Trends in recombinant protein use in animal production

Recombinant technologies have made possible the production of a broad catalogue of proteins of interest, including those used for animal production. The most widely studied proteins for the animal sector are those with an important role in reproduction, feed efficiency, and health. Nowadays, mammalian cells and fungi are the preferred choice for recombinant production of hormones for reproductive purposes and fibrolytic enzymes to enhance animal performance, respectively. However, the development of low-cost products is a priority, particularly in livestock. The study of cell factories such as yeast and bacteria has notably increased in the last decades to make the new developed reproductive hormones and fibrolytic enzymes a real alternative to the marketed ones. Important efforts have also been invested to developing new recombinant strategies for prevention and therapy, including passive immunization and modulation of the immune system. This offers the possibility to reduce the use of antibiotics by controlling physiological processes and improve the efficacy of preventing infections. Thus, nowadays different recombinant fibrolytic enzymes, hormones, and therapeutic molecules with optimized properties have been successfully roduced through cost-effective processes using microbial cell factories. However, despite the important achievements for reducing protein production expenses, alternative strategies to further reduce these costs are still required. In this context, it is necessary to make a giant leap towards the use of novel strategies, such as nanotechnology, that combined with recombinant technology would make recombinant molecules affordable for animal industry.info:eu-repo/semantics/publishedVersio

Longitudinal Study of Fecal Microbiota in Calves with or without Diarrhea Episodes before Weaning

Animal production is searching for ways to reduce antimicrobial use, and the best way is to avoid their use by maintaining the health of the animals. The microbiota is involved in the host health, and when the fecal microbiota was analyzed in calves that developed or not diarrhea, differences linked to the health status were detected. While changes in the fecal microbiota were observed with time (during the first 2 months of age) in all the calves, the microbiota from the healthy animals presented an earlier stabilization and some changes in low abundant bacteria, which may play a role in the subsequent health status of the animals. Bacteria classified in the families Coriobacteriaceae and Phyllobacteriaceae, and the bacterium Epulopiscium were found in the core of the microbiota of the healthy calves (calves that did not have diarrhea) possibly with a protective probiotic effect. On the other hand, several bacteria, such as Lachnospira, Neisseria and Solibacillus, were found only in the core of the microbiota obtained from calves that had diarrhea, indicating that they could be linked to a higher predisposition to suffer diarrhea. These results can help in the development of new probiotics to promote gut health in calves. The microbiota plays an important role in the development of diarrhea in pre-weaned calves. The characterization of the fecal microbiota in health and disease can be critical to unravel the bacterial dynamics associated with diarrhea and help with its prevention and control. In this study, we aimed to detect changes in the fecal microbiota of calves that experienced early-life diarrhea episodes. Fecal samples were taken from calves remaining healthy and calves with an episode of diarrhea during the study. We sampled at arrival (12 days of age) and after one and two months of life; also, at the time of the diarrhea episode for the diarrheic calves (day 17). Samples were processed to extract total DNA, submitted to 16S rRNA gene sequencing, and bioinformatically analyzed to infer the bacterial populations. Microbiota changes through time were reported for both groups. However, we detected an earlier stabilization in the healthy group. Moreover, we detected changes within low abundant taxa that may play a role in the subsequent health status of the animals. The fecal microbiota of healthy and diarrheic calves showed different dynamics in the diversity through time that may be the reflections of the variations within low-abundant taxa

Using compositional mixed-effects models to evaluate responses to amino acid supplementation in milk replacers for calves

The consequences of supplementing Lys, Met, and Thr in milk replacers (MR) for calves have been widely studied, but scarce information exists about potential roles of other AA (whether essential or not). The effects on growth performance of supplementation of 4 different AA combinations in a mixed ration (25.4% crude protein and 20.3% fat) based on skim milk powder and whey protein concentrate were evaluated in 76 Holstein male calves (3 ± 1.7 d old). The 4 MR were as follows: CTRL with no AA supplementation; PG, supplying additional 0.3% Pro and 0.1% Gly; FY, supplying additional 0.2% Phe and 0.2% Tyr; and KMT, providing additional 0.62% Lys, 0.22% Met, and 0.61% Thr. All calves were fed the same milk allowance program and were weaned at 56 d of study. Concentrate intake was limited to minimize interference of potential differences in solid feed intake among treatments. Animals were weighed weekly, intakes recorded daily, and blood samples obtained at 2, 5, and 7 wk of study to determine serum urea and plasma AA concentrations. Plasma AA concentrations were explored using compositional data analysis, and their isometric log-ratio transformations were used to analyze their potential influence on ADG and serum urea concentration using a linear mixed-effects model. We detected no differences in calf performance and feed intake. Plasma relative concentration of the AA supplemented in the KMT and PG treatments increased in their respective treatments, and, in PG calves, a slight increase in the proportion of plasma Gly, Glu, and branched-chain AA was also observed. The proportions of plasma branched-chain AA, His, and Gln increased, and those of Thr, Arg, Lys, and Glu decreased with calves' age. A specific log-contrast balance formed by Arg, Thr, and Lys was found to be the main driver for lowering serum urea concentrations and increasing calf growth. The use of compositional mixed-effects models identified a cluster formed by the combination of Arg, Thr, and Lys, as a potential AA to optimize calf growth.Peer ReviewedPostprint (published version

