Microbiological and hydrogeological assessment of groundwater in southern Italy.

This study represents the first investigation of microbiological groundwater pollution as a function of aquifer type and season for the Apulia region of southern Italy. Two hundred and seven wells were randomly selected from those monitored by the Regional Agency for Environmental Protection for emergency use. Both compulsory (Escherichia coli, Total Coliform, and Enterococci) and optional (Pseudomonas aeruginosa, Salmonella spp., Heterotrophic Plate Count at 37 and 22 °C) microbiological parameters were assessed regularly at these wells. Groundwater from only 18 of the 207 (8.7 %) wells was potable; these all draw from karst-fissured aquifers. The remaining 189 wells draw from karst-fissured (66.1 %) or porous (33.9 %) aquifers. Of these, 82 (43.4 %) tested negative for Salmonella spp. and P. aeruginosa, while 107 (56.6 %) tested positive for P. aeruginosa (75.7 %), Salmonella spp. (10.3 %), or for both Salmonella spp. and P. aeruginosa (14 %). A logistic regression model shows that the probability of potable groundwater depends on both season and aquifer type. Typically, water samples were more likely to be potable in autumn-winter than in spring-summer periods (odds ratio, OR = 2.1; 95 % confidence interval, 95 % CI = 1.6-2.7) and from karst-fissured rather than porous aquifers (OR = 5.8; 95 % CI = 4.4-7.8). Optional parameters only showed a seasonal pattern (OR = 2.6; 95 % CI = 1.7-3.9). Clearly, further investigation of groundwater microbiological aspects should be carried out to identify the risks of fecal contamination and to establish appropriate protection methods, which take into account the hydrogeological and climatic characteristics of this region.

Susceptibility to echinocandins of Candida spp. strains isolated in Italy assessed by European Committee for Antimicrobial Susceptibility Testing and Clinical Laboratory Standards Institute broth microdilution methods.

BACKGROUND: The echinocandins are recommended as first-line therapy for Candida species infections, but drug resistance, especially among Candida glabrata, is becoming more frequent. We investigated the antifungal susceptibility of anidulafungin, caspofungin, and micafungin against 584 isolates of Candida spp. (bloodstream, other sterile sites) collected from patients admitted to an Italian university hospital between 2000 and 2013. The susceptibility was evaluated using the broth microdilution method according to both the European Committee for Antimicrobial Susceptibility Testing (EUCAST EDef 7.2) and the Clinical Laboratory Standards Institute (CLSI M27-A3). The echinocandin susceptibilities were assessed on the basis of the species-specific clinical breakpoints proposed by the EUCAST version 6.1 and CLSI M27-S4 documents. The two methods were comparable by assessing essential agreement (EA), categorical agreement (CA), and Spearman's correlation analysis (rho, r). RESULTS: The modal minimum inhibitory concentrations (MICs; µg ⋅ mL (-1)) values by both methods (EUCAST/CLSI) for anidulafungin, caspofungin, and micafungin for each species were, respectively, as follows: C. albicans, 0.03/0.12, 0.016/0.5, and 0.016/0.008; C. parapsilosis complex, 2/1, 2/2, and 2/1; C. tropicalis, 0.06/0.12, 0.06/0.12, and 0.06/0.12; C. glabrata complex, 0.03/0.25, 0.06/0.12, and 0.03/0.06; C. guilliermondii, 2/1, 2/2, and 2/2; and C. krusei, 0.06/0.12, 0.12/0.5, and 0.06/0.12. The overall resistance rates for EUCAST/CLSI were as follows: anidulafungin, 2.5/0.9%; caspofungin, breakpoint not available/3.8%; micafungin, 2.7/1.5%. Candida glabrata complex was the least susceptible to all three echinocandins, and the percentages of resistant isolates by EUCAST/CLSI were as follows: anidulafungin, 13.5/2.7%; caspofungin, breakpoint not available/16.2%; micafungin, 18.9/13.5%. The overall EA was 93 % for micafungin, 92% for anidulafungin, and 90% for caspofungin. The CA was >90% for all organism-drug combinations with the exception of C. glabrata and anidulafungin (89%). Spearman's rho for EUCAST/CLSI was 0.89 (p < 0.001) for caspofungin, 0.85 (p < 0.001) for anidulafungin, and 0.83 for micafungin (p < 0.001). CONCLUSIONS: Independent of the procedure applied, no alarming resistance to the tested agents was found, although a reduced susceptibility was detected for C. glabrata complex. The EUCAST and CLSI methods produce similar MICs, indicating that using one method or the other should not result in susceptibilities different enough to affect treatment decisions.

