Characterization of an ecto-ATPase of Tritrichomonas foetus.

In this work, we describe the ability of living Tritrichomonas foetus to hydrolyze extracellular ATP. The addition of MgCl(2) to the assay medium increased the ecto-ATPase activity in a dose-dependent manner. At 5mM ATP, half maximal stimulation of ATP hydrolysis was obtained with 0.46mM MgCl(2). The ecto-ATPase activity was also stimulated by MnCl(2) and CaCl(2), but not by SrCl(2). The Mg(2+)-dependent ATPase presents two apparent K(m) values for Mg-ATP(2-) (K(m1)=0.03 mM and K(m2)=2.01 mM). ATP was the best substrate for this enzyme, although other nucleotides such as ITP, CTP, UTP also produced high reaction rates. GTP produced a low reaction rate and ADP was not a substrate for this enzyme. The Mg(2+)-dependent ecto-ATPase activity was insensitive to inhibitors of other ATPase and phosphatase activities, such as oligomycin, sodium azide, bafilomycin A(1), ouabain, furosemide, vanadate, molybdate, sodium fluoride and levamizole. The acid phosphatase inhibitors (vanadate and molybdate) inhibited about 60-70% of the Mg(2+)-independent ecto-ATPase activity, suggesting that the ATP hydrolysis measured in the absence of any metal divalent could, at least in part, also be catalyzed by an ecto-phosphatase present in this cell. In order to confirm the observed Mg(2+)-dependent activity as an ecto-ATPase, we used an impermeant inhibitor, 4,4'-diisothiocyanostylbene-2',2'-disulfonic acid (DIDS) as well as suramin, an antagonist of P(2) purinoreceptors and inhibitor of some ecto-ATPases. These two reagents inhibited the Mg(2+)-dependent ATPase activity in a dose-dependent manner. This ecto-ATPase was stimulated by more than 90% by 50mM D-galactose. Since previous results showed that D-galactose exposed on the surface of host cells is involved with T. foetus adhesion, the Mg(2+)-dependent ecto-ATPase may be involved with cellular adhesion and possible pathogenicity.     

Influence of edge and topography on the vegetation in an Atlantic Forest remnant in northeastern Brazil

Like edge effects, variations in topography can influence the physiognomy and structural patterns of the vegetation due to the high dynamics encountered in inclined areas. We evaluated the effect of edges on the structure and physiognomy of the arboreal and woody understory communities in patches of an Atlantic Forest remnant in two distinct topographic positions. Four study situations were laid out (slope edge, top edge, slope interior, and top interior), with ten 10 × 10 m plots each to sample the arboreal components (circumference at breast height ≥15 cm), and ten 5 × 5 m plots (in the corner of each arboreal component plot) to sample the woody understory (circumference at ground level ≥3 cm and with circumference at breast height <15 cm). We investigated physiognomic and structural variables of edge and interior environments in similar topographic positions on both slope and top (level) areas of the same remnant forest. Physiognomic and structural differences between edge and interior vegetation were noted, and these were found to be still more evident when different topographic positions were considered. Physiognomic and structural variations between the edge and interior vegetation were more notable on slopes than on hill tops, indicating greater dynamics in areas with greater topographic inclinations.     

In vitro and in vivo characterization of p-amino-phenethyl-m-trifluoromethylphenyl piperazine (PAPP), a novel serotonergic agonist with anthelmintic activity against Haemonchus contortus, Teladorsagia circumcincta and Trichostrongylus colubriformis

The neurotransmitter serotonin (5-hydroxy tryptamine or 5HT) regulates key physiological processes in nematodes such as locomotion and feeding. PAPP (p-amino-phenethyl-m-trifluoromethylphenyl piperazine) is a known agonist of the 5-HT(1Hc) receptor of the barber pole worm, Haemonchus contortus. In this study, PAPP was highly active against L3-stage larvae of H. contortus and Trichostrongylus colubriformis in an in vitro larval migration assay, with EC50 values of 9.36 and 11.8 microM, respectively, that were comparable to levamisole (10.2 microM) and superior to pyrantel (55.39 microM). When administered orally or subcutaneously to nematode infected gerbils, PAPP provided >99% efficacy against H. contortus and >98% efficacy against Teladorsagia circumcincta at 100 mg/kg, comparable to levamisole at 10 mg/kg. Drug titration revealed significant activity down to 50 mg/kg against these two species. Spectrum was limited, however, with somewhat lower efficacy (83%) in T. colubriformis infected gerbils at 100 mg/kg. Oral delivery of hydrochloride, acetate and phosphate salts of PAPP to nematode infected gerbils did not result in an increase in either potency or spectrum. The finding that PAPP exhibits significant anthelmintic activity suggests that the nematode-specific serotonergic system is a viable target for future anthelmintic discovery.     

