Presence of sub-epithelial lymphoid tissues in the teat of ewe-lambs and adult ewes

In the first part of the study, 24 clinically healthy teats from non-lactating ewe-lambs were examined bacteriologically and histologically. No bacteria were isolated from any of these teats; lymphocytes were observed in teat cisterns of six teats (25%) from three ewes. In the second part, 87 teats from adult ewes were examined; their origin was from lactating mammary glands with no bacteria isolated (n = 23), from glands after lactation with no bacteria isolated (n = 25), from lactating glands with bacteria isolated (n = 22) or from glands after lactation with bacteria isolated (n = 17). The salient histological feature was sub-epithelial leucocytic infiltration. In teat cisterns, lymphocytes were the predominant cell type and in teat ducts, lymphocytes and neutrophils were seen in equal proportions. Sub-epithelial lymphoid nodules, some with germinal centers, were detected in 43 (49%) teats; their majority was observed at the border between teat duct and teat cistern. Presence of bacteria was significantly associated with presence of leucocytic activity (P < 0.001) and with presence of lymphoid nodules (P = 0.032). We conclude that the presence of induced sub-epithelial lymphoid tissue at the border between teat duct and teat cistern appears to be important in protecting the mammary gland during the early stages of bacterial invasion.     

A Study of the Development of Endotoxin-induced Inflammation in the Bovine Teat

Endotoxin-induced local inflammation was studied by frequent samplings in a bovine teat cistern model, which provides a unique possibility for in vivo studies of reactions in the teat without interference from the mammary gland. A rapid inflammatory response of rather short duration was elicited after endotoxin administration. An initial increase in the concentrations of bovine serum albumin and N-acetyl-β-D-glucosaminidase, indicating a disturbance in the epithelial integrity, was observed between 1 and 1.5 h post infusion (p.i.). Approximately 0.5 h later, the first influx of leukocytes, mainly neutrophils, appeared. The neutrophils tended to enter the teat cistern in several peaks occurring between 2.5 and 5 h p.i.. The sampling procedure decreased the accumulation of cells by approximately 40%, which was probably due to the removal of inflammatory mediators at an early stage. The parallel use of 2 teats instead of 1 had no major influence on the inflammatory process. This teat cistern model and the experimental procedure used should be suitable for further studies of the development of local inflammation.     

An observational study investigating the association of ultrasonographically assessed machine milking-induced changes in teat condition and teat-end shape in dairy cows

Mechanical forces during machine milking induce changes in teat condition which can be differentiated into short-term and long-term changes. Machine milking-induced short-term changes in teat condition (STC) are defined as tissue responses to a single milking and have been associated with the risk of new intramammary infection. Albeit, their association with teat characteristics, such as teat-end shape, has not been investigated by rigorous methods. The primary objective was to determine the association of STC, as measured by ultrasonography, with teat-end shape. The second objective was to describe possible differences in the recovery time of teat tissue after machine milking among teats with different teat-end shapes. Holstein cows (n=128) were enrolled in an observational study, housed in free-stall pens with sand bedding and milked three times a day. Ultrasonography of the left front and right hind teat was performed after teat preparation before milking (t−1), immediately after milking (t₀) and 1, 3, 5 and 7 h after milking (t₁, t₃, t₅, t₇). The teat tissue parameters measured from ultrasound scans were teat canal length, teat-end diameter, teat-end diameter at the midpoint between the distal and proximal end of the teat canal, teat wall thickness, and teat cistern width. Teat-end shape was assessed visually and classified into three categories: pointed, flat and round. Multivariable linear regression analyses showed differences in the relative change of teat tissue parameters (compared with t−1) at t₀ among teats with different teat-end shapes, with most parameters showing the largest change for round teats. The premilking values were reached (recovery time) after 7 h in teats with a pointed teat-end shape, whereas recovery time was greater than 7 h in teats with flat and round teat-end shapes. Under the same liner and milking machine conditions, teats with a round teat-end shape had the most severe short-term changes. The results of this observational study indicated that teat-end shape may be one of the factors that contribute to the severity of STC.     

