BACKGROUND: Macrophages may play a prominent role in defense of the bovine mammary gland, and their functionality is necessary for successful eradication of bacterial pathogens. In contrast to necrosis, however, apoptosis has not yet been studied in macrophages from bovine mammary glands. Therefore, the aim of this study was to confirm the occurrence of apoptosis in macrophages from resting heifer mammary glands and during the inflammatory response. METHODS: Inflammatory response was induced by phosphate buffered saline (PBS) and by lipopolysaccharide (LPS). Resident macrophages (RESMAC) were obtained before and inflammatory macrophages (INFMAC) 24, 48, 72 and 168 hours after inducing inflammatory response in mammary glands of unbred heifers. Cell samples were analyzed for differential counts, apoptosis and necrosis using flow cytometry. RESULTS: Populations of RESMAC and INFMAC contained monocyte-like cells and vacuolized cells. Apoptosis was detected differentially in both morphologically different types of RESMAC and INFMAC and also during initiation and resolution of the inflammatory response. In the RESMAC population, approximately one-tenth of monocyte-like cells and one-third of vacuolized cells were apoptotic. In the INFMAC population obtained 24 h after PBS treatment, approximately one-tenth of monocyte-like cells and almost one-quarter of vacuolized cells were apoptotic. At the same time following LPS, however, we observed a significantly lower percentage of apoptotic cells in the population of monocyte-like INFMAC and vacuolized INFMAC. Moreover, a higher percentage of apoptotic cells in INFMAC was detected during all time points after PBS in contrast to LPS. Comparing RESMAC and INFMAC, we observed that vacuolized cells from populations of RESMAC and INFMAC underwent apoptosis more intensively than did monocyte-like cells. CONCLUSIONS: We conclude that apoptosis of virgin mammary gland macrophages is involved in regulating their lifespan, and it is involved in the resolution process of the inflammatory response.
- MeSH
- Apoptosis MeSH
- Cells, Cultured MeSH
- Lipopolysaccharide Receptors metabolism MeSH
- Lipopolysaccharides pharmacology MeSH
- Macrophages cytology metabolism MeSH
- Mammary Glands, Animal cytology drug effects pathology MeSH
- Gene Expression Regulation MeSH
- Cattle MeSH
- Cell Survival physiology MeSH
- Inflammation chemically induced MeSH
- Animals MeSH
- Check Tag
- Cattle MeSH
- Female MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
Enterotoxigenic Staphylococcus aureus in raw milk poses a potential health hazard to consumers, and the identification of such strains should be used as part of a risk analysis of milk and milk products. The primary purpose of this study was to investigate the occurrence of enterotoxigenic S. aureus strains in raw milk supplied for dairy processing in the Czech Republic. A further aim was to compare the production of staphylococcal enterotoxins (SEs) with the presence of the corresponding genes. This was undertaken using multiplex polymerase chain reaction (PCR) and reversed passive latex agglutination (RPLA). Out of 440 bulk tank milk samples from 298 dairy herds, 70 proved positive for S. aureus (15.9%). Staphylococcal enterotoxin genes (ses) were detected in 39 (55.7%) isolates. The genes most commonly detected were sei (38.6%), seg (31.4%) and sea (27.1%). Genes seb, seh, sed, sej and sec were observed in 10%, 4.3%, 2.9%, 2.9% and 1.4% of strains respectively. Genes see and sel did not occur. The most frequently detected genotypes were seg, sei at 11.4%; sea at 10.0%; and sea, seg, sei at 8.6%. Toxin production was observed in nine (12.9%) S. aureus isolates. Seven strains were detected as SEB- (10%) and two as SED- (2.9%) producing. A relatively high number (32%) of discrepancies between the results with multiplex PCR and RPLA assays was obtained, particularly on account of SEA. Nineteen strains were sea positive by PCR but SEA negative by RPLA, and one strain was sec positive and SEC negative. The results of both methods were identical concerning SEB and SED. It was concluded that detection of ses by PCR was a useful additional tool to support identification of enterotoxigenic strains.
- MeSH
- Gene Amplification MeSH
- Antigens, Bacterial analysis genetics MeSH
- DNA, Bacterial genetics chemistry MeSH
- Enterotoxins analysis genetics MeSH
- Genotype MeSH
- Food Contamination analysis MeSH
- Latex Fixation Tests MeSH
- Humans MeSH
- Milk MeSH
- Colony Count, Microbial MeSH
- Polymerase Chain Reaction methods MeSH
- Food Microbiology MeSH
- Prevalence MeSH
- Base Sequence MeSH
- Cattle MeSH
- Consumer Product Safety MeSH
- Staphylococcus aureus isolation & purification metabolism MeSH
- Animals MeSH
- Check Tag
- Humans MeSH
- Cattle MeSH
- Female MeSH
- Animals MeSH
- Geographicals
- Czech Republic MeSH
The aim of the study was to evaluate the effect of selected temperatures on viability (apoptosis and necrosis) of bovine blood neutrophil granulocytes (neutrophils) in vitro. The following temperatures were tested: -80, -20, 4, 23, 37 degrees C. Heparinised bovine blood was incubated for 1, 4 and 24 h under respective temperature. Apoptosis and necrosis of neutrophils were detected by light microscopy, transmission electron microscopy (TEM) and flow cytometry (FCM). From selected temperatures, 4 degrees C impaired the neutrophil viability least. The proportion of apoptotic and necrotic neutrophils amounted to (mean +/- SD) 5.25 +/- 3.53% and 0.83 +/- 0.38%; 7.09 +/- 2.07% and 1.64 +/- 0.50%; 35.39 +/- 12.53% and 5.46 +/- 1.46%; after 1, 4 and 24 h incubation, respectively. The temperature (4 degrees C) is the best alternative for short-term storage.
