Frontier Approach to Study the Relationship between the Hygienic Quality of Bulk Tank Sheep Milk and Technical Efficiency of the Coagulation Process.

Sheep milk from local breeds is important for the production of high-quality cheeses throughout the Mediterranean region, such as Manchego cheese in Spain. To maintain sustainable and efficient production, it is necessary to reach a better understanding of how the composition and hygiene of the milk affect the coagulation process, with the aim of optimizing production yield. This study implemented a stochastic production frontier function to estimate the potential production of curd and efficiency using data from the four seasons of a study of 77 Manchega sheep farms. The Cobb-Douglas production frontier model was estimated using the maximum likelihood estimation method. The results showed that the content of protein, lactose, and fat exhibited increasing returns to scale, with protein content being the most significant factor for curd production. Approximately half of the inefficiency was due to factors related to the technological properties and the hygiene of the milk. The pH, curd firmness, and concentration of lactic acid bacteria improved the efficiency of coagulation, while the concentration of spores of lactate-fermenting Clostridium spp., Pseudomonas spp., staphylococci, and catalase-negative gram-positive cocci favored the inefficiency of the coagulation process. To date, this is the first study to evaluate the effect of different factors, such as microbial groups, milk composition, and technological properties, on the efficiency of the coagulation process in dairy sheep.

Associations between Milk Coagulation Properties and Microbiological Quality in Sheep Bulk Tank Milk.

This study conducted a seasonal analysis of bulk tank milk from 77 sheep farms to establish relationships between the concentration of major microbial groups and milk coagulation properties. The investigated milk traits included composition (pH, fat, casein, lactose), coagulation properties (curd firmness: A60-, rennet clotting time: RCT-, curd firming time: k20-, curd yield: CY-), and somatic cell score (SCS). The main microbial groups analyzed were total mesophilic bacteria (SPC), thermodurics (THERMO), psychrotrophs (PSYCHRO), Pseudomonas spp. (PSEUDO), lactic acid bacteria (LAB), catalase-negative gram-positive cocci (GPCNC), Escherichia coli (ECOLI), coliforms other than Escherichia coli (COLI), coagulase-positive staphylococci (CPS), coagulase-negative staphylococci (CNS), and spores of lactate-fermenting Clostridium (BAB). Mixed linear models were used to explore associations between coagulation properties and the aforementioned variables. Results demonstrated that incorporating microbial loads into the models improves their fit and the relative quality of the outcomes. An important seasonality is demonstrated by an increase in CY and A60, along with a decrease in RCT and k20 during autumn and winter, contrasting with spring and summer. BAB concentration resulted in a reduction of A60 and an increase in RCT, whereas SPC concentration led to an enhancement of A60 and a reduction in RCT. An increase in GPCNC concentration was associated with an increase in k20 and a decrease in CY.

Can microbiology help to make aviation more sustainable?

Production of sustainable aviation fuels (SAFs) using microbes still requires huge research efforts to fulfill the needs of aviation, both in the biological utilization of raw materials as well as in the biological processes to convert these materials (oils, sugars, aromatic compounds and others) into SAFs. However, we should also be aware of the microbiology constraints that, in some cases, will not allow us to reach the commercial level and that, by creating false expectations we will harm the credibility of microbiologists. However, in our opinion microbiologists can and should continue to find new avenues for producing SAFs, and for evaluating the advantages and feasibility of their production. This last step will require a close collaboration between researchers and industry.

Evolution of psychopathology, purpose in life, and moral courage in healthcare workers during the COVID-19 pandemic: a longitudinal study.

