Search

Mastitis affects a high proportion of dairy cows and is still one of the greatest challenges faced by the dairy industry. Staphylococcal bacteria remain the most important cause of mastitis worldwide. We investigated how distinct staphylococcal species evade some critical host defense mechanisms, which may dictate the establishment, severity, and persistence of infection and the outcome of possible therapeutic and prevention interventions. Thus, the present study investigated variations among distinct bovine-associated staphylococci in their capability to resist phagocytosis and to trigger respiratory burst activity of blood and milk polymorphonuclear neutrophil leukocytes (PMNL) in dairy cows. To do so, PMNL of 6 primiparous and 6 multiparous dairy cows were used. A collection of 38 non-aureus staphylococci (NAS) and 12 Staphylococcus aureus were included. The phagocytosis and intracellular reactive oxygen species (ROS) production by blood and milk PMNL were analyzed by flow cytometry. Phagocytosis, by both blood and milk PMNL, did not differ between S. aureus and NAS as a group, although within-NAS species differences were observed. Staphylococcus chromogenes (a so-called milk-adapted NAS species) better resisted phagocytosis by blood PMNL than the so-called environmental (i.e., Staphylococcus fleurettii) and opportunistic (i.e., Staphylococcus haemolyticus) NAS species. Otherwise, S. haemolyticus was better phagocytosed by blood PMNL than S. aureus, S. fleurettii, and S. chromogenes. No influence of the origin of the isolates within the staphylococci species in the resistance to phagocytosis by blood and milk PMNL was found. Overall, both S. aureus and NAS did not inhibit intracellular ROS production in blood and milk PMNL. Non-aureus staphylococci induced fewer ROS by milk PMNL than S. aureus, which was not true for blood PMNL, although species-specific differences in the intensity of ROS production were observed. Staphylococcus chromogenes induced more blood PMNL ROS than S. fleurettii and S. haemolyticus, and as much as S. aureus. Conversely, S. chromogenes induced fewer milk PMNL ROS than S. aureus. The origin of the isolates within the staphylococci species did not affect the ROS production by blood and milk PMNL. In conclusion, our study showed differences in staphylococci species in evading phagocytosis and triggering ROS production, which may explain the ability of some staphylococci species (i.e., S. aureus and S. chromogenes) to cause persistent infection and induce inflammation.

This study aimed to evaluate the antimicrobial activity of clove (Syzygium aromaticum), citronella (Cymbopogon winteranius) and melaleuca (Melaleuca alternifolia) essential oils (EOs) on CoPS. Of the 67 cerumen samples collected, 17 CoPS strains were isolated. Using the microdilution method in a 96-well plate, EOs were added at a maximum concentration of 10% and minimum of 0.04%. It is concluded that the EOs of clove and citronella demonstrate antimicrobial activity against strains of CoPS isolated from canine external otitis. At the concentrations used, melaleuca essential oil did not show any bacteriostatic or bactericidal effects.

The purpose of this study was to evaluate the effect of protected niacin and yeast rich in chromium on the productive and metabolic performance of dairy cows under thermal stress. 46 lactating Holstein cows were divided into four treatments: protected niacin (GN), yeast rich in chromium (GCr), niacin+chromium (GNCr) and control (GC). Were measured: milk production, SCC, MUN, NEFA, BHB, glucose, cholesterol, insulin, cortisol, T4 and T3. There was an 8% increase in milk production in the GNCr. The use of these elements is an alternative for producers who need to maintain production during periods of thermal stress.

Backgrounds The present study explored the viability of bovine milk macrophages, their intracellular production of reactive oxygen and nitrogen species (RONS), and their phagocytosis of Staphylococcus aureus, as well as the profile of lymphocytes, from healthy udder quarters and udder quarters infected by Corynebacterium bovis. The study included 28 healthy udder quarters from 12 dairy cows and 20 udder quarters infected by C. bovis from 10 dairy cows. The percentages of macrophages and lymphocytes were identified by flow cytometry using monoclonal antibodies. Macrophage viability, RONS production, and S. aureus phagocytosis were evaluated by flow cytometry. Results Milk samples from quarters infected with C. bovis showed a lower percentage of macrophages but an increased number of milk macrophages per mL and a higher percentage of macrophages that produced intracellular RONS and phagocytosed S. aureus. No effect of C. bovis infection on macrophage viability was found. Udder quarters infected by C. bovis showed a higher percentage of T cells and CD4+ T lymphocytes, but no effect was found on the percentage of CD8+ CD4− T, CD8− CD4− T, or B lymphocytes. Conclusions Thus, our results corroborate, at least in part, the finding that intramammary infections by C. bovis may offer protection against intramammary infections by major pathogens.

The objective of this study was to evaluate the metabolic profile, the activity of blood neutrophils, and the immunoglobulin G levels of primiparous and multiparous sheep (Lacaune breed) during the first 30 days after lambing. Fifteen primiparous ewes (GPR) and 15 multiparous ewes (GPM) were used. Evaluations were performed on the days of lambing and at three, seven, fifteen and thirty days post-lambing. In general, the basal and bactericidal activity of neutrophils were lower in GPR than in GPM. Phagocytosis was greater in the primiparous sheep in the initial moments after lambing. Non-esterified fatty acid concentrations were highest for GPM, indicating a negative energy balance in this group. The results obtained in this study allow us to conclude that primiparous sheep have higher neutrophil phagocytosis, while this cell type has higher bactericidal activity in multiparous sheep. Multiparous ewes presented with higher lipomobilization due to maintenance needs and higher milk production.

