Lack of negative effects of the biological control agent Duddingtonia flagrans on soil nematodes and other nematophagous fungi.

The possible environmental effects of the massive use of Duddingtonia flagrans for controlling sheep nematodes were evaluated in two regions. Non-supplemented faeces and faeces from sheep supplemented with D. flagrans were deposited three times on pasture plots and samples were collected 7 and 14 days post-deposition. Samples were cultured in agar-water (2%) with Panagrellus spp. to recover D. flagrans and other nematophagous fungi, and soil nematodes were extracted using Baermann funnels and counted. No significant differences in the populations of soil nematodes and fungi colonizing sheep faeces (P > 0.05) were observed between supplemented and non-supplemented groups, except in one sample. The topsoil in contact with the faeces was sampled 1-4 months post-deposition, revealing that, with one exception, D. flagrans did not persist in soil beyond 2 months post-deposition. Duddingtonia flagrans does not affect faecal colonization by other fungi and soil nematodes and, once deployed on pasture, does not survive for long periods in the environment.

Large deep-sea zooplankton biomass mirrors primary production in the global ocean.

The biological pump transports organic carbon produced by photosynthesis to the meso- and bathypelagic zones, the latter removing carbon from exchanging with the atmosphere over centennial time scales. Organisms living in both zones are supported by a passive flux of particles, and carbon transported to the deep-sea through vertical zooplankton migrations. Here we report globally-coherent positive relationships between zooplankton biomass in the epi-, meso-, and bathypelagic layers and average net primary production (NPP). We do so based on a global assessment of available deep-sea zooplankton biomass data and large-scale estimates of average NPP. The relationships obtained imply that increased NPP leads to enhanced transference of organic carbon to the deep ocean. Estimated remineralization from respiration rates by deep-sea zooplankton requires a minimum supply of 0.44 Pg C y-1 transported into the bathypelagic ocean, comparable to the passive carbon sequestration. We suggest that the global coupling between NPP and bathypelagic zooplankton biomass must be also supported by an active transport mechanism associated to vertical zooplankton migration.

Efficacy of some anthelmintics on an ivermectin-resistant strain of Haemonchus contortus in sheep.

Following evidence of reduced efficacy of ivermectin in a field population of Haemonchus contortus in Brazil, this strain of the parasite was submitted to a controlled anthelmintic test. Eighty worm-free lambs were randomly split into two groups of 40. Each lamb in the first group was infected with 5000 third stage larvae (L3) of the ivermectin-resistant strain; the remaining 40 lambs were each infected with 5000 L3 of a H. contortus strain of known susceptibility to the major groups of anthelmintic compounds used in sheep. On Day 28 post-infection, each group was subdivided according to egg counts and at random into four sub-groups of ten lambs, each of which was treated with albendazole (ABZ) at 3.8 mg kg-1, levamisole (LEV) at 7.5 mg kg-1 or ivermectin (IVM) at 0.2 mg kg-1, or was left as untreated control. At slaughter, 7 days later, all the anthelmintics reduced worm burdens in animals infected with the susceptible strain (ABZ 98.9%, LEV and IVM 100%). By contrast, in the lambs infected with the ivermectin-resistant strain, LEV was 99.8% effective, but ABZ reduced worm counts by only 14.7% and IVM by only 10.4%. Interestingly at necropsy on Day 7 post-treatment, the egg counts in the resistant strain treated with ABZ had been reduced by 92.5%, although worm counts were reduced by only 14.7%.(ABSTRACT TRUNCATED AT 250 WORDS)

Isoenzyme analysis of Haemonchus contortus resistant or susceptible to ivermectin.

Three different strains of Haemonchus contortus (susceptible to ivermectin, S-IVM; selected for resistance to ivermectin, R-IVM; a multiple resistant strain, i.e. resistant to benzimidazole and ivermectin, R-IVM/SA) were examined for isoenzyme variation by starch gel electrophoresis. Using stains for seven enzymes separated in five different buffer systems, no differences in the electrophoretic mobility could be detected between any of the strains. Results demonstrate a low level of enzyme variation in H. contortus and no differences in enzyme electrophoretic profile between IVM-sensitive and IVM-resistant parasites. Differences between the ivermectin-sensitive and both ivermectin-resistant strains were observed with the propionyl esterases and although some of the differences are probably associated with benzimidazole resistance, others are associated with resistance to ivermectin. The three strains of H. contortus are generally identical; however, differences between all strains of H. contortus and a strain of Dictyocaulus viviparus were detected.

Model predictions for anthelmintic resistance amongst Haemonchus contortus populations in southern Brazil.

