Effects of feeding pasteurized waste milk or saleable milk to calves on weight, health and fecal Escherichia coli antimicrobial resistance.

The aim of this study was to compare the effects of feeding pasteurized waste milk or saleable milk to calves on weight, health and emergence of antimicrobial resistance in Escherichia coli strains isolated from those calves. An experimental study under field conditions on a commercial pasture-based Argentinian dairy farm was carried out. Forty Holstein calves were assigned randomly to either pasteurized waste milk (PWM) or non-pasteurized saleable milk (SM). The antimicrobial agents (AM) used on the farm, both to treat or prevent diseases, were recorded. The passive immunity level, calf live weight, AM presence in milk, clinical examination of calves, and E. coli isolation and identification, were performed. A total of 258 E. coli strains were isolated from fecal samples (132 isolates from SM calves and 126 from PWM calves at six sampling times). All E. coli isolated were used to perform AM susceptibility tests (disc diffusion and agar dilution). No differences were observed between groups in health parameters, average daily gain or prevalence of resistant E. coli strains to any AM evaluated throughout the study. Peaks of trimethoprim, sulfamethoxazole and enrofloxacin minimum inhibitory concentration (MIC) were observed at 30 d in E. coli from both groups of calves, whilst additional peaks to tetracyclin and ampicillin were observed only in SM calves. All MIC apart from gentamicin decreased at 75 and 90 d of age (during the weaning period). Gentamicin MIC behaved differently, having no peaks and increasing at 90 d only in PWM group. In conclusion, we found no evidence that emergence of antibiotic resistance is related to the consumption of pasteurized waste milk.

Antimicrobial resistance of Streptococcus uberis isolated from bovine mastitis: Systematic review and meta-analysis.

Streptococcus uberis is one of the most common pathogens associated with bovine mastitis, commonly treated with antimicrobials (AM), favoring the appearance of antimicrobial resistance (AMR). The objective of this work was to determine the proportion of phenotypic AMR among S. uberis isolated worldwide from bovine intramammary infections between the years 1983-2022, and to assess the variables associated by means of a systematic review and metanalysis. Sixty articles were eligible for quantitative review. Ninety-four independent studies were obtained. The antimicrobials evaluated in more S. uberis strains were penicillin (21,987 strains), oxacillin (21,727 strains), erythromycin (20,013 strains), and ampicillin (19,354 strains). Most of the studies included in this meta-analysis were from Europe (44), followed by America (25), Africa (10), Asia (10), and Oceania (5). Among the included articles, 22 were published from 1983 to 2006, 23 from 2007 to 2012, 25 from 2013 to 2015, and the remaining 24 after 2016. Penicillin, erythromycin, and tetracycline were the antimicrobials with >25 studies. Therefore, the following analyses were performed only for these antimicrobials, presenting a high heterogeneity index (I2). The variability observed for penicillin and tetracycline was only explained, partially, by continent of origin. The variability observed for erythromycin was not explained by any of the potential explanatory variables included in this study. The S. uberis proportion of resistance to antimicrobials is highly variable and probably influenced by many factors other than those studied in this meta-analysis, where it was not possible to inform a unique average proportion of resistance.

Teat-apex colonizer Bacillus from healthy cows antagonizes mastitis-causing Staphylococcus aureus biofilms.

Staphylococcus aureus is the most frequent causal agent of bovine mastitis, which is largely responsible for milk production losses worldwide. The pathogen's ability to form stable biofilms facilitates intramammary colonization and may explain disease persistence. This virulence factor is also highly influential in the development of chronic intramammary infections refractory to antimicrobial therapy, which is why novel therapies that can tackle multiple targets are necessary. Since udder microbiota have important implications in mastitis pathogenesis, they offer opportunities to develop alternative prophylactic and therapeutic strategies. Here, we observed that a Bacillus strain from the teat apex of lactating cows was associated to reduce colonization by S. aureus. The strain, identified as Bacillus sp. H21, was able to antagonize in-formation or mature S. aureus biofilms associated to intramammary infections without affecting cell viability. When exploring the metabolite responsible for this activity, we found that a widespread class of Bacillus exopolysaccharide, levan, eliminated the pathogenic biofilm under evaluated conditions. Moreover, levan had no cytotoxic effects on bovine cellular lines at the biologically active concentration range, which demonstrates its potential for pathogen control. Our results indicate that commensal Bacillus may counteract S. aureus-induced mastitis, and could therefore be used in novel biotechnological strategies to prevent and/or treat this disease.

