Development of a novel molecular tool for the rapid assessment of the biodiversity changes of benthic nematodes assemblages. Mares Conference Marine Ecosystems Helath and conservation

The molecular profiling system was developed using directed terminal-restriction fragment length polymorphism (dT-RFLP) to characterize soil nematode assemblages by relative abundance of feeding guilds and validation by comparison to traditional morphological method. The good performance of these molecular tools applied to soil nematodes assemblages create an opportunity to develop a novel approach for rapid assessment of the biodiversity changes of benthic nematodes assemblages of marine and estuarine sediments. The main aim of this research is to combine morphological and molecular analysis of estuarine nematodes assemblages, to establish a tool for fast assessment of the biodiversity changes within habitat recovery of Zostera noltii seagrass beds; and validate the dT-RFLP as a high-throughput tool to assess the system recovery. It was also proposed to develop a database of sequences related to individuals identified at species level to develop a new taxonomic reference system. A molecular phylogenetic analysis of the estuarine nematodes has being performed. After morphological identification, barcoding of 18S rDNA are being determined for each nematode species and the results have shown a good degree of concordance between traditional morphology-based identification and DNA sequences. The digest strategy developed for soil nematodes is not suitable for marine nematodes. Then five samples were cloned and sequenced and the sequence data was used to design a new dT-RFLP strategy to adapt this tool to marine assemblages. Several solutions were presented by DRAT and tested empirically to select the solution that cuts most efficiently, separating the different clusters. The results of quantitative PCR showed differences in nematode density between two sampling stations according the abundance of the nematode density obtained by the traditional methods. These results suggest that qPCR could be a robust tool for enumeration of nematode abundance, saving time.

Proposal and Validation of a Scale of Composite Measure Reactivity Score to Characterize the Reactivity in Horses During Handling

This study aimed to identify the parameters related to the expression of the reactivity in horses during handling and based on that proposed and validated a scale of composite measure reactivity score to characterize horse's reactivity. To this end, the first stage (S1) proposed the scale and the second (S2) validated it. In S1, 364 Lusitano horses were evaluated, 188 were adult breeding mares (4–12 years old), and 176 were foals (males/females, aged from 2 months to 2 years). During hooves trimming, vermifuge application, palpation scores were assigned to behaviors of movement, ears and eyes position, breathing, vocalization, and urination. A response parameter called reactivity was attributed to each animal, ranging from score 1 (nonreactive/calm) to score 4 (very reactive/aggressive). The verification of the possible parameters (age, behavior), which explains the response parameter (reactivity), was taken using ordinal proportional odds model. Movement, breathing, ears and eyes position, vocalization, and age appear to explain the reactivity of horses during handling (P < .01). Therefore, based on these parameters, it was possible to propose two scales of composite measure reactivity score: one to characterize the mares and another the foals. On S2, the proposed scale was validated by the simultaneous application of Forced Human Approach Test, another commonly used test to evaluate the reactivity in horses, with a correlation of 0.97 (P < .05). The assessment of the reactivity of horses during handling by a composite measure reactivity score scale is valid, and easy to apply, without disrupting daily routine and override the impact of individual differences.

Physicochemical and microbial analysis of feces from horses fed diets containing citrus pulp.

This study aimed to evaluate the e ect of diets containing increasing levels of citrus pulp on the physic-chemical and microbiological characteristics of horses feces. Five mares, at an average age of 3.5 years old and body weight of 492 ± 44.5 kg were arranged in a 5 x 5 Latin Square. The experimental diet consisted of 60% coast-cross hay and 40 % of concentrate with increasing levels of citrus pulp (0, 7, 14, 21, and 28 %). To determine the fecal pH, samples were collected directly from the oor, immediately after defecation, in the rst feces the day at 07:00 a.m., and color and fecal consistency were evaluated. For microbiological analysis, an aliquot was reserved in plastic bags, frozen, and sent to the microbiological laboratory for further analysis. Lactic acid bacteria were counted for Lactobacillus spp. and Streptococcus spp. from fecal samples under anaerobic conditions. The diet produced di erences (P<0.05) in feces consistency 98% had normal and rm stools, while 2% had loose ruminant-type feces. We observed no di erence (P<0.05) for color, verifying 100% of greenish feces, normal for equines. There was no e ect (P>0.05) on pH and on the number of Lactobacillus spp. and Streptococcus spp. The inclusion of up to 28% citrus pulp concentrates for horses did not promote change in the physio-chemical characteristics and on the population of lactic acid-producing bacteria in feces.