Roberto L. Caccuri

Capsule-negative Staphylococcus aureus induces chronic experimental mastitis in mice.

ABSTRACT Staphylococcus aureus capsular polysaccharides (CP) have been shown to enhance staphylococcal virulence in numerous animal models of infection. Although serotype 5 CP (CP5) and CP8 predominate among S. aureus isolates from humans, most staphylococcal isolates from bovines with mastitis in Argentina are capsule negative. This study was designed to evaluate the effects of CP5 and CP8 expression on the pathogenesis of experimental murine mastitis. Lactating mice were challenged by the intramammary route with one of three isogenic S. aureus strains producing CP5, CP8, or no capsule. Significantly greater numbers of acapsular mutant cells were recovered from the infected glands 12 days after bacterial challenge compared with the encapsulated strains. Histopathological analyses revealed greater polymorphonuclear and mononuclear leukocyte infiltration and congestion in the mammary glands of mice infected with the encapsulated strains compared with the acapsular mutant, and the serotype 5 strain elicited more inflammation than the serotype 8 strain. In vitro experiments revealed that the acapsular S. aureus strain was internalized by MAC-T bovine epithelial cells in significantly greater numbers than the CP5- or CP8-producing strain. Taken together, the results suggest that S. aureus lacking a capsule was able to persist in the murine mammary gland, whereas encapsulated strains elicited more inflammation and were eliminated faster. Loss of CP5 or CP8 expression may enhance the persistence of staphylococci in the mammary glands of chronically infected hosts.

Attenuation and Persistence of and Ability To Induce Protective Immunity to a Staphylococcus aureus aroA Mutant in Mice

ABSTRACT Staphylococcus aureus is the most important etiological agent of bovine mastitis, a disease that causes significant economic losses to the dairy industry. Several vaccines to prevent the disease have been tested, with limited success. The aim of this study was to obtain a suitable attenuated aro mutant of S. aureus by transposon mutagenesis and to demonstrate its efficacy as a live vaccine to induce protective immunity in a murine model of intramammary infection. To do this, we transformed S. aureus RN6390 with plasmid pTV1ts carrying Tn917. After screening of 3,493 erythromycin-resistant colonies, one mutant incapable of growing on plates lacking phenylalanine, tryptophan, and tyrosine was isolated and characterized. Molecular characterization of the mutant showed that the affected gene was aroA and that the insertion occurred 756 bp downstream of the aroA start codon. Complementation of the aroA mutant with a plasmid carrying aroA recovered the wild-type phenotype. The mutant exhibited a 50% lethal dose (1 × 106 CFU/mouse) higher than that of the parental strain (4.3 × 104 CFU/mouse). The aroA mutant showed decreased ability to persist in the lungs, spleens, and mammary glands of mice. Intramammary immunization with the aroA mutant stimulated both Th1 and Th2 responses in the mammary gland, as ascertained by reverse transcription-PCR, and induced significant protection from challenge with either the parental wild-type or a heterologous strain isolated from a cow with mastitis.

Effect of Melatonin on Changes in Locomotor Activity Rhythm of Syrian Hamsters Injected with Beta Amyloid Peptide 25–35 in the Suprachiasmatic Nuclei

Abstract1. Alzheimers disease is associated with circadian rhythm disturbances, probably because of beta amyloid-induced neuronal damage of hypothalamic suprachiasmatic nuclei (SCN).2. Since there is no published study on the circadian consequences of injecting beta amyloid peptide in experimental animals, one objective of the present study was to examine circadian locomotor activity in Syrian hamsters injected with beta amyloid peptide 25–35 into both SCN.3. Because one of the proposed therapies for circadian alterations in dementia is the administration of melatonin, a chronobiotic agent with antioxidant properties, the preventive effect of melatonin on the circadian changes produced by beta amyloid microinjection into SCN was also assessed.4. Wheel running activity was recorded by using the Dataquest III system in male golden hamsters kept under 14:10 light–dark photoperiods. Animals received microinjections of beta amyloid peptide 25–35 (100 μM solution, 1 μL) or saline in each SCN. Only those animals with neuronal lesions larger than 10% of SCN after beta amyloid injection were considered for further analysis.5. To assess the effect of melatonin on beta-amyloid peptide activity, melatonin was given in the drinking water (25 μg/mL) starting 15 days in advance to the microinjection of beta amyloid peptide into SCN.6. Beta amyloid-treated hamsters exhibited a significant phase advance of onset of running activity of about 22 min as compared to saline-injected animals. They also showed a significantly greater variability in onset time of wheel running activity, mainly evident from 6 to 15 days of treatment.7. Melatonin administration in the drinking water prevented the phase advance of onset time and the increased variability of onset time brought about by beta amyloid peptide.8. The results support the existence of a neuroprotective effect of melatonin on beta amyloid-induced circadian changes in hamsters.

