Overall, the number of serological tests for CanL (n=80,309) performed in the three areas of Italy is reported in Table 1, being higher in southern Italy (37.7% of all tests performed) than in central Italy (31%), and northern Italy (24.7%). Conversely, the overall number of tests requested for the diagnosis of HW throughout Italy (n=10,324) was significantly lower (11.3% of all the requested serological tests) than for CanL, with a relative high number in northern Italy (51.7% of all tests performed) and the lowest in southern Italy (Table 1).
Results showed a significant difference in CanL prevalence among the different study macroareas (χ2=218.564, d.f.=2, p<0.0001), with an overall prevalence of 28.2% in southern Italy and islands, 29.68% in central Italy and 21.62% in northern Italy (Table 1; Fig. 2). The impact of the study year on CanL prevalence in Italy was also significant (χ2=559.846, d.f.=10, p<0.0001), with values >30% in 2011, 2012, 2014 and 2015. HW prevalence showed significant differences among the study macroareas (χ2=114.879, d.f.=2, p<0.0001), being of 2.83% in northern Italy, 7.75% in central Italy and 4.97% in southern Italy and islands. On the whole Italian territory, HW prevalence significantly varied over years (χ2=108.401, d.f.=10, p<0.0001), gradually increasing from 0.77% in 2009 to values ranging from 5.19-8.47% in 2016-2017 (Table 1; Fig. 3).
A detailed analysis over CanL and HW prevalence trend over time in the three Italian macroareas was provided by assessing the impact of the study year and region on CanL and HW prevalence. In northern Italy, CanL prevalence showed significant differences over the study years (χ2=286.277, d.f.=10, p<0.0001), being highest in 2012 and 2014 (i.e., 33.51% and 27.27%, respectively) (Fig. 2). Notably, significant variations in CanL prevalence among the various northern Italy regions were recorded (χ2=190.657, d.f.=7, p<0.0001), with the highest values in Piedmont, Trentino Alto Adige, Valle d’Aosta, and Friuli Venezia Giulia (28.93%, 27.59%, 27.40% and 27.17%, respectively). The study year and region significantly impacted HW prevalence in northern Italy (χ2=56.954, d.f.=10, p<0.0001; χ2=40.555, d.f.=7, p<0.0001, respectively), being highest prevalence rate observed in 2016 (7.20%) (Fig. 3). Highest prevalence values were retrieved in Val d’Aosta (11.36%), Trentino Alto Adige (7.41%) and Piedmont (6.29%), while the lowest regional prevalence was in Veneto (2.12%).
In central Italy, both the study year and region led to significant differences in CanL prevalence (χ2=371.252, d.f.=10, p<0.0001; χ2=609.769, d.f.=5, p<0.0001, respectively). It was higher than 30% in 2012, 2014, 2015 and 2016 (Fig. 2), with the highest values in Lazio (38.52%), followed by Umbria (35.61%) and Abruzzo (34.10%). HW prevalence was also affected by the study year (χ2=55.333, d.f.=10, p<0.0001), showing values >10% in 2016 (i.e., 12.1%) and 2017 (i.e, 13.32%) (Fig. 3). Significant differences in the prevalence of HW were noted among central Italy (χ2=80.975, d.f.=4, p<0.0001, respectively) with the highest values recorded in Tuscany (11.48%) and Marche (7.84%).
In southern Italy and islands, CanL prevalence was significantly different among the study years (χ2=201.963, d.f.=10, p<0.0001), with values ranging from 24.55% (2017) to 50.66% (2011) (Fig. 2). CanL prevalence in this macroarea also showed significant differences among regions (χ2=642.949, d.f.=6, p<0.0001) with the highest values recorded in Molise (54.26%), Sicily (50.18%) and Sardinia (38.34%). On the other hand, the study region did not play a significant role impacting HW prevalence, even if a trend was observed (χ2=10.723, d.f.=6, p=0.10, respectively). Indeed, the largest number of analysed samples was from Apulia (n = 1608) followed by Basilicata (n = 80) and Sardinia (n = 70). Considering regions with sample size >40, the highest prevalence rates were from Sardinia and Apulia (10%, and 4.73%, respectively). The effect of the study year was not significant (χ2=16.145, d.f.=10, p=0.136) due to the limited number of samples examined in the years 2009-2014 (i.e., n=117), at variance with the larger samplings done in 2015-2019 (i.e., n=1,729).
Contingency results obtained analysing the separate datasets for each study macroarea are given in the Supplementary Online Material Table S1. Overall, observed changing distribution patterns of CanL and HW in Italy were not random or due to a biased sampling over the different areas and years (CanL, macroarea: χ2=486.62, d.f.=2, p<0.0001; year: χ2= 827.903, d.f.=10, p<0.0001, respectively; HW, macroarea: χ2=104.545, d.f.=2, p<0.0001; year: χ2=99.070, d.f.=10, p<0.0001, respectively). Serological CanL results had a similar cumulative prevalence throughout the Italian territory (Fig. 2), showing an annual slight decrease in all the regions, from 2015 to 2018. CanL was widely distributed throughout the Italian peninsula, with a number of positive animals >400 in central (Lazio and Tuscany) and southern (Apulia and Basilicata) regions as well as in both Islands (Sardinia and Sicily). In addition provinces with >300 positive samples were in northern regions (i.e., Turin, Piedmont, and Vicenza Veneto) (Fig. 4).
The highest prevalence of HW was registered in the central regions (i.e., Tuscany and Lazio) followed by southern and northern regions with annual variation patterns throughout the observation period (Fig. 3). An overall lower number of samples was positive for HW compared to CanL, with >10 positive cases recorded in three provinces from the northern Italy (i.e., Turin, Piedmont; Genova, Liguria; and Belluno, Veneto). The province with the highest number of HW cases in central Italy (i.e., Florence, Tuscany) was surrounded by provinces with more than 20 positive samples (i.e., Bologna, Emilia Romagna; Arezzo and Prato, Tuscany), was recorded in central Italy. The largest number of positive samples in southern Italy was recorded in the Apulia region (i.e., Brindisi and Lecce provinces). No data was available for four provinces of the North (i.e., Biella, Piedmont; Lecco Lombardia; Piacenza, Emilia Romagna; and Verona, Veneto), for two provinces of southern Italy (i.e., Isernia, Molise; Vibo-Valentia, Calabria), and many provinces of the Islands (i.e., Nuoro and Oristano, Sardinia; Agrigento, Caltanissetta and Ragusa, Sicily) (Fig. 5).