Silver nanoparticles are lethal to the ciliate model Tetrahymena and safe to the pike silverside Chirostoma estor.

Ciliate ectoparasites are one of the most important groups of pathogens in fish culture, and the traditional treatments are sometimes harmful to the fish and the environment. Thus, the search for novel compounds that are effective at low concentrations and safe for fish are necessary to optimise treatments in aquaculture. The antiprotozoal capacity of silver nanoparticles (AgNPs) against the ciliate Tetrahymena has been documented; however, their toxicity may vary with the synthesis methodology and nanoparticle size. The objectives of this study were a) to evaluate the acute toxicity in vitro of two AgNPs (Argovit™ and UTSA) on Tetrahymena sp., a biological model for ciliated ectoparasites of fish and b) to test the safety of lethal and higher doses of UTSA AgNPs for ciliates on the fish C. estor. Light microscopy and scanning electron microscopy (SEM) were used to determine whether AgNPs affected the structure of the cell surface of Tetrahymena. The mortality, histopathological alterations and metagenomics of the fish were used to determine the major effects of UTSA AgNPs. In Tetrahymena, the median lethal concentration (LC50) for Argovit™ was 2501 ± 1717 ng/L at 15 min and 796 ± 510 ng/L at 60 min, while the LC50 for UTSA AgNPs was 4 ± 2 and 1 ± 0.6 ng/L at 15 min and 60 min, respectively. A concentration of 3300 ng/L Argovit™ and 10.6 ng/L UTSA AgNPs for 15 and 60 min, respectively, was 100% effective against Tetrahymena. After 60 min of exposure to 0.25 and 0.50 ng/L UTSA AgNPs, the number of cilia significantly reduced, there were small holes on the cell surface, and the cellular membrane was ruptured. In fish exposed to lethal (10.6 ng/L) and higher (31.8 and 95.4 ng/L) doses of UTSA, the AgNPs did not affect fish survival after 96 h, and there were no signs of histopathological damage or gut microbial changes. This study is the first report on microscopic and ultrastructural changes in Tetrahymena after exposure to significantly low concentrations of UTSA AgNPs with antiprotozoal efficacy without evidence of harmful effects on fish. These results provide the basis for further studies of both pet aquarium and commercial fish that may validate these findings at a larger experimental scale, taking into account AgNPs bioaccumulation, safety for human consumption and environmental impact.

Bacterial symbionts of Bathymodiolus mussels and Escarpia tubeworms from Chapopote, an asphalt seep in the Southern Gulf of Mexico.

Chemosynthetic life was recently discovered at Chapopote, an asphalt hydrocarbon seep in the southern Gulf of Mexico. Preliminary morphological analyses indicated that one tubeworm and two mussel species colonize Chapopote. Our molecular analyses identified the tubeworm as Escarpia sp., and the mussels as Bathymodiolus heckerae and B. brooksi. Comparative 16S rRNA analysis and FISH showed that all three species harbour intracellular sulfur-oxidizing symbionts highly similar or identical to those found in the same host species from northern Gulf of Mexico (nGoM). The mussels also harbour methane-oxidizing symbionts, and these shared highly similar to identical 16S rRNA sequences to their nGoM conspecifics. We discovered a novel symbiont in B. heckerae, which is closely related to hydrocarbon-degrading bacteria of the genus Cycloclasticus. In B. heckerae, we found key genes for the use of aromatic compounds, and its stable carbon isotope values were consistently higher than B. brooksi, indicating that the novel symbiont might use isotopically heavy aromatic hydrocarbons from the asphalt seep. This discovery is particularly intriguing because until now only methane and reduced sulfur compounds have been shown to power cold-seep chemosynthetic symbioses. The abundant hydrocarbons available at Chapopote would provide these mussel symbioses with a rich source of nutrition.

Species-specific primers for the identification of the ectomycorrhizal fungus Tuber macrosporum Vittad.

Limitations in the use of morphological traits to identify ectomycorrhizae have led to the development of species-specific molecular markers. Herein, we report a PCR-based technique for the reliable molecular identification of the ectomycorrhizal fungus Tuber macrosporum Vittad. Species-specific primers were designed from an alignment of internal transcribed spacer rDNA sequences from Tuber spp. and from the most common ectomycorrhizal contaminants found in the root systems of truffle-infected plants. The primers were tested for selective amplification using both different truffles and different ectomycorrhizae and were found to identify T. macrosporum successfully. The application of the primers in certifying the quality of truffle-inoculated seedlings is discussed.