A computer model developed to study Ostertagia circumcincta resistance to anthelmintics in UK sheep flocks has been adapted for use with Haemonchus contortus under southern Brazilian conditions. The model simulates the effect of different anthelmintic control regimens on the year-to-year pattern of resistance in breeding ewes. The nematode control regimen most used by Brazilian sheep farmers was found to increase the frequency of genes which confer resistance from approximately 3% to 14% in an H. contortus population over a 20 year period. The effect of early versus late season anthelmintic treatment was investigated. This indicated that early season treatment would select for resistance rapidly, whereas late season treatments would not, owing to large numbers of untreated parasites accumulating at the beginning of the season. A model which can predict the development of anthelmintic resistance in parasites of ewes is a valuable tool in the understanding of the effect of different strategies on nematode control programmes and merits further consideration.

Use of reseeded pastures as an aid in the control of gastrointestinal nematodes.

The degree of nematode infectivity of a previously contaminated pasture which was reseeded after harvesting of a soya bean crop was determined by use of four tracer lambs and four tracer calves. The eight tracers were put to graze for 14 days and housed for another 14 days before being slaughtered, when faeces and the intestinal tracts were examined for worm eggs and parasites. With the exception of one animal, which was found to be harbouring 20 Cooperia punctata, the tracers had no adult or immature nematodes. The implications of the use of such pastures are discussed in relation to their possible role in parasite control and anthelmintic resistance in Brazil.

Laboratory selection for ivermectin resistance in Haemonchus contortus.

A strain of Haemonchus contortus, susceptible to ivermectin (IVM), was submitted to a selection procedure using IVM at 0.02 mg kg-1 during nine successive passages in lambs. Treatment reduced worm egg counts (epg) by more than 99% up to the seventh passage, but thereafter there was no significant reduction in epg. The efficacy of ivermectin against the ninth selected generation compared with the original susceptible strain was assessed. The results showed that animals infected with the selected strain and treated with IVM did not show any significant reduction (P > 0.05) in epg compared with a 99.9% reduction in the susceptible group. At necropsy, post-treatment worm burdens confirmed that the selection pressure applied to the original strain had been successful in stimulating resistance to IVM.

Experiments on anthelmintic control of Fasciola hepatica in Brazil.

Two separate field trials involving naturally infected cattle were carried out on two farms known to have a history of Fasciola hepatica infection. On the first farm, 15 animals per group were allocated as follows: G1, triclabendazole (TCBZ) four times a year; G2, TCBZ twice a year (May and September); G3, untreated control. All groups grazed together and after 3.5 years the animals were slaughtered and their livers examined by federal meat inspectors who condemned 100% of livers in G3 and 8.3% in G2 owing to the presence of lesions of fasciolosis. In G1 no livers were condemned. Significant differences in weight gains were not detected and fluke counts remained at low levels in the treated groups. Also, in the control group, egg counts started to decrease when animals were 2 years old. On the second farm, groups of 20 animals were treated as follows: G1, TCBZ three times a year (May, September and December); G2, TCBZ twice a year (May and September); G3, nitroxynil twice a year (May and September); G4, rafoxanide twice a year (May and September); G5, untreated controls. All animals were weighed and faecal samples examined at approximately 28-day intervals. During the period of the study, larger weight gains were detected in the TCBZ treated groups than in the others. TCBZ treatment kept F. hepatica egg counts at a lower level for longer periods than the other drugs and significant differences in weight gains were only obtained between the group receiving TCBZ three times a year and the control group.

The prevalence of anthelmintic resistance in nematode parasites of sheep in southern Latin America: general overview.

South America has a large population of small ruminants. Currents estimates are approximately 100 million sheep and 23 million goats. A large percentage of these flocks are raised in the humid tropics/sub-tropics. Nematode parasitism is singly the most important disease of these animals and typically farmers resort to frequent anthelmintic treatment in attempts to control this problem. Because of this reliance on drugs, price consideration is an important determinant in a farmer's choice of anthelmintic. In some circumstances, this has led to unethical practices of drug adulteration and substitution, or the offering to farmers of cheap, sub-standard generic products. The prevalence of anthelmintic resistance has not been investigated in any widespread sense in South America, although some of the first reports of resistance were made in southern Brazil almost 30 years ago. The following series of papers outline surveys conducted in the humid topics/sub-tropics of southern Latin America to assess the significance of resistance to the broad-spectrum anthelmintic groups in nematode parasites of sheep flocks.

The prevalence of anthelmintic resistance in nematode parasites of sheep in southern Latin America: Brazil.

This survey was conducted in the southern Brazilian state of Rio Grande do Sul and involved 182 farms located in 26 counties. In addition to the three major broad-spectrum anthelmintic groups (viz. benzimidazole, levamisole and ivermectin) the combination benzimidazole and levamisole and the H. contortus specific anthelmintic, closantel, were tested by the faecal egg count reduction method for the prevalence of anthelmintic resistance. Resistance was found to be 90%, 84%, 13%, 73% and 20%, respectively. This is a crisis situation. Immediate, drastic action needs to be implemented, otherwise the sheep industry in this region (approx. 10 million head) will soon face a lack of any effective anthelmintics with the inevitable consequences of major restructuring or abandonment.

