Invasive Candidiasis in Brescia, Italy: Analysis of Species Distribution and Antifungal Susceptibilities During Seven Years.

The aims of this study were to evaluate the epidemiology of nosocomial candidemia in a large teaching hospital in Brescia, Italy, and the in vitro antifungal susceptibility of isolates. We analyzed 196 isolates causing fungemia in patients admitted in our hospital, between January 2009 and December 2015. Strains were identified by VITEK 2 and MALDI-TOF MS. MICs were determined by Sensititre Yeast OneTM. The resistance was defined by using the revised CLSI breakpoints/epidemiological cutoff values to assign susceptibility or wild type to systemic antifungal agents. Most infections were caused by Candida albicans (60%), Candida parapsilosis (15%), Candida glabrata (12%) and Candida tropicalis (6%). The susceptibility rate for fluconazole was 96.5%. Non-Candida species isolates exhibited full susceptibilities to echinocandins according to CLSI breakpoints. Amphotericin B demonstrated excellent activity against all Candida species. Local epidemiological and antifungal susceptibility studies are necessary in order to improve empirical treatment guidelines.

Genotyping analyses of tuberculosis transmission among immigrant residents in Italy.

We used DNA fingerprinting to analyse tuberculosis (TB) epidemiology in immigrant patients living in two major northern Italian urban areas. The study population included 1999 TB patients (1500 Italian-born and 499 immigrants). Univariate and multivariate logistic regression models were used to identify risk factors related to clustering similar proportions of immigrant and Italian-born patients (46%) had infection with TB strains that belonged to genetic clusters. This supports the hypothesis that the disease in foreign patients is more likely to have arisen from reactivation of latent infection acquired in the country of origin than from recent transmission. Gender, age, human immunodeficiency virus infection and drug resistance were not significantly linked to TB clustering. Risk factors associated with strain clustering were country of origin (Somalia, adjusted OR (AOR) 3.19, p 0.017; Peru, AOR 2.86, p 0.014; and Senegal, AOR 2.60, p 0.045) and city of residence. Immigrant status in the larger urban area was an independent risk factor for infection with clustered TB, as reinforced by a subanalysis of the Senegalese group. In conclusion, variations in TB transmission were observed among immigrants from different countries and even within national groups, where living conditions have been found to exert a profound impact. These results emphasize the importance of improving social integration of immigrant subjects in order to limit risks of TB transmission in developed countries.

Zygomycosis in Italy: a survey of FIMUA-ECMM (Federazione Italiana di Micopatologia Umana ed Animale and European Confederation of Medical Mycology).

The aims of the study were to analyze the clinical and epidemiological characteristics and treatments for patients who developed zygomycosis enrolled in Italy during the European Confederation of Medical Mycology of medical mycology survey. This prospective multicenter study was performed between 2004 and 2007 at 49 italian Departments. 60 cases of zygomycosis were enrolled: the median age was 59.5 years (range 1-87), with a prevalence of males (70%). The majority of cases were immunocompromised patients (42 cases, 70%), mainly hematological malignancies (37). Among non-immunocompromised (18 cases, 30%), the main category was represented by patients with penetrating trauma (7/18, 39%). The most common sites of infection were sinus (35%) with/without CNS involvement, lung alone (25%), skin (20%), but in 11 cases (18%) dissemination was observed. According to EORTC criteria, the diagnosis of zygomycosis was proven in 46 patients (77%) and in most of them it was made in vivo (40/46 patients, 87%); in the remaining 14 cases (23%) the diagnosis was probable. 51 patients received antifungal therapy and in 30 of them surgical debridement was also performed. The most commonly used antifungal drug was liposomal amphotericin B (L-AmB), administered in 44 patients: 36 of these patients (82%) responded to therapy. Altogether an attributable mortality rate of 32% (19/60) was registered, which was reduced to 18% in patients treated with L-AmB (8/44). Zygomycosis is a rare and aggressive filamentous fungal infection, still associated with a high mortality rate. This study indicates an inversion of this trend, with a better prognosis and significantly lower mortality than that reported in the literature. It is possible that new extensive, aggressive diagnostic and therapeutic procedures, such as the use of L-AmB and surgery, have improved the prognosis of these patients.