Improvement of bovine pestiviral diagnosis by the development of a cost-effective method for detecting viral RNA in fresh specimens and samples spotted in filter papers / Optimización del diagnóstico de Pestivirus bovinos mediante el desarrollo de una metodología económica para detectar ARN viral en muestras frescas y fijadas en papeles de filtro

Abstract Bovine pestiviruses are the causative agents of bovine viral diarrhea, a disease thatcauses severe economic losses in cattle. The aim of this study was to improve their diagnosisby developing a RT-qPCR to detect bovine pestiviruses A, B and H; and to set up a protocolfor collecting, shipping and preserving bovine pestiviral RNA on filter papers. The developedRT-qPCR showed high sensitivity in detecting these viruses in different matrices: viral stocks,semen and serum samples. With regard to the possibility of using the technique to test serumpools, it was possible to identify a positive serum sample within a pool containing 30 sera.In addition to evaluating the qPCR from fresh samples, the use of filter papers to sow bovinesamples was analyzed. The sampling method on two different filter papers using bovine blooddrops was a useful alternative for diagnostic purposes and allowed to preserve pestiviral RNAfor up to 12 months under refrigeration.

Resumen Los Pestivirus bovinos son los agentes causales de la diarrea viral bovina, una enfermedad que genera importantes pérdidas económicas en el ganado vacuno. El objetivo de este trabajo fue mejorar su diagnóstico mediante el desarrollo de una RT-qPCR para detectar los Pestivirus bovinos A, B y H y disenar un protocolo de recolección, envío y conservación de ARN viral en papeles de filtro. La RT-qPCR desarrollada demostró alta sensibilidad en la detección de estos virus en diferentes matrices: stock viral, suero y semen. Respecto de la posibilidad de usar la técnica para testear pools de suero, fue posible identificar un suero positivo dentro de un pool compuesto por 30 sueros. Además de evaluar la qPCR en muestras frescas, se analizó el uso de papeles de filtro para sembrar muestras de bovinos. La metodología de toma de muestras en dos tipos de papeles de filtro usando gotas de sangre fue una alternativa útil para el diagnóstico y permitió conservar ARN viral por hasta 12 meses a temperaturas de refrigeración.

In vitro evaluation of ginsenoside Rg1 immunostimulating effect in bovine mononuclear cells.

The aim of this study was to evaluate the immunomodulatory effect of ginsenoside Rg1 on mammary secretion and peripheral blood mononuclear cells (MSMC and PBMC, respectively). The mRNA expression of TLR2, TLR4 and selected cytokines were evaluated on MSMC after Rg1 treatment. Also, TLR2 and TLR4 protein expression was evaluated on MSMC and PBMC after Rg1 treatment. Phagocytic activity and capacity, ROS production and MHC-II expression were evaluated on MSMC and PBMC after Rg1 treatment and co-culture with Staphylococcus aureus strain 5011. Rg1 induced mRNA expression of TLR2, TLR4, TNF-α, IL-1ß, IL-6 and IL-8 in groups treated with different concentrations and at different times in MSMC, and induced TLR2 and TLR4 protein expression in MSMC and PBMC. Rg1 increased phagocytic capacity and ROS production in MSMC and PBMC. Rg1 increased MHC-II expression by PBMC. However, Rg1 pre-treatment had no effect on cells co-cultured with S. aureus. In conclusion, Rg1 was able to stimulate several sensing and effector activities in these immune cells.

Improvement of bovine pestiviral diagnosis by the development of a cost-effective method for detecting viral RNA in fresh specimens and samples spotted in filter papers.

