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Biomedical subjects

R Murgia

Publications and source records attributed to R Murgia.

At least 19 recordsLinked to original sources
PubMed2004

Induction of cystic forms by different stress conditions in Borrelia burgdorferi.

Cystic forms of Borrelia burgdorferi might represent a low metabolic activity state or phase of B. burgdorferi cells that allows the spirochete to survive in a hostile environment until conditions are favourable to multiply again. In this study we evaluated the rate of cyst formation induced by oxidative stress, pH variations, and heating, reconversion of cysts to vegetative forms, and some aspects of their metabolic activity. We observed cyst formation in the presence of extreme pH values, and at high temperature, but the best production of cystic forms was observed in the presence of H2O2. When transferred to BSK II medium, the cystic forms reconverted to spirochetes in relation to their age and type of induction treatment. Furthermore, we demonstrated a low metabolic activity of cystic forms by measuring amino acid incorporation. Overall, these data suggest that the phenomenon of conversion to cysts by B. burgdorferi provides a limited survival potential. This short-term survival, however, gives borreliae an additional chance to overcome unfavourable environmental conditions.

Amino Acids↗
PubMed2003

Acarological risk of exposure to agents of tick-borne zoonoses in the first recognized Italian focus of Lyme borreliosis.

Acarological risk was calculated as the probability of encountering at least one host-seeking Ixodes ricinus tick infected by the pathogen Borrelia burgdorferi sensu lato, in 100 m transects in the province of Genoa, Italy. The seasonal pattern of I. ricinus was studied using generalized estimating equations (GEE) with negative binomial error, to consider overdispersion of tick counts and repeated sampling of the same dragging sites from April 1998 to March 1999. Prevalence of infection by B. burgdorferi s.l. was evaluated by PCR and hybridization with genospecies-specific probes. Acarological risk (R) peaked in April (R = 0.2, 95% CI 0.13-0.26) and November (R = 0.29, 95% CI 0.10-0.46). Borrelia garinii and B. valaisiana were the most common genospecies at our study site suggesting a major role of birds as reservoirs. DNA from Anaplasma phagocytophilum, the agent of granulocytic ehrlichiosis in humans and animals, was amplified from an adult I. ricinus.

Anaplasma↗
PubMed2001

Conversion of Borrelia garinii cystic forms to motile spirochetes in vivo.

Cystic forms (also called spheroplasts or starvation forms) and their ability to reconvert into normal motile spirochetes have already been demonstrated in the Borrelia burgdorferi sensu lato complex. The aim of this study was to determine whether motile B. garinii could develop from cystic forms, not only in vitro but also in vivo, in cyst-inoculated mice. The cysts prepared in distilled water were able to reconvert into normal motile spirochetes at any time during in vitro experiments, lasting one month, even after freeze-thawing of the cysts. Motile spirochetes were successfully isolated from 2 out of 15 mice inoculated intraperitoneally with cystic forms, showing the infectivity of the cysts. The demonstrated capacity of the cysts to reconvert into motile spirochetes in vivo and their surprising resistance to adverse environmental conditions should lead to further studies on the role and function of these forms in Lyme disease.

Animals↗
PubMed2001

Evidence of involvement of the mannose receptor in adhesion of Borrelia burgdorferi to monocyte/macrophages.

The mannose receptor (MR) plays an important role in the recognition of some pathogens in nonopsonic phagocytosis and in antigen presentation to T cells. We found that Borrelia burgdorferi, the agent of Lyme borreliosis, adheres to monocyte-derived macrophages and to rat MR-transfected cells but not to untransfected cells. Antibodies to MR and sugars such as mannose, mannan, fucose, and some lectins significantly lowered the adhesion, confirming participation of the MR in the binding.

Bacterial Adhesion↗
PubMed2001

Isolation and characterization of Borrelia burgdorferi sensu lato strains in an area of Italy where Lyme borreliosis is endemic.