Activation of goat primordial follicles in vitro: Influence of alginate and ovarian tissue

The present study aimed to evaluate the effect of three culture systems on caprine primordial follicle activation in vitro: follicles cultured either in the isolated form within alginate (Isolated follicles + Alginate treatment), or enclosed in ovarian tissue (in situ), with or without alginate (Fragment + Alginate, and Fragment alone treatments, respectively). After culture, the Isolated follicles + Alginate treatment presented a percentage of morphologically normal follicles (MNF) similar to both the non-cultured control and the Fragment Alone treatments. Nevertheless, Fragment + Alginate treatment showed a significant reduction in the number of MNF when compared to the other treatments. Regarding follicle development, our results showed that regardless of the alginate, the presence of ovarian tissue limited primordial follicle activation during in vitro culture. Remarkably, the Isolated primordial follicle + Alginate treatment was the only one that significantly promoted follicle activation and increased both follicle and oocyte diameters during IVFC, pointing out a higher cell proliferation. In conclusion, the presence of ovarian tissue with or without alginate limited follicle development (activation) after culture. Nevertheless, when primordial follicles were isolated and encapsulated in alginate they presented suitable survival rates, higher rates of follicle activation and continued to grow throughout the culture period.     

Evaluation of irrigation systems in the irrigated area of Chanza (Huelva)

In the irrigated area of Chanza (Huelva, Spain) there are about 1500 ha of strawberries under localized irrigation. A survey, covering 12% the farms and 20% of the area, was performed in order to characterize the farms, their water use, the cultivation techniques and crop yields and the localized irrigation systems used. Results of this survey were employed to select 20 farms for the evaluation of the irrigation systems using the procedure of Merriam and Keller (1978) modified according to local conditions. Results of the survey and evaluations of the strawberry crops are presented and the unitary water use determined. The conclusions summarize the main problems detected as well as their causes and possible solutions.     

An Earlier Uterine Environment Favors the In Vivo Development of Fresh Pig Morulae and Blastocysts Transferred by a Nonsurgical Deep-uterine Method

This study aimed to evaluate the effect of recipient-donor estrous cycle synchrony on recipient reproductive performance after nonsurgical deep-uterine (NsDU) embryo transfer (ET). The transfers (N=132) were conducted in recipients sows that started estrus 24 h before (–24 h; N=9) or 0 h (synchronous; N=31), 24 h (+24 h; N=74) or 48 h (+48 h; N=18) after the donors. A total of 30 day 5 morulae or day 6 blastocysts (day 0=onset of estrus) were transferred per recipient. The highest farrowing rates (FRs) were achieved when estrus appeared in recipients 24 h later than that in the donors (81.1%), regardless of the embryonic stage used for the transfers. The FR notably decreased (P<0.05) when recipients were –24 h asynchronous (0%), synchronous (61.3%) or +48 h asynchronous (50%) relative to the donors. No differences in litter size (LS) and piglet birth weights were observed among the synchronous and +24 h or +48 h asynchronous groups. While a +24 h asynchronous recipient was suitable for transfers performed with either morulae (FR, 74.3%; LS, 9.2 ± 0.6 piglets) or blastocysts (FR, 84.6%; LS, 9.8 ± 0.6 piglets), a + 48 h asynchronous recipient was adequate for blastocysts (FR, 87.5%; LS, 10.4 ± 0.7 piglets) but not for morulae (FR, 30.0%; LS, 7.3 ± 2.3 piglets). In conclusion, this study confirms the effectiveness of the NsDU-ET technology and shows that porcine embryos tolerate better a less advanced uterine environment if they are nonsurgically transferred deep into the uterine horn.     