The objective was to evaluate the influence of clinical (HC) and subclinical (HSC) hypocalcemia on the energetic metabolism and innate immune response of cows. The samples were divided into groups: Group 1: 74 samples (control), Group 2: 142 samples (HSC) and Group 3: 6 samples (HC). There was no difference in BHB and immune response. In G2 higher values of AGNE were detected on the day of delivery. In G3 higher levels of AGNE were observed on the 15th day postpartum in relation to the 30th day. Higher values of glucose were detected in G3 compared to G1 and G2. Thus, hypocalcemia did not cause alterations in the immune response but altered the energetic metabolism during the transition period.

Background : Bovine leukemia virus (VLB) is an oncogenic deltaretrovirus associated with the development of persistent lymphocytosis (LP) and lymphosarcomas in cattle. LP is characterized by chronic elevation of the number of circulating lymphocytes, in the case of B lymphocytes. Several studies have described functional changes in various leukocyte populations in both blood and milk in VLB-infected animals. The impact of some chronic diseases of low lethality is aggravated by the emergence of comorbidities.The objective of the present study was to evaluate the oxidative metabolism and neutrophil phagocytosis of bovines of the Holtein breed naturally infected with the bovine leukemia virus (VLB). Materials, Methods & Results : In this study, 20 cows were divided into three groups: (NG) seven non-seroreagent animals for VLB and without hematological alterations; (GAL) eight seroreagent animals for VLB and without hematological alterations; and (GLP) five seroreagent animals for VLB with persistent lymphocytosis (LP). The oxidative metabolism of neutrophils was determined by the tetrazolium nitroblast reduction test stimulated or not with Zymosan particles. The percentage of neutrophils that phagocytosed Zymosan particle (s) was also evaluated. The data were initially evaluated for normality and homoscedasticity by the Shapiro-Wilk test. Then the ANOVA test followed by the Student-Newman-Keuls test was applied for the comparison between the NG, GAL and GLP animals. Comparison between the NG animals and the seroreagent animals for the VLB (GVLB) was also performed through the unpaired Student's t - test . The value of P < 0.05 was considered significant. No significant differences were observed in oxidative neutrophil metabolism in stimulated and non-stimulated samples with Zymosan particles nor in the percentage of neutrophils that phagocytosed Zymosan particle (s) among the three experimental groups. However, as no differences were observed between the seroreagent animals for VLB with and without LP, we chose to divide the animals into only two experimental, non-seroreagent and seroreagent groups for VLB. Thus, when non-seroreagent animals for the VLB were compared with the seroreagent animals for the VLB, which corresponds to the GAL and GLP animals, a significant difference was observed in relation to the oxidative metabolism by neutrophils stimulated with Zymosan particles. Discussion : Some viral diseases are often associated with increased susceptibility to new infections and several studies have evaluated the role of peripheral blood mononuclear cells in VLB infection, but few studies have investigated neutrophil function. Some authors, when evaluating phagocytic capacity and oxidative metabolism, respectively, of blood leukocytes from VLB-infected animals, observed that VLB-infected animals displaying LP had lower phagocytic capacity and lower production of Reactive Oxygen Species (ROS). Some studies have shown that oxygen consumption by neutrophils was higher in experimentally infected sheep by VLB after 15 weeks of challenge, but this species is not a natural host of the virus, since transmission does not occur between sheep and cattle and the pathogenesis of infection by VLB is more acute in sheep, a result of the lower latency period for LP development. Other authors, when evaluating the interference of VLB in milk leukocytes, concluded that VLB-infected animals show lower intensity of intracellular ROS production by flow cytometry in VLB-infected animals, especially animals expressing LP, despite the fact that percentage of milk neutrophils that produced ROS did not differ between groups. It can be concluded that VLB interferes in neutrophilic function with possible implications for the health of VLB-infected animals and may favor secondary infections.

Despite the vast literature on postpartum mastitis in cattle, there is a demand for and lack of studies on the cellularity dynamics and cellular populations in milk and the occurrence of mastitis in ewes during the postpartum period. Thus, the aim of the present study was to evaluate the dynamics of cellularity in milk/colostrum and of mastitis in primiparous and multiparous Lacaune ewes during early lactation. For this purpose, 492 milk/colostrum samples were collected from the udder halves of 20 primiparous ewes and 21 multiparous ewes at the following six time points: day of parturition (T1) and one day (T2), three days (T3), seven days (T4), 15 days (T5), and 30 days (T6) after parturition. The milk and colostrum samples were subjected to microbiological examination, microscopic and automatic somatic cell counts (SCCs), differential somatic cell counts, and the California Mastitis test (CMT). The multiparous ewes had a higher prevalence of intramammary infections than the primiparous ewes during the postpartum period. Non-aureus staphylococci bacteria were the main mastitis etiological agents isolated in the present study. Furthermore, one of the most important findings of our study was that 35% of the udder halves of the primiparous ewes were infected at parturition, suggesting potential concerns for mastitis control programs. Finally, the SCC did not appear to be a good indicator of inflammation during early lactation because this measure was influenced by the parity and the sampling period. Moreover, the predictive values of the CMT and neutrophil counts were slightly better than those of the SCC during the immediate postpartum period.

