Estimation of Mineral and Trace Element Profile in Bubaline Milk Affected with Subclinical Mastitis

The milk samples from buffaloes of Murrah breed at mid lactation stage, reared at an organised dairy farm, were screened for subclinical mastitis based on bacteriological examination and somatic cell count following International Dairy Federation criteria. Milk samples from subclinical mastitis infected and healthy buffaloes were analysed to evaluate physicochemical alterations in terms of protein, fat, pH, electrical conductivity, chloride, minerals (sodium, potassium and calcium) and trace elements (iron, zinc, copper and selenium). In the present study, protein, fat, zinc, iron, calcium and selenium content was significantly lower (P < 0.001), while pH and electrical conductivity were significantly higher in mastitic milk as compared to normal milk. Concentration of electrolytes mainly sodium and chloride significantly increased with higher somatic cell count in mastitic milk and to maintain osmolality; potassium levels decreased proportionately. Correlation matrix revealed significantly positive interdependences of somatic cell count with pH, electrical conductivity, sodium and chloride. However, protein, fat, calcium and potassium were correlated negatively with elevated somatic cell count in mastitic milk. It is concluded that udder infections resulting in elevated somatic cells may alter the mineral and trace element profile of milk, and magnitude of changes may have diagnostic and prognostic value.     

Effect of Disodium Cromoglycate on Exercise-induced Asthma

In eight patients with exercise-induced asthma, disodium cromoglycate was found to produce a definite inhibition of the post-exercise fall in forced expiratory volume in one second. This effect may be part of the cause for the subjective improvement experienced with this drug.     

Incidence of Subclinical Mastitis and Prevalence of Major Mastitis Pathogens in Organized Farms and Unorganized Sectors

Subclinical mastitis (SCM) represents a major proportion of the burden of mastitis. Determining somatic cell count (SCC) and electrical conductivity (EC) of milk are useful approaches to detect SCM. In order to correlate grades of SCM with the load of five major mastitis pathogens, 246 milk samples from a handful of organized and unorganized sectors were screened. SCC (>5 × 10⁵/mL) and EC (>6.5 mS/cm) identified 110 (45 %) and 153 (62 %) samples, respectively, to be from SCM cases. Randomly selected SCM-negative samples as well as 186 samples positive by either SCC or EC were then evaluated for isolation of five major mastitis-associated bacteria. Of the 323 isolates obtained, 95 each were S. aureus and coagulase-negative staphylococci (CoNS), 48 were E. coli and 85 were streptococci. There was no association between the distribution of organisms and (a) the different groups of SCC, or (b) organised farms and unorganised sectors. By contrast, there was a significant difference in the distribution of CoNS, and not other species, between organized farms and unorganized sectors. In summary, bacteria were isolated irrespective of the density of somatic cells or the type of farm setting, and the frequency of isolation of CoNS was higher with organized farms. These results suggest the requirement for fine tuning SCC and EC limits and the higher probability for CoNS to be associated with SCM in organized diary sectors, and have implications for the identification, management and control of mastitis in India.     

The Electrical Conductivity of Bovine Milk in Mastitis Diagnosis

The electrical conductivity (EC) of milk is mainly a function of the electrolyte concentration in the milk and therefore raised in mastitis. The present investigation was aimed at elaborating, if possible, a diagnostic model for screening purposes based on EC determinations and consistent with the diagnostic procedures and interpretations commonly used in laboratory milk diagnosis in the Nordic countries (Klastrup 1975). According to this diagnosis (here called reference diagnosis) cell numbers above 300,000/ml (cell count or the corresponding CMT-score) in foremilk quarter samples during the main part of the lactation period and significantly above the lowest value on within-udder comparison during late lactation are considered indicative of mastitis and bacteriological examinations are made when called for.     

Phenylbutazone and Flunixin Meglumine Fail to Show Beneficial Effects on Bovine Subclinical Mastitis

Inflammation is caused by the release of chemical mediators from tissues and migra-tory cells. These mediators include leuco-trienes and prostaglandins (PGs), which are generated from arachidonic acid (Vane & Botting 1987). Elevated concentrations of arachidonic acid metabolites have been found in experimental and spontaneous ma-stitis, which indicates that these substances may have a role in the pathophysiologic process of mastitis (Anderson et al. 1985, Atroshietal 1986).     

