"Ozonated water for parasitic control in buffalo farms (OZO-PAR)"

Dairy water buffalo (Bubalus bubalis) farming plays an important role in the economy of several countries, including Italy, as buffalo milk is almost exclusively used for the production of mozzarella cheese. However, in intensive farming systems, infection of water buffaloes with intestinal protozoa, such as Giardia, Cryptosporidium and Eimeria, might threaten the profitability and sustainability of milk production. These infections have constantly increased over the years through contamination of water, feed and environment by the infective stages of these parasites, e.g., Giardia cysts and Cryptosporidium/Eimeria oocysts. In particular, well water used for daily activities on the farm (e.g., watering the animals, cleaning the premises, etc.) could represent an important source of infection. Even conventional chlorination programs, at the dosages used in the official treatment plans, are not effective against intestinal protozoa. Sanitation of drinking water (using products such as ammonia, chlorine dioxide, hydrogen dioxide and ozone) for livestock animals has been suggested as a useful strategy to be directed at reducing and/or preventing the transmission of the infective (oo)cysts to the animals. Therefore, the adoption of appropriate control strategies against intestinal protozoa is a considerable challenge for water buffalo farms worldwide. Although metaphylactic approaches have been used successfully to control infections by intestinal protozoa in ruminant farms, reinfections are very common, thus requiring repeated treatments that, in turns, might increase the potential for developing drug resistance as well as contributing to the dispersion of antiparasitic drugs into the environment. In light of these concerns, the need to introduce eco-friendly and alternative strategies to control intestinal protozoa infections is a considerable challenge for preserving the health and welfare of water buffalo farms. The general aim of this industrial PhD project was to introduce an innovative system for water ozone production to be used as a sanitation strategy to control intestinal protozoa (Giardia and Eimeria) infections of water buffaloes in the Campania region of southern Italy. The specific aims presented in this thesis were: i) presenting the epidemiological scenario of intestinal protozoa in water buffaloes in the Campania region of southern Italy; ii) selecting a sensitive technique to detect Giardia cysts; iii) setting up an innovative ozone generator system; iv) conducting a series of in vitro tests to assess the effect of the ozone treatment on Giardia cysts and Eimeria oocysts. Chapter 1 provides a general overview of Eimeria spp. and Giardia duodenalis infections in livestock. In this chapter basic knowledge related to the taxonomy, life cycle, epidemiology and pathogenesis of Eimeria spp. and G. duodenalis are reported. Moreover, the different coprological, serological and molecular techniques used in veterinary medicine are described. In the face of this knowledge, control of both protozoa represents the principal challenge in water buffalo farms. Currently, conventional metaphylactic treatments are associated with increased resistance, thus, eco- friendly, alternative strategy to control intestinal protozoa are indispensable tools to reduce the risk of transmission of protozoal infections in water buffalo farms. Chapter 2 provides an overview of the ozone gas, its general properties and current application in water sanitation treatment. This chapter details the chemical and physical properties of ozone, its strong oxidizing power and environmental factors (e.g., pH, temperature) limiting its working power. Moreover, in this chapter, the main industrial methods to generate ozone are described, by specifying advantages and disadvantages. For my thesis, here is described the use of ozone in water treatment as an alternative sanitizer. Chapter 3 shows the results of an epidemiological investigation of Eimeria spp. in water buffaloes in Southern Italy. Compared with cattle, there is limited scientific knowledge about the health of water buffaloes so updated data on parasitic infections (as eimeriosis) is an interesting challenge in this species. Furthermore, the published studies on eimeriosis in large ruminants in Italy are few and focused mainly on treatment while the epidemiological data in Europe are scarce, not updated, and focused only on cattle. For this purpose, parasitological data on eimeriosis from a 10-year surveillance were analysed. The results shows that Eimeria spp., in the same way as Giardia spp., is a persistent and complex problem in water buffalo farms, and control strategies need to be implemented on farms. Chapter 4 investigates a sensitive and cost-effective technique for the detection of Giardia cysts in faecal samples. While copromicroscopic techniques are well-established methods for the detection of Eimeria oocysts, some concerns still apply to the use of copromicroscopic methods for the detection of Giardia cysts. For this purpose, immunoassays and FLOTAC techniques were compared for diagnosing Giardia spp. infection. The results from the cost-effectiveness analysis, in combination with the sensitivity and specificity of the FLOTAC technique, suggest that the FLOTAC technique can be used in the routine diagnosis of Giardia spp. infection in animals. Chapter 5 describes activities carried out in the UK, at the University of Bristol and the Draper Biotech Limited (DBL), the latter being an industrial company specialized in air and water purification systems using ozone. Moreover, the training in the UK included visits to poultry and cattle farms designed by the industrial company as the experimental setting to study the effect of ozonated drinking water in vivo. Moreover, in the Aberdeen Angus farm (Salisbury, UK) I participate to instal the farm box pipefitting ozone generator. In this chapter the main part of the farm box is detailed. Furthermore, once in Italy, some preliminary in vitro tests using a well-water medium were performed to evaluate the effect of water ozonation (using an electrolytic cell within polycrystalline diamond electrode) on the viability of Eimeria oocysts and Giardia cysts collected from water buffaloes. Moreover, a preliminary in vivo test was performed in water buffalo calves to evaluate Eimeria oocyst output reduction and performance, like weight gain from the antiprotozoal treatment with ozonated drinking water. The results of the preliminary in vitro and in vivo studies on the effect of ozonated water on Eimeria and Giardia suggested that ozone could be a promising eco-friendly tool to control protozoa infections in water buffalo farms. Chapter 6 reports the results of a proof-of-concept study aimed at evaluating the effect of ozone on the viability of Eimeria oocysts and Giardia cysts. For this purpose, in vitro tests were performed to determine the minimum concentrations of O3 (mg/l) and the contact times (minutes) necessary to inactivate Eimeria oocysts and G. duodenalis cysts. The results of the present study showed different values of efficacy of ozonated water on the viability of Eimeria oocysts (33.0%) and G. duodenalis cysts (96.3%) isolated from water buffaloes. Eimeria spp. and Giardia duodenalis represent a persistent and complex problem in water buffaloes that impair their health, welfare, and production. Currently, conventional metaphylactic treatments are associated with increased resistance. The sanitation of drinking water for livestock animals could be a useful eco-friendly, alternative strategy to control the diffusion of both protozoa in intensive buffalo farms