Detection of Ureaplasma biovars and polymerase chain reaction-based subtyping of Ureaplasma parvum in women with or without symptoms of genital infections.

Ureaplasma parvum colonises human mucosal surfaces, primarily in the urogenital and respiratory tracts, causing a wide spectrum of diseases, from non-gonococcal urethritis to pneumonitis in immunocompromised hosts. Although the basis for these diverse clinical outcomes is not yet understood, it has been suggested that only certain strains of these micro-organisms are disease-associated. The aim of this study was to determine the distribution of Ureaplasma biovars and U. parvum serovars and to estimate their possible association with age, absence of lactobacilli, clinical symptoms and antibiotic resistance. DNA was extracted by endocervical, vaginal and urethral samples obtained from 158 women positive for U. urealyticum by culture and were biotyped by polymerase chain reaction (PCR) targeting the multiple-banded gene. Parvo biovar (biovar 1) was found in 136 (86%) and T960 biovar (biovar 2) in 22 (14%) patients. Among the different serovars of U. parvum, we found that serovar 3/14 was present maximally in the 21-25-year-old age group, while T960 biovar was distributed with quite similar frequency in women of 26-30 and >40 years of age. In this study, U. parvum serovar 3/14 and T960 biovar were found to be significantly associated with symptomatic patients and a loss of lactobacilli, while, on the contrary, U. parvum serovar 6 was significantly correlated with asymptomatic women and normal vaginal flora. The most active antibiotic for the majority of Ureaplasma isolates was tetracycline. These preliminary data show the possibility of distinguishing between the more or less virulent strains of Ureaplasma, with important consequences for therapeutic treatment.

Clustering of tuberculosis among senegalese immigrants in Italy.

OBJECTIVE: To study clustered Mycobacterium tuberculosis isolates as an indicator of recent TB transmission in a small urban setting in Italy, and to determine associated risk factors. METHODS: M. tuberculosis strains isolated between 1991 and 1997 were characterised by IS6110 restriction fragment length polymorphism (RFLP) analysis. RESULTS: One hundred and ninety-five isolates were available for RFLP analysis, which revealed 163 different patterns. Available cases were represented by 137 Italians (70%), 32 Senegalese (17%), and 26 other foreign-born cases (13%). A unique fingerprint pattern was found in 143 cases (73.3%), while 52 strains (26.7%) were grouped into 20 clusters. Nineteen cases (10%) were resident in the same quarter of Brescia with a high density of Senegalese immigrants (Area A). An increased probability of yielding clustered M. tuberculosis strains was associated with residence in Area A (OR 3.87, 95%CI 1.42-10.56; P = 0.02) and being Senegalese (OR = 5.96, 95%CI 1.48-23.97; P = 0.005). In the logistic regression analysis, being Senegalese was independently associated with yielding a clustered M. tuberculosis strain. CONCLUSIONS: Our results demonstrate a clustering of TB cases among Senegalese immigrants and suggest that RFLP analysis may be used to identify geographical areas where efforts can be targeted to interrupt TB transmission.

Identification of Mycobacterium xenopi by gas chromatography.

For the purposes of the following study we cultured 32 strains of Mycobacterium xenopi isolated from clinical specimens and several strains of other slowly growing mycobacteria. The cultures were grown in liquid medium and then analysed--after saponification, methylation, extraction with organic solvent and washing of the organic phase--using a highly sensitive manual gas-liquid chromatographic assay for the determination of secondary alcohol 2-OH-docosanol. The percentage of this compound was compared with that previously measured in strains of Mycobacterium xenopi grown on solid medium. The presence of this specific alcohol was always apparent, even though its quantity was lower than that obtained by growing mycobacteria on solid medium. The absence of interference peaks around the compound was checked by analyzing strains of other slowly growing mycobacteria in the same conditions.

Gas chromatographic assay of cellular fatty acids and alcohols for the identification of Mycobacterium species.