Between 1993 and 1998, we isolated Borrelia burgdorferi sensu lato from 55 of the 119 patients with clinically diagnosed Lyme borreliosis who were admitted to "San Martino" Hospital in Belluno, Veneto, an Adriatic region in northeastern Italy where Lyme borreliosis is endemic. Upon hospitalization, all patients presented erythema migrans. Isolates were typed using ribosomal DNA PCR-restriction fragment length polymorphism (RFLP) analysis of the rrfA-rrlB intergenic spacer. Of the 41 isolates typed, 37 belonged to Borrelia afzelii, 2 to Borrelia garinii, and 2 to B. burgdorferi sensu stricto. Pulsed-field gel electrophoresis, performed on 21 strains (13 new isolates and 8 controls), revealed different RFLP patterns within the B. garinii and B. afzelii strains; among the five B. garinii strains and the 12 B. afzelii strains, three or two different RFLP patterns were identified, according to the restriction enzyme used. The protein patterns of the new isolates confirmed their genotypic classification and revealed the level of expression of some immunodominant proteins like OspA and other characteristic Osps. These findings constitute the first report of such a high recovery rate of B. burgdorferi from patients in a very restricted area in Italy; they also indicate the predominance of the genospecies B. afzelii in the study area and the heterogeneity of the circulating strains.

Adult↗
PubMed2001

Sensitivity of Borrelia and Leptospira to Quinupristin-dalfopristin (Synercid) in vitro.

In vitro activity of Quinupristin-dalfopristin (Synercid) against seventeen isolates of Borrelia burgdorferi and two representatives of Leptospira spp. was investigated. MICs ranged from 0.03 to 0.125 for B. burgdorferi and 0.125-0.25 microg/ml for Leptospires. Time killing studies carried out with 2 MIC demonstrated U 3 log(10)-unit killing after 72 h, showing a significant activity against spirochetes, though at a lower level than other antibiotics in use in the therapy of Lyme disease and leptospirosis.

Anti-Bacterial Agents↗
PubMed1999

Comparable effects of flickering and steady patterns of light adaptation on photomechanical responses of cones in amphibian (Xenopus laevis) retina.

The effects of two distinct patterns of light stimulus, steady and flicker, on cone photomechanical movements (PMMs) in the Xenopus laevis retina were investigated. For both patterns studied, the effects on PMMs were assessed by quantitative analysis of the cone positions in the outer retina. Steady light adaptation was found to be equally effective as flicker in causing cone contractions. This was unlike the situation previously found in the cyprinid fish retina, in which flickering light was significantly more effective than steady. This difference could be related to the light-evoked response characteristics and circuitry of dopaminergic retinal neurones in the two vertebrate classes. The role of dopamine and other possible neuromodulator(s) in light adaptive control of vertebrate retinae is discussed.

Adaptation, Ocular↗
PubMed1999

Comparative bacteriostatic and bactericidal activities of cefodizime against Borrelia burgdorferi sensu lato.

The MIC and MSC (minimum spirocheticidal concentration) and killing rate for Borrelia burgdorferi, the etiological agent of Lyme disease, were assessed for cefodizime in comparison with ceftriaxone, minocycline, azithromycin, roxithromycin, and ciprofloxacin. The range of cefodizime MICs was greater than those of azithromycin and roxithromycin but comparable to those of ceftriaxone and minocycline. The MSCs were 1 to 2 dilutions higher than the MICs of all of the tested compounds. The killing curves of cefodizime and ceftriaxone showed parallel courses. In conclusion, cefodizime exerted an activity comparable to that of ceftriaxone against B. burgdorferi.

Borrelia burgdorferi Group↗
PubMed1998

Surface receptors of neutrophils towards B. burgdorferi.

The spirochetal agent of Lyme borreliosis, Borrelia burgdorferi, is able to induce an infection which develops in three stages: an early, localized infection, disseminated infection and a third stage, chronic infection, which probably indicates that a protected niche has been established in one or more tissues, where the spirochetes persist even if a specific immune response has been initiated. During the first stage, immediately after their entry into the host tissue, B. burgdorferi meet the motile phagocytic cells, neutrophils and monocytes; this is followed by consequent phagocytosis and killing. Although the rate and mechanism of this killing is not entirely clear, there is evidence that phagocytosis by both neutrophils and monocytes proceeds even in the absence of specific antibodies. We have demonstrated in both neutrophils and CHO Mac-1 (CR3 integrin) transfected cells, that one phagocyte receptor which is involved in B. burgdorferi adhesion in non osponic phagocytosis is the CR3 complement receptor known as integrin alpha m beta 2. Both recognition domains of the integrin, the iC3b site and the COOH terminal lectin site, bind to B. burgdorferi. Data presented here show that inhibition of adhesion on CR3 Mac-1 transfected cells and neutrophils is induced by mannose as well as by N-acetyl-D-glucosamine, sugars known to be specific inhibitors of the COOH terminal lectin-site of the integrin CR3. The inhibitory effect was serum complement independent. On the contrary, monoclonal antibody VIM12 directed towards the lectin domain not only failed to inhibit but improved adhesion, suggesting that, as a consequence of the binding, the integrin becomes more receptive to B. burgdorferi attachment at the I domain. Pretreatment of the borrelias with NalO4 eliminated adhesion, suggesting that the sugar residue/s recognized by CR3 is located on the bacteria.

