1,721,100 research outputs found
Helicobacter pylori: ancient human host but always new pathogen. Towards new therapeuticals approaches
No full textHelicobacter pylori (H. pylori) is a microaerophilic spiral bacterium that has been associated with the pathogenesis of active and chronic gastritis, peptic ulcer, and gastric carcinoma. In 1994, H. pylori was classified as a type one carcinogen, and it appears to be responsible for 5.5% of all cancers worldwide. Antibiotic therapy has been successfully used against H. pylori but with increasing drug resistance, failure to eradicate the bacterium is more common. Although H. pylori is considered a non-invasive pathogen, an intracellular location of the bacterium both in vitro and in vivo has been widely reported. Therefore, new drugs should present activity against both the extracellular and intracellular microorganisms. Development of drugs from natural sources is receiving attention because of their potential to delay the onset of resistance and keep the pathogenic strains drug sensitive for longer time. Artemisinin extracted from Artemisia annua, has been used in traditional Chinese medicine for over two millennia. Different artemisinin derivatives have been developed. Artemisinin and its derivatives possess antimicrobial, antileishmanial, and antitumor activities. Our study showed that artemisinin derivatives were active against extracellular and the intracellular H. pylori suggesting consideration for treatment of H. pylori infection. Finally, several clinical extra-gastric diseases, have been reported. For some of these, the pathogenic mechanism is clear. Guidelines suggest that, after H. pylori infection determination, the eradication antibiotic therapy is recommended. Of 51 antibiotics in clinical development, less than 10% have chance to be used. So, new therapeutical approaches for the management of H. pylori infection are required
Susceptibility to vancomycin of enterococci isolated from dairy products
No full textThe antibiotic vancomycin is often used in clinical therapy against multiple antibiotic-resistant bacteria. Twenty strains of enterococci, either Enterococcus faecium or Ent. faecalis, isolated from different cheeses were tested for resistance to vancomycin in liquid medium. Minimum inhibitory concentration (MIC) values ranged from less than 1 to 4 μg ml-1. A 399 bp intragenic fragment of the vanA gene was not observed after amplification of total DNA of the strains. It seems, therefore, that resistance to vancomycin, which is a common trait for enterococci of nosocomial origin, is not widespread among dairy isolates
Mutation in Serratia marcescens AmpC beta-lactamase producing high-level resistance to ceftazidime and cefpirome
No full textStarting from a clinical isolate of Serratia marcescens that produced a chromosomally encoded AmpC beta -lactamase inducibly, we isolated by stepwise selection two laboratory mutants that showed high levels of resistance to some cephalosporins. The 98R mutant apparently overproduced the unaltered beta -lactamase constitutively, but the 520R mutant produced an altered enzyme, also constitutively. Ceftazidime and cefpirome MICs for the 520R mutant were much higher (512 and 64 μg/ml, respectively) than those for the 98R mutant (16 and 16 μg/ml, respectively). Yet the MICs of cephaloridine and piperacillin for the 520R mutant were four- to eightfold lower than those for the 98R mutant. Cloning and sequencing of the ampC alleles showed that in the 520R mutant enzyme, the Thr64 residue, about two turns away from the active-site serine, was mutated to isoleucine. This resulted in a >1,000-fold increase in the catalytic efficiency (kcat/Km) of the mutated AmpC enzyme toward ceftazidime, whereas there was a >10-fold decrease in the efficiency of the mutant enzyme toward cefazolin and cephaloridine. The outer membrane permeability of the 520R strain to cephalosporins was also less than in the 98R strain, and the alteration of the kinetic properties of the AmpC enzyme together with this difference in permeability explained quantitatively the resistance levels of both mutant strains to most agents studie
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Epidemiologia molecolare di ceppi di Clostridium difficile isolati in reparti geriatrici
No full textClostridium difficile (Cd) è un microrganismo anaerobio, sporigeno, gram positivo. Si può trovare come commensale nella flora intestinale dell’uomo. C.d e’ il principale agente eziologico di diarrea associata a terapia antibiotica acquisita in ambiente ospedaliero. I ceppi patogeni producono tossine (A e B) con effetti citotossici a carico dell’epitelio intestinale. Alcuni ceppi produttori di tossina binaria, sono causa di infezioni più severe. A tutt’oggi non è noto il ruolo specifico di questa tossina.
Nei pazienti istituzionalizzati con età superiore ai 60 anni aumenta il rischio di acquisizione dell’infezione in relazione alle patologie concomitanti, alle modifiche della flora intestinale, alla pressione selettiva degli antibiotici, all’esposizione all’ambiente ospedaliero. Scopo del nostro lavoro è stato quello di effettuare una tipizzazione molecolare dei ceppi di Cd isolati da pazienti ospedalizzati presso ASP Pio Albergo Trivulzio. Nell’analisi di campioni sequenziali si è voluto verificare se le infezioni ricorrenti fossero dovute a recidive o reinfezioni
Helicobacter pylori : ureA, cagA and vacA expression during conversion to the coccoid form
No full textAs viability of coccoid forms of Helicobacter pylori can only be verified by demonstrating the integrity of the DNA and active protein synthesis, we analysed the expression of ureA, cagA, vacA genes after prolonged incubation in a liquid medium. Exponentially growing and ageing phase cultures were used. Our results showed that, although the coccoid forms had decreased DNA and RNA levels after 31 days, they were not degraded and still expressed the urease, cytotoxic island and vacuolating toxin genes. Coccoid forms are therefore viable and may act as a transmissible agent that plays a crucial role in disease relapses after antibiotic therap
Molecular identification of Candida lusitaniae using a PCR-REA method
No full textA pair of primers selected from the costant region of the ERG11 gene of C. albicans, was used to amplified a 350 bp segment from the genomic DNA of C. lusitaniae strain. The PCR product was cloned in plasmid pCR-TOPO (Invitrogen) and sequenced (GenBank submitted). The sequence of the fragment was compared with the published gene sequences of C. albicans, C. tropicalis, C. krusei, C. glabrata, C. guillermondi, C. dubliniensis, C. parapsilosis and C. kefyr. The homology range observed was 62% to 97% among all species.