Cow teat skin, a potential source of diverse microbial populations for cheese production

The diversity of the microbial community on cow teat skin was evaluated using a culture-dependent method based on the use of different dairy-specific media, followed by the identification of isolates by 16S rRNA gene sequencing. This was combined with a direct molecular approach by cloning and 16S rRNA gene sequencing. This study highlighted the large diversity of the bacterial community that may be found on teat skin, where 79.8% of clones corresponded to various unidentified species as well as 66 identified species, mainly belonging to those commonly found in raw milk (Enterococcus, Pediococcus, Enterobacter, Pantoea, Aerococcus, and Staphylococcus). Several of them, such as nonstarter lactic acid bacteria (NSLAB), Staphylococcus, and Actinobacteria, may contribute to the development of the sensory characteristics of cheese during ripening. Therefore, teat skin could be an interesting source or vector of biodiversity for milk. Variations of microbial counts and diversity between the farms studied have been observed. Moreover, Staphylococcus auricularis, Staphylococcus devriesei, Staphylococcus arlettae, Streptococcus bovis, Streptococcus equinus, Clavibacter michiganensis, Coprococcus catus, or Arthrobacter gandavensis commensal bacteria of teat skin and teat canal, as well as human skin, are not common in milk, suggesting that there is a breakdown of microbial flow from animal to milk. It would then be interesting to thoroughly study this microbial flow from teat to milk.     

Infrared Thermography and Ultrasonography to Indirectly Monitor the Influence of Liner Type and Overmilking on Teat Tissue Recovery

Eight Danish Holstein cows were milked with a 1-mm thick specially designed soft liner on their right rear teat and a standard liner mounted under extra high tension on their left rear teat. Four of the animals were overmilked for 5 min. Rear teats were subjected to ultrasound examination on the first day and to infrared thermography on the second day. Teats were submersed in ethanol 20 min post-milking on the second day. Ultrasonography measurements showed that teat canal length increased by 30–41% during milking. Twenty minutes after milking, teats milked with modified standard liners still had elongated teat canals while teats milked with the soft liner were normalized. Overmilking tended to increase teat wall thickness. Approximately 80% of variability in teat canal length, from before teat preparation to after milking, could be explained by changes during teat preparation. Thermography indicated a general drop in teat temperature during teat preparation. Teat temperature increased during milking and continued to increase until the ethanol challenge induced a significant drop. Temperatures approached pre-challenge rather than pre-milking temperatures within 10 minutes after challenge. Teat temperatures were dependent on type of liner. Mid-teat temperatures post-challenge relative to pre-teat preparation were dependent on overmilking. Thermography and ultrasound were considered useful methods to indirectly and non invasively evaluate teat tissue integrity.     

A sentinel function for teat tissues in dairy cows: dominant innate immune response elements define early response to E. coli mastitis

Escherichia coli intramammary infection elicits localized and systemic responses, some of which have been characterized in mammary secretory tissue. Our objective was to characterize gene expression patterns that become activated in different regions of the mammary gland during the acute phase of experimentally induced E. coli mastitis. Tissues evaluated were from Fürstenburg’s rosette, teat cistern (TC), gland cistern (GC), and lobulo-alveolar (LA) regions of control and infected mammary glands, 12 and 24 h after bacterial (or control) infusions. The main networks activated by E. coli infection pertained to immune and inflammatory response, with marked induction of genes encoding proteins that function in chemotaxis and leukocyte activation and signaling. Genomic response at 12 h post-infection was greatest in tissues of the TC and GC. Only at 24 h post-infection did tissue from the LA region respond, at which time the response was the greatest of all regions. Similar genetic networks were impacted in all regions during early phases of intramammary infection, although regional differences throughout the gland were noted. Data support an important sentinel function for the teat, as these tissues responded rapidly and intensely, with production of cytokines and antimicrobial peptides.     

Nuclear bodies in bovine mammary gland in different physiological states

The frequency of nuclear bodies was determined in epithelial cells of mammary ductules or alveoli, milk cistern and teat sinus of virgin heifers, non-lactating cows and cows in early lactation. The determined frequency was within the range 12–16% with the single exception of the epithelial cells of secretory alveoli of lactating cows, in which the frequency was 3 %. These data support the concept that nuclear bodies are normal nuclear organelles which can respond to altered physiological conditions.     