Antibiotic susceptibility of Legionella pneumophila strains isolated from hospital water systems in Southern Italy.

OBJECTIVES: The purpose of this study was to describe the susceptibility of environmental strains of Legionella spp. to 10 antimicrobials commonly used for legionellosis therapy. A study of environmental strains could be useful to timely predict the onset of antibiotic resistance in the environment before it is evidenced in clinical specimens. METHODS: The minimum inhibitory concentrations (MICs) of 100 environmental Legionella pneumophila (Lpn) strains belonging to serogroups (sgs) 1, 6, 8, and 10 were tested using the E-test methodology on buffered charcoal yeast extract agar supplemented with α-ketoglutarate. The most frequent sgs were selected from those obtained during microbiological surveillance conducted in 2014 in a hospital in Southern Italy. The MICs were read after 2 days of incubation at 35 °C in a humidified atmosphere without CO2. RESULTS: All isolates were inhibited by low concentrations of fluoroquinolones and macrolides. Rifampicin was the most active drug against the isolates in vitro. All Lpn isolates were inhibited by the following drugs (in decreasing order of their MICs): doxycycline>tigecycline>cefotaxime. The MICs of azithromycin, ciprofloxacin, levofloxacin, moxifloxacin, and tigecycline were significantly lower for Lpn non-sg 1 than Lpn sg 1 isolates. CONCLUSIONS: Susceptibility testing of Legionella strains to appropriate antibiotics should be performed often to evaluate the possible emergence of resistance, to improve the outcomes of patients, and to reduce the direct costs associated with hospitalization.

Detection of Listeria monocytogenes in ready-to-eat foods sampled from a catering service in Apulia, Italy.

OBJECTIVE: Listeria monocytogenes is currently considered a relevant emerging food-borne pathogen. In particular, the European Centre for Disease Prevention and Control (ECDC) illustrates its widespread presence in different foods. In the present article, L. monocytogenes prevalence was estimated in cooked ready-to-eat foods sampled from a catering service in a Apulia city, southern Italy. METHODS: The study was carried out from January to June 2014 in according to Regulation (EC) No. 852/2004, and ISO 11290-1:1996/Amd.1:2004 methods. RESULTS: Listeria spp. was isolated in 8.3% of the samples: L. monocytogenes was identified with the highest prevalence in potato gateau (66.6%), followed by rice dishes (11.1%), Listeria innocua was isolated from potato purea (11.1%) and cooked vegetables (11.1%). CONCLUSIONS: These preliminary results confirm the diffusion of the microorganism in ready-to-eat products; therefore, strategies aimed at protecting the consumers should be adopted. First of all, correct hygiene procedures should be followed and then microbiological tests should be implemented in order to early detect Listeria spp. (not only LM) contamination in cooked foods.

Mold contamination in a controlled hospital environment: a 3-year surveillance in southern Italy.