Ten mycobacterial species obtained from 141 cultures isolated from clinical specimens were studied. The cultures were grown on solid medium and then analysed-after saponification, methylation, extraction with organic solvent and washing of the organic phase--by capillary gas-liquid chromatography for fatty acid and secondary alcohol composition. The absence of secondary alcohols was characteristic of M. genavense, M. tuberculosis and the following Mycobacterium species with specific branched-chain fatty acids allowing their direct identification: M. gordonae, M. kansasii and M. marinum. The presence of secondary alcohols was characteristic of M. avium, M. phlei, M. scrofulaceum, M. terrae and M. xenopi. In the case of M. xenopi direct identification was made possible by the presence of a specific alcohol.

Mycobacterium genavense in AIDS patients, report of 24 cases in Italy and review of the literature.

Mycobacterium genavense is a frequently missed agent of disseminated disease in AIDS patients. The increasing frequency with which such organism is being isolated in Italy suggested a comparison of local survey with data reported in literature. Isolates presumed to belong to the species M. genavense were centralized and identified by means of genomic sequencing and/or HPLC analysis of cell wall mycolic acids; clinical data were obtained from relevant patients' record and collected using a proper questionnaire. In 24 cases in which this organism has been isolated in Italy M. genavense was grown, prevalently from blood, in liquid medium after an average of six weeks of incubation. In overwhelming majority, patients were males, presented other opportunistic diseases and were characterized by very low CD4+ counts (average 23/microl); most frequent symptoms were fever, anemia and weight loss. All but two patients, who died before the mycobacterial infection was diagnosed, were treated with at least three drugs; the mean survival was close to one year. A review of literature reports revealed a wide overlapping of clinical and microbiological features.

Tuberculosis due to drug-resistant Mycobacterium bovis in pregnancy.

We describe the management practices adopted in a case of pulmonary and extra-pulmonary tuberculosis caused by an isoniazid/pyrazinamide resistant strain of Mycobacterium bovis in a 26-week pregnant woman. She was initially treated with rifampin, isoniazid and ethambutol, pre-term delivery was induced and streptomycin was then added to the regimen. Screening of the new-born revealed no signs of either disease or infection. Isoniazid prophylaxis was not administered and the new-born was vaccinated and isolated from the mother for two months; however she continued to be fed with her mother's milk for the whole period.

Long term treatment with clarithromycin for cryptosporidiosis and emergence of drug resistant disseminated infection due to Mycobacterium avium: case report.

Macrolide resistance in disseminated Mycobacterium avium infection is of major concern in AIDS patients as these drugs represent the main component of combination therapy. Clarithromycin and azithromycin should not be used alone for the treatment and prophylaxis of the disease because of the risk of selecting resistant strains. We report the case of a clarithromycin resistant disseminated M. avium infection in an AIDS patient, acquired after long term monotherapy with clarithromycin for the treatment of cryptosporidiosis.

Detection and identification of Mycobacterium avium in the blood of AIDS patients by the polymerase chain reaction.

One hundred fifty-three blood samples from patients positive for the human immunodeficiency virus (HIV) were analyzed by polymerase chain reaction (PCR) to detect the presence of Mycobacterium avium. Samples were collected from patients who also had blood cultures performed by a radiometric method. Blood samples were centrifuged on a Ficoll-Hypaque gradient to purify peripheral blood mononuclear cells. The purified cells were washed and incubated with a resin, boiled to release mycobacterial DNA, and then amplified. Polymerase chain reaction products were detected by a nonisotopic method. A 123 base-pair (bp) insertion sequence, namely IS6110, from Mycobacterium tuberculosis complex was also included in the reaction as an internal control of Taq polymerase activity to exclude the presence of enzyme inhibitors. This IS6110 fragment can be distinguished from the 383 bp target product on ethidium bromide-stained agarose gel and may also be used in a colorimetric assay. Such results were compared with the results of culture and indicated that the assay is as sensitive as bacteriological methods, though faster.

Characterization of an isolate of the newly described species Mycobacterium interjectum.