The amplicon obtained from C. lusitaniae was therefore subjected to REA using Sau3A, HincII and Not I according to a precedent study in which restriction enzyme analysis (REA) of PCR product was used to identify Candida strains at the species level.
The REA pattern obtained allowed to differenziate C. lusitaniae from the other species of Candida except from C. glabrata whose patterns were very similar. Thus we tested another restriction enzyme, RsaI, with all species of Candida previously described. The results showed that only C. lusitaniae and C. krusei had a recognition site for this enzyme but the patterns were different from each other . The use of Rsa I for PCR-REA, in addition to the other enzyme, makes the identification of C. lusitaniae possible.
An early and rapid identification of this species in clinical specimens is of great importance because C. lusitaniae is an emerging human pathogen that can cause severe infections in immunocompromised hosts
Identificazione molecolare di Candida lusitaniae
No full textCandida lusitaniae è un patogeno emergente, causa di gravi infezioni nell’ospite immunocompromesso, e presenta una ridotta sensibilità all’amfotericina B. Una rapida e precoce identificazione di tale specie nei campioni clinici riveste, pertanto, una notevole importanza. L’utilizzazione di metodiche molecolari può costituire una valida alternativa ai convenzionali metodi colturali.
Allo scopo sono stati utilizzati primers selezionati dalla regione costante del gene ERG11 di Candida albicans per amplificare un segmento di 350 bp dal DNA genomico di C. lusitaniae. L’amplicone ottenuto è stato clonato, sequenziato e comparato con le sequenze geniche di C. albicans, C. dubliniensis, C. glabrata, C. guillermondii, C. kefyr, C. krusei, C. parapsilosis e C. tropicalis, riscontrando un’omologia del 62% - 97%. L’amplicone è stato successivamente sottoposto a digestione (Restriction Enzyme Analysis) con le seguenti endonucleasi Sau3A, HincII e NotI, che in un precedente lavoro avevano consentito di differenziare le 7 principali specie patogene di Candida.
I pattern elettroforetici ottenuti consentivano di differenziare C. lusitaniae dalle altre specie di Candida ad eccezione di C. glabrata, i cui pattern risultavano pressocchè identici. L’aggiunta di un quarto enzima di restrizione RsaI differenziava C. lusitaniae da C. glabrata.
Sono attualmente in corso saggi di PCR su campioni simulati (urine e sangue) per valutarne l’eventuale utilizzazione diretta sui campioni clinici
Tipizzazione molecolare di ceppi di Aspergillus fumigatus isolati da pazienti con fibrosi cistica
No full textI pazienti affetti da Fibrosi Cistica (CF) sono esposti al rischio di infezioni delle basse vie respiratorie, in genere gli agenti causali sono Staphylococcus aureus e Pseudomonas aeruginosa con tutte le problematiche connesse alla formazione di biofilm e, soprattutto, alla farmaco-resistenza dei ceppi isolati. La colonizzazione e/o una superinfezione con funghi possono rappresentare un ulteriore complicanza infettiva, in particolare se si tratta di funghi filamentosi. Scopo del nostro lavoro è stato quello di verificare se i pazienti con CF fossero colonizzati da un genotipo predominante di Aspergillus fumigatus, il fungo filamentoso di più frequente isolamento dalle secrezioni delle basse vie respiratorie in tale categoria di pazienti.
Ventisette ceppi di A. fumigatus sono stati sottoposti a RAPD (Random Amplified Polymorphic DNA) con due differenti primers (NS3 and R108) ed a RFLP (Restriction Fragment Lenght Polymorphism) con AfutI. La combinazione di più sistemi di tipizzazione molecolare ha consentito di poter discriminare tra i ceppi e di dimostrare che diversi genotipi erano presenti nei nostri pazienti.
Il contemporaneo riscontro di un ceppo con una ridotta sensibilità all’itraconazolo ci ha spinti ad indagare il coinvolgimento di meccanismi molecolari di resistenza valutando la presenza di eventuali mutazioni nei geni (cyp51A e cyp51B) che codificano per l’enzima bersaglio degli azoli e l’espressione delle pompe di efflusso. L’analisi di sequenza dei geni cyp51A e cyp51B non ha rilevato mutazioni puntiformi significative correlabili al fenotipo resistente così come l’espressione delle pompe di efflusso. Sono in corso esperimenti di valutazione della diversa espressione delle pompe di efflusso in questo ed in ceppi con fenotipo sensibile dopo esposizione a concentrazioni sub-mic di itraconazolo
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