Bovine pestiviruses are the causative agents of bovine viral diarrhea, a disease that causes severe economic losses in cattle. The aim of this study was to improve their diagnosis by developing a RT-qPCR to detect bovine pestiviruses A, B and H; and to set up a protocol for collecting, shipping and preserving bovine pestiviral RNA on filter papers. The developed RT-qPCR showed high sensitivity in detecting these viruses in different matrices: viral stocks, semen and serum samples. With regard to the possibility of using the technique to test serum pools, it was possible to identify a positive serum sample within a pool containing 30 sera. In addition to evaluating the qPCR from fresh samples, the use of filter papers to sow bovine samples was analyzed. The sampling method on two different filter papers using bovine blood drops was a useful alternative for diagnostic purposes and allowed to preserve pestiviral RNA for up to 12 months under refrigeration.

Differential immune response to two Staphylococcus aureus strains with distinct adaptation genotypes after experimental intramammary infection of dairy cows.

The aim of this study was to evaluate and compare the ability of two S. aureus strains with different adaptation genotypes (low and high) to the bovine mammary gland (MG) to establish an intramammary infection (IMI) and induce an immune response after an experimental challenge in lactating cows. Two isolates (designated 806 and 5011) from bovine IMI with different genotypic profiles, harboring genes involved in adherence and biofilm production, belonging to different capsular polysaccharide (CP) type, accessory gene regulator (agr) group, pulsotype (PT) and sequence type/clonal complex (ST/CC) were selected. Strains 806 and 5011 were associated with low (nonpersistent-NP) and high (persistent-P) adaptation to the MG, respectively. Strain 806 (NP) was characterized as agr group II, cap5 positive and ST350; strain 5011 (P) agr group I, cap8 positive and CC188. Three groups of clinically healthy cows, 4 cows/treatment group, were inoculated by the intramammary route with strain 806 (NP), strain 5011 (P) and pyrogen-free saline solution. All mammary quarters challenged with strain 806 (NP) developed mild clinical mastitis between 1 and 7 d post inoculation (pi). Quarters challenged with strain 5011 (P) developed a persistent IMI; bacteria were recovered from milk from d 7 pi and up to d 56 pi. In quarters inoculated with strain 806 (NP) the inflammatory response induced was greater and earlier than the one induced by strain 5011 (P), since a somatic cell count (SCC) peak was observed at d 2 pi, while in quarters inoculated with strain 5011 (P) no variations in SCC were observed until d 4 pi reaching the maximum values at d 14 pi; indicating a lower and delayed initial inflammatory response. The highest levels of nitric oxide (NO) and lactoferrin (Lf) detected in milk from quarters inoculated with both S. aureus strains coincided with the highest SCC at the same time periods, indicating an association with the magnitude of inflammation. The high levels of IL-1ß induced by strain 806 (NP) were associated with the highest SCC detected (d 2 pi); while quarters inoculated with strain 5011 (P) showed similar IL-1ß levels to those found in control quarters. In quarters inoculated with strain 806 (NP) two peaks of IL-6 levels on d 2 and 14 pi were observed; while in quarters inoculated with strain 5011 (P) IL-6 levels were similar to those found in control quarters. The strain 806 (NP) induced a higher total IgG and IgG1 response; while strain 5011 (P) generated a higher IgG2 response (even against the heterologous strain). The present study demonstrated that S. aureus strains with different genotype and adaptability to bovine MG influence the local host immune response and the course and severity of the infectious process.

A safe and effective vaccine against bovine leukemia virus.