The objective of the present study was to evaluate different methods for indirectly diagnosing mastitis during the postpartum period. These methods were: automatic and microscopic somatic cell counting (SCC), the California Mastitis Test (CMT) and Somaticell®. A total of 538 milk samples from 34 cows were used. These were collected at six times: day of parturition (M1) and 3 (M2), 7 (M3), 15 (M4), 21 (M5) and 30 (M6) days after parturition. Automatic and microscopic SCC, CMT and Somaticell® were all able to detect mastitis during the immediate postpartum period (up to 3 days postpartum). However, higher cut-off values should be applied to automatic and microscopic SCC. The negative score (score 0) of CMT was considered to be the best cut-off point at all times. Moreover, the values found using the Somaticell® test should not be used to presume the automatic SCC values, since there are discrepancies between the values of Somaticell® and automatic and microscopic SCC. It can be concluded that the different methods evaluated here to milk cellularity can be applied for diagnosing bovine mastitis, even during the immediate postpartum period, when there is greater cellularity, such as in the colostrum and transition milk.

RESUMO: A contagem de células somáticas (CCS) é um parâmetro amplamente utilizado para monitorar a saúde do úbere e a qualidade do leite, porém não diferencia as distintas populações leucocitárias. Portanto, a diferenciação das populações celulares no leite pode aprimorar o diagnóstico da mastite bovina. Dessa forma, o objetivo do presente trabalho foi avaliar as diferentes técnicas de contagem diferencial de leucócitos no leite para diagnosticar precisamente a mastite. Para tal, foram utilizadas 31 vacas da raça holandesa preta e branca em lactação (124 quartos mamários). Foram empregadas a contagem automática de células somáticas, e a contagem diferencial de leucócitos pelas técnicas de citocentrifugação, contagem diferencial de leucócitos por esfregaço direto, e citometria de fluxo com a utilização de anticorpos monoclonais específicos para identificação de cada população leucocitária. Os resultados demonstraram correlação positiva e significativa entre a proporção de leucócitos polimorfonucleares pelas diferentes técnicas e a contagem automática de células somáticas, sendo observada uma correlação discretamente mais forte com a citometria de fluxo. Além disso, foi demonstrado que os macrófagos são a população predominante no leite oriundo de glândula mamária com baixa CCS. Observaram-se também diferenças na proporção das distintas populações leucocitárias entre as distintas técnicas, resultado da possível subjetividade do examinador na contagem diferencial de leucócitos pelas técnicas de citocentrifugação e contagem microscópica direta por esfregaços, o que reforça que a citometria de fluxo pode ser uma ferramenta confiável no controle e diagnóstico da mastite. ABSTRACT: Milk somatic cell count (SCC) is the basis of mastitis and milk quality control programs, however it not differentiate the distinct leukocyte populations which in turn can improve the diagnosis of mastitis. Thus, the present study aimed to evaluate different techniques used to measure the distinct leukocyte populations in milk in attempt to improve the diagnosis of mastitis. Here, milk samples from 31 dairy cows (124 quarter milk samples) were used. The differential leukocytes count was determined by cytocentrifugation, direct microscopy smears, and monoclonal antibodies by flow cytometry. The automatic SCC was also performed. The results showed a positive and significant correlation between the proportion of polymorphonuclear leukocytes determined by all techniques and automatic cell count; although a discrete higher correlation between flow cytometry and automatic SCC was found. Furthermore, the present study reinforces the idea that macrophages were the predominant cell type in mammary gland with low SCC. The proportion of each leukocyte population differ among techniques, probably due to the subjectivity of the examiner in the evaluation of the differential leukocyte counts by cytocentrifugation and direct microscopy smears, which emphasize that flow cytometry can be a useful and feasible tool in the diagnosis and control of mastitis.

The immune response capacity of the mammary gland plays a major role to determine if mastitis will or not be established. Thus, we hypothesize that a better understanding of polymorphonuclear neutrophil leukocyte (PMN) function will elucidate mechanisms that will improve our knowledge of how we could avoid an inflammatory process by increasing the immune capacity of the cow, and even further, to search for a tool to diagnose mastitis or a possible way to select and identify non-susceptible animals. The present study utilized 112 quarters from 28 Holstein dairy cows that were divided into quarters milk samples with somatic cell count (SCC) 2×105 cells mL-1 (n=40). The percentages of milk PMNs and the levels of intracellular reactive oxygen species (ROS) and the phagocytosis of Staphylococcus aureus by milk neutrophils were evaluated by flow cytometry. Our results showed a higher percentage of neutrophils in quarter milk samples with high SCC (P=0.0003), and this group also had a significantly higher percentage of neutrophils that produced ROS (P=0.008). On the other hand, the phagocytosis intensity of S. aureus by milk neutrophils was higher in quarters with low SCC (P=0.003), suggesting a better mammary gland immunity against invading pathogens. Analyzing the results of the predictive values of the measured PMN functions, they cannot be used isolated as a good diagnosis test since none of them had a satisfactory sensitivity and specificity values, which was also confirmed by the Youden index values being far from one. In conclusion, the assessment of milk bovine neutrophil functions could improve our understanding of the cellular basis of mastitis. Although, the intracellular ROS production and S. aureus phagocytosis by milk neutrophil did not have high predictive values to detect intramammary infections, our results strengthen the idea that that poor bovine mammary gland neutrophil phagocytic ability may be associated with high SCC, and might be considered to identify susceptible dairy cows to mastitis.

