Expression profile of genes associated with mastitis in dairy cattle

In order to characterize the expression of genes associated with immune response mechanisms to mastitis, we quantified the relative expression of the IL-2, IL-4, IL-6, IL-8, IL-10, IFN-γ and TNF- α genes in milk cells of healthy cows and cows with clinical mastitis. Total RNA was extracted from milk cells of six Black and White Holstein (BW) cows and six Gyr cows, including three animals with and three without mastitis per breed. Gene expression was analyzed by real-time PCR. IL-10 gene expression was higher in the group of BW and Gyr cows with mastitis compared to animals free of infection from both breeds (p < 0.05). It was also higher in BW Holstein animals with clinical mastitis (p < 0.001), but it was not significant when Gyr cows with and without mastitis were compared (0.05 < p < 0.10). Among healthy cows, BW Holstein animals tended to present a higher expression of all genes studied, with a significant difference for the IL-2 and IFN- γ genes (p < 0.001). For animals with mastitis no significant difference in gene expression was observed between the two breeds. These findings suggest that animals with mastitis develop a preferentially cell-mediated immune response. Further studies including larger samples are necessary to better characterize the gene expression profile in cows with mastitis.     

pH Modulation of Efflux Pump Activity of Multi-Drug Resistant Escherichia coli: Protection During Its Passage and Eventual Colonization of the Colon

Background

Resistance Nodulation Division (RND) efflux pumps of Escherichia coli extrude antibiotics and toxic substances before they reach their intended targets. Whereas these pumps obtain their energy directly from the proton motive force (PMF), ATP-Binding Cassette (ABC) transporters, which can also extrude antibiotics, obtain energy from the hydrolysis of ATP. Because E. coli must pass through two pH distinct environments of the gastrointestinal system of the host, it must be able to extrude toxic agents at very acidic and at near neutral pH (bile salts in duodenum and colon for example). The herein described study examines the effect of pH on the extrusion of ethidium bromide (EB).

Methodology/Principal Findings

E. coli AG100 and its tetracycline induced progeny AG100_TET that over-expresses the acrAB efflux pump were evaluated for their ability to extrude EB at pH 5 and 8, by our recently developed semi-automated fluorometric method. At pH 5 the organism extrudes EB without the need for metabolic energy (glucose), whereas at pH 8 extrusion of EB is dependent upon metabolic energy. Phe-Arg β-naphtylamide (PAβN), a commonly assumed inhibitor of RND efflux pumps has no effect on the extrusion of EB as others claim. However, it does cause accumulation of EB. Competition between EB and PAβN was demonstrated and suggested that PAβN was preferentially extruded. A K_m representing competition between PAβN and EB has been calculated.

Conclusions/Significance

The results suggest that E. coli has two general efflux systems (not to be confused with a distinct efflux pump) that are activated at low and high pH, respectively, and that the one at high pH is probably a putative ABC transporter coded by msbA, which has significant homology to the ABC transporter coded by efrAB of Enterococcus faecalis, an organism that faces similar challenges as it makes its way through the toxic intestinal system of the host.     

Exposure of persons accompanying patients in nuclear medicine departments.

Patient who receives a radiopharmaceutical remains radioactive for hours or even days and so represents a potential hazard to other individuals. Thus, in nuclear medicine departments, apart from the personnel who are occupationally exposed to ionizing radiation, protection rules must be observed for members of the public, in particular persons accompanying patients. The purpose of this work was to evaluate the absorbed dose levels, which persons accompanying patients are exposed to. Integral dose studies were carried out in three nuclear medicine departments, in Brazil. Dose levels are presented, and issues are discussed with regard to the presence of members of the public in waiting areas of such departments.     

Histological study and immunohistochemical location of cytoskeletal proteins in the testis and epididymis of the three species of lizards of the family Leiosauridae (Reptilia: Squamata)

The objective of the study was to histologically describe the structure of the testis and epididymis, identifying the GAGs in them. Furthermore, the distribution of smooth muscle alpha-actin, desmin, and vimentin is described of E. bilineatus, E. perditus, and U. vautieri, through immunohistochemical method. Two adult males of each species of the lizards were collected in southeastern Brazil. In the testicular albuginea, the presence of neutral glycoproteins was detected. In E. bilineatus, the epididymal duct epithelium is simple, composed of secretory cells, cylindrical in shape and acidophilic cytoplasm. In the testis of the three lizards there was a positive immunolocation accentuated for α-SMA in the dense connective tissue of the tunica albuginea. A moderate reaction was observed in the connective tissue albuginous and in the layer of smooth muscle fibers in a circumferential arrangement of the epididymal duct. The immunolocation for desmin was accentuated in the fibers of the testicular albuginea of E. bilineatus, while in U. vautier and E. perditus was moderate. In the epididymis, only in E. perditus and E. bilineatus positive immunoreaction was verified. In the three lizards there were no observations of immunostaining for vimentin with the antibody utilized.     