Animals↗
PubMed1998

Detection of HGE agent-like Ehrlichia in Ixodes ricinus ticks in northern Italy by PCR.

Little is known about the distribution of Human Granulocytic Ehrlichiosis (HGE) in Europe and even less is known in Italy, where no case of clinically documented HGE has been reported. In a previous study we reported the presence of Ehrlichia DNA in Ixodes ricinus ticks from Central Italy. By the use of an Ehrlichia-specific PCR we found that 24% of the ticks were positive. Furthermore, we demonstrated a simultaneous coinfection of the same tick by both, Borrelia burgdorferi and Ehrlichia phagocytophila. Since the Friuli-Venezia Giulia region (North-east of Italy) is endemic for Lyme borreliosis (LB) and the geographic distribution of HGE usually overlaps that of LB, we decided to carry out a survey concerning the presence of Ehrlichia spp. in a recreational area near Trieste where the presence of Lyme borreliosis and Borrelia burgdorferi sensu lato in ticks is well demonstrated. Ticks were analyzed in pools (because a low infection rate was expected): eleven samples out of 93 were found positive by Ehrlichia-PCR. Subsequent sequence analysis of some of the positive PCR products revealed a high homology with the HGE agent Ehrlichia (only one base substitution in almost 450 bp sequenced). These findings add new and interesting data on the Ehrlichia epidemiology in Italy. By now we have demonstrated the presence of two distinct granulocytic ehrlichiae in Italiyn ticks by the aid of a PCR-based analysis: Ehrlichia phagocytophila in Central Italy and an HGE-like Ehrlichia in the north-eastern Italy, in a region close to Slovenia where the first reported case of HGE in Europe occurred.

Animals↗
PubMed1998

Rate of infection of Ixodes ricinus ticks with Borrelia burgdorferi sensu stricto, Borrelia garinii, Borrelia afzelii and group VS116 in an endemic focus of Lyme disease in Italy.

A study to evaluate the natural rate of infection of Ixodes ricinus with Borrelia burgdorferi sensu lato was carried out in an endemic focus of Lyme disease in the Trieste area in northern Italy. Two-hundred and twenty-seven ticks collected in ten different stations were tested individually for the presence of the spirochetes using polymerase chain reaction techniques able to identify both Borrelia burgdorferi sensu lato and the four genospecies (Borrelia burgdorferi sensu stricto, Borrelia garinii. Borrelia afzelii and group VS116). Multiple infection of individual ticks was found. The infection rate ranged from 0-70%. Infection of Ixodes ricinus with Borrelia burgdorferi group VS116 was found for the first time in Italy in both a high and a low endemic focus of Lyme disease.

Animals↗
PubMed1998

Coiling phagocytosis discriminates between different spirochetes and is enhanced by phorbol myristate acetate and granulocyte-macrophage colony-stimulating factor.

The mechanisms involved in coiling phagocytosis are not yet known, and it is not even clear whether this phenomenon is either an incidental event or a specific response. Therefore, the phagocytic uptake of Borrelia burgdorferi and other spirochetes by human monocytes in vitro was used to investigate the involvement of both sides--microbes and phagocytes--in coiling phagocytosis. As seen with electron microscopy, morphologically similar Borrelia, Leptospira and Treponema strains induced markedly different frequencies of coiling phagocytosis. The monocytes used coiling phagocytosis for both live (motile) and killed (nonmotile) B. burgdorferi, but pseudopod coils were observed neither with fragmented B. burgdorferi nor with cell-free supernatant from B. burgdorferi cultures. Investigation of the relationship of coiling phagocytosis with other pseudopod-based cellular mechanisms revealed that the use of bioreagents that inhibit conventional phagocytosis also inhibited coiling phagocytis but did not affect membrane ruffling. Bioreagents that increase membrane ruffling did not affect phagocytosis of B. burgdorferi, except for granulocyte-macrophage colony-stimulating factor and phorbol myristate acetate, which increased coiling phagocytosis selectively. These results demonstrate that coiling phagocytosis is not induced by microbial motility, viability, or a certain morphology and that it is not a random event. Rather, it is a selective uptake mechanism actively driven by the phagocytes. However, whether coiling phagocytosis represents an independent alternative to conventional phagocytosis or, alternatively, a fault in conventional phagocytosis remains to be determined.