This study examined neutrophil and monocyte functions and the blood lymphocyte profile of naturally BLV-infected cows with or without persistent lymphocytosis (PL). The percentage of neutrophils and monocytes that phagocytosed Staphylococcus aureus was lower in BLV-infected dairy cows, particularly those with PL. The relative percentage of CD44+ monocytes and neutrophils and CD11b expression by neutrophils was also lower in BLV-infected dairy cows with PL. A correlation between the percentage of CD11b+ neutrophils and that produced reactive oxygen species (ROS) was found. Furthermore, the percentage of CD44+ monocytes was positively correlated with the percentage of monocytes that phagocytosed S. aureus and the same phenomenon was observed for neutrophils. In BLV-infected dairy cows, particularly those with PL, inhibition of monocyte and neutrophil apoptosis was observed. Additionally, the percentage of neutrophils producing ROS was lower in BLV-infected cows with PL, in contrast to higher intensity of intracellular production of ROS by monocytes. The result from the lymphocyte immunophenotyping of BLV-infected cows with PL was an increase in B cells, mainly B CD5+ CD11b+, due to the apoptosis inhibition. In conclusion, this study provides novel insight into the implications of BLV infection for cattle, which can include the dysfunction of blood monocytes and neutrophils.

An important question about intramammary infections that is still debated in the literature is the independence or interdependence of the quarters of dairy cows. The present study sought to explore milk neutrophil function and the milk lymphocyte profile of uninfected quarters from uninfected and infected (one infected quarter per cow) udders to evaluate interdependence of the quarters. Thus, 32 (8 cows) and 18 (6 cows) uninfected quarters from uninfected and infected udders were used, respectively. Using flow cytometry, we evaluated the percentage of milk neutrophils and their expression of adhesion molecules L-selectin (CD62L), β2-integrin (CD11b), and an endothelial-selectin ligand (CD44); levels of intracellular reactive oxygen species (ROS); phagocytosis of Staphylococcus aureus by milk neutrophils; and neutrophil viability. Furthermore, we assessed the percentage of B-cell (CD21(+)) and T-lymphocyte subsets (CD3(+)/CD4(+)/CD8(-), CD3(+)/CD8(+)/CD4(-), CD3(+)/CD4(+)/CD25(-), CD3(+)/CD4(+)/CD25(+), and CD3(+)/CD4(-)/CD25(-)) using flow cytometry with monoclonal antibodies. The infected quarter did not affect somatic cell count or the percentage of neutrophils in the neighboring uninfected quarters. Furthermore, the infected quarter did not influence neutrophil viability, intracellular reactive oxygen species production, or phagocytosis of S. aureus by milk neutrophils. Conversely, the expression of adhesion molecules CD11b, CD62L, and CD44 by milk neutrophils differed between uninfected quarters from infected versus uninfected udders. The lymphocyte subsets did not differ between groups, except for a higher percentage of B cells in uninfected quarters from infected udders than in those from uninfected udders. Thus, our study strongly supports the hypothesis of interdependence of quarters based on the influence of infection on both the percentage of B cells and the expression of adhesion molecules by milk neutrophils in the neighboring uninfected quarters.

The exact influence of caprine arthritis encephalitis virus (CAEV) infection on blood and milk polymorphonuclear leukocytes (PMNLs) and monocyte/macrophages of goats remains unclear. Thus, the present study sought to explore the blood and milk PMNL and monocyte/macrophage functions in naturally CAEV-infected goats. The present study used 18 healthy Saanen goats that were segregated according to sera test outcomes into serologically CAEV negative (n=8; 14 halves) and positive (n=10; 14 halves) groups. All milk samples from mammary halves with milk bacteriologically positive outcomes, somatic cell count ≥2×106cellsmL-1, and abnormal secretions in the strip cup test were excluded. We evaluated the percentage of blood and milk PMNLs and monocyte/macrophages, the viability of PMNLs and monocyte/macrophages, the levels of intracellular reactive oxygen species (ROS) and the nonopsonized phagocytosis of Staphylococcus aureus and Escherichia coli by flow cytometry. In the present study, a higher percentage of milk macrophages (CD14+) and milk polymorphonuclear leukocytes undergoing late apoptosis or necrosis (Annexin-V+/Propidium iodide+) was observed in CAEV-infected goats; we did not find any further alterations in blood and milk PMNL and monocyte/macrophage functions. Thus, regarding our results, the goats naturally infected with CAEV did not reveal pronounced dysfunctions in blood and milk polymorphonuclear leukocytes and monocytes/macrophages.