BACKGROUND: Environmental monitoring of airborne filamentous fungi is necessary to reduce fungal concentrations in operating theaters and in controlled environments, and to prevent infections. The present study reports results of a surveillance of filamentous fungi carried out on samples from air and surfaces in operating theaters and controlled environments in an Italian university hospital. METHODS: Sampling was performed between January 2010 and December 2012 in 32 operating theaters and five departments with high-risk patients. Indoor air specimens were sampled using a microbiological air sampler; Rodac contact plates were used for surface sampling. Fungal isolates were identified at the level of genera and species. RESULTS: Sixty-one samples (61/465; 13.1%) were positive for molds, with 18 from controlled environments (18/81; 22.2%) and 43 (43/384; 11.2%) from operating theaters. The highest air fungal load (AFL, colony-forming units per cubic meter [CFU/m(3)]) was recorded in the ophthalmology operating theater, while the pediatric onco-hematology ward had the highest AFL among the wards (47 CFU/m(3)). The most common fungi identified from culture of air specimens were Aspergillus spp. (91.8%), Penicillium spp., (6%) and Paecilomyces spp. (1.5%). During the study period, a statistically significant increase in CFU over time was recorded in air-controlled environments (p = 0.043), while the increase in AFL in operating theaters was not statistically significant (p = 0.145). Molds were found in 29.1% of samples obtained from surfaces. Aspergillus fumigatus was the most commonly isolated (68.5%). CONCLUSIONS: Our findings will form the basis for action aimed at improving the air and surface quality of these special wards. The lack of any genetic analysis prevented any correlation of fungal environmental contamination with onset of fungal infection, an analysis that will be undertaken in a prospective study in patients admitted to the same hospital.

Invasive fungal infections in patients with hematologic malignancies (aurora project): lights and shadows during 18-months surveillance.

The aim of this multicenter prospective study was to evaluate the incidence of invasive fungal infections (IFIs) in adult and pediatric patients with hematologic malignancies, involving nine nosocomial facilities in Southern Italy over a period of 18 months. Furthermore, results of an environmental microbial surveillance routinely carried out in some of the enrolled hospitals are reported. A total of 589 onco-hematological patients were enrolled and 27 IFIs were documented. The main infections were caused by yeasts, more than filamentous fungi (overall incidence of 2.7% and 1.9%, respectively). The yeasts were mainly represented by Candida spp. (87.5%), all isolated by blood cultures; C. parapsilosis was the most common species. Among mould infections, the most frequent site was the lung, with regard to aspergillosis (81.8%). In six of the 10 patients with suspected aspergillosis, the diagnosis was made by the detection of galactomannan and (1,3)-ß-d-glucan antigens. The microbiological surveillance carried out on 156 air, 312 water and 312 surface samples revealed low environmental contamination: Alternaria alternata was the only fungus isolated from two surface samples. Our data, especially the low occurrence of filamentous fungi, suggest a particular local epidemiology. Further studies are needed to confirm this microbiological trend in onco-hematological patients in Southern Italy, the results of which might be helpful to improve the management of these patients.

Diagnostic performance of 1→3-ß-d-glucan in neonatal and pediatric patients with Candidemia.

Fungal sepsis is one of the major problems in neonatal and pediatric care unit settings. The availability of new diagnostic techniques could allow medical practitioners to rapidly identify septic patients and to improve their outcome. The aim of this study was to evaluate the performance of the 1→3-ß-d-glucan (BDG), individually and in comparison with the Candida mannan (CM) antigen, in ten preterm infants and five onco-haematological pediatric patients with Candida bloodstream infections already proven by positive culture. The serum levels of BDG were >80 pg/mL on the same day as a positive blood culture in all examined patients, while CM antigen was negative in the patients with C. parapsilosis fungemia and in one further case due to C. albicans. These results suggest that a regular monitoring of serum circulating antigens (i.e., 1→3-ß-d-glucan) combined with other microbiological and clinical information, may allow earlier and accurate diagnosis. However, further studies are necessary to confirm its usefulness in routine clinical practice.

Candidemia in the Neonatal Intensive Care Unit: A Retrospective, Observational Survey and Analysis of Literature Data.