The Effect of Post Milking Teat Dip and Suckling on Teat Skin Condition,Bacterial Colonisation,and Udder Health

The teat skin of cows was scored (1: smooth as silk; 2: smooth; 3: slightly rough; 4: rough; 5: cracked; and 6: sores) by trained technicians who moved their fingers down the barrel of the teat with a light touch. Technicians ranked the same population of teats in the same rank order, but their mean values differed by half a score which probably could be related to the skin condition of their own fingers. Half udder experiments were carried out for 6 months at 4 farms with 35 to 52 cows each. A postmilking teat spray with 10% glycerol improved teat skin condition (p<0.10) compared with no teat spray. A postmilking teat spray with 120 ppm chlorine dioxide did not influence teat skin condition compared with no teat spray. No differences in udder health could be proven between treatments. Control studies revealed that 10% glycerol as an emollient of a postmilking teat spray improved teat skin condition within 3 weeks from being slightly rough to being smooth for lactating cows (p<0.05) but not for dry cows having smooth teat skin. Neither glycerol nor chlorine dioxide influenced absolute number of bacteria on teat skin after a challenge with Streptococcus uberis and Staphylococcus aureus but half life of S. aureus on unsprayed teats was longest (p = 0.05). Suckling made teat skin more rough than machine milking. Nevertheless, suckling lowered the number of esculin positive bacteria on the teat skin. We concluded that the condition of healthy teat skin (scores 1–4) has no influence on bacterial colonisation in the absence of cracks and sores (scores 5–6).     

Characterization of bacterial populations recovered from the teat canals of lactating dairy and beef cattle by 16S rRNA gene sequence analysis

Bovine mastitis is of major concern to the dairy industry worldwide. The bovine teat canal is the primary route through which pathogens enter the mammary gland. The microflora of the teat canals of dairy and beef cattle was investigated by analysis of 16S rRNA gene sequences. The goal was to examine the global difference between dairy cattle, which are sanitized on a regular basis, and beef cattle, which receive little sanitary management. A diverse population of 16S rRNA gene sequences was recovered from both the dairy and the beef herd samples, with diversity higher in the beef sample. Analysis revealed the presence of 90 operational taxonomic units (OTUs) among 156 sequences, with 45 OTUs in the dairy sample and 53 OTUs in the beef sample. Only eight OTUs were common to both samples. Members of the classes Clostridia and Bacilli dominated both samples, followed by Actinobacteria and Proteobacteria. The dairy sample contained a cluster (20/80 clones) of Staphylococcaceae members, seven of which were identifiable as coagulase-negative Staphylococcus species. The beef sample was dominated by members of the genus Clostridia, many of which have not been previously cultured. The results suggest that the microorganisms present in the bovine teat canal are more diverse than previously described.     

The Interaction of Glycerol and an lodophor Teat Dip

In vivo tests confirmed that significantly more Staphylococcus aureus can be recovered by swabbing from bovine teats dipped in an iodophor glycerol mixture than from teats dipped in an iodophor alone. In contrast no reduction in activity was detected when these materials were compared in a suspension test nor when they were used to treat excised pieces of teat skin. The recovery of Staph. aureus from teat skin by swabbing was shown to be greater in the presence of glycerol alone. It is concluded, therefore, that the addition of glycerol to an iodophor does not reduce bactericidal activity but its presence on teat skin increases the recovery of bacteria surviving disinfectant treatment. The significance of these results for the testing of teat disinfectants is discussed.     

Relationships among udder characteristics,milk yield and,non-yield traits in Frizarta dairy sheep

In the present study the relationships between five body structural and nine udder morphological traits with milk yield and non-yield traits, i.e. milk contents, somatic cell count and body weight in n = 171 Frizarta ewes were investigated. Positive correlations of moderate to high magnitude were observed among body measurements ranging from 0.25 (body length-chest width) to 0.61 (chest width-chest girth). High correlations were observed between udder width and udder circumference (r = 0.69) as well as between udder circumference at base and middle (r = 0.62). Udder height was found to be positively correlated with cistern depth (r = 0.32), udder width (r = 0.40) and udder circumference (r = 0.42) as well as with teat measurements (0.19 and 0.25 with teat length and teat circumference, respectively). Cistern depth was positively and highly correlated with teat angle (r = 0.64). The correlation between teat length and circumference was also high (r = 0.57). Teat angle was found to be negatively correlated with teat measurements (r = −0.39 and −0.23 for teat length and teat circumference, respectively). Principal components analysis revealed four components that jointly explained 67% of the variance of all the body and the udder morphological traits. The first principal component interpreted as the body dimensions was found to be associated with body weight of ewes. The second component interpreted as global and rear measurements of the udder was significant for milk yield traits. The third and fourth principal components were interpreted as the udder type and teat dimensions, respectively. Ewes’ body weight could be adequately predicted by chest girth, body length, chest width and withers height. Test day milk yields were found to be associated with udder circumference and udder width as well as with udder height and teat length. None of the udder measurements and/or teat characteristics was found to be correlated with milk content(s) and/or somatic cell count. New statistical techniques can be successfully implemented in investigation of relationships between various traits in dairy sheep.     