Lyophilized Streptomyces aureofaciens spores

The viability of mutant strains of Streptomyces aureofaciens (106 spores/0.2ml) and tetracycline production remained stable for stored lyophilized spores. The same mutant produced (9375±1584) and (12336±1023) g/g of tetracycline (p<0.01), respectively for spores lyophilized in the media: (i) control skim milk with added sucrose and sodium glutamate, and (ii) synthetic polyvinylpyrrolidone plus sucrose, peptone and sodium glutamate.     

Development of NADPH-Diaphorase in the Avian Retina: Regulation by Calcium Ions and Relation to Nitric Oxide Synthase

Abstract: Nitric oxide plays an important role as an intercellular messenger in the CNS. In the present work we measured NADPH-diaphorase activity, which is considered to be a marker of cells producing nitric oxide, in homogenates of the developing chick retina. The enzyme activity can be detected beginning in 8-day-old embryonic retinas with no further quantitative variations throughout development. Arginine analogues inhibit ∼65% of the activity in embryonic retinas and 50% in posthatched retinas. The enzyme is stimulated 50% by 2 m M calcium chloride in retinas from 8 to 14 embryonic days, but this effect decreases to 20% in 17-day embryonic retinas and practically disappears in posthatched animals. The stimulation by calcium is completely blocked by arginine analogues. The decrease in enzyme activity at posthatched retinas is not due to stimulation by endogenous calcium or the presence of insufficient amounts of calmodulin, because addition of EGTA or calmodulin, respectively, did not restore the stimulation to levels observed at embryonic stages. Inhibition of NADPH-diaphorase activity by N ^G-nitro- l -arginine or l - N ^G-(iminoethyl)ornithine is concentration dependent with IC₅₀ values of ∼1 m M at all stages studied. However, in the presence of calcium, the inhibition by both analogues is shifted to the left and is apparently biphasic at all developmental stages, including in posthatched animals, with IC₅₀ values in the low micromolar range. NADPH-diaphorase was also detected by histochemistry in specific groups of cells in the early embryonic retina and in subsets of amacrine and ganglion cells, as well as in photoreceptors, in more developed retinas. The results indicate that different isoforms of nitric oxide synthase are present in the chick retina and that a calcium-dependent isoform is predominant in early periods of development.     

The specificity of viral and bacterial sialidases for alpha(2-3)- and alpha(2-6)-linked sialic acids in glycoproteins

The anomeric specificity of six sialidases (Vibrio cholerae, Arthrobacter ureafaciens, Clostridium perfringens, Newcastle disease virus, fowl plague virus and influenza A2 virus sialidases) was assessed with sialylated antifreeze glycoprotein, ovine submandibular gland glycoprotein and alpha 1-acid glycoprotein, resialylated specifically in alpha(2-3) or alpha(2-6) linkage with N-acetylneuraminic acid or N-glycolylneuraminic acid using highly purified sialyltransferases. The rate of release of sialic acid from these substrates was found to correlate well with the specificity observed earlier with the same sialidases using small oligosaccharide substrates, i.e., alpha(2-3) glycosidic linkages are hydrolyzed faster than alpha(2-6) linkages, with the exception of the enzyme from A. ureafaciens. Sialidase activity was higher with N-acetylneuraminic acid when compared with N-glycolylneuraminic acid. The studies also showed that the core oligosaccharide and protein structure in glycoproteins may influence the rate of release for different glycosidic linkages.     

Null allele in a human polymorphism restricted to the placenta: call for a search

The identification of a null allele in a human genetic system restricted to the placenta is a great challenge for two reasons: the impossibility of carrying out family studies and the unviability of sample recollections because the placenta itself is a disposable universe. Thus, in addition to reporting the finding of a null phenotype of placental alkaline phosphatase in a dark mulatto newborn from a black mixed population of Bahia, Brazil, here we present other evidence for the presence of the ALPP*Q0 allele with considerably high frequency in this population.