Response to ivermectin treatment of parasitic stages of Haemonchus contortus resistant or susceptible to ivermectin.

Two groups of 33 helminth-naive lambs were infected with 5,000 L3 of an ivermectin-resistant or -susceptible strain of Haemonchus contortus (groups R and S). On days 6, 10, 16, and 21 postinfection, 5 animals from each group were chosen at random and orally treated with 0.2 mg/kg of ivermectin. On each occasion, 2 randomly selected lambs from each group were also killed to determine the number and stage of development of the worms present at the time of treatment. These necropsies revealed that by day 6 early and late fourth-stage larvae were present, whereas on day 10 the early fifth stage had been reached; by days 16 and 21 all worms had reached the adult stage. Necropsies on day 28 postinfection revealed that although animals treated at day 6 had 26.3% fewer worms than the controls, there was no significant difference (P greater than 0.05) between worm burdens from any of the animals infected with the R strain and treated at different times after infection when compared with the untreated controls. With ivermectin significant reductions were obtained in the worm burdens of the animals infected with the susceptible strain; these were reduced by 96% when treatment was given on day 6 against fourth-stage larvae and 98.9% when the drug was given on day 21 against adult stages. From these results it is clear that resistance to ivermectin in this strain of H. contortus is present as early as the fourth larval stage.

Survival and development of ivermectin-resistant or susceptible strains of Haemonchus contortus under field and laboratory conditions.

The free-living development of three strains of Haemonchus contortus was studied in two experiments. Day 21 faecal samples were collected from lambs infected with either a susceptible strain, a laboratory-selected ivermectin (IVM) resistant strain or a South African field strain showing multiple anthelmintic resistance, which included IVM. No eggs hatched in samples cultured at 4 or 10 degrees C. At 22 degrees C the laboratory-selected strain showed the highest rate of development while at 27 degrees C the susceptible strain produced the highest yield of third stage larvae (L3): at both temperatures the field strain showed the lowest percentage development to L3. The second experiment was a field study carried out in southern Brazil. Faeces containing either an IVM-susceptible or an IVM-resistant strain of H contortus were placed in two series of grass plots during each of three summer months. Soil subsequently yielded more larvae than did grass suggesting migration or mechanical transport into the soil. For plots contaminated during the first two months there was no significant difference in recovery rate between the two strains (P > 0.05). When contamination occurred during the third month, the IVM-resistant strain produced significantly higher recovery rates (P < 0.05) from both pasture and soil.

Response of Corriedale and Crioula Lanada sheep to artificial primary infection with Haemonchus contortus.

Clinical, parasitological and biochemical parameters were evaluated in Corriedale and Crioula Lanada sheep after a single experimental infection with Haemonchus contortus. Ten 4-month-old worm-free lambs, of each breed, were infected with 200 L3 H. contortus per kg live weight and four uninfected animals of each breed were used as controls. Every week, the animals were weighed and blood and faecal samples were collected for measurement of packed cell volume (PCV), total serum protein (TSP) and albumin (ALB), and the number of eggs per gram of faeces (EPG), respectively. Twelve weeks after infection, the animals were slaughtered. The worm burden was determined and samples of the abomasal mucosa were processed for determination of the number of eosinophils, mast cells and globule leukocytes. No significant differences in PCV, TSP, ALB, parasite burden or the cell populations of the abomasal mucosa were observed between breeds, but Crioula lambs had a lower EPG count. The comparison of the infected groups with their respective controls revealed significant alterations in PCV, TSP and ALB in the Corriedale lambs and in PCV, TSP, ALB and the density of eosinophils and mast cells in the Crioula lambs.

[The neurobiology of sleep: Cajal and present-day neuroscience]. / Neurobiología del sueño: Ramón y Cajal y la neurociencia actual.

AIMS: We briefly describe the most significant findings obtained recently concerning the sleep-waking cycle in comparison to the studies conducted by Cajal on the same subject. DEVELOPMENT: This paper includes a short biographical sketch of Santiago Ramón y Cajal, with special emphasis on his importance within the framework of neuroscience. Cajal represents the decisive turning point in neurobiological studies, with the discovery of the synapse and his law of dynamic polarization. We conduct a short survey of the current knowledge about the phases of sleep and oneiric phenomena, based on their anatomo-physiological foundations. We present a summary of the history of the subject and analyze the contributions made by Cajal to this field, i.e. his study of the associative cortices, which are essential in memory processes and related to the mechanisms governing the sleep-waking cycle. For Cajal the fine anatomy of the thalamus must be considered in relation to the specificity of its connections an idea that is still completely valid today. He did not observe any projections of the thalamic reticular nucleus towards the cerebral cortex, a fact that has been corroborated using modern-day techniques. He spoke of the involvement of neuroglia in the attentional and sleep processes, which is so, although not quite in the way Cajal thought. He considered the production of dreams to be based on intimate neural mechanisms, which is still so. He also studied other brain structures related with the regulation of the sleep waking cycle, although avoiding any specific mention of the mechanisms controlling such a cycle. Furthermore, he conducted self-observation studies with a high degree of insight. CONCLUSIONS: Cajal studied the phenomena of attention and sleep in an objective manner and contributed a number of significant interpretations, some of which are now somewhat outdated while others are still wholly valid today.