Enhanced Expression of Adenovirus Encoding rhEPO Assisted by BAPTA

Infection efficiency is the key issue for gene delivery using adenovirus vector and usually unsatisfactory. In this study, recombinant adenoviruses encoding recombinant human EPO were prepared using the Adeasy system, and injected into the mammary gland of goats via the teat canal. BAPTA was used to treat the mammary gland to facilitate adenoviruses infection compared with EGTA. Milk serum was collected from the infected mammary gland and characterized by ELISAs and Western blotting. Expression level of rhEPO from the group treated by BAPTA was higher than that treated by EGTA.     

Summer mastitis experimentally induced by Hydrotaea irritans exposed to bacteria

Abstract. Summer mastitis is an acute suppurative bacterial infection of the udder in heifers and dry cows. To ascertain the possible role of flies in the transmission of the disease, experimental exposures of recipient heifers to Hydrotaea irritans previously exposed to bacteria were carried out. Flies were allowed to feed on secretions from clinical cases of summer mastitis. The pathogens present were the bacteria Staphylococcus aureus, Streptococcus dysgalactiae, Actinomyces pyogenes , Stuart-Schwan cocci, Peptococcus indolicus, Fusobacterium necrophorum and Bacterioides species. The teats of eight heifers were exposed to flies with verified pathogen content. Two teats of each animal were deliberately damaged before fly exposure. One teat was cut, another pricked with insect needles to mimic insect bites. Two of the heifers developed summer mastitis in the quarters where teats had been cut. The bacterial species isolated from these quarters corresponded to those that had previously been fed to the flies. For the first time, it is now demonstrated that H.irritans is capable of transmitting summer mastitis pathogens and so causing summer mastitis in recipient heifers. Lesions on the teat orifice may be a predisposing factor in the development of the disease.     

Fatty acids in the milk of goats after cessation of lactation

Regular hand milking of six goats was discontinued after 32-35 weeks of lactation. A few days after milking out ceased, the concentration of triglyceride in peripheral blood plasma increased. Over a period of weeks, the concentration of triglyceride in small samples of fluid taken from the teat canal fell gradually. Lipase activity of the milk fluid was temporarily reduced shortly after milking out ended, but, despite this, its concentration of free fatty acids increased. It is suggested that free fatty acids are released during clearance of milk triglyceride from residual fluid in the mammary gland after cessation of lactation.     

Engineering Disease Resistant Cattle

Mastitis is a disease of the mammary gland caused by pathogens that find their way into the lumen of the gland through the teat canal. Mammary gland infections cost the US dairy industry approximately $2 billion dollars annually and have a similar impact in Europe. In the absence of effective treatments or breeding strategies to enhance mastitis resistance, we have created transgenic dairy cows that express lysostaphin in their mammary epithelium and secrete the antimicrobial peptide into milk. Staphylococcus aureus, a major mastitis pathogen, is exquisitely sensitive to lysostaphin. The transgenic cattle resist S. aureus mammary gland challenges, and their milk kills the bacteria, in a dose dependent manner. This first step in protecting cattle against mastitis will be followed by introduction of other genes to deal with potential resistance issues and other mastitis causing organisms. Care will be taken to avoid altering milk’s nutritional and manufacturing properties. Multi-cistronic constructs may be required to achieve our goals as will other strategies possibly involving RNAi and gene targeting technology. This work demonstrates the possibility of using transgenic technology to address disease problems in agriculturally important species. The U.S. Government's right to retain a non-exclusive, royalty-free license in and to any copyright is acknowledged.     

The effects of concanavalin A on milk secretion and mammary metabolism in the goat

The effects of concanavalin A on the rate of milk secretion and the concentration of metabolites in milk were studied following intramammary injection of the lectin via the teat canal into one mammary gland of lactating goats. Concanavalin A decreased milk secretion from the treated gland, reduced the concentrations of phosphoenolpyruvate, nucleoside diphosphate and 2-oxoglutarate in milk and increased the concentrations of glucose, galactose, glycerol, L-lactate, pyruvate, isocitrate and citrate. The changes in the concentrations of the metabolites in milk are discussed in relation to biochemical changes occurring in the mammary gland during the suppression of milk secretion. It is suggested that, when lactose synthesis and secretion is decreased, substantial metabolism of glucose via glycolysis occurs.     