Short communication: Recombinant mammary serum amyloid A3 as a potential strategy for preventing intramammary infections in dairy cows at dryoff

Mammary serum amyloid A3 (M-SAA3) has shown potential in stimulating innate immunity during intramammary infections, at calving and at dryoff. In this study, we produced recombinant caprine M-SAA3 to test its ability to reduce intramammary infections with Staphylococcus aureus, Streptococcus uberis, Streptococcus dysgalactiae, and Escherichia coli, which are all common mastitis-producing pathogens. Recombinant production of M-SAA3 (followed by lipopolysaccharide removal to avoid lipopolysaccharide-nonspecific stimulation of the immune system) was successfully achieved. Mammary serum amyloid A3 stimulated the expression of IL-8 in a dose-dependent manner in primary mammary cultures. Although a direct killing effect on Staph. aureus by M-SAA3 was not detected, this acute phase protein was able to reduce Staph. aureus, Strep. uberis, and Strep. dysgalactiae infections by up to 50% and induced a reduction in E. coli counts of 67%. In general, the best concentration of caprine M-SAA3 for inhibiting infections was the lowest concentration tested (10 μg/mL), although higher concentrations (up to 160 μg/mL) increased its antimicrobial potential against some pathogens.info:eu-repo/semantics/acceptedVersio

Pre-calving Intravaginal Administration of Lactic Acid Bacteria Reduces Metritis Prevalence and Regulates Blood Neutrophil Gene Expression After Calving in Dairy Cattle

Metritis affects up to 40% of dairy cows and it is usually treated with antibiotics. In spite of their advantages, there is an increased concern about antibiotic resistance leading to the research of alternative methods. The aim of this study was to evaluate the effects of a combination of lactic acid bacteria (LAB) on the prevalence of metritis and modulation of endometrial and neutrophil inflammatory markers in dairy cows. One hundred and thirty-five cows were enrolled 3 week before calving and randomly assigned to three treatments. Treatment groups were: (1) two intravaginal doses of LAB/wk during 3 week pre-calving (vaginal, n = 45); (2) an intra-uterine dose, once 1 d after calving (uterine, n = 44); and (3) no intervention (CTRL, n = 45). Metritis was defined as body temperature > 39.5°C and purulent vaginal discharge (> 50% pus), and diagnosed 6 d after calving. Blood samples were taken at d −14, −10, −7, −4, +1, +3, +6, and +14 relative to calving for non-esterified fatty acids (NEFA) analysis. At d −10, +1, +3, and +6 neutrophils were isolated from blood for gene expression analysis by RT-qPCR. Endometrium biopsies were taken from 30 cows, 15 from CTRL and 15 from the uterine group at d +1, +3, and +6 after calving for pro-inflammatory markers analysis by NanoString®. Vaginal treatment reduced metritis prevalence (6/45) up to 58% compared with CTRL group (14/45), but there was no difference between the uterine and CTRL group. Uterine and vaginal treatments reduced blood neutrophil gene expression. Expression of pro-inflammatory markers in the endometrium did not differ between uterine and CTRL cows. Metritic cows expressed more C-X-C motif chemokine ligand 8 (CXCL8) and interleukin 1 beta (IL1B) at d 3 than healthy cows, whereas healthy cows expressed more CXCL8 at d 1 relative to calving in the endometrium. This study shows a promising potential of LAB probiotics as a preventive treatment against metritis in dairy cows

Pre-calving intravaginal administration of lactic acid bacteria reduces metritis prevalence and regulates blood neutrophil gene expression after calving in dairy cattle