Introduction: Almost 2 years and five infection waves after the COVID-19 pandemic started, healthcare workers continued dealing with the pandemic situation and facing the health consequences and the mental health disorders it caused. This study aimed to evaluate the onset and progression of psychopathology as well as the role of predictor variables such as purpose in life and moral courage among healthcare workers during this time. Materials and methods: This was a longitudinal prospective study carried out with 45 Spanish healthcare workers who answered two questionnaires, the first questionnaire in April-May 2020 (T1) and the second questionnaire in September-October 2021 (T2). Results: Although 29.5% of the sample considered that their mental health had improved over this time, almost half of them (47.7%) said it had not changed, while 22.7% reported a decline in their mental health from the first time they were asked. Specifically, 46.8% presented anxiety, 23.4% depression, and 42.6% acute stress at T1, and 38.3% had anxiety, 17% depression, and 27.7% post-traumatic stress disorder at T2. Despite this, there were no differences between T1 and T2 anxiety scores (p = 0.53), although there was a decrease in depression (p = 0.03) and acute stress (p = 0.02) scores. Predictor variable outcomes such as purpose in life (p = 0.88) and moral courage (p = 0.86; p = 0.38) did not change over time, but when modelling the data, purpose in life predicted psychopathology at T1, which in turn affected the psychopathology results at T2. Conclusion: This study showed that, although psychopathology decreased over the months, its prevalence remained high. Even though the purpose in life predicted psychopathology at T1, it seems that once the psychopathology is established (T2), the factors that would improve it would be different from the protective factors that prevented its establishment, which become secondary.

Neural correlates of cocaine-induced conditioned place preference in the posterior cerebellar cortex.

Introduction: Addictive drugs are potent neuropharmacological agents capable of inducing long-lasting changes in learning and memory neurocircuitry. With repeated use, contexts and cues associated with consumption can acquire motivational and reinforcing properties of abused drugs, triggering drug craving and relapse. Neuroplasticity underlying drug-induced memories takes place in prefrontal-limbic-striatal networks. Recent evidence suggests that the cerebellum is also involved in the circuitry responsible for drug-induced conditioning. In rodents, preference for cocaine-associated olfactory cues has been shown to correlate with increased activity at the apical part of the granular cell layer in the posterior vermis (lobules VIII and IX). It is important to determine if the cerebellum's role in drug conditioning is a general phenomenon or is limited to a particular sensory modality. Methods: The present study evaluated the role of the posterior cerebellum (lobules VIII and IX), together with the medial prefrontal cortex (mPFC), ventral tegmental area (VTA), and nucleus accumbens (NAc) using a cocaine-induced conditioned place preference procedure with tactile cues. Cocaine CPP was tested using ascending (3, 6, 12, and 24 mg/kg) doses of cocaine in mice. Results: Compared to control groups (Unpaired and Saline animals), Paired mice were able to show a preference for the cues associated with cocaine. Increased activation (cFos expression) of the posterior cerebellum was found in cocaine CPP groups and showed a positive correlation with CPP levels. Such increases in cFos activity in the posterior cerebellum significantly correlated with cFos expression in the mPFC. Discussion: Our data suggest that the dorsal region of the cerebellum could be an important part of the network that mediates cocaine-conditioned behavior.

Wild Ungulates Constitute the Basis of the Diet of the Iberian Wolf in a Recently Recolonized Area: Wild Boar and Roe Deer as Key Species for Its Conservation.

The Iberian wolf (Canis lupus signatus) is recolonizing historical distribution areas after decades of absence. As in other human-dominated landscapes, finding a balance to protect this species by favoring recolonization and mitigating human-wildlife conflicts is a challenge. Since wolves are often generalist opportunistic predators, we studied their diet composition in central Spain to evaluate the consumption of domestic ungulates and provide reliable data that could help local authorities to deal with the current wolf-cattle ranchers conflict and coexistence. Diet composition (% prey occurrence, % prey ingested biomass) was analyzed through the identification of prey hairs present in 671 scats collected between 2017 and 2021. The wolves fed more on wild ungulates (82% occurrence) than domestic ones (18%). Wild boar (Sus scrofa, 44% occurrence) and roe deer (Capreolus capreolus, 35%) were the most consumed prey. The wolves positively selected these two species. The wolves' diets varied between seasons, years, and forest regions, but a diet based on wild ungulates predominated over domestic ones. Food niche breadth showed variations depending on seasons and years. Preserving the availability and diversity of wild ungulates may favor reducing livestock attacks and would be an achievable goal that would help to conserve this species and reduce conservation conflicts.