The aim of this study was to investigate the relationship between clinical findings and bacterial isolation in milk samples of meat-producing ewes. The study was conducted in 17 commercial flocks and 550 udder halves from suckling Santa Ines ewes. Initially, the clinical examination of the mammary glands and teats was performed by visual inspection and palpation of the teats and udder halves; then a scoring system was devised for all the findings. After that, the strip cup test and the California mastitis test (CMT) were performed. Then, milk samples for somatic cell counts (SCCs) and bacteriological analyses were collected. Staphylococci bacteria were the main etiological agent isolated in the present study. Upon investigation of the correlations between bacterial isolation and the clinical findings, only the presence of teat injury, pendulous udder, and alterations in the palpation of the teat were associated with bacterial isolation. A significant correlation between bacteriologically positive milk samples and CMT and SCC was also found. Thus, some clinical findings appeared as a risk factor for bacteriologically positive milk samples and can be used as a tool in mastitis control programs. However, a complete and extensive diagnosis, an appropriate therapy, and an efficient mastitis control program will require the combination of clinical examination, microbiological tests, and SCC.

The most acceptable criteria for diagnosing bovine intramammary infections include results of bacteriological culture and measures of inflammation. Therefore, information on the diagnostic characteristics of the procedures used to identify infected quarters is required. Thus, this study was designed to evaluate a set of criteria to classify the infectious status of an udder at the quarter (single and duplicate milk samples) and cow (composite milk sample) levels, and to compare the infectious status with somatic cell counts (SCCs) of the samples. Here, the SCC thresholds determined by receiver operating characteristic curve analysis had a higher Youden index using mammary quarter duplicate milk samples as the gold standard for testing compared with single quarter and composite milk samples, especially for samples for which at least one of the duplicates was microbiologically positive, regardless of the mastitis pathogen isolated. The kappa coefficient for bacteriological results of the single quarter milk samples (single S1 and S2) was 0.85±0.019, indicating that single quarter milk sampling can be useful in mastitis control programs. Therefore, the use of composite milk samples to detect mastitis pathogens may be limited to the detection of major pathogens, given their predictive values. Thus, our findings suggest that the milk SCCs and microbiological examinations, although regarded as the most reliable indicators of ongoing mastitis, should be used in an integrated manner in mastitis control programs. Furthermore, the accuracy of single, duplicate and composite microbiological analyses to diagnosis mastitis should be considered for its implications in mastitis control strategies. Resumo: Os critérios mais aceitáveis para o diagnóstico das infecções intramamárias em bovinos incluem tanto os resultados da cultura bacteriológica e dos indicadores de inflamação. Portanto, a informação sobre os procedimentos mais adequados a serem utilizados para identificação dos quartos infectados é necessária. Assim, o objetivo do presente estudo foi avaliar um conjunto de critérios para identificação da infecção intramamária em bovinos pelo exame microbiológico (amostras individuais de leite simples ou em duplicata, e amostras de leite compostas), e comparar o isolamento do patógeno nas amostras de leite coletadas por distintos critérios com a contagem de células somáticas (CCS). Os valores de corte da CCS determinados pela curva de característica de operação do receptor demonstraram que a coleta de amostras de leite em duplicata apresentou o maior valor do índice de Youden, especialmente quando considerou-se o quarto mamário infectado se pelo menos uma das amostras de leite da duplicata apresentou resultado bacteriológico positivo independentemente do patógeno isolado. O coeficiente kappa dos resultados do exame microbiológico das amostras de leite individuais (amostra simples S1 e S2) foi de 0,85±0,019, indicando que a coleta de amostras de leite individual, ou seja, a coleta de uma amostra de leite por quarto mamário, pode ser utilizada nos programas de controle de mastite. Por outro lado, a coleta de amostras de leite compostas para detectar patógenos causadores de mastite deve ser limitada à detecção dos patógenos principais, considerando os valores preditivos encontrados no presente estudo. Portanto, os resultados do presente estudo indicam que a CCS e o exame microbiológico do leite, embora considerados como os critérios mais aceitos para o diagnóstico da mastite, devem ser utilizados de forma integrada em programas de controle de mastite. Além disto, os critérios de coleta de amostras de leite para o diagnóstico da mastite pelo exame microbiológico e seus valores preditivos devem ser considerados nos programas de controle de mastite.

One of the main concerns of natural orifice surgery is the local and systemic impact on physiology. Few studies have compared natural orifice transluminal endoscopic surgery (NOTES) with other surgical modalities. Most studies are based on systemic variables such as postoperative serum cytokines, with conflicting results. Surgical trauma induces an early inflammatory response, release of cytokines, and local leukocyte activation and oxidative burst. Major surgical trauma is related to impairment of phagocytic function and an increase in production of active oxygen species by phagocytes. The aim of this study was to evaluate the impact of transgastric peritoneoscopy on peritoneal innate immune response compared with laparoscopy and laparotomy in swine. Thirty-four male Sus scrofa domesticus swine were assigned to four groups: transgastric peritoneoscopy (13), laparoscopy (7), laparotomy (7), and sham procedure (7). Twenty-four hours after the procedure, peritoneal fluid cells were harvested by peritoneal washing after necropsy. Flow cytometry analysis of labeled S. aureus and E. coli phagocytosis by peritoneal neutrophils and macrophages was blindly performed. Oxidative burst activity measured by H2O2 production under different challenges was also evaluated. Total operative time varied between all groups. The transgastric, laparoscopy, and laparotomy groups required 56, 17.2, and 40.3 min of mean operative time, respectively (p