Mycoplasmosis: Experimental Mastitis. Demonstration of Antibody in Milk

According to several authors specific antibodies can be demonstrated in serum of cows with mycoplasma mastitis. For practical reasons, and for pathogenetic studies, it would be of interest if presence of antibody in milk could be established. To elucidate this and to examine the pathogenicity of the type strain, PG 11, of M. bovigenitalium, the experiment described below was conducted.     

Study on Correlation of Maedi-Visna Virus (MVV) with Ovine Subclinical Mastitis in Iran

Maedi-Visna is an important slow viral disease of sheep leading to progressive pneumonia, encephalitis and mastitis. Udder is one of the organs affected by MVV. Despite the fact that in Iran Maedi-Visna is known since 2000, to the authors’ knowledge correlation of subclinical mastitis and infection with MVV has not been assayed. In this study 50 milk samples from 10 flocks in East Azerbaijan Province of Iran were tested. None of the animals exhibited any clinical signs of the disease. Forty samples were collected from CMT positive ewes and ten were taken from CMT negative ewes. Milk samples were analyzed using PCR targeting gag sequence. Presence of provirus DNA was detected in one sample from CMT negative and seven samples from CMT positive ewes. These data demonstrate that 16.5 % of sheep with subclinical mastitis were infected to MVV. Thus this virus can be considered one of the main pathogenic agents of mastitis and can be dramatically transmitted to lambs by milk.     

Multimeric α-Lactalbumin from Human Milk Induces Apoptosis through a Direct Effect on Cell Nuclei

A fraction from human milk containing spf-multimer α-lactalbumin (MAL) induces apoptosis in tumor cells and immature cells but spares mature cells. The mechanism of apoptosis induction and the molecular basis for the difference in susceptibility between tumor cells and healthy cells have not been defined. In this study we examined the interaction of MAL with different cellular compartments, using confocal microscopy and subcellular fractionation. MAL was shown to accumulate in the nuclei of sensitive cells rather than in the cytosol, the vesicular fraction, or the ER–Golgi complex. Nuclear uptake occurred rapidly in cells that were susceptible to the apoptosis-inducing effect, but not in nuclei of resistant cells. Nuclear uptake was through the nuclear pore complex and was critical for the induction of DNA fragmentation, since inhibition of nuclear uptake with WGA rescued digitonin-permeabilized cells from induction of DNA fragmentation. Ca²⁺was required for MAL-induced DNA fragmentation but nuclear uptake of MAL was independent of Ca²⁺. This way MAL differs from most previously described agents in that it crosses the plasma membrane and cytosol, and enters cell nuclei where it induces DNA fragmentation through a direct effect at the nuclear level.     

Effect of Disodium Cromoglycate on Exacerbations of Asthma Produced by Hyperventilation

The prophylactic inhalation of disodium cromoglycate lessens the exacerbations which occur in some asthmatic patients after voluntary hyperventilation. Reasons are given for considering this to be a possible mechanism for certain types of emotional exacerbation of the disease and for disodium cromoglycate giving sypmtomatic relief of asthma disproportionate to the improvement in routine measurements of ventilatory capacity. It is suggested that one clinical indication for giving disodium cromoglycate is to observe whether a fall in FEV₁ occurs after two minutes hyperventilation.     

Recurrent Exposure to Subclinical Lipopolysaccharide Increases Mortality and Induces Cardiac Fibrosis in Mice

Background

Circulating subclinical lipopolysaccharide (LPS) occurs in health and disease. Ingesting high fatty meals increases LPS that cause metabolic endotoxemia. Subclinical LPS in periodontal disease may impair endothelial function. The heart may be targeted as cardiac cells express TLR4, the LPS receptor. It was hypothesized that recurrent exposure to subclinical LPS increases mortality and causes cardiac fibrosis.

Methods

C57Bl/6 mice were injected with intraperitoneal saline (control), low dose LPS (0.1 or 1 mg/kg), or moderate dose LPS (10 or 20 mg/kg), once a week for 3 months. Left ventricular (LV) function (echocardiography), hemodynamics (tail cuff pressure) and electrocardiograms (telemetry) were measured. Cardiac fibrosis was assessed by picrosirius red staining and LV expression of fibrosis related genes (QRT-PCR). Adult cardiac fibroblasts were isolated and exposed to LPS.