Rectus Femoris Cross-Sectional Area and Phase Angle asPredictors of 12-Month Mortality in Idiopathic Pulmonary Fibrosis Patients.

BACKGROUND: The value of the phase angle (PhA), measured via bioelectrical impedance analysis (BIA), could be considered a good marker of the cell mass and the cell damage of a patient; however, there are new techniques, such as muscle ultrasonography, that allow the quantity and quality of muscle to be assessed in a minimally invasive way. The aim of this study is to determine the prognostic value of morphofunctional techniques in the prognosis of mortality in patients with idiopathic pulmonary fibrosis (IPF). METHODS: This multicenter, cross-sectional study included 86 patients with idiopathic pulmonary fibrosis with a mean age of 71 years, 82.7% of whom were male. The nutritional risk of the patients was assessed by means of questionnaires, such as the Subjective Global Assessment (SGA), and non-invasive functional techniques, including BIA, nutritional ultrasound, and hand grip strength (HGS). Statistical analysis of the sample was performed using JAMOVI version 2.3.22. RESULTS: Correlations were made between the RF-CSA techniques with PhA (r = 0.48, p < 0.001), BCM (r = 0.70, p < 0.001), SMI (r = 0.64, p < 0.001), and HGS (r = 0.54, p < 0.001). The cut-off points for 12-month mortality were PhA = 4.5° (AUC = 0.722, sensitivity of 72.7% and specificity of 66.6%), BCM = 28.8 kg (AUC = 0.609, sensitivity of 32.4% and specificity of 100.0%), RF-CSA = 3.00 cm2 (AUC = 0.857, sensitivity of 64.4% and specificity of 100.0%), 6MMW = 420 m (AUC = 0.830, sensitivity of 63.27% and specificity of 100.0%), and TUG = 7.2 s (AUC = 0.771, sensitivity of 100.0% and specificity of 56.67%). In addition, a multivariate analysis was performed with RF-CSA, HR = 8.11 (1.39-47.16, p = 0.020), and PhA of 6.35 (1.29-31.15, p = 0.023), taking into account age, sex, and BMI to determine mortality. Finally, a Kaplan-Meier survival analysis was conducted with low or normal values for classical parameters (GAP and T6MM) and new parameters (PhA, BCM, RF-CSA, and TUG). CONCLUSION: RF-CSA and PhA were shown to be good prognostic markers of 12-month mortality and could, therefore, be useful screening tools to complement the nutritional assessment of IPF patients.

GHG Emissions from Dairy Small Ruminants in Castilla-La Mancha (Spain), Using the ManleCO2 Simulation Model.

The first goal of this work was the description of a model addressed to quantify the carbon footprint in Spanish autochthonous dairy sheep farms (Manchega group), foreign dairy sheep farms (foreigners group: Lacaune and Assaf breeds), and Spanish autochthonous dairy goat farms (Florida group). The second objective was to analyze the GHG emission mitigation potential of 17 different livestock farming practices that were implemented by 36 different livestock farms, in terms of CO2e per hectare (ha), CO2e per livestock unit (LU), and CO2e per liter of fat- and protein-corrected milk (FPCM). The study showed the following results: 1.655 kg CO2e per ha, 6.397 kg CO2e per LU, and 3.78 kg CO2e per liter of FPCM in the Manchega group; 12.634 kg CO2e per ha, 7.810 CO2e kg per LU, and 2.77 kg CO2e per liter of FPCM in the Foreigners group and 1.198 kg CO2e per ha, 6.507 kg CO2e per LU, and 3.06 kg CO2e per liter of FPCM in Florida group. In summary, purchasing off-farm animal feed would increase emissions by up to 3.86%. Conversely, forage management, livestock inventory, electrical supply, and animal genetic improvement would reduce emissions by up to 6.29%, 4.3%, 3.52%, and 0.8%, respectively; finally, an average rise of 2 °C in room temperature would increase emissions by up to 0.62%.

