2.2. Experimental assays
The experiment was conducted between April and September 2021, at the Federal Institute of Paraíba (IFPB), Sousa campus, Paraíba, northeastern Brazil, which is located at latitude 06º 50' 22" S, longitude 38º 17' 42" W. The region is characterized by a rainy season from January to May, during which 98.6% of the annual rainfall occurs; and a dry season, in the other months (INMET, 2010).
Twenty female sheep, aged between 12 and 18 months, Santa Inês breed, were used. Before the beginning of the experiment, the animals were dewormed according to the protocol described by Vilela et al. (2012).
After confirmation that zero EPG had been achieved, the animals were randomly divided into two groups (treated and control), with ten animals each. Each group was placed in a 1.2 ha paddock, composed of native Caatinga pasture, predominantly Senna obtusifolia L., Mimosa tenuiflora, Caesalpinia pyramidalis, Croton sonderianus, Malva sp., Zizyphus joazeiro, Digitaria horizontalis, Mimosa caesalpiniifolia and Cnidoscolus phyllacanthus. The paddocks were naturally infested with helminth larvae, due to the previous grazing of four sheep over a 15-day period (1875 ± 900 EPG; Haemonchus sp. 86%; Trichostrongylus spp. 10% and Oesophagostomum sp. 4%). Throughout the experimental period, each group remained in its respective paddock.
The animals in each group received daily protein-energy commercial food in the proportion of 0.5% body weight, along with mineral salt and water ad libitum. In the treated group, each animal received 1 g of the Bioverm® product for each 10 kg of live weight, daily, together with the commercial feed. In the control group, the animals received the feed without Bioverm®. In order to avoid deaths among the animals, salvage treatments were applied, consisting of the anthelmintic 5% levamisole hydrochloride, in any situations in which the animals presented hematocrit lower than 16%.
Fecal samples were monthly collected from all the animals to measure egg counts per gram of feces (EPG - Gordon and Whitlock, 1939), and perform fecal cultures (Roberts and O’Sullivan, 1950). Infective larvae were morphologically identified (Keith, 1953). Blood samples were monthly collected to determine the packed cell volume (PCV) percentage (Ferreira Neto et al., 1981). Animal weights were estimated on an analytical balance. Furthermore, to determine the levels of environmental infestation by infective larvae, five samples of approximately 500 g of leaf mass from the pasture were collected monthly from the paddocks of both groups, in different places of each paddock, in accordance to Vilela et al. (2018). The dry matter was weighed and the infective larvae were quantified, using an optical microscope, to determine the values of L3/kg of dry matter.
Data on maximum and minimum temperature, rainfall and relative humidity were recorded at a meteorological station in Sousa, Paraíba, Brazil.