Previous attempts to develop a vaccine against bovine leukemia virus (BLV) have not been successful because of inadequate or short-lived stimulation of all immunity components. In this study, we designed an approach based on an attenuated BLV provirus by deleting genes dispensable for infectivity but required for efficient replication. The ability of the vaccine to protect from natural BLV infection was investigated in the context of dairy productive conditions in an endemic region. The attenuated vaccine was tested in a farm in which the prevalence rose from 16.7% in young cattle at the beginning of the study to more than 90% in adult individuals. Sterilizing immunity was obtained in 28 out of 29 vaccinated heifers over a period of 48 months, demonstrating the effectiveness of the vaccine. As indicated by the antiviral antibody titers, the humoral response was slightly reduced compared to wild-type infection. After initial post-vaccination bursts, the proviral loads of the attenuated vaccine remained most frequently undetectable. During the first dairy cycle, proviral DNA was not detected by nested-PCR in milk samples from vaccinated cows. During the second dairy cycle, provirus was sporadically detected in milk of two vaccinated cows. Forty-two calves born from vaccinated cows were negative for proviral DNA but had antiviral antibodies in their peripheral blood. The attenuated strain was not transmitted to sentinels, further supporting the safety of the vaccine. Altogether, these data thus demonstrate that the vaccine against BLV is safe and effective in herd conditions characterized by a very high incidence. This cost-effective approach will thus decrease the prevalence of BLV without modification of production practices. After facing a series of challenges pertaining to effectiveness and biosafety, the vaccine is now available for further large-scale delivery. The different challenges and hurdles that were bypassed may be informative for the development of a vaccine against HTLV-1.

Detection by multiplex PCR of Mycoplasma species associated with dairy cattle in Argentina / Detección por PCR multiplex de especies de Mycoplasma asociadas a ganado lecheroen Argentina

Abstract There is scarce information about the frequency and epidemiological and clinicalfeatures associated with the presence of Mycoplasma spp. in Argentine dairy herds. The objec-tives of this study were to develop a multiplex PCR for identifying M. bovis and M. canadenseand to describe the frequency of Mycoplasma spp. isolated from clinical samples submitted to adiagnostic laboratory. Of a total of 1548 samples from intramammary infections, bulk tank milkand biological fluids, 38 Mycoplasma isolates were obtained. M. bovis, M. canadense, M. cali-fornicum and M. leachii were detected by using two multiplex PCRs, confirming their presencein clinical conditions in dairy cattle. The techniques used in the present study can be usefulto broaden the knowledge about Mycoplasma infections in cattle, since the search for theseorganisms is not usually included in routine diagnoses.

Resumen Existe poca información sobre la frecuencia, así como las características epidemi-ológicas y clínicas asociadas con la presencia de Mycoplasma en los rodeos lecheros argentinos.Los objetivos de este estudio fueron desarrollar una PCR multiplex para identificar M. bovis yM. canadense y describir la frecuencia de especies de Mycoplasma aisladas de muestras clíni-cas enviadas a un laboratorio de diagnóstico. De un total de 1.548 muestras de infeccionesintramamarias, leche de tanque de frío y fluidos biológicos, se obtuvieron 38 aislamientos de Mycoplasma. Mediante 2 PCR multiplex se detectaron M. bovis, M. canadense, M. californicumy M. leachii, confirmando su presencia en síndromes clínicos en ganado lechero. Las técnicasutilizadas en el presente estudio pueden ser útiles para ampliar el conocimiento sobre las infec-ciones por Mycoplasma en bovinos, ya que la búsqueda de estos organismos no suele incluirseen los diagnósticos de rutina.

Detection by multiplex PCR of Mycoplasma species associated with dairy cattle in Argentina.

There is scarce information about the frequency and epidemiological and clinical features associated with the presence of Mycoplasma spp. in Argentine dairy herds. The objectives of this study were to develop a multiplex PCR for identifying M.bovis and M.canadense and to describe the frequency of Mycoplasma spp. isolated from clinical samples submitted to a diagnostic laboratory. Of a total of 1548 samples from intramammary infections, bulk tank milk and biological fluids, 38 Mycoplasma isolates were obtained. M. bovis, M. canadense, M.californicum and M.leachii were detected by using two multiplex PCRs, confirming their presence in clinical conditions in dairy cattle. The techniques used in the present study can be useful to broaden the knowledge about Mycoplasma infections in cattle, since the search for these organisms is not usually included in routine diagnoses.

Involvement of matrix metalloproteinases and their inhibitors in Staphylococcus aureus chronically infected bovine mammary glands during active involution.