Milk somatic cell counts are the basis of mastitis and milk quality control programs. Milk somatic cell counts for goats, in which neutrophils comprise the main cell type, are higher than those for sheep and cows. In cows, macrophages are the major cell type present in milk, as is the case for sheep. Milk secretion in small ruminants is apocrine in nature, and cytoplasmic particles, similar in size to milk somatic cells, are normal constituents of their milk, although concentrations of cytoplasmic particles are much higher in milk from goats than from ewes. This fact has led to the development of cell counting procedures that are specific for deoxyribonucleic acid to enable accurate milk somatic cell counts to be obtained. Furthermore, milk somatic cell counts in small ruminants are influenced by noninfectious factors such as lactation stage and parity. However, mastitis is the main factor that leads to an increase in somatic cell count due to the response to infection. There is some evidence that physiological variations in the viability, number and proportions of each leukocytes subpopulation may be related to mammary gland immune status, which is associated with periods of increased susceptibility to mastitis. The main etiological agent of mastitis in small ruminants is coagulase-negative staphylococci, and special attention should be paid to this group of bacteria. Unlike in cows, novobiocin-sensitive coagulase-negative staphylococci cause a significant increase in milk somatic cell counts. Finally, the somatic cell count and the California Mastitis Test represent valuable tools for mastitis screening and assessing the disease prevalence, but their predictive values are better in ewes than in goats.

O objetivo do presente trabalho foi avaliar a frequência e número absoluto de linfócitos B e das subpopulações de linfócitos T em bovinos infectados pelo vírus da leucemia bovina (BLV) com distintos perfis leucocitários conhecidos como não leucêmicos (AL) e com linfocitose persistente (LP). Deste modo, 15 animais foram selecionados e divididos uniformemente em três grupos (negativo, AL e LP). A infecção pelo vírus da BLV foi detectada por imunodifusão em ágar gel e por ensaio imunoenzimático. A quantificação das populações de linfócitos B e T foi determinada por citometria de fluxo utilizando anticorpos monoclonais. Os resultados do presente estudo apontaram para aumento da população de linfócitos B, e também das células CD5+ e CD11b+, que geralmente são alvo da infecção pelo vírus da BLV, nos animais com LP. Consequentemente pode-se observar redução da porcentagem de linfócitos T, T CD4+ no sangue periférico, e de linfócitos T, T CD4+ e T CD8+ nas células mononucleares do sangue periférico isoladas por gradiente de centrifugação. No entanto, nenhuma alteração no número absoluto de linfócitos T, T CD4+ e T CD8+ no sangue periférico foi encontrada nos animais manifestando LP. Entretanto, encontrou-se correlação alta e significativa entre o número absoluto de linfócitos T e T CD8+ e o número absoluto de linfócitos B no sangue periférico nos animais manifestando LP, não encontrando a mesma correlação com os linfócitos T CD4+. Além disso, não foram observadas correlações significativas entre o número absoluto de linfócitos T, T CD4+ e T CD8+ e o número absoluto de linfócitos B no sangue periférico nos animais AL e negativos.The aim of the present trial was to determine the frequencies and absolute number of B and T lymphocytes subpopulations in bovine leukemia virus (BLV)-infected dairy cows with distinct lymphocyte profile known as non-leukemic (AL) and persistent lymphocytosis (PL). Thus, 15 animals were selected and divided uniformly in three groups (negative, AL, PL). The BLV infection was detected by agar gel immunodiffusion and enzyme-linked immunosorbent-assay. The lymphocytes subsets were evaluated using monoclonal antibodies by flow cytometry. The results of the present study pointed out to an increase in B lymphocytes, and also an augment in CD5+ and CD11b+ cells in animals showing PL. Consequently, it can be observed a decrease in the percentage of T cells subsets in these animals. Conversely, no significant alterations in the absolute number of the T lymphocytes, T CD4+ cells and T CD8+ lymphocytes were found in BLV-infected dairy cows with PL. Therefore, the correlation between the absolute numbers of B- and T cell subsets in the peripheral blood applied to each group showed a significant and positive strong correlation between numbers of B cells and T cells or T CD8+ cells in the PL animals, although the same cannot be predicted for T CD4+ lymphocytes. No such correlation was encountered for the AL and negative-control animals.

O objetivo do presente estudo foi avaliar a capacidade de liberação de peróxido de hidrogênio (H2O2) por fagócitos oriundos de glândulas mamárias bovinas sadias e infectadas. Desse modo, 73 amostras de leite provenientes das glândulas mamárias foram classificadas em sadias e infectadas de acordo com a cultura bacteriológica e a contagem de células somáticas (CCS). Após o isolamento das células do leite, procedeu-se à contagem diferencial de leucócitos e determinação da liberação de H2O2 pela oxidação da solução de vermelho fenol. Foi observada menor liberação de H2O2 pelos fagócitos oriundos dos quartos mamários infectados, assim como houve correlação negativa entre a liberação de H2O2 por fagócitos e a CCS (r=-0,34; P=0,0025), e a porcentagem de neutrófilos (r=-0,24; P=0,04). Além disso, houve tendência de menor liberação de H2O2 pelos fagócitos estimulados por forbol 12-miristato 13-acetato nas glândulas mamárias infectadas. Entretanto, observou-se maior liberação de H2O2 pelos fagócitos em 1mL de leite nos quartos mamários infectados, ao considerar a CCS mL-1. Pode-se concluir que fagócitos de quartos mamários infectados apresentaram menor liberação de H2O2, o que indica menor capacidade microbicida. Por outro lado, observou-se maior liberação de H2O2 pelos fagócitos em 1mL de leite nos quartos infectados, fato que pode contribuir com o maior recrutamento de leucócitos para a glândula mamária e/ou a persistência do processo inflamatório.

