Our study provides a reliable estimate of the burden of the disease in a large patient population (4951 patients), that underwent C. difficile toxin testing through a 6-year period in 5 urban Italian hospitals. Data from our study showed that the rate of CDI among patients undergoing C. difficile toxin testing is on average 0.84 per 10,000 patient-days, and that this frequency significantly increased during the years in the study, with the peak (i.e. 2.3 per 10,000 patient-days) in 2011.
Limited information on the burden of CDI in Italy is available. Italian studies mainly focus on ribotyping of collected strains or antimicrobial susceptibility testing . The rate of C. difficile infection in Italy is still uncertain. Italian data on the incidence of CDI are derived solely from a recent European survey and a single-hospital study [13, 14]. As such, the overall number of patients tested is small (<1000).
Two European studies on nosocomial CDI conducted in 2005 and 2008 reported an incidence of CDI of 2.45 per 10,000 patient-days and 4.1 per 10,000 patient-days respectively [12, 13]. However both reported a wide range in incidence amongst different countries.
In the European hospital survey, 5 Italian hospitals reported a mean CDI rate per hospital of 3.6 per 10,000 patient-days . This represents a four-fold higher rate compared with our average incidence rate over the period examined, but only slightly higher if compared to the 2011 rate alone (2.3 CDI episodes per 10,000 patient-days). However, the Italian population analyzed in the European survey is limited and is approximately one tenth compared to our study. Furthermore in some Italian hospitals participating in the survey, cytotoxicity test and enzyme immunoassay for C. difficile-specific glutamate dehydrogenase were also perfomed . These test may have a higher diagnostic sensitivity. In addition, unlike in the European survey, where the CDI incidence was evaluated in a one-month period, we evaluated CDI incidence and trend over a 6-year period.
Interestingly, our trend is different from that described in the US by Lucado et al. , that reported that in the second half of the last decade the incidence rate appeared blunted. Our rising trend appears to be more similar to the first half of last decade in the Lucado study, thus hypothesizing that a delay of CDI diffusion in our region could have accounted for such a difference.
It is also of interest that in our study, there is a progressive increase in patients undergoing C. difficile toxin testing in the 6-year period, and a parallel progressive increase of CDI rates during the years. Increased testing could be partly related to a higher antibiotic consumption in the hospitals considered in the study, and partly to a developing CDI awareness among healthcare professionals. In recent years there has been an increasing usage of antimicrobials in the hospitals of Lazio Region, i.e. up to 34% increase from 2007 to 2009 . A similar increase was found in Spain by Asensio et al., and was associated with the increase of CDI prevalence rates .
Recently a new variable has been proposed for epidemiological studies on CD: C. difficile testing intensity (CDTI). CDTI was defined as the total number of CD test performed, normalized to the total number of patients with at least 1 CD test recorded. The authors explored this variable with the hypothesis that part of the increase in CDI incidence is attributable to false positive related to repeated testing . As a limitation of our study, data on CDTI are not available in our study population.
Our works demonstrate that the > 80 year-old age class display the highest percentage of CDI episodes, and overall the highest rates were reported in patients older than 60 years. This finding reflects similar data from previous studies conducted in the US  and Spain , where significant increasing trends between 2000 and 2003 and 1997–2005, respectively, were reported in both the older age groups with a larger increase in those aged ≥ 65 than in those aged 45–64 [19, 20].
Concerning the distribution of positivity by ward, in a survey conducted in Canadian hospitals 53% of CDI occurred in medical wards (including medicine unit, long-term-care facility, oncology/hematology unit and transplant unit), 23% occurred in surgical wards and 10% in intensive care units , which is comparable to our findings.
Our study has some limitations. First, due to the retrospective design of the study, the clinical pictures of the patients are not available. Secondly, data on C. difficile testing intensity (CDTI) are not available in our study population. CDTI has been proposed as a possible factor associated with the incidence of hospital acquired CDI. CDTI, defined as the total number of CD test performed, normalized to the total number of patients with at least 1 CD test recorded, reflects the frequency of multiple test recorded for a single patient, presumably in response to an initially negative result . This variable could limit the possible bias deriving from repeat testing.
Another potential limitation could be an improved sensitivity in laboratory procedures over time, leading to an increased CDI detection. However, the participating hospitals used the enzyme immunoassays for A/B toxins for the study period, and no more sensitive/specific diagnostic tool (i.e. 2-step algorithm including glutamate dehydrogenase testing and molecular biology) was adopted during the study period.
Data concerning antimicrobial consumption in the surveyed hospitals during the study period are lacking. We were unable to assess the rate of CDI episodes per patient-days by ward, due to the unavailability of the number of admissions per ward. Finally, data analised according to subspecialty are not available either.