Vibrio virulence genes in fishes collected from estuarine waters in Italy.

AIMS: The aim of this study was to investigate the presence of Vibrio vulnificus and potentially pathogenic strains of Vibrio parahaemolyticus in mullets collected from estuarine environment in Italy. METHODS AND RESULTS: Two hundred and ninety-five mullets were analysed by culture using the selective medium thiosulfate citrate bile salt sucrose agar, during a monitoring period of 2 years (2008-2009). Presumptive Vibrio colonies were initially identified by using biochemical tests, and strains identified as V. parahaemolyticus and V. vulnificus were subsequently examined by PCR for the presence of species-specific and virulence genes (toxR, trh, tdh and vvh). V. parahaemolyticus was found in 55% (162/295) of fishes and V. vulnificus in 1% (3/295) with a higher presence in summer months. The trh+/tdh- strains were detected in 16% (47/295) of samples and only one strain resulted trh+/tdh+. One of the V. parahaemolyticus trh+ strains isolated belonged to the O1:KUT (K untypeable), a serotype recently associated to gastroenteritis in Italy. CONCLUSIONS: This is the first report demonstrating a high percentage of potential pathogenic V. parahaemolyticus trh+ strains in estuarine fishes of the Mediterranean area. SIGNIFICANCE AND IMPACT OF THE STUDY: These findings indicate the potential human health risk associated with the presence of pathogenic Vibrio spp. in wild fishes.

Evaluation of Adenovirus and E. coli as indicators for human enteric viruses presence in mussels produced in La Spezia Gulf (Italy).

AIMS: The purpose of this work was to verify whether E. coli is a good indicator of viral contamination in mussels and Adenovirus could represent a better alternative as indicator organism of viral presence to guarantee consumer health protection. METHODS AND RESULTS: Eighty samples of mussels from La Spezia Gulf were analysed for E. coli, Salmonella, Adenovirus, Norovirus and hepatitis A virus with cultural and biomolecular tests. The results of bacterial parameters showed E. coli within the law's limits and the absence of Salmonella. Twelve samples were positive for Adenovirus presence, one for Norovirus genogroup II and two for hepatitis A virus. None of these positive mussels was found to be contaminated with more than one virus at the same time. CONCLUSION: This study showed that there was not a direct correlation between the presence of human pathogenic viruses and bacterial indicators. SIGNIFICANCE AND IMPACT OF THE STUDY: Both E. coli and Adenovirus cannot be considered valid substitutes for the direct research of human pathogenic viruses in mussels. To improve consumer health protection, the European Commission will provide standardized methods for Norovirus and hepatitis A virus detection as soon as possible.

Legionella in industrial cooling towers: monitoring and control strategies.

AIMS: Legionella contamination of industrial cooling towers has been identified as the cause of sporadic cases and outbreaks of legionellosis among people living nearby. To evaluate and control Legionella contamination in industrial cooling tower water, microbiological monitoring was carried out to determine the effectiveness of the following different disinfection treatments: (i) continuous chlorine concentration of 0.01 ppm and monthly chlorine shock dosing (5 ppm) on a single cooling tower; (ii) continuous chlorine concentration of 0.4 ppm and monthly shock of biocide P3 FERROCID 8580 (BKG Water Solution) on seven towers. METHODS AND RESULTS: Legionella spp. and total bacterial count (TBC) were determined 3 days before and after each shock dose. Both strategies demonstrated that when chlorine was maintained at low levels, the Legionella count grew to levels above 10(4) CFU l(-1) while TBC still remained above 10(8 )CFU l(-1). Chlorine shock dosing was able to eliminate bacterial contamination, but only for 10-15 days. Biocide shock dosing was also insufficient to control the problem when the disinfectant concentration was administered at only one point in the plant and at the concentration of 30 ppm. On the other hand, when at a biocide concentration of 30 or 50 ppm was distributed throughout a number of points, depending on the plant hydrodynamics, Legionella counts decreased significantly and often remained below the warning limit. Moreover, the contamination of water entering the plant and the presence of sediment were also important factors for Legionella growth. CONCLUSIONS: For effective decontamination of outdoor industrial cooling towers, disinfectants should be distributed in a targeted way, taking into account the possible sources of contamination. SIGNIFICANCE AND IMPACT OF THE STUDY: The data of the research permitted to modify the procedure of disinfection for better reduce the water and aerosol contamination and consequently the exposure risk.

Study of the viral removal efficiency in a urban wastewater treatment plant.

Municipal and agricultural wastewater contain a variety of microorganisms and in particular enteric viruses. For the reuse of this treated wastewater it is important to ensure the efficiency of purification treatments and disinfection practices, that have often been insufficient to lower the viral load below the risk level. For this reason, for the routine analysis of recycled waters, the research into pathogenic viruses (e.g. HAV) and classical bacterial parameters (E. coli, enterococci and Salmonella) has to be associated with specific viral indicators such as somatic coliphages, adenovirus and TTV. The results of environmental monitoring, carried out in a wastewater treatment plant, showed the presence of adenovirus DNA in 100% of collected samples and TTV DNA in 95% (19/20) of raw sewage and in 85% (17/20) of the exit samples, while HAV was detected only in 2 samples over 40 (5%). The quantitative analysis has revealed an average reduction of 2 log for adenovirus and 1.58 log for TTV. The bacterial indicators were reduced by 1.74 log and 1.99 log respectively for E. coli and enterococci, while for somatic coliphages an average reduction of 2.2 log was observed. No significant correlation was shown between these parameters, confirming their inadequacy for the virological risk assessment. However the results of adenovirus confirm it as the best indicator to evaluate the efficacy of wastewater depuration plant in eliminating viruses.