The phenotypic features of a clinical isolate of the new species Mycobacterium interjectum, identified on the basis of its 16S rRNA gene sequence, are compared with those of the type strain. The differentiation of M. interjectum from Mycobacterium gordonae or Mycobacterium scrofulaceum is not achievable on the basis of phenotypic traits usually tested for mycobacterial speciation, but it can be reached by 16S rRNA gene sequencing or by high performance liquid chromatography (HPLC) of cell wall mycolic acids. The former reveals sequence identity with the signature region of the type species, and the latter yields a profile which is easily differentiated from those of the other two species. The unique HPLC profile of M. interjectum is reported here for the first time and so are the MICs of a wide spectrum of drugs.

Diagnosis of bronchopulmonary infections by quantification of microflora.

The quantification of bacteria and fungi in sputum or bronchoaspirate is of clinical value for the diagnosis of respiratory tract infections. We have developed an easy method to count the micro-organisms in patients with respiratory tract infections. This consists of the quantification of micro-organisms by subsequent streakings of a calibrated loop on agar. The correlation between microbiological quantitative data and the clinical status of patients with lower respiratory tract infections is discussed. The data seem to indicate that certain bacteria present in sputum or bronchoaspirate above a certain concentration may be responsible for lower respiratory tract infections. In patients with immunological disorders or chronic pathologies even lower concentrations of micro-organisms in bronchial secretions probably are enough to cause infections. The advantage of this counting method of the microbic species from the respiratory tract consists of their quantification: thus we can attribute an etiological role to a high concentration of the germs, while micro-organisms at low concentrations are probably contaminants. By this method isolated colonies are obtained after 12-18 hours. The bacterial quantification, by respiratory samples examination of the same patient in the following days, allows us to evaluate the efficacy of antibacterial therapy, producing a reduction of bacterial concentration.

Rapid methods for microbial diagnosis of mycobacterial infections.

The Authors describe their experience in rapid diagnosis of mycobacterial infections using a combination of a radiometric blood culture (Bactec 13 A) and a nucleic acid hybridization system (Gen probe, Accuprobe) to detect and identify Mycobacteria. They found out that a high number of septicaemias in HIV positive patients are due to Mycobacterium avium, while in HIV negative subjects Mycobacterium tuberculosis is the most frequent mycobacterium.

Antibacterial activity of the new carbapenem meropenem (SM-7338) against clinical isolates.

The in vitro antibacterial activity of the new carbapenem antibiotic meropenem (SM-7338) against 567 clinical isolates was evaluated. SM-7338 exhibited activity against a broad spectrum of organisms, including aerobes and anaerobes, and was superior to the other beta-lactam drugs tested (piperacillin, cefotaxime, ceftazidime, ceftriaxone, cefoxitin). SM-7338 was more active than imipenem, gentamicin and amikacin against Enterobacter cloacae and Pseudomonas aeruginosa. SM-7338 was less potent than imipenem against staphylococci and enterococci, but the activity of the two antibiotics against anaerobes was similar. SM-7338 and imipenem showed a high bactericidal activity at a concentration of 2-4 x MIC.

Rufloxacin (MF-934): in vitro and in vivo antibacterial activity.

Rufloxacin (MF-934) is a new quinolone which shows in vitro antibacterial activity against E. coli, Salmonella, Klebsiella, Proteus and Staphylococcus spp. Lower in vitro activity was observed with Pseudomonas, Serratia, Enterobacter and the streptococci group D. The antimicrobial activity of MF-934 in vitro is higher than that of nalidixic acid but lower than that of ciprofloxacin, ofloxacin, pefloxacin or norfloxacin. The protective effects of MF-934 in systemic infections in mice are lower than those of ciprofloxacin and ofloxacin. In respiratory infections in rats and in subcutaneous infections in guinea-pigs, the protective effects of MF-934 are of the same order as ciprofloxacin and ofloxacin. This is probably due to the pharmacokinetic properties of MF-934, i.e. the long half-life and tissue concentrations higher than plasma levels.