The aim of this study was to characterize the protein expression of matrix metalloproteinase-2 (MMP-2) and -- 9 and their inhibitors (TIMP-1 and -2) in mammary tissue of dairy cows with naturally occurring chronic S. aureus intramammary infections (IMI) during active involution. Moreover, the gelatinolytic activity of MMP-2 and -9 in mammary secretions was evaluated. Cows in late lactation that were either uninfected or with chronic naturally acquired S. aureus IMI were included in this study. Protein expression of MMP-2 and -9 in mammary tissues was significantly higher in S. aureus-infected than uninfected quarters at day 14 and 21 of involution. Protein expression of TIMP-1 and -2 was significantly higher in S. aureus-infected than uninfected quarters at day 7, 14 and 21 of involution. The MMP-2/TIMP-1, MMP-2/TIMP-2, MMP-9/TIMP-1 and MMP-9/TIMP-2 ratios were significantly higher in S. aureus-infected compared with uninfected quarters at day 14 of involution. The MMP-2 activity was significantly higher in mammary secretions from S. aureus-infected compared with uninfected quarters at day 1, 2, 7 and 14 of involution. The MMP-9 activity was significantly higher in mammary secretions from infected quarters compared with uninfected quarters at day 7, 14 and 21 of involution. The increased expression of MMP-2 and -9 in mammary tissue as well as the high levels of activity observed in mammary secretion from infected quarters compared with uninfected quarters during active involution, strongly suggests that these gelatinases could contribute to degradation of mammary tissue components during chronic S. aureus IMI. The MMPs/TIMPs imbalance could lead to greater proteolysis and potentially more damage to mammary tissue in S. aureus-infected quarters.

Efficacy of Panax ginseng extract combined with cephalexin as a dry cow therapy.

Our objective was to evaluate the efficacy of intramammary administration, at drying-off, of a Panax ginseng extract (PGe) combined with cephalexin (Ceph) on the post-calving bacteriological cure rate of pre-existing intramammary infections (IMI) and on the occurrence of new IMI during the dry period. In addition, milk yield and somatic cell count (SCC) in the post-treatment lactation were evaluated. One hundred and eight late-lactation cows were randomly divided into two experimental groups and were treated at drying-off with Ceph alone or PGe combined with Ceph.Cure rates for IMI present at drying-off were similar for both treatments (OR = 0.95, 95% CI = 0.33-2.74). Cure rates for Staphylococcus aureus were lower (OR = 15.4, 95% CI = 1.66-142.52) in quarters treated with PGe + Ceph than in those treated with Ceph alone. Intramammary infusion of PGe + Ceph at drying-off had no effect on preventing new dry period IMI (OR = 0.75, 95% CI = 0.38-1.51), compared with infusion of Ceph alone. Milk production and SCC in the ensuing lactation were not affected by PGe + Ceph treatment. In conclusion, addition of PGe to dry cow therapy did not show any advantage over the use of dry cow therapy alone.

Antimicrobial resistance of Staphylococcus aureus isolated from bovine mastitis: Systematic review and meta-analysis.

Staphylococcus aureus, one of the main contagious mastitis pathogens worldwide, is characterized for causing chronic intramammary infections that respond poorly to antimicrobial therapy, disseminating within the herd leading to high economic losses. The aim of this study was to determine the prevalence of phenotypic resistance to antimicrobial agents among S. aureus collected worldwide in the context of bovine intramammary infections between the years 1969-2020. A systematic review was performed according to Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA). One hundred and fifty-five articles were eligible for quantitative review. Most of studies included in this meta-analysis were from Europe (88), followed by Asia (56), Latin America (39), Africa (32), North America (26), and Oceania (8). The highest overall prevalence of resistant S. aureus was against penicillin (pestimate 0.451, CI95 % 0.415-0.487), followed by clindamycin, erythromycin, and gentamycin (p-estimate = 0.149, 0.085, and 0.069, respectively). Ceftiofur and cephalotin presented the lowest overall prevalence of antimicrobial resistance (AMR, p-estimate = 0.020 and 0.015, respectively). The AMR to almost all the antimicrobials evaluated presented an increasing pattern over time, more apparent from 2009 onwards. The antimicrobials with a higher increase in their AMR prevalence over time were clindamycin, gentamycin, and oxacillin. Africa, Asia and Latin America were the continents with higher AMR to most compounds included in this study. No differences in AMR were detected regarding the clinical origin of the isolates (subclinical vs clinical mastitis) for almost all antibiotics evaluated. Differences in the method for testing AMR (disc diffusion method vs minimum inhibitory concentration) and type of study design for monitoring AMR were detected underscoring the importance of these variables as critical factors to enable comparisons for evaluating emergence of AMR.