Burnout, psychopathology and purpose in life in healthcare workers during COVID-19 pandemic.

Background: The COVID-19 pandemic has led to a significant increase in the workload of healthcare workers that, together with the risks associated with exposure to this new virus, has affected their mental health. Objectives: The objective of the current study was to determine the prevalence of psychopathology and burnout syndrome in healthcare workers and the predictive role of purpose in life and moral courage in this relationship. Methods: A cross-sectional study was carried out in 115 Spanish healthcare workers. Results: Participants with burnout had higher anxiety (p = 0.001), depression (p < 0.001), post-traumatic stress (p = 0.01) and alcohol consumption (p = 0.03) levels. The different components of burnout (emotional fatigue and despersonalization) were associated with the occurrence of anxiety (OR = 0.31) and depression (OR = 0.26), respectively. A strong purpose in life decreased emotional fatigue (OR = -0.39) depersonalization (OR = -0.23) scores, increased personal accomplishment (OR = 0.52), subsequently reducing burnout levels (OR = -0.45). Conclusions: Purpose in life was most strongly related to decreased levels of burnout. Furthermore, an association between anxiety, depression and the components of burnout was found.

Effect of intensity and duration of anthropic noises on European mink locomotor activity and fecal cortisol metabolite levels.

Human activities involving noise emission can affect wild animals. European mink was exposed to road noise and human voice playbacks to analyze how sound intensity level and duration of both noises altered the time that individuals were active and if their fecal cortisol metabolite (FCM) levels varied. A Hierarchical Analysis Cluster was performed to establish 2 mink groups with respect to both noise source type: short duration/low intensity (SL) and long duration/high intensity (LH). We performed general linear mixed models to evaluate the variation in locomotor activity duration (s) and FCM (nanogram per gram) levels, respectively. The results showed both road noise and human voices decreased locomotor activity duration in SL more sharply compared with LH, and human voices were the triggers that induced the most pronounced response to both exposure conditions. FCM (ng/g) levels increased in SL compared with LH during road noise while the opposite happened during human voices. Differences based on sex and age of individuals were observed. In conclusion, noise characteristics given by the sound type determined the variations in locomotor activity duration while noise exposure level determined the variations in FCM (ng/g) levels. Attention should be paid to noisy activities (e.g., recreational activities for visitors in protected natural areas) and loud groups of people to conserve wildlife, especially noise sensitive species.

The Role of Two Linear ß-Glucans Activated by c-di-GMP in Rhizobium etli CFN42.

Bacterial exopolysaccharides (EPS) have been implicated in a variety of functions that assist in bacterial survival, colonization, and host-microbe interactions. Among them, bacterial linear ß-glucans are polysaccharides formed by D-glucose units linked by ß-glycosidic bonds, which include curdlan, cellulose, and the new described Mixed Linkage ß-Glucan (MLG). Bis-(3',5')-cyclic dimeric guanosine monophosphate (c-di-GMP) is a universal bacterial second messenger that usually promote EPS production. Here, we report Rhizobium etli as the first bacterium capable of producing cellulose and MLG. Significant amounts of these two ß-glucans are not produced under free-living laboratory conditions, but their production is triggered upon elevation of intracellular c-di-GMP levels, both contributing to Congo red (CR+) and Calcofluor (CF+) phenotypes. Cellulose turned out to be more relevant for free-living phenotypes promoting flocculation and biofilm formation under high c-di-GMP conditions. None of these two EPS are essential for attachment to roots of Phaseolus vulgaris, neither for nodulation nor for symbiotic nitrogen fixation. However, both ß-glucans separately contribute to the fitness of interaction between R. etli and its host. Overproduction of these ß-glucans, particularly cellulose, appears detrimental for symbiosis. This indicates that their activation by c-di-GMP must be strictly regulated in time and space and should be controlled by different, yet unknown, regulatory pathways.