Results

LPS injections transiently increased heart rate and blood pressure (<6 hours) and mildly decreased LV function with full recovery by 24 hours. Mice tolerated weekly LPS for 2–3 months with no change in activity, appearance, appetite, weight, blood pressure, LV function, oximetry, or blood chemistries. Mortality increased after 60–90 days with moderate, but not low dose LPS. Arrhythmias occurred a few hours before death. LV collagen fraction area increased dose-dependently from 3.0±0.5% (SEM) in the saline control group, to 5.6±0.5% with low dose LPS and 9.7±0.9% with moderate dose LPS (P<0.05 moderate vs low dose LPS, and each LPS dose vs control). LPS increased LV expression of collagen Iα1, collagen IIIα1, MMP2, MMP9, TIMP1, periostin and IL-6 (P<0.05 moderate vs low dose LPS and vs control). LPS increased α-SMA immunostaining of myofibroblasts. LPS dose-dependently increased IL-6 in isolated adult cardiac fibroblasts.

Conclusions

Recurrent exposure to subclinical LPS increases mortality and induces cardiac fibrosis.     

Evaluation of a Cow-Side Test for Detection of Gram-Negative Bacteria in Milk from Cows with Mastitis

A modified Limulus amebocyte lysate (LAL) cow-side test was evaluated under field conditions. The principle of the test is to visualize reactions between test components and endotoxin from the cell wall of Gram-negative bacteria. The practical purpose is to detect such bacteria in mastitic milk. Secretions from 789 udder quarters with clinical mastitis were examined by the LAL-test. Parallel quarter milk samples were sent to a mastitis laboratory for microbiological examination. Eleven veterinary surgeons in three veterinary districts in Norway performed the field investigations. Results of the LAL-test and culture agreed in 93% of all milk samples tested, agreement measured by kappa being 0.63. The sensitivity of the test in detecting Gram-negative bacteria was 63%, while the specificity was 97%. The predictive value of a positive test result was 70%, the figure being somewhat higher (75%) when the material was limited to milk samples without antibiotic residues. The predictive value of a negative test result was 95%. The LAL-test is considered to constitute a valuable cow-side test for the veterinary practitioner, aiding the selection of antibacterial drug of choice for the initial treatment of clinical mastitis.     

Effect of Alpha-receptor Blocking Drugs and Disodium Cromoglycate on Histamine Hypersensitivity in Bronchial Asthma

Twenty patients with extrinsic type bronchial asthma are shown to have a significant fall in vital capacity (V.C.) and forced expiratory volume in 1 second (F.E.V.₁) after an intravenous infusion of 50μg. of histamine dihydrochloride. In 10 of these subjects the fall in V.C. and F.E.V.₁ produced by intravenous histamine is inhibited by the alpha-receptor blocking drugs phentolamine and phenoxybenzamine injected before the histamine test. The inhalation of disodium cromoglycate in 10 subjects is also shown to inhibit the fall in V.C. and F.E.V.₁ produced by the intravenous infusion of histamine. It is suggested that bronchial smooth muscle in asthmatic subjects has alpha-adrenergic receptor sites, and that a possible mechanism for the action of disodium cromoglycate is that it stabilizes the cell membrane, thereby altering calcium ion transport.     

Bacterial Community Profiling of Milk Samples as a Means to Understand Culture-Negative Bovine Clinical Mastitis

Inflammation and infection of bovine mammary glands, commonly known as mastitis, imposes significant losses each year in the dairy industry worldwide. While several different bacterial species have been identified as causative agents of mastitis, many clinical mastitis cases remain culture negative, even after enrichment for bacterial growth. To understand the basis for this increasingly common phenomenon, the composition of bacterial communities from milk samples was analyzed using culture independent pyrosequencing of amplicons of 16S ribosomal RNA genes (16S rDNA). Comparisons were made of the microbial community composition of culture negative milk samples from mastitic quarters with that of non-mastitic quarters from the same animals. Genomic DNA from culture-negative clinical and healthy quarter sample pairs was isolated, and amplicon libraries were prepared using indexed primers specific to the V1–V2 region of bacterial 16S rRNA genes and sequenced using the Roche 454 GS FLX with titanium chemistry. Evaluation of the taxonomic composition of these samples revealed significant differences in the microbiota in milk from mastitic and healthy quarters. Statistical analysis identified seven bacterial genera that may be mainly responsible for the observed microbial community differences between mastitic and healthy quarters. Collectively, these results provide evidence that cases of culture negative mastitis can be associated with bacterial species that may be present below culture detection thresholds used here. The application of culture-independent bacterial community profiling represents a powerful approach to understand long-standing questions in animal health and disease.     