Metritis affects up to 40% of dairy cows and it is usually treated with antibiotics. In spite of their advantages, there is an increased concern about antibiotic resistance leading to the research of alternative methods. The aim of this study was to evaluate the effects of a combination of lactic acid bacteria (LAB) on the prevalence of metritis and modulation of endometrial and neutrophil inflammatory markers in dairy cows. One hundred and thirty-five cows were enrolled 3 week before calving and randomly assigned to three treatments. Treatment groups were: (1) two intravaginal doses of LAB/wk during 3 week pre-calving (vaginal, n = 45); (2) an intra-uterine dose, once 1 d after calving (uterine, n = 44); and (3) no intervention (CTRL, n = 45). Metritis was defined as body temperature > 39.5◦C and purulent vaginal discharge (> 50% pus), and diagnosed 6 d after calving. Blood samples were taken at d −14, −10, −7, −4, +1, +3, +6, and +14 relative to calving for non-esterified fatty acids (NEFA) analysis. At d −10, +1, +3, and +6 neutrophils were isolated from blood for gene expression analysis by RT-qPCR. Endometrium biopsies were taken from 30 cows, 15 from CTRL and 15 from the uterine group at d +1, +3, and +6 after calving for pro-inflammatory markers analysis by NanoString®. Vaginal treatment reduced metritis prevalence (6/45) up to 58% compared with CTRL group (14/45), but there was no difference between the uterine and CTRL group. Uterine and vaginal treatments reduced blood neutrophil gene expression. Expression of pro-inflammatory markers in the endometrium did not differ between uterine and CTRL cows. Metritic cows expressed more C-X-C motif chemokine ligand 8 (CXCL8) and interleukin 1 beta (IL1B) at d 3 than healthy cows, whereas healthy cows expressed more CXCL8 at d 1 relative to calving in the endometrium. This study shows a promising potential of LAB probiotics as a preventive treatment against metritis in dairy cows.info:eu-repo/semantics/publishedVersio

Potential of MMP-9 based nanoparticles at optimizing the cow dry period : pulling apart the effects of MMP-9 and nanoparticles

Altres ajuts: ICREA Academia awardThe cow dry period is a non-milking interval where the mammary gland involutes and regenerates to guarantee an optimal milk production in the subsequent lactation. Important bottlenecks such as the high risk of intramammary infections complicate the process. Antibiotics have been routinely used as a preventive treatment but the concerns about potential antibiotic resistance open a new scenario in which alternative strategies have to be developed. Matrix metalloproteinase-9 (MMP-9) is an enzyme able to degrade the extracellular matrix, triggering the involution and immune function of cow mammary gland. We have studied the infusion into the mammary gland of MMP-9 inclusion bodies as protein-based nanoparticles, demonstrating that 1.2 mg of MMP-9 enhanced the involution and immune function of the cow mammary gland. However, the comparison of the effects triggered by the administration of an active and an inactive form of MMP-9 led to conclude that the response observed in the bovine mammary gland was mainly due to the protein format but not to the biological activity of the MMP-9 embedded in the inclusion body. This study provides relevant information on the future use of protein inclusion bodies in cow mammary gland and the role of MMP-9 at dry-off