Surrounded by challenges: The simulated presence of competitors and predators modulates perianal secretion marking behaviour in the European mink (Mustela lutreola).

Animals face a variety of daily challenges to their reproduction and survival that can detect in time through to the environment cues. By using an individual focal sampling, we evaluated the variations in the time devoted to the perianal secretion marking behaviour (PSMB) of European mink when they were exposed to the simulated presence of conspecifics and two potential predators. Model results indicated that males and adult individuals dedicated more time to PSMB than females and subadults. The presence of conspecifics increased PSMB time only in adult and males, probably as an intrasexual territorial competence response. The heightened decrease in PSMB time in presence of a dog suggests an innate response due to the detection of volatile substances from faeces of carnivorous. In addition, simulated conspecific presence increased PSMB in absence of odours (control) and with owl faeces. However, when facing dog faeces, the simulated conspecific presence had no effects on PSMB. Thus, minks seem to prioritize the imminent risk of predation to avoid being detected. The stimulation of PSMB in captivity by simulated cues from conspecifics and potential predators could be useful to facilitate the reintroduction of individuals into nature, as well as their adaptation and survival.

Behavioral responses of the European mink in the face of different threats: conspecific competitors, predators, and anthropic disturbances.

Prey species assess the risk of threat using visual, olfactory, and acoustic cues from their habitat. Thus, they modify their behavior in order to avoid encounters with competitors, predators, and human disturbances that endanger their fitness. European mink (Mustela lutreola) is a critically endangered species that can be preyed upon by larger carnivores and displaced by dominant conspecifics to areas of lower quality, e.g., near to more anthropized localities which may be noisier. In this study, the behavioral responses of 24 European mink were evaluated by conducting an experiment in which the presence of a conspecific competitor was simulated with a visual cue (mirror) and the presence of predators (terrestrial and aerial) with odorous cues. Additionally, they were also exposed to potential sources of anthropic disturbance with acoustic cues (road traffic noise and human voices). Our results showed that European mink were hidden for longer periods of time due to the presence of conspecifics and being exposed to the fecal odors of a terrestrial predator such as dog, but especially when they were exposed to anthropic noises. In the presence of a conspecific, the females and the subadults were the ones who remained hidden for the longest time. As well, they were hidden for longer periods of time due to the presence of conspecifics but in combination with dog feces and anthropic sounds did not induce variations in the response, as both by themselves already triggered an increase in the time they spent hiding. The vigilance model showed the effects of the same factors as the hiding model, but with antagonistic effects in the case of vigilance time which decreased during anthropic noises exposition. Finally, we want to highlight that European mink showed an innate response favorable to all three types of threats, but attention should be focused on human disturbances-as they trigger the most extreme responses-which may affect the rate of survival of this threatened species.

Pharmacokinetics and Pharmacodynamics of Cefepime in Adults with Hematological Malignancies and Febrile Neutropenia after Chemotherapy.

Patients with chemotherapy-induced febrile neutropenia (CIFN) may have changes in the pharmacokinetics (PK) compared to patients without malignancies or neutropenia. Those changes in antibiotic PK could lead to negative outcomes for patients if the therapy is not adequately adjusted to this. In this, open-label, non-randomized, prospective, observational, and descriptive study, a PK model of cefepime was developed for patients with hematological neoplasms and post-chemotherapy febrile neutropenia. This study was conducted at a cancer referral center, and study participants were receiving 2 g IV doses of cefepime every 8 h as 30-min infusions. Cefepime PK was well described by a two compartment model with a clearance dependent on a serum creatinine level. Using Monte Carlo simulations, it was shown that continuous infusions of 6g q24h could have a good achievement of PK/PD targets for MIC levels below the resistance cut-off point of Enterobacteriaceae. According to the simulations, it is unnecessary to increase the daily dose of cefepime (above 6 g daily) to increase the probability of target attainment (PTA). Cumulative fraction of response (CFR) using interment dosing was suboptimal for empirical therapy regimens against K. pneumoniae and P. aeruginosa, and continuous infusions could be used in this setting to maximize exposure. Patients with high serum creatinine levels were more likely to achieve predefined PK/PD targets than patients with low levels.