Dopamine Oxidation Alters Mitochondrial Respiration and Induces Permeability Transition in Brain Mitochondria

Both reactive dopamine metabolites and mitochondrial dysfunction have been implicated in the neurodegeneration of Parkinson's disease. Dopamine metabolites, dopamine quinone and reactive oxygen species, can directly alter protein function by oxidative modifications, and several mitochondrial proteins may be targets of this oxidative damage. In this study, we examined, using isolated brain mitochondria, whether dopamine oxidation products alter mitochondrial function. We found that exposure to dopamine quinone caused a large increase in mitochondrial resting state 4 respiration. This effect was prevented by GSH but not superoxide dismutase and catalase. In contrast, exposure to dopamine and monoamine oxidase-generated hydrogen peroxide resulted in a decrease in active state 3 respiration. This inhibition was prevented by both pargyline and catalase. We also examined the effects of dopamine oxidation products on the opening of the mitochondrial permeability transition pore, which has been implicated in neuronal cell death. Dopamine oxidation to dopamine quinone caused a significant increase in swelling of brain and liver mitochondria. This was inhibited by both the pore inhibitor cyclosporin A and GSH, suggesting that swelling was due to pore opening and related to dopamine quinone formation. In contrast, dopamine and endogenous monoamine oxidase had no effect on mitochondrial swelling. These findings suggest that mitochondrial dysfunction induced by products of dopamine oxidation may be involved in neurodegenerative conditions such as Parkinson's disease and methamphetamine-induced neurotoxicity.     

Hexylresorcinol Induces Chromosome Aberrations in Mouse Peripheral Blood Cells

Hexylresorcinol has been demonstrated to induce chromosome aberrations in eukaryotic cells at doses of 0.5, 0.05, and 0.005 mg/g body weight. The metabolic transformation of hexylresorcinol in mice decreases its genotoxic effect. The mutagenic effect is retained for three days only after the administration of the highest dose of hexylresorcinol (0.5 mg/g); during the first two days, lower doses are also genotoxic. Therefore, hexylresorcinol doses lower than 0.5 mg/g body weight are metabolized within two days to the extent precluding the expression of the cytotoxic effect. After a single administration to mice, exogenous hexylresorcinol is transformed at a rate of 0.0025–0.025 mg/day.__________Translated from Genetika, Vol. 41, No. 8, 2005, pp. 1045–1048.Original Russian Text Copyright © 2005 by Margulis, Ozhiganova, Bushmanova, Kolpakov, Il’inskaya.     

Antibodies Against Staphylococcus Aureus Nuclease in Bulk Milk As An Indicator of Mastitis in Dairy Herds

Titres of staphylococcal antinucleases and cell counts in bulk milk samples were compared as indicative criteria of the mastitis situation in dairy herds. The correlation coefficients between the prevalence rate of mastitis and the antinuclease titre and cell count, respectively, were of the same order of magnitude. The incidence of clinical mastitis showed a better correlation to the antinuclease titre than to the cell count. When cell counts and antinuclease titres were combined a more reliable selection of herds with a possible mastitis problem was achieved. The antinuclease test is consequently recommended as a supplement to cell counting when screening bulk milk.     

Effect of Cromoglycate on Gas Changes, During Bronchial Challenge by UNCDW in Children with Asthma

Eighteen asthmatic children were challenged with ultrasonically nebulized cold distilled water (UNCDW). Blood gas composition was monitored transcutaneously (tcpO(2) and tcpCO(2)) during and after the challenge. Assuming as basal the response to this UNCDW test, nine children (Group A) were then chosen at random to inhale cromoglycate by aerosol delivery for 8 days. Nine children (Group B), acting as a control, inhaled saline for 8 days. At the end of this therapy, each child repeated the UNCDW test. Statistical analysis with t-test for paired data was used to compare the results of each child to both tests. Mean basal tcpO(2) and tcpCO(2) were all within the expected normal range. In all children, both mean tcpO(2) and tcpCO(2) were reduced during and after UNCDW inhalation. Mean tcpCO(2) values during the challenge were significantly (p < 0.001) lower than the corresponding steady state 2 rain after the UNCDW challenge, with a mean drop of -7% (2.1 S.D.). Mean tcpO(2) values remained significantly decreased (p < 0.001) from the fifth mitt of the UNCDW challenge to the end of the observation period, with a mean drop of -20% (15.5 S.D.). After treatment with cromoglycate (Group A), the mean tcpCO(2) values during UNCDW did not change significantly from those ofsteady state conditions: -0.8% (0.5 S.D.); whereas mean tcpO(2) values decreased by -4% (4.9 S.D.). The control children treated with saline (Group B) showed mean tcpCO(2) and tcpO(2) values which were significantly different (p < 0.001) from those of the steady state conditions: mean drop of tcpCO(2), -6% (4.2 S.D.); mean drop of tcpO(2), -20% (4.7 S.D.). In conclusion, it emerges that: UNCDW induces nonspecific broncho-constriction in asthmatic children with a typical drop of tcpCO(2) and tcpO(2); the treatment with cromoglycate normalizes the time course of tcpCO(2) (hyper-reactivity) and reduces dramatically the drop of tcpO(2) time course (hyper-responsivity) during and after the UNCDW test.     