The pattern of faecal egg output in lambs infected with a multiple resistant strain of Haemonchus contortus after treatment with albendazole.

Worm-free lambs (n = 6) were infected with 5,000 L3 of an ivermectin/benzimidazole/closantel resistant strain of Haemonchus contortus. On Day 21 post infection animals were allocated, according to worm egg counts, to 2 groups of 3 animals each and on Day 29 one group was treated with albendazole at 3.8 mg kg-1, while animals in the other group remained untreated as controls. All animals were slaughtered on Day 44 when the treated group was found to have 2,083 +/- 1,718 worms and the controls 2,783 +/- 633, a reduction of 25.2% in worm burdens. The pattern of egg output was monitored and it was found that 2 d after dosing, worm egg counts had been suppressed by 89% in the treated group; thereafter the egg counts increased, but never returned to the pretreatment levels and by Day 15 post treatment, they were still 41.1% lower in the treated group than in the controls.

Large mesopelagic fishes biomass and trophic efficiency in the open ocean.

With a current estimate of ~1,000 million tons, mesopelagic fishes likely dominate the world total fishes biomass. However, recent acoustic observations show that mesopelagic fishes biomass could be significantly larger than the current estimate. Here we combine modelling and a sensitivity analysis of the acoustic observations from the Malaspina 2010 Circumnavigation Expedition to show that the previous estimate needs to be revised to at least one order of magnitude higher. We show that there is a close relationship between the open ocean fishes biomass and primary production, and that the energy transfer efficiency from phytoplankton to mesopelagic fishes in the open ocean is higher than what is typically assumed. Our results indicate that the role of mesopelagic fishes in oceanic ecosystems and global ocean biogeochemical cycles needs to be revised as they may be respiring ~10% of the primary production in deep waters.

Eight-channel ultrasonic device for non-invasive quality evaluation in packed milk.

An eight-channel ultrasonic detecting device for microbiological quality evaluation of packed liquid foods is presented in this paper. This device makes possible a non-invasive detection of the microbial growth in liquid foods with no need to open the carton-based packages where they are contained. Thermal and humidity stabilization are required inside the measuring chamber. The changes in the liquid media produced by the microorganism growth induce variations in the ultrasonic propagation parameters giving a non-invasive evidence of the developing contamination. For that purpose, the amplitude and time of flight of an ultrasonic 800kHz tone burst travelling through an UHT milk pack are analysed. Inoculated and sterile packs were tested to evaluate the performance of this new non-invasive ultrasonic microbiological quality sensor.

Movement of zymosterol, a precursor of cholesterol, among three membranes in human fibroblasts.

Where examined, cholesterol is synthesized in the endoplasmic reticulum; however, its precursor, zymosterol, is found mostly in the plasma membrane. The novel implication of these disparate findings is that zymosterol circulates within the cell. In tracing its movements, we have now established the following: (a) in human fibroblasts, zymosterol is converted to cholesterol solely in the rough ER. (b) Little or no zymosterol or cholesterol accumulates in the rough ER in vivo. (c) Newly synthesized zymosterol moves to the plasma membrane without a detectable lag and with a half-time of 9 min, about twice as fast as cholesterol. (d) The pool of radiolabeled zymosterol in the plasma membrane turns over rapidly, faster than does intracellular cholesterol. Thus, plasma membrane zymosterol is not stagnant. (e) [3H]Zymosterol pulsed into intact cells is initially found in the plasma membrane. It is rapidly internalized and is then converted to [3H] cholesterol. Half of the [3H]cholesterol produced returns to the plasma membrane within 30 min of the initial [3H]zymosterol pulse. (f) Nascent zymosterol accumulates in a buoyant sterol-rich intracellular membrane before it reaches the plasma membrane. This membrane also acquires nascent cholesterol, exogenous [3H]zymosterol pulsed into intact cells, and [3H]cholesterol synthesized from the exogenous [3H] zymosterol. These results suggest that at least one sterol moves rapidly and in both directions among the rough endoplasmic reticulum, a sterol-rich intracellular membrane bearing nascent cholesterol, and the plasma membrane.