Borrelia burgdorferi Group↗
PubMed1998

Elastase is the only human neutrophil granule protein that alone is responsible for in vitro killing of Borrelia burgdorferi.

Phagocytosis of Borrelia burgdorferi by human polymorphonuclear leukocytes triggers oxygen-dependent and -independent mechanisms of potentially cidal outcome. Nevertheless, no factor or process has yet been singled out as being borreliacidal. We have studied the B. burgdorferi-killing ability of the myeloperoxidase-H2O2-chloride system and that of primary and secondary granule components in an in vitro assay. We found that neither secondary granule acid extracts nor the chlorinating system could kill these microorganisms, while primary granule extracts were effective. The Borrelia-killing factor was purified to homogeneity and demonstrated to be elastase. Its cidal activity was found to be independent of its proteolytic activity.

Amino Acid Sequence↗
PubMed1997

Oligonucleotides specific for pathogenic and saprophytic leptospira occurring in water.

Sets of primers specific for both pathogenic (SPL) and saprophytic (SSL) Leptospira were designed from ribosomal 16S genes (rrs) available in databases. They were used as two sets of primer pairs for the PCR amplification of known pathogenic and saprophytic strains. It was possible to identify pathogenic strains by the use of SPL primers and saprophytic ones by SSL primers. Serovars from L. meyeri, of controversial pathogenicity status, confirmed the heterogeneity of the species representatives in this respect. Serovars ranarum, sofia and perameles were amplified by SPL and not SSL. Conversely, serovar semaranga was amplified by SSL and not SPL. In order to use SPL primers for the detection of pathogenic leptospires from a natural water environment, we set up an additional semi-nested PCR by employing a second internal primer which succeeded in detecting as few as 5 pathogenic leptospires per ml of water.

DNA, Bacterial↗
PubMed1997

Integrin CR3 mediates the binding of nonspecifically opsonized Borrelia burgdorferi to human phagocytes and mammalian cells.

Like other pathogens, the spirochete Borrelia burgdorferi, the agent of Lyme disease, possesses multiple pathways for cell binding; adhesion to phagocytic cells is of particular interest since it reportedly occurs even in the absence of specific antibodies. This study sets out to investigate how B. burgdorferi binds to human polymorphonuclear leukocytes (PMNs) when an exogenous complement is added and how the CR3 complement receptor, known as Mac-1 or alpha(m)beta2 integrin, is involved in the binding process. Experiments performed on PMNs and CHO Mac-1-expressing cells demonstrate that binding is inhibited by monoclonal anti-iC3b site antibodies, fibrinogen, and N-acetyl-D-glucosamine. These findings, which are not present with non-Mac-transfected CHO cells, indicate that the integrin alpha(m)beta2 acts as a receptor for spirochetes in nonimmune phagocytosis; furthermore, binding occurs on different domains of the CD11b subunit, involving the iC3b site and the lectin domain. The interaction of B. burgdorferi with alpha(m)beta2 integrin adds a novel pathway to Borrelia-phagocyte binding; not only does this binding affect the early stages of phagocytosis, but also it can influence the effector intracellular mechanisms which are activated by the beta2 integrin, as are the cytotoxic mechanisms.

Animals↗
PubMed1996

Leptospira interrogans and Leptospira peptidoglycans induce the release of tumor necrosis factor alpha from human monocytes.

Elevated plasma concentrations of the cytokine tumor necrosis factor alpha (TNF alpha) have been observed in patients affected by leptospirosis. In this study we found that a preparation of peptidoglycan of Leptospira interrogans, serovar copenhageni, was able to induce the release of TNF alpha from peripheral blood mononuclear cells. TNF alpha induction occurred in a dose dependent manner and was not affected by the endotoxin inhibitor polymixin B. This is the first report on induction of TNF alpha release by a peptidoglycan of spirochetes. Our findings are consistent with existing clinical data and provide a potential mechanism for TNF alpha production.

Biological Assay↗