Farming Practices Influence Antibiotic Resistance and Biogenic Amine Capacity of Staphylococci from Bulk Tank Ewe's Milk.

Staphylococci are one of the main microorganisms responsible for intramammary infections in sheep, causing important economic losses for farmers and eventually health problems in humans, especially by the consumption of dairy products made with raw milk containing toxic compounds, such as biogenic amines or antibiotic resistant bacteria. This study aimed to check the presence and safety of staphylococci in bulk tank ewe's milk from different farms, and to determine the relationship between the presence of these staphylococci and farming practices, by applying nonlinear canonical correlation models (OVERALS). Two-hundred and fifty-nine staphylococci from milk samples from eighteen farms were genotyped and representative isolates of the major clusters were identified as belonging to Staphylococcus (S.) aureus, S. epidermidis, S. arlettae, S. lentus, S. simulans, and S. chromogenes species. Identified isolates were assayed in terms of their safety, by evaluating resistance to antimicrobial drugs and the aminobiogenic capacity, using both phenotypic and genetic assays. Antibiotic resistance phenotypic assay revealed that 82.9% were resistant to some antibiotics, although in the genotypic assay only the genes tetM, ermB, ermC, and grlA were detected. Fifty-three percent were high biogenic amine (BA) producers, being putrescine the most produced amine. A lowered risk of finding antibiotic-resistant and BA-producing staphylococci is related to some farming methods such as enrolling in a breeding program, use of good farming practices, postdipping teat disinfection, hygienic livestock housing, or periodic check of the milking machine.

Dominant Yeast Community in Raw Sheep's Milk and Potential Transfers of Yeast Species in Relation to Farming Practices.

Yeasts are always present in any type of cheese, as well as in the factories where it is produced. However, the role of the yeast community in the cheese making process, as well as the routes of contamination used by yeast species to contaminate milk from the dairy farm environment, are not well known. The objectives of this study were to broaden the knowledge of the dominant yeast community in Manchega sheep's milk and to assess the contamination routes of the yeast species depending on the farm practices. Milk, teat surface (collected from ten ewes per farm), feed, and air (collected in milking parlours and livestock housing) samples were collected from 12 typical farms in Castilla-La Mancha, Spain with differences in farming practices, and the yeast species were identified using DNA sequencing methods. To evaluate whether certain farming practices have an effect on the distribution of species of yeast in the milk samples, a mixed model was used. The results showed that most of the dominant yeast species (mainly belonging to the genus Candida) found in milk were also found in the other samples, indicating a microbial transfer from the farm environment to the milk. Furthermore, the statistical model showed that factors influencing yeast counts in milk were the presence of yeasts in the milking parlour, the use of silage, and the frequency of acid treatment for cleaning the milking machines. In conclusion, milk contamination from the yeast species present in the dairy farm environment is related to certain farming practices such as the use of silage and the daily use of acid in the cleaning of the milking machines, which favours the presence of desirable microbiota in milk.

Screening of c-di-GMP-Regulated Exopolysaccharides in Host Interacting Bacteria.

Bacterial exopolysaccharides (EPS) often confer a survival advantage by protecting the cell against abiotic and biotic stresses, including host defensive factors. They are also main components of the extracellular matrix involved in cell-cell recognition, surface adhesion and biofilm formation. Biosynthesis of a growing number of EPS has been reported to be regulated by the ubiquitous second messenger c-di-GMP, which promotes the transition to a biofilm mode of growth in an intimate association with the eukaryotic host. Here we describe a strategy based on the combination of an approach to artificially increase the intracellular level of c-di-GMP in virtually any gram-negative bacteria with a high throughput screening (HTS) for the identification of monosaccharide composition and carbohydrate fingerprinting of novel EPS, or modified variants, that can be involved in host-bacteria interactions.