The study was designed to assess the effects of in vitro selenium addition on intracellular hydrogen peroxide production by neutrophils from the milk and blood of dairy cows. Blood from 10 dairy cows and 20 milk samples from five dairy cows were incubated with 0 mg (control) or 10μM of sodium selenite. Then, milk and blood neutrophils were submitted for evaluation of intracellular hydrogen peroxide production by flow cytometry using 2',7'-dichlorofluorescein diacetate as a probe. The selenium status of the animals was evaluated by determination of the blood glutathione peroxidase activity. The results of the present work showed that in vitro selenium supplementation leads to an enhancement in intracellular hydrogen peroxide production, which indicates an improvement in the bactericidal effects of blood and milk neutrophils even in cows with a selenium-adequate status. Thus, the present study showed that in vitro Se supplementation leads to an enhancement in intracellular hydrogen peroxide production, indicating an improvement in the bactericidal effects of blood and milk neutrophils in cows with Se-adequate status.

Para a avaliação funcional de monócitos de bovinos infectados pelo vírus da leucose enzoótica bovina (LEB), foram coletadas amostras de sangue de 10 vacas com sorodiagnóstico negativo (SN), 10 com sorodiagnóstico positivo e que manifestavam linfocitose persistente (LP), e 10 com sorodiagnóstico positivo alinfocitóticas (AL). Os monócitos foram separados por gradiente de densidade e aderência em placa, submetidos aos testes de viabilidade por exclusão do azul de tripan, fagocitose de partículas de Zymosan, espraiamento em lamínula de vidro e quantificação da liberação de peróxido de hidrogênio (H2O2) e de óxido nítrico (ON). Monócitos de animais com LP apresentaram os menores índices de viabilidade (P

The purpose of the present trial was to compare the percentages of necrotic and apoptotic polymorphonuclear leukocytes (PMNL) in goat milk with low and high somatic cell count (SCC). Twenty eight milk samples were collected from 20 lactating goats, determined to be negative in bacteriological examination, and divided in three groups, according to their SCC: samples with SCC lower than 500 × 10 3 cells/mL; between 500 and 1500 × 10 3 cells/mL; and higher than 1500 × 10 3 cells/mL. SCC was performed in an automatic somatic cell counter. Apoptosis and necrosis were quantified using dual-color flow cytometry with fluorescein labeled annexin-V and propidium iodide (PI). Results of the present study showed a significant positive correlation between the percentage of the viable PMNL and milk SCC ( r = 0.495, P = 0.008), as well as a significant negative correlation between apoptotic PMNL and milk SCC ( r = −0.486, P = 0.009). Results also pointed out lower PMNL viability rates due to higher apoptosis rates in milk samples with SCC lower than 5 × 10 5 cells/mL.

Several factors can affect bovine mammary gland health and although bacterial mastitis is the most studied and reported cause, viral infections may also have negative effects on bovine udder health. Viral infections can indirectly damage the papillary duct of the teat, and induce or exacerbate signs of bovine mastitis due to viral-induced immunosuppressive effects that may lead to a greater susceptibility to bacterial mastitis and even intensify the severity of established bacterial infections. Some viruses (Bovine alphaherpesvirus 2, cowpox, pseudocowpox, foot-and-mouth disease, vesicular stomatitis and papillomavirus) affect the integrity of the udder skin, leading to teat lesions, favoring the entry of mastitis-causing pathogens. It is therefore possible that the association between mastitis and viruses is underestimated and may, for example, be associated with negative bacterial culture results. Few milk samples are tested for the presence of viruses, mainly because of the more laborious and expensive procedures required. Furthermore, samples for virus testing would require specific procedures in terms of collection, handling and storage. Thus, there is a knowledge gap in regard to the actual impact of viruses on bovine udder health. Despite the fact that serum anti-virus antibodies can be detected, there is not enough evidence to confirm or exclude the effect of viruses on udder health. However, any expectation of milk production from healthy animals should consider the possible impact of viral infections in mastitis development and not underestimate the importance of actions to diagnose and control the disease. Therefore, the purpose of this review is to describe the association of diagnosis and control of viral diseases and their effect on bovine udder health. Diversos fatores podem afetar a saúde da glândula mamária bovina e embora a mastite bacteriana seja a causa mais estudada e relatada, as infecções virais também podem ter efeitos negativos sobre a saúde da glândula mamária bovina. De forma indireta as infecções virais podem danificar o ducto papilar do teto ou ainda, induzir ou agravar a mastite bovina, devido aos seus efeitos imunossupressores que podem levar a uma maior susceptibilidade para casos de mastite bacteriana e até mesmo intensificar a severidade das infecções bacterianas já estabelecidas. Alguns vírus (Alphaherpesvirus bovino 2, cowpox, pseudocowpox, febre aftosa, estomatite vesicular e papilomavírus) afetam a integridade da pele do úbere, levando a lesões no teto, favorecendo a entrada de patógenos causadores de mastite. Portanto é possível que a associação entre mastites e viroses seja subestimada e podem, por exemplo, estar associada às amostras de diagnóstico de mastite bacteriana com resultados de cultura bacteriana negativa. Além disso, as amostras de leite de vacas com mastite não são coletadas, tratadas e armazenadas adequadamente para pesquisa de vírus que requer cuidados específicos, além de um diagnóstico mais trabalhoso e caro. Desse modo, há lacunas aserem preenchidas quanto ao real impacto das viroses sobre a saúde da glândula mamária bovina. Apesar da possibilidade de detecção de anticorpos séricos contra os vírus, não existem evidências suficientes para incluir ou excluir os efeitos das viroses a saúde da glândula mamária bovina. Porém, a expectativa de produção de leite por meio de animais saudáveis deve considerar os possíveis impactos das infecções virais no desenvolvimento de mastites, não se pode, portanto, subestimar a importância de ações para o diagnóstico e controle das mesmas. Sendo assim, o propósito desta revisão é descrever as relações entre o diagnóstico de controle das doenças virais e seus potenciais impactos sobre a saúde da glândula mamária bovina.