One-year monthly monitoring of Torque teno virus (TTV) in river water in Italy.

Torque teno virus (TTV) is prevalent worldwide in general populations but at present is not related with any specific pathology. Its presence in faeces and its remarkable environmental stability suggest the possibility of using it as an indicator of faecal contamination in the environment. To evaluate the waterborne spread of TTV and its possible relationship with human pathogen enteric viruses, water samples were collected monthly for a year (May 2004-April 2005) from a river receiving the effluent of the treatment plant of the city of Pisa, concentrated and assayed with bimolecular tests (PCR, RT-PCR). TTV was detected in three samples (25%) while 16% of samples were positive for enteroviruses, 33% for rotaviruses, 8% for noroviruses genotype 1 and 25% for noroviruses genotype 2. Only two TTV samples (June and January) were also positive for rotavirus and norovirus, respectively. The detection of TTV in water confirmed its possible faecal-oral route of transmission but data are still insufficient to draw conclusions about the role of TTV as a viral indicator.

Epidemiological surveillance of human enteric viruses by monitoring of different environmental matrices.

In the aim of studying possible relations between viruses detected in clinical specimens and the ones found in different environmental matrices, in the period May 2004 to April 2005, the collection of faecal samples from gastroenteritis cases and the monthly monitoring of raw and treated wastewater, river water, seawater and mussels were carried out. The viruses considered for environmental monitoring were adenovirus, rotavirus, enterovirus, norovirus, hepatitis A virus (HAV) and Torque teno virus (TTV): they were searched for with PCR and RT-PCR and confirmed by gene sequencing. Faecal coliforms and somatic coliphages' counts were also determined. The surveillance of case detected 45 positive faecal samples out of 255 (17.6%) while 35 of 56 environmental samples (62.5%) resulted positive for at least one of the considered viruses. The detection of the same viral strain in the faeces of gastroenteritis cases and in water was possible for adenovirus and rotavirus, which were also predominant in environmental matrices; thus they could be considered as a reference for risk assessment.

Outpatient management of acute promyelocytic leukemia after consolidation chemotherapy.

The feasibility and safety of outpatient management of acute promyelocytic leukemia (APL) during the aplastic phase after intensive consolidation chemotherapy, the incidence and types of complications requiring readmission to hospital, and the number of hospital days spared by this policy have been prospectively evaluated. After chemotherapy administration, patients were evaluated on an ambulatory basis. In the event of any complication they referred to the Emergency Unit (EU) of our Department dedicated to outpatients with hematologic diseases. Forty patients with APL observed over a 4 year period were eligible for intensive chemotherapy. After the achievement of complete remission they received a total of 104 consolidation courses and in 98 instances they were followed on an ambulatory basis. There were 41 cases (42%) of rehospitalization for fever (40 cases) or severe anemia (one case). Only one patient died due to a brain hemorrhage. Streptococcus viridans was the organism most frequently isolated from blood. Empiric once-a-day antibacterial therapy with ceftriaxone and amikacin was effective in 87% of the cases and made possible early discharge in 28% of the cases to continue the antibiotic therapy on an outpatient setting. Patients were managed out of the hospital for 76% of the post-consolidation neutropenia period. Thanks to the availability of an EU specifically dedicated to outpatients with hematologic diseases, out-hospital management of APL patients after consolidation therapy appeared to be safe, well accepted, potentially cost-saving, and contributed to saving the risk of developing severe nosocomial infections.

Detection of antigenemia by enzyme-linked immunosorbent assay in horses with experimental Ehrlichia risticii infection.

Four horses were inoculated with Ehrlichia risticii contained in either infected murine P388 D1 cells or heparinized blood from an infected horse. All 4 horses produced serum antibody, plasma antigen, and clinical signs of the disease. An enzyme-linked immunosorbent assay was used to detect antibody in the serum and was also used in conjunction with an anti-E. risticii monoclonal antibody to detect antigenemia. These laboratory and clinical findings were correlated to determine the efficiency of the antigen detection method for discerning E. risticii infection.

Detection of humoral antigen and antibody by enzyme-linked immunosorbent assay in horses with experimentally induced Ehrlichia equi infection.

An enzyme-linked immunosorbent assay (ELISA) was used to detect antigen in plasma and antibody in serum of 3 horses inoculated with Ehrlichia equi. Clinical signs, including rectal temperature, were correlated with the antigen and antibody detection. ELISA was very efficient in detection of serum antibody. Antigen detection using monoclonal antibodies to E. equi and ELISA should be considered as a diagnostic method.