A New Cage-Like Particle Adjuvant Enhances Protection of Foot-and-Mouth Disease Vaccine.

Foot-and-Mouth Disease (FMD) is an acute viral disease that causes important economy losses. Vaccines with new low-cost adjuvants that stimulate protective immune responses are needed and can be assayed in a mouse model to predict their effectiveness in cattle. Immunostimulant Particle Adjuvant (ISPA), also known as cage-like particle adjuvant, consisting of lipid boxes of dipalmitoyl-phosphatidylcholine, cholesterol, sterylamine, alpha-tocopherol, and QuilA saponin, was shown to enhance protection of a recombinant vaccine against Trypanosoma cruzi in a mouse model. Thus, in the present work, we studied the effects on the magnitude and type of immunity elicited in mice and cattle in response to a vaccine based on inactivated FMD virus (iFMDV) formulated with ISPA. It was demonstrated that iFMDV-ISPA induced protection in mice against challenge and elicited a specific antibody response in sera, characterized by a balanced Th1/Th2 profile. In cattle, the antibody titers reached corresponded to an expected percentage of protection (EPP) higher than 80%. EPP calculates the probability that livestock would be protected against a 10,000 bovine infectious doses challenge after vaccination. Moreover, in comparison with the non-adjuvanted iFMDV vaccine, iFMDV-ISPA elicited an increased specific T-cell response against the virus, including higher interferon gamma (IFNγ)+/CD8+ lymphocyte production in cattle. In this work, we report for first time that an inactivated FMDV serotype A vaccine adjuvanted with ISPA is capable of inducing protection against challenge in a murine model and of improving the specific immune responses against the virus in cattle.

Capacity of two Staphylococcus aureus strains with different adaptation genotypes to persist and induce damage in bovine mammary epithelial cells and to activate macrophages.

The aim of this study was to evaluate and compare the ability to adhere/internalize, persist, and induce damage in mammary epithelial cells (MAC-T) of two Staphylococcus aureus strains with different adaptation genotypes (low and high) to the bovine mammary gland (MG). Also, the phagocytic and bactericidal capacity induced after the interaction between macrophages, isolated from mammary secretion, of both S. aureus strains was evaluated. Two isolates (designated 806 and 5011) from bovine intramammary infection (IMI) harboring genes involved in adherence and biofilm production, belonging to different capsular polysaccharide (CP) type, accessory gene regulator (agr) group, pulsotype (PT) and sequence type/clonal complex (ST/CC). Strains 806 and 5011 were associated with low (nonpersistent-NP) and high (persistent-P) adaptation to the MG, respectively. Strain 5011 (P), agr group I, cap8 positive and strong biofilm producer showed higher capacity to adhere/internalize in MAC-T compared with strain 806 (NP), characterized as agr group II, cap5 positive and weak biofilm producer. Strain 5011(P) could be recovered from MAC-T lysates up to 72 h pi; while strain 806 (NP) could be recovered only at 4 h pi. Strain 5011 (P) showed greater capacity to induce apoptosis compared with strain 806 (NP) at 4, 24 and 48 h pi. Macrophages infected with strain 5011 (P) showed a greater phagocytic capacity and higher percentage of intracellular reactive oxygen species (ROS) production than strain 806 (NP). No viable bacteria were isolated from macrophages lysates stimulated with any of the S. aureus strains at 2, 4, 8 and 24 h pi. The knowledge of the molecular profile of the S. aureus strains causing bovine mastitis in a herd could become a tool to expose the most prevalent virulence gene patterns and advance in the elucidation of the pathogenesis of chronic mastitis.