- MeSH
- Apoptosis MeSH
- Radiation Injuries, Experimental MeSH
- Financing, Organized MeSH
- Cryopreservation methods veterinary MeSH
- Necrosis pathology MeSH
- Neutrophils pathology MeSH
- Leukocyte Count MeSH
- Flow Cytometry MeSH
- Cattle MeSH
- Temperature MeSH
- Microscopy, Electron, Transmission MeSH
- Cell Survival MeSH
- Animals MeSH
- Check Tag
- Cattle MeSH
- Female MeSH
- Animals MeSH
This work characterizes macrophage morphological features during initiation and resolution of an inflammatory response by the bovine mammary gland. The study has been carried out in 20 mammary glands of five virgin heifers by using light microscopy of natural and stained cells and by using transmission electron microscopy (TEM). The inflammatory reaction was induced by an intramammary administration of phosphate-buffered saline (PBS). It has been found that both the initial as well as the resolution phases of the inflammatory reaction are characteristic of the presence of various morphologically different macrophage forms. During the initial phase of the inflammatory response, the major proportion of the macrophage population consisted of monocyte-like macrophages, which represented newly migrated cells. These macrophages were 12-15 mum in size, with spherical or ovoidal shapes, and contained homogenous, fine-granular cytoplasm rich in Golgi complexes, numerous mitochondria, and no lysosomes. The nuclei of the macrophages were kidney-shaped, and surrounded by dark chromatin along the peripheries. Macrophages with phagocytosed apoptotic neutrophils in the cytoplasm were detected already during the initial phase. These macrophages reached the highest proportion 48-72 h after the influx induction and participated in the resolution of the inflammatory reaction. Other cells, also detected during the resolution of the inflammatory reaction, were vacuolized macrophages that formed the largest cells in the lavages of the mammary glands and that were structurally characteristic for the presence of vacuoles in the cytoplasm. In TEM the macrophage vacuoles formed both phagolysosomes with residues of pre-digested material of phagocytosed apoptotic neutrophils and vacuoles that were less electon-dense. Morphologically different forms of macrophages reflected their real-time functions in the inflammation process.
- MeSH
- Phagocytes ultrastructure MeSH
- Financing, Organized MeSH
- Macrophages ultrastructure MeSH
- Microscopy veterinary MeSH
- Mammary Glands, Animal immunology pathology MeSH
- Cattle Diseases pathology MeSH
- Neutrophils ultrastructure MeSH
- Cattle MeSH
- Microscopy, Electron, Transmission veterinary MeSH
- Inflammation pathology veterinary MeSH
- Animals MeSH
- Check Tag
- Cattle MeSH
- Female MeSH
- Animals MeSH
The objective was to clarify the association between bulk tank milk somatic cell count (BTSCC) and total bacterial count (BTTBC) and coliform bacteria count (BTCBC) in a large set of data based on the currently accepted legal limit of BTSCC=400,000/ml. We analysed the database obtained from one of four laboratories offering routine estimation of microbiological indicators and counts of somatic cells in bulk tank milk samples in the Czech Republic during the year 2003 (74,174; 73,921 and 33,020 records of BTSCC, BTTBC and BTCBC estimations, respectively, in milk from 2,769 suppliers). Raw data of BTSCC (with arithmetic mean 220,000/ml; 95th percentile=502,000/ml; 99th percentile=784,000/ml) indicated that the BTSCC limit was exceeded in 12% of samples. BTSCC did not sufficiently reflect the hygiene status of particular producing herds because correlation coefficients between bulk tank milk somatic cell score (BTSCS) and log BTTBC or log BTCBC were low. Categorization of herds according to the percentage of records exceeding the BTSCC limit gave significantly higher correlation coefficients for the association between this characteristic and log BTTBC or log BTCBC (r=0.84 and r=0.68, respectively). The percentage of records exceeding the BTSCC limit was a useful tool to highlight problem herds kept in inadequate hygienic conditions in primary milk production. Likewise, the value of BTSCS>5 seemed to be a useful tool for the discrimination of problem herds.
- MeSH
- Enterobacteriaceae isolation & purification MeSH
- Financing, Organized MeSH
- Hygiene MeSH
- Laboratories standards MeSH
- Mastitis, Bovine microbiology MeSH
- Dairying standards MeSH
- Milk cytology microbiology standards MeSH
- Cell Count veterinary MeSH
- Colony Count, Microbial veterinary MeSH
- Cattle MeSH
- Animals MeSH
- Check Tag
- Cattle MeSH
- Female MeSH
- Animals MeSH
- Publication type
- Comparative Study MeSH
- Geographicals
- Czech Republic MeSH