Tight junctions as targets of infectious agents

The epithelial barrier is a critical border that segregates luminal material from entering tissues. Essential components of this epithelial fence are physical intercellular structures termed tight junctions. These junctions use a variety of transmembrane proteins coupled with cytoplasmic adaptors, and the actin cytoskeleton, to attach adjacent cells together thereby forming intercellular seals. Breaching of this barrier has profound effects on human health and disease, as barrier deficiencies have been linked with the onset of inflammation, diarrhea generation and pathogenic effects. Although tight junctions efficiently restrict most microbes from penetrating into deeper tissues and contain the microbiota, some pathogens have developed specific strategies to alter or disrupt these structures as part of their pathogenesis, resulting in either pathogen penetration, or other consequences such as diarrhea. Understanding the strategies that microorganisms use to commandeer the functions of tight junctions is an active area of research in microbial pathogenesis. In this review we highlight and overview the tactics bacteria and viruses use to alter tight junctions during disease. Additionally, these studies have identified novel tight junction protein functions by using pathogens and their virulence factors as tools to study the cell biology of junctional structures.     

Modified pore canals in the cuticle of Gammarus (Crustacea : Amphipoda); A study by scanning and transmission electron microscopy

A novel type of pore canal is described from the cuticle of three species of Gammarus. Each canal passes from the epidermis vertically through the endocuticle and exocuticle, and in the most distal layers of the latter is slightly expanded. Before entering the epicuticle the canal narrows, forming a neck the base of which is encircled by an electron-dense collar. Several tubular structures arise from the collar and pass distally into the reticular innermost regions of the epicuticle. Within the neck and just below its opening at the cuticle surface, a rod-like structure is inserted; this protrudes a short distance from the pore. Each pore canal is connected to many necks; the openings of the latter are aligned in rows over the surface, the openings and rows being about 0.15 and 1.0 μm apart, respectively. Changes in the pore and canal contents are visible and their significance is discussed.     

Effects of amount of milk, milk flow and access to a rubber teat on cross-sucking and non-nutritive sucking in dairy calves

The aim of this study was to test the effects of different amounts of milk, flow rate of milk, and access to a teat after milk intake on non-nutritive sucking on an empty teat and on cross-sucking on other calves in Bos taurus dairy calves. An additional aim was to test if calves prefer to perform non-nutritive sucking on a teat with the taste of milk or on a clean teat. First experiment involved 11 groups of three calves during gradual decrease from 5 to 2.5 and 1 to 0l of whole milk per meal twice daily. Milk flow was either 0.5 or 1l/min, and the teat buckets were either left or removed after milk intake. When the calves were fed with 5l of milk, non-nutritive sucking and cross-sucking decreased (P<0.001), compared to 1 and 2.5l. Removing the empty teat after milk intake increased cross-sucking (P<0.001). Slow milk flow reduced non-nutritive sucking after milk intake (P<0.05), but did not influence cross-sucking. Duration of recorded milk intake was similar with fast flow and 5l of milk, compared to slow flow and 2.5l of milk. The latter combination increased non-nutritive sucking and cross-sucking, suggesting that amount of milk is more important than duration of milk intake.Second experiment was carried out on 12 individually kept calves. The calves received 2.5l of milk either in an open bucket, in a rubber teat bucket, or half/half in both buckets. When not used for feeding, the open bucket and the teat bucket were filled and emptied again to make them 'tasty' with some drops of milk. The tasty teat bucket was either removed or present after milk intake. The calves had always access to a second clean teat bucket. Source of milk had no influence on recorded behaviour after milk intake. The tasty teat was sucked for twice as long as the clean one when both teats were present (P<0.001). When the tasty teat was removed, the calves increased sucking on the clean teat (P<0.001), but decreased total time spent sucking (P<0.001). We conclude that a high amount of milk and the presence of the teat bucket after milk intake reduces cross-sucking, and that a clean teat is less effective in encouraging calves to suck than a teat with taste of milk.     

Ultrastructure of the protonephridia of Seison annulatus (Rotifera)

Summary Each of the two protonephridial systems of Seison annulatus consists of three sections which are separated by cell borders with septate junctions: (a) a terminal syncytium with eight terminal organs and a capillary canal, (b) a canal syncytium which is divided into a multiciliary canal region and a main canal region, and (c) a nephroporus cell. The terminal syncytium is branched and linked twice to the canal syncytium. The supporting structure of each filtration barrier is a hollow cylinder which is perforated by pores and lacks microvilli (pillars). A protonephridial spine is situated in the multiciliary canal region and stabilizes the neck region. The ored, hollow cylinder and the protonephridial spine are new characteristics for the Rotifera.