Plasma Antimullerian Hormone as a Predictor of Ovarian Antral Follicular Population in Bos indicus (Nelore) and Bos taurus (Holstein) Heifers

In Bos taurus cattle, antimullerian hormone (AMH) has been demonstrated to have a high degree of correlation with ovarian antral follicle count and the number of healthy follicles and oocytes. To document the correlation between the plasma concentration of AMH and follicular number in Bos indicus and Bos taurus heifers, Nelore (Bos indicus, n = 16) and Holstein heifers (Bos taurus, n = 16) had their ovarian follicular waves synchronized. After synchronization, ovarian antral follicular population (AFP) was evaluated three times at 60‐day (d) intervals (T‐120 d, 120 days before plasma AMH determination; T‐60 d, 60 days before; and T0, at the time of plasma AMH determination). The plasma AMH concentration was positively correlated with the number of ovarian follicles on the day of the follicular wave emergence in Bos indicus (Nelore) and Bos taurus (Holstein) heifers at each evaluation time (p < 0.05). The AFP was higher in Bos indicus (Nelore) than in Bos taurus (Holstein) heifers (p < 0.05). Similarly, the AMH concentration was higher in Bos indicus (Nelore) than in Bos taurus (Holstein) heifers (p < 0.0001). When heifers were classified as to present high or low AFP according to the mean of the AFP within each genetic group, high‐AFP heifers presented a greater (p < 0.0001) AMH concentration than low‐AFP heifers, regardless of the genetic group. In conclusion, the AFP is positively correlated with plasma AMH concentration in both Bos indicus (Nelore) and Bos taurus (Holstein) heifers. Furthermore, Bos indicus (Nelore) heifers presented both greater plasma AMH concentrations and AFP than Bos taurus (Holstein) heifers.     

Bacteria associated with winter mortalities in laboratory-reared common dentex (Dentex dentex L.)

A pathological study was conducted on laboratory‐reared juvenile common dentex (Dentex dentex) suffering trickling and continuous mortalities. During a 3‐month period (October–December), water temperature, clinical signs and mortalities were recorded. Moribund or dead fish were examined for bacteria and parasites. Bacteria were isolated from head kidney and external ulcers, and samples from the gills, intestine, stomach, trunk kidney, gall bladder and liver were taken for histology. Cumulative mortality reached 73%, and 80% of fish examined were positive for bacteria (102 isolates). Vibrio splendidus was the most prevalent in head kidney (59.7%) and ulcers (88.9%), and it was frequently isolated as pure culture (74.7% and 100% respectively). This is the first report of this bacterium in association with mortality in common dentex. Vibrio scophthalmi was the second most prevalent bacterium (29.2%) and accounted for 63.6% of the mixed infections with V. splendidus, V. harveyi, Pseudoalteromonas spp. and other species were rarely isolated. No parasites were found in histological sections. Bacteria were frequently observed in the lumen of the intestine and stomach, destroying the epithelium. A cellular reaction was suggested by the high numbers of rodlet cells (RC) in the intestinal epithelium, and the abundance of eosinophilic granular cells (EGC) in the intestinal lamina propria. In the stomach, vacuolized cells containing unidentified debris, sometimes of crystalloid appearance, were very common. The high prevalence of V. splendidus in pure cultures and the absence of other aetiological agents suggest that V. splendidus is substantially involved in the registered mortalities, although the implication of V. scophthalmi and even of some nutritional factors cannot be discounted.     

Segmentation of Somatic Cells in Goat Milk Using Color Space CIELAB

Somatic cell counts （SCCs） levels indicate the occurrence of infections in goat udders and are related to the productivity of goat milk, cheese and yoghurt. This work presents a segmentation method for counting somatic cells in goat milk images, intending to detect an infection known as mastiffs, which is the major cause of loss in dairy farming. The image segmentation procedure is devised by using the lab color space and the watershed transform. A large number of samples under variable preparation conditions are treated with the proposed method. A comparison between manual and the proposed technique is presented. Promising results indicates that video-microscopy systems may be employed to develop automated SCC for goat milk.