Factors associats a un brot de malaltia boca-mà-peu en una llar d’infants

Malaltia boca-mà-peu; Brot de malaltia boca-mà-peu; Brot; Llar d’infants; Mesures de controlEnfermedad boca-mano-pie; Brote de enfermedad boca-mano-pie; Brote; Guardería; Medidas de controlHand, foot, and mouth disease; Outbreak of hand, foot, and mouth disease; Outbreak; Kindergarten; Control measuresIntroducció. La malaltia boca-mà-peu (MBMP) és una infecció comuna, altament contagiosa i amb una incidència molt més alta en infants menors de 5 anys. Se’n va fer un estudi amb l’objectiu de descriure els factors associats a un brot de MBMP que va tenir lloc en una llar d’infants. Mètodes. Es va dur a terme un estudi epidemiològic de cohorts. Es va dissenyar una enquesta en la qual es van recollir dades de filiació, del diagnòstic de la MBMP i dels símptomes associats. Les enquestes epidemiològiques es van realitzar per via telefònica amb els pares. Les dades es van introduir al programa Epi Info versió 7.2.2.6 per fer-ne l’anàlisi. Es va considerar l’existència de diferències estadísticament significatives amb un valor p<0,05 i es va calcular el risc relatiu amb un interval de confiança del 95%. Resultats. La taxa d’atac entre els alumnes va ser del 29% (18/62). El primer cas va iniciar símptomes el 27 d’abril i el darrer, l’11 de maig. En la inspecció sanitària inicial es van establir les mesures de control. L’aula on es va enregistrar el major nombre d’afectats va ser la de 0-1 anys, amb una taxa d’atac del 73,3% (11/15) i va ser en aquesta aula on va aparèixer el primer cas del brot. La mitjana d’edat dels casos va ser de 19,8 mesos amb una desviació estàndard de 7,1 i en els no casos de 29,4 mesos amb una desviació estàndard de 8,2 (p<0,01). Conclusions. L’actuació ràpida en la instauració de les mesures de control pot haver contribuït a limitar el nombre de casos de MBMP i la durada del brot.Introducción. La enfermedad boca-mano-pie (EBMP) es una infección común, altamente contagiosa y con una incidencia mucho más alta en niños menores de 5 años. Se hizo un estudio con el objetivo de describir los factores asociados a un brote de EBMP que tuvo lugar en una guardería. Métodos. Se llevó a cabo un estudio epidemiológico de cohortes. Se diseñó una encuesta en la que se recogieron datos de filiación, del diagnóstico de la EBMP y de los síntomas asociados. Las encuestas epidemiológicas se realizaron por vía telefónica con los padres. Los datos se introdujeron en el programa Epi Info versión 7.2.2.6 para su análisis. Se consideró la existencia de diferencias estadísticamente significativas con un valor p <0,05 y se calculó el riesgo relativo con un intervalo de confianza del 95%. Resultados. La tasa de ataque entre los alumnos fue del 29,03% (18/62). El primer caso inició síntomas el 27 de abril y el último, el 11 de mayo. En la inspección sanitaria inicial se establecieron las medidas de control. El aula donde se registró el mayor número de afectados fue la de 0-1 años, con una tasa de ataque del 73,3% (11/15) y fue en esta aula donde apareció el primer caso del brote. La media de edad de los casos fue de 19,8 meses con una desviación estándar de 7,1 y en los no casos, de 29,4 meses con una desviación estándar de 8,2 (p <0,01). Conclusiones. La actuación rápida en la instauración de las medidas de control pudo haber contribuido a limitar el número de casos de MBMP y la duración del brote.Introduction: Hand, foot, and mouth disease (HFMD) is a common, highly contagious infection and its incidence is much higher in children under 5 years of age. The objective of this study was to describe the factors associated with an outbreak of HFMD that took place in a nursery. Methods. An epidemiological cohort study was carried out. A questionnaire was designed to collect data related to affiliation, diagnosis of HFMD and associated symptoms. An epidemiological survey was conducted by telephone with these children’s parents. Data were introduced in the Epi Info version 7.2.2.6 programme for analysis. The existence of statistically significant differences was considered with a p <0.05 and the risk ratio was calculated with its confidence interval of 95%. Results. The attack rate among the children was 29.03% (18/62). The first case started symptoms on April 27 and the last on May 11. In the preliminary health inspection control measures were established. The class group aged 0-1 presented the highest number of children affected, with an attack rate of 73.3% (11/15) and where the first case of the outbreak appeared. The mean age of the cases was 19.8 months with a standard deviation of 7.1 and 29.4 months in non-cases with a standard deviation of 8.2 (p <0.01). Conclusions. A rapid action in establishing control measures could have helped to limit the number of cases and the duration of the outbreak

A new approach to obtain pure and active proteins from Lactococcus lactis protein aggregates

The production of pure and soluble proteins is a complex, protein-dependent and time-consuming process, in particular for those prone-to-aggregate and/or difficult-to-purify. Although Escherichia coli is widely used for protein production, recombinant products must be co-purified through costly processes to remove lipopolysaccharide (LPS) and minimize adverse effects in the target organism. Interestingly, Lactococcus lactis, which does not contain LPS, could be a promising alternative for the production of relevant proteins. However, to date, there is no universal strategy to produce and purify any recombinant protein, being still a protein-specific process. In this context and considering that L. lactis is also able to form functional protein aggregates under overproduction conditions, we explored the use of these aggregates as an alternative source of soluble proteins. In this study, we developed a widely applicable and economically affordable protocol to extract functional proteins from these nanoclusters. For that, two model proteins were used: mammary serum amyloid A3 (M-SAA3) and metalloproteinase 9 (MMP-9), a difficult-to-purify and a prone-to-aggregate protein, respectively. The results show that it is possible to obtain highly pure, soluble, LPS-free and active recombinant proteins from L. lactis aggregates through a cost-effective and simple protocol with special relevance for difficult-to-purify or highly aggregated proteins