Effect of a Hot Milk Drink on Movements During Sleep

The effects on sleep of a hot drink of milk and Horlicks were compared with those of hot water taken before retiring by medical student volunteers. Horlicks reduced the number of small movements made during sleep.     

l-Tyrosine Induces DNA Damage in Brain and Blood of Rats

Mutations in the tyrosine aminotransferase gene have been identified to cause tyrosinemia type II which is inherited in an autosomal recessive manner. Studies have demonstrated that an excessive production of ROS can lead to reactions with macromolecules, such as DNA, lipids, and proteins. Considering that the l-tyrosine may promote oxidative stress, the main objective of this study was to investigate the in vivo effects of l-tyrosine on DNA damage determined by the alkaline comet assay, in brain and blood of rats. In our acute protocol, Wistar rats (30 days old) were killed 1 h after a single intraperitoneal l-tyrosine injection (500 mg/kg) or saline. For chronic administration, the animals received two subcutaneous injections of l-tyrosine (500 mg/kg, 12-h intervals) or saline administered for 24 days starting at postnatal day (PD) 7 (last injection at PD 31), 12 h after the last injection, the animals were killed by decapitation. We observed that acute administration of l-tyrosine increased DNA damage frequency and damage index in cerebral cortex and blood when compared to control group. Moreover, we observed that chronic administration of l-tyrosine increased DNA damage frequency and damage index in hippocampus, striatum, cerebral cortex and blood when compared to control group. In conclusion, the present work demonstrated that DNA damage can be encountered in brain from animal models of hypertyrosinemia, DNA alterations may represent a further means to explain neurological dysfunction in this inherited metabolic disorder and to reinforce the role of oxidative stress in the pathophysiology of tyrosinemia type II.     

Mucosal Immunization of Lactating Female Rhesus Monkeys with a Transmitted/Founder HIV-1 Envelope Induces Strong Env-Specific IgA Antibody Responses in Breast Milk

We previously demonstrated that vaccination of lactating rhesus monkeys with a DNA prime/vector boost strategy induces strong T-cell responses but limited envelope (Env)-specific humoral responses in breast milk. To improve vaccine-elicited antibody responses in milk, hormone-induced lactating rhesus monkeys were vaccinated with a transmitted/founder (T/F) HIV Env immunogen in a prime-boost strategy modeled after the moderately protective RV144 HIV vaccine. Lactating rhesus monkeys were intramuscularly primed with either recombinant DNA (n = 4) or modified vaccinia virus Ankara (MVA) poxvirus vector (n = 4) expressing the T/F HIV Env C.1086 and then boosted twice intramuscularly with C.1086 gp120 and the adjuvant MF59. The vaccines induced Env-binding IgG and IgA as well as neutralizing and antibody-dependent cellular cytotoxicity (ADCC) responses in plasma and milk of most vaccinated animals. Importantly, plasma neutralization titers against clade C HIV variants MW965 (P = 0.03) and CAP45 (P = 0.04) were significantly higher in MVA-primed than in DNA-primed animals. The superior systemic prime-boost regimen was then compared to a mucosal-boost regimen, in which animals were boosted twice intranasally with C.1086 gp120 and the TLR 7/8 agonist R848 following the same systemic prime. While the systemic and mucosal vaccine regimens elicited comparable levels of Env-binding IgG antibodies, mucosal immunization induced significantly stronger Env-binding IgA responses in milk (P = 0.03). However, the mucosal regimen was not as potent at inducing functional IgG responses. This study shows that systemic MVA prime followed by either intranasal or systemic protein boosts can elicit strong humoral responses in breast milk and may be a useful strategy to interrupt postnatal HIV-1 transmission.