Risk factors of S. aureus intramammary infection in pre partum dairy heifers under grazing conditions and molecular characterization of isolates from heifers and cows.

The aims of the research reported here were to identify potential risk factors associated with the presence of Staphylococcus aureus intramammary infection (IMI) in pre partum dairy heifers on 17 dairy farms from three provinces of Argentina and to characterize, at molecular level, isolates from those heifers and lactating cows from two selected herds. A total of 1474 heifers and 4878 lactating cows were studied. The prevalence of Staphylococcus aureus IMI in the heifers, heifers at quarter level and lactating cow mammary quarters was 14.41, 4.82, and 14.65%, respectively. Univariate analysis showed the key variables associated with S. aureus IMI presence in the heifers were: S. aureus IMI prevalence in cows of the lactating herd, the time calves stayed with their dam after birth, the calf rearing system, the place of rearing (own farm or other dairy farm) and fly control on the farm. None of the variables included in the multivariable analysis was associated with the presence of S. aureus IMI in the pre partum heifers, probably due to low variability among management practices used by the farms for rearing the heifer calves. At the molecular level, S. aureus isolates were grouped into three main PFGE clusters and several genotypes within the clusters. Isolates from mammary secretion of pre partum heifers and milk of lactating cows comprised different PFGE clusters in both herds, although two exceptions occurred. The absence of gene fnbpB, which codifies for a virulence factor protein involved in cell invasion by S. aureus, was significantly more frequent in pre partum heifer secretion isolates than in isolates from lactating cow milk. These results suggest that, under these management conditions, isolates from mammary secretions of pre partum heifers do not originate from the milk of lactating cows, but rather other sources to which the heifer is exposed.

BLV: lessons on vaccine development.

Vaccination against retroviruses is a challenge because of their ability to stably integrate into the host genome, undergo long-term latency in a proportion of infected cells and thereby escape immune response. Since clearance of the virus is almost impossible once infection is established, the primary goal is to achieve sterilizing immunity. Besides efficacy, safety is the major issue since vaccination has been associated with increased infection or reversion to pathogenicity. In this review, we discuss the different issues that we faced during the development of an efficient vaccine against bovine leukemia virus (BLV). We summarize the historical failures of inactivated vaccines, the efficacy and safety of a live-attenuated vaccine and the economical constraints of further industrial development.

Effects of chronic Staphylococcus aureus infection on immunological parameters and functionality of macrophages isolated from bovine mammary secretions.

The aim of this study was to characterize the effects of chronic S. aureus intramammary infection (IMI) on local innate and adaptive immune response during active involution. Cows in late lactation that were either uninfected or with chronic naturally acquired S. aureus IMI were included in this study. The levels of interleukin (IL)-1ß, IL-6 and IL-4 were significantly higher in mammary secretions of S. aureus-infected quarters compared with uninfected at d 7, 14 and 21 of involution. Lactoferrin (Lf), total IgG and S. aureus specific IgG1 levels were significantly lower in mammary secretions of infected quarters compared with uninfected during the first three weeks of involution. The amount of intracellular reactive oxygen species (ROS) produced per macrophage, was significantly higher in mammary secretions of infected quarters compared with uninfected at d 14 post drying off. Nitrite production was significantly higher in phagocytes from infected mammary secretions compared with uninfected at d 7 and 14 post drying off. Chronic S. aureus IMI altered normal secretion composition during bovine mammary gland involution. The high IL-1ß and IL-6 levels and increased functionality of macrophages in mammary secretions of infected quarters could be a result of the chronic inflammatory environment triggered by the presence of viable bacteria in mammary tissue. The lower levels of total and S. aureus specific antibodies and other immune factors in mammary secretion during this period may reduce the natural defense potential of the gland contributing to S. aureus persistence.