Capsule Expression and Genotypic Differences among Staphylococcus aureus Isolates from Patients with Chronic or Acute Osteomyelitis

ABSTRACT There is ample evidence that Staphylococcus aureus capsular polysaccharide (CP) promotes virulence. Loss of capsule expression, however, may lead to S. aureus persistence in a chronically infected host. This study was conducted to determine the relative prevalence of nonencapsulated S. aureus in patients with chronic and acute osteomyelitis. Only 76/118 (64%) S. aureus isolates from patients with osteomyelitis expressed CP, whereas all 50 isolates from blood cultures of patients with infections other than osteoarticular infections expressed CP (P = 0.0001). A significantly higher prevalence of nonencapsulated S. aureus was found in patients with chronic osteomyelitis (53%) than in those with acute osteomyelitis (21%) (P = 0.0046). S. aureus isolates obtained from multiple specimens from five of six patients with chronic osteomyelitis exhibited phenotypic (expression of CP, α-hemolysin, β-hemolysin, slime, and the small-colony variant phenotype) and/or genotypic (pulsed-field gel electrophoresis and spa typing) differences. Nonencapsulated S. aureus was recovered from at least one specimen from each chronic osteomyelitis patient. Fourteen isolates obtained from two patients with acute osteomyelitis were indistinguishable from each other within each group, and all produced CP5. In conclusion, we demonstrated that nonencapsulated S. aureus is more frequently isolated from patients with chronic osteomyelitis than from those with acute osteomyelitis, suggesting that loss of CP expression may be advantageous to S. aureus during chronic infection. Our findings on multiple S. aureus isolates from individual patients allow us to suggest that selection of nonencapsulated S. aureus is likely to have occurred in the patient during long-term bone infection.

Host Response to a dam Mutant of Salmonella enterica Serovar Enteritidis with a Temperature-Sensitive Phenotype

ABSTRACT The temperature-sensitive dam mutant strain of Salmonella enterica serovar Enteritidis SD1 is highly attenuated and induces innate and protective immunity in mice. SD1 activates NF-κB and induces gamma interferon secretion. Early interaction of the SD1 mutant with intestinal epithelial cells was associated with ruffling of enterocytes. Invading bacteria were found inside Peyers patches after inoculation.

Neuroprotective Effect of Melatonin on Glucocorticoid Toxicity in the Rat Hippocampus

Dexamethasone has a neurotoxic action on rodent hippocampus. The objective of this study was to examine the extent of neuroprotection exerted by melatonin on that neurotoxic effect. A group of 24 rats received 9 daily subcutaneous injections of 0.5 mg/kg of dexamethasone. Half of them received 25 μg/ml of melatonin in the drinking water for 10 days. Controls included rats injected with vehicle or rats injected with vehicle plus melatonin in the drinking water. At the end of treatment, the brains were processed for a morphometric analysis, the results being expressed as percent number of ab- normal hipoccampal neurons (defined as necrotic cells) per field. Melatonin decreased by 77 % the effect of dexametha- sone (p< 0.001). A laterality of neurotoxic effect of dexamethasone was apparent in rats that did not receive melatonin (percent of necrotic cells in left and right hippocampus: 32.0 ± 4.4 and 19.6 ± 1.9 %, respectively, p< 0.01). The results indicate a protective effect of melatonin on glucocorticoid neurotoxicity in the rat hippocampus.

Long-lasting astrocyte reaction to persistent Junin virus infection of rat cortical neurons.

Summary. Immunoperoxidase labeling was performed in histological sections from rat brain harvested during acute (10–30 days), clinically inapparent (90–270 days) and late (450–540 days) stages of Junin virus-induced neurological disease. In frontoparietal cortex, count of viral antigen (+) neurons peaked during the acute period (27.7±6.8), dropped within the intermediate (4.8±4.0 to 1.4±1.1) and increased (7.6±4.3) at the onset of the late neurological syndrome. In infected vs. control rats, the number of GFAP (+) astrocytes maximized during the acute stage (19±4 vs. 11±5), and from the end of the intermediate (27±5 vs. 21±5) up to the late (37±7 vs. 26±6) periods. In turn, surface density of GFAP (+) material in infected samples peaked at 0.196±0.066, while it failed to exceed 0.090±0.043 in controls. Both astrocyte hypertrophy relapsing into chronicity, as depicted by surface density, and astrocyte hyperplasia preceding the onset of the late neurological syndrome, support their pathogenic contribution to disease expression.

Are astrocytes executive cells within the central nervous system

Experimental evidence suggests that astrocytes play a crucial role in the physiology of the central nervous system (CNS) by modulating synaptic activity and plasticity. Based on what is currently known we postulate that astrocytes are fundamental, along with neurons, for the information processing that takes place within the CNS. On the other hand, experimental findings and human observations signal that some of the primary degenerative diseases of the CNS, like frontotemporal dementia, Parkinsons disease, Alzheimers dementia, Huntingtons dementia, primary cerebellar ataxias and amyotrophic lateral sclerosis, all of which affect the human species exclusively, may be due to astroglial dysfunction. This hypothesis is supported by observations that demonstrated that the killing of neurons by non-neural cells plays a major role in the pathogenesis of those diseases, at both their onset and their progression. Furthermore, recent findings suggest that astrocytes might be involved in the pathogenesis of some psychiatric disorders as well.