Epidemiology and spatial distribution of Echinococcus granulosus in sheep and goats slaughtered in a hyperendemic European Mediterranean area

Background: Cystic echinococcosis (CE) is a worldwide parasitic zoonosis caused by the larval stage of Echinococcus granulosus sensu lato affecting livestock, particularly sheep and goats. However, often this parasitosis is underestimated. For this reason, this study aimed to evaluate the epidemiological features and spatial distribution of CE in sheep and goats slaughtered in a hyperendemic Mediterranean area. Methods: A survey was conducted in the Basilicata region (southern Italy) from 2014 to 2019. A total of 1454 animals (1265 sheep and 189 goats) from 824 farms were examined for hydatid cyst detection by visual inspection, palpation and incision of target organs. All the CE cysts were counted and classified into five morphostructural types (unilocular, multiseptate, calcified, caseous and hyperlaminated). Molecular analysis was performed on 353 cysts. For spatial analysis, a kriging interpolation method was used to create risk maps, while clustering was assessed by Moran’s I test. Results: CE prevalence of 72.2% (595/824) and 58.4% (849/1454) was observed at the farm and animal levels, respectively, with higher values in sheep (62.9%) than goats (28.0%). The liver and lungs were the most frequently infected organs in both sheep and goats. Most of recovered cysts were of the calcified and multiseptate morphotypes. All the isolates were identified as E. granulosus sensu stricto (genotypes G1–G3). Spatial distribution showed a moderate clustering of positive animals. Conclusion: The findings of this study can be used to better understand the eco-epidemiology of echinococcosis and to improve CE surveillance and prevention programs in regions highly endemic for CE.Fil: Bosco, Antonio. Università degli Studi di Napoli Federico II; ItaliaFil: Alves, Leucio Camara. Universidad Federal Rural Pernambuco; BrasilFil: Cociancic, Paola. Università degli Studi di Napoli Federico II; Italia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Centro de Estudios Parasitológicos y de Vectores. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Centro de Estudios Parasitológicos y de Vectores; ArgentinaFil: Amadesi, Alessandra. Università degli Studi di Napoli Federico II; ItaliaFil: Pepe, Paola. Università degli Studi di Napoli Federico II; ItaliaFil: Morgoglione, Maria Elena. Università degli Studi di Napoli Federico II; ItaliaFil: Maurelli, Maria Paola. Università degli Studi di Napoli Federico II; ItaliaFil: Ferrer Miranda, Edyniesky. Universidad Federal Rural Pernambuco; BrasilFil: Santoro, Kleber Régis. Universidad Federal Rural Pernambuco; BrasilFil: Nascimento Ramos, Rafael Antonio. Universidad Federal Rural Pernambuco; BrasilFil: Rinaldi, Laura. Università degli Studi di Napoli Federico II; ItaliaFil: Cringoli, Giuseppe. Università degli Studi di Napoli Federico II; Itali

Constraints of using historical data for modelling the spatial distribution of helminth parasites in ruminants

Dicrocoelium dendriticum is a trematode that infects ruminant livestock and requires two different intermediate hosts to complete its lifecycle. Modelling the spatial distribution of this parasite can help to improve its management in higher risk regions. The aim of this research was to assess the constraints of using historical data sets when modelling the spatial distribution of helminth parasites in ruminants. A parasitological data set provided by CREMOPAR (Napoli, Italy) and covering most of Italy was used in this paper. A baseline model (Random Forest, VECMAP®) using the entire data set was first used to determine the minimal number of data points needed to build a stable model. Then, annual distribution models were computed and compared with the baseline model. The best prediction rate and statistical output were obtained for 2012 and the worst for 2016, even though the sample size of the former was significantly smaller than the latter. We discuss how this may be explained by the fact that in 2012, the samples were more evenly geographically distributed, whilst in 2016 most of the data were strongly clustered. It is concluded that the spatial distribution of the input data appears to be more important than the actual sample size when computing species distribution models. This is often a major issue when using historical data to develop spatial models. Such data sets often include sampling biases and large geographical gaps. If this bias is not corrected, the spatial distribution model outputs may display the sampling effort rather than the real species distribution