O eritrograma é uma avaliação que representa um auxílio ao clínico no diagnóstico de afecções que acometem os animais domésticos. No entanto, estudos hematológicos em bubalinos são escassos, havendo poucas informações na literatura a respeito de valores de referência dos constituintes sanguíneos desta espécie. Assim, este trabalho teve como objetivo a análise do eritrograma de búfalos de diferentes faixas etárias da raça Murrah, machos e fêmeas. Os animais em estudo foram distribuídos em quatro grupos experimentais, de acordo com as idades: Grupo 1: animais com idade entre o nascimento e 3 meses (n=15); Grupo 2: animais com idade entre 4 e 6 meses (n=50); Grupo 3: animais com idade entre 7 e 12 meses (n=50); e Grupo 4: animais com idade superior a 12 meses (n=50). Como resultados das análises foram encontrados os seguintes valores médios, no Grupo 1: 7,9x106 hemácias/mL de sangue (He); 13,0 g/dL de hemoglobina (Hb); hematócrito (Ht) de 38,9%; Volume Corpuscular Médio (VCM) de 49,0 fl; Concentração Hemoglobínica Corpuscular Média (CHCM) de 33,6 %, e Hemoglobina Corpuscular Média (HCM) de 16,4 pg; no Grupo 2: He: 7,1 x10(6)/mL; Hb: 12,5 g/dL; Ht: 36,8%; VCM: 52,4 fl; CHCM: 33,9%, e HCM: 17,8 pg; no Grupo 3: He: 7,9 x10(6)/mL; Hb: 12,0 g/dL; Ht: 33,8%; VCM: 43,1 fl; CHCM: 35,4%, e HCM: 15,34 pg; e no Grupo 4: He: 6,7 x10(6)/mL; Hb: 11,7 g/dL; Ht: 34,4%; VCM: 53,4 fl; CHCM: 34,4%. e HCM: 17,4 pg. A análise estatística dos resultados encontrados para as diferentes faixas etárias permitiu concluir que ocorreram variações determinadas pela evolução da idade, caracterizadas por diminuição do número médio de hemácias (He), da concentração de hemoglobina (Hb) e do valor do hematócrito (Ht). Os índices hematimétricos apresentaram variações significativas no G3.

International audience; AbstractThe effects of bovine leukemia virus (BLV) on the immune response have been extensively investigated; however, its effects on mammary gland immunity are only speculative. Although BLV has a tropism for B cells, it can affect both adaptive and innate immunities because these systems share many effector mechanisms. This scenario is the basis of this investigation of the effects of BLV on mammary gland immunity, which is largely dependent upon neutrophilic functions. Thus, the present study sought to examine neutrophilic functions and the lymphocyte profile in the milk of naturally BLV-infected cows. The viability of the milk neutrophils and the percentage of milk neutrophils that produced reactive oxygen species (ROS) or phagocytosed Staphylococcus aureus were similar between BLV-infected and BLV-uninfected dairy cows. Furthermore, the expression of CD62L and CD11b by the milk neutrophils and the percentage of milk neutrophils (CH138+ cells) that were obtained from the udder quarters of the BLV-infected cows were not altered. Conversely, the median fluorescence intensity (MFI) representing intracellular ROS production and the phagocytosis of S. aureus, the expression of CD44 by the milk neutrophils and the percentage of apoptotic B cells were lower in the milk cells from BLV-infected dairy cows, particularly those from animals with persistent lymphocytosis (PL). The lymphocyte subsets were not different among the groups, with the exception of the percentage of CD5−/CD11b− B cells, which was higher in the milk cells from BLV-infected cows, particularly those with PL. Thus, the present study provides novel insight into the implications of BLV infection for mammary gland immunity.

Com o objetivo de avaliar a relação entre o exame físico da glândula mamária de ovelhas com o leite, 292 ovelhas da raça Santa Inês foram examinadas. As amostras de leite foram submetidas ao exame bacteriológico, ao CMT e a CCS (automática e microscópica). Variações especificamente identificadas por inspeção da mama apresentaram alterações de celularidade observadas pelo CMT (p