We evaluated the epidemiology of Candida bloodstream infections in the neonatal intensive care unit (NICU) of an Italian university hospital during a 9-year period as a means of quantifying the burden of infection and identifying emerging trends. Clinical data were searched for in the microbiological laboratory database. For comparative purposes, we performed a review of NICU candidemia. Forty-one candidemia cases were reviewed (overall incidence, 3.0 per 100 admissions). Candida parapsilosis sensu stricto (58.5%) and C. albicans (34.1%) were the most common species recovered. A variable drift through years was observed; in 2015, 75% of the cases were caused by non-albicans species. The duration of NICU hospitalization of patients with non-albicans was significantly longer than in those with C. albicans (median days, 10 versus 12). Patients with non-albicans species were more likely to have parenteral nutrition than those with C. albicans (96.3% versus 71.4%). Candida albicans was the dominant species in Europe and America (median, 55% and 60%; resp.); non-albicans species predominate in Asia (75%). Significant geographic variation is evident among cases of candidemia in different parts of the world, recognizing the importance of epidemiological data to facilitate the treatment.

Invitro antifungal susceptibilities of Candida species to liposomal amphotericin B, determined using CLSI broth microdilution, and amphotericin B deoxycholate, measured using the Etest.

The antifungal susceptibilities of 598 isolates of Candida spp. (bloodstream and other sterile sites) to liposomal amphotericin B (L-AmB) versus amphotericin B (AmB) were determined. MICs were calculated using the Clinical and Laboratory Standards Institute broth microdilution (M27-A3) method for L-AmB and the Etest method for AmB. The MIC50/MIC90 (µg ml-1) values for L-AmB broth microdilution and AmB Etest were 0.25/1 and 0.19/0.5, respectively. The overall essential agreement (±2 dilutions) was 91.5 %, ranging from 37.5 % (Candida lusitaniae) to 100 % (Candida glabrata and Candida krusei). Categorical agreement between the two methods was categorized based on a previously published breakpoint (susceptible/resistant MIC cut-off of 1 µg ml-1). The overall categorical agreement at the 48 h reading was 97.3 %, ranging from 72.7 % (C. krusei) to 100 % (Candida albicans). Major and very major discrepancies occurred in 2.3 and 0.3 %, respectively. Spearman's ρ was 0.48 (P<0.0001). These results demonstrate the utility of the AmB Etest as a surrogate marker to predict the sensibility and resistance of Candida spp. to L-AmB and thus to support its use in antifungal treatment.

Molecular Identification and Echinocandin Susceptibility of Candida parapsilosis Complex Bloodstream Isolates in Italy, 2007-2014.

The Candida parapsilosis group encompasses three species: C. parapsilosis, C. orthopsilosis, and C. metapsilosis. Here, we describe the incidence and echinocandin susceptibility profile of bloodstream isolates of these three species collected from patients admitted to an Italian university hospital from 2007 to 2014. Molecular identification of cryptic species of the C. parapsilosis complex was performed using polymerase chain reaction amplification of the gene encoding secondary alcohol dehydrogenase, followed by digestion with the restriction enzyme BanI. Minimum inhibitory concentrations were determined using the broth microdilution method according to European Committee for Antimicrobial Susceptibility Testing (EUCAST EDef 7.2) and Clinical Laboratory Standards Institute (CLSI M27-A3) guidelines, and the results were compared with those obtained using the E-test and Sensititre methods. Of the 163 C. parapsilosis complex isolates, 136 (83.4%) were identified as C. parapsilosis, and 27 (16.6%) as C. orthopsilosis. The species-specific incidences were 2.9/10,000 admissions for C. parapsilosis and 0.6/10,000 admissions for C. orthopsilosis. No resistance to echinocandins was detected with any of the methods. The percent essential agreement (EA) between the EUCAST and E-test/Sensititre methods for anidulafungin, caspofungin, and micafungin susceptibility was, respectively, as follows: C. parapsilosis, 95.6/97.8, 98.5/88.2, and 93.4/96.3; C. orthopsilosis, 92.6/92.6, 96.3/77.8, and 63.0/66.7. The EA between the CLSI and E-test/Sensititre methods was, respectively, as follows: C. parapsilosis, 99.3/100, 98.5/89.0, and 96.3/98.5; C. orthopsilosis, 96.3/92.6, 100/81.5, and 92.6/88.9. Only minor discrepancies, ranging from 16.9% (C. parapsilosis) to 11.1% (C. orthopsilosis), were observed between the CLSI and E-test/Sensititre methods. In conclusion, this epidemiologic study shows a typical C. parapsilosis complex species distribution, no echinocandin resistance, and it reinforces the relevance of using commercially available microbiological methods to assess antifungal susceptibility. These data improve our knowledge of the national distribution of species of the psilosis group, as there are very few studies of these species in Italy.

Candida Bloodstream Infections in Italy: Changing Epidemiology during 16 Years of Surveillance.

Although considerable progress has been made in the management of patients with invasive fungal infections, Candida bloodstream infections are still widespread in hospital settings. Incidence rates vary geographically, often because of different patient populations. The aim of the present study was to describe the epidemiology of candidemia, to analyze the trend of species distribution, and to measure the in vitro susceptibility to antifungal drugs in a university Italian hospital from 1998 to 2013. The antifungal susceptibility for all Candida isolates was evaluated by broth microdilution assay (CLSI M27-A3 document). Of 394 episodes of candidemia, the average incidence was 3.06/10,000 admissions. C. albicans and non-albicans Candida species caused 44.2% and 55.8% of the episodes, respectively. C. parapsilosis (62.2%) was the most common non-albicans. C. albicans predominated in almost all departments whereas C. parapsilosis was found in adult and paediatric oncohaematology units (34.8% and 77.6%, resp.). Overall, mortality occurred in 111 (28.2%) patients. Death occurred most often in intensive care units (47.1%) and specialist surgeries (43.7%). Most of the isolates were susceptible to antifungal drugs, but there was an upward trend for azole (P < 0.05). In conclusion, this study emphasizes the importance of monitoring local epidemiologic data and the diversity of patient groups affected.

In vitro activities of amphotericin B deoxycholate and liposomal amphotericin B against 604 clinical yeast isolates.

We determined the in vitro antifungal activity of liposomal amphotericin B (L-AmB) against 604 clinical yeast isolates. Amphotericin B deoxycholate (D-AmB) was tested in parallel against all the isolates. Susceptibility testing was performed according to the Clinical and Laboratory Standards Institute (CLSI) M27-A3 method. Overall, L-AmB was highly active against the isolates (mean MIC, 0.42 µg ml(-1); MIC90, 1 µg ml(-1); 97.2 % of MICs were ≤1 µg ml(-1)) and comparable to D-AmB (mean MIC, 0.48 µg ml(-1); MIC90, 1 µg ml(-1); 97.3 % of MICs were ≤1 µg ml(-1)). The in vitro activity of D-AmB and L-AmB was correlated (R(2) = 0.61; exp(b), 2.3; 95 % CI, 2.19-2.44, P<0.001). Candida albicans (mean MICs of D-AmB and L-AmB, 0.39 µg ml(-1) and 0.31 µg ml(-1), respectively) and Candida parapsilosis (mean MICs of D-AmB and L-AmB, 0.38 µg ml(-1) and 0.35 µg ml(-1), respectively) were the species most susceptible to the agents tested, while Candida krusei (currently named Issatchenkia orientalis) (mean MICs of D-AmB and L-AmB, 1.27 µg ml(-1) and 1.13 µg ml(-1), respectively) was the least susceptible. The excellent in vitro activity of L-AmB may have important implications for empirical treatment approaches and support its role in treatment of a wide range of invasive infections due to yeasts.