Prediction of gestational age by ultrasonic fetometry in llamas (Lama glama) and alpacas (Lama pacos).

Fetal biparietal diameter (BPD) and thorax height (TH) were measured by ultrasound during intrauterine growth in pregnant llamas (Lama glama) and alpacas (Lama pacos). The goal was to establish representative curves that allows estimation of gestational age (GA) from real-time ultrasonic measurements of these fetal structures at any stage of gestation. Llamas and alpacas were mated under controlled conditions. Ultrasound exams were conducted to determine pregnancy status 1 month later. Measurements of fetal BPD and TH were conducted from the second month of pregnancy until term. Observation and assessment of fetal TH was difficult during the last 3 months of pregnancy, specially in llamas. Regression curves were calculated from the data as a function of GA, with the best fit represented by the following equations: llama GA=(BPD-0.002399)43.02293,r=0.98,P<0.001; llama GA=(TH-0.07137)46.94485, r=0.95,P<0.001; alpaca GA=(BPD-0.11376)47.23287, r=0.98,P<0.001; alpaca GA=(TH-0.36436)52.87663, r=0.96,P<0.001, where GA was measured in days and BPD and TH in centimeters. Results indicate that ultrasonic measurement of these fetal biometric variables constitute a valuable tool to estimate GA at any stage of pregnancy in these domestic South American camelids.

Plasma progesterone in alpaca (Lama pacos) during pregnancy, parturition and early postpartum.

Plasma progesterone concentration during pregnancy and the early postpartum period was measured by radioimmunoassay in five alpacas (Lama pacos), of the Huacaya breed, whose pregnancy length had a mean of 344.8 +/- 4.4 days. Concentration of progesterone increased (P < 0.001) from low premating values (0.11 +/- 0.06 nmol/l) to greater values at 30 days of pregnancy (8.05 +/- 1.13 nmol/l) and remained high up to 2 months of pregnancy. A slight transitory decline was observed between 3 to 7 months. Concentration of plasma progesterone dropped markedly during the 72 h before parturition especially, at the day of parturition.

Entamoeba histolytica: identification of functional Gs and Gi proteins as possible signal transduction elements in the interaction of trophozoites with fibronectin.

Trophozoites of Entamoeba histolytica adhere to several components of the extracellular matrix. Binding is mediated by specific receptors identified in the parasite surface. Interaction of trophozoites with FN induces the formation of special adhesion structures that are dynamic cytoskeleton membrane complexes and facilitate both adhesion and substrate degradation. The process requires activation of signaling pathways in which PLC, IP3, Ca2-, and PKC participate. These observations, and recent experiments showing increments in cAMP in the trophozoites during the interaction with FN, suggest that FN receptors in the amebic surface could be coupled to G-proteins. We report here that trophozoite plasma membrane peptides of 92, 49, 42, 37, and 21 kDa are ADP-ribosylated by Vibrio cholerae and Bordetella pertussis toxins. Three of them are also recognized by antibodies prepared against the alpha-subunit of Gs-and Gi-proteins. Adenylyl cyclase activity detected in isolated membranes was strongly stimulated by treatment with the toxins. Forskolin (an agonist of the enzyme) and FN also induced increments in the enzymatic activity. Live amebas incubated with the toxins showed enhanced adhesion to FN substrates and a striking reorganization of polymerized actin. The actin rearrangement is reminiscent of the one induced by either forskolin or dibutyril cyclic AMP treatment. Our present data show the presence and the functionality of Gs- and Gi-like proteins and their apparent activation during in vitro interaction of amebas with FN and complement previous observations indicating the operation of signal transduction mechanisms in E. histolytica.

Early pregnancy diagnosis in alpaca (Lama pacos) and llama (Lama glama) by ultrasound.

An ultrasonography study of early pregnancy diagnosis was carried out in 19 alpacas and 12 llamas, after controlled matings. The aim was to determine the earliest gestational age at which pregnancy diagnosis by transrectal ultrasonography could be achieved, and to generate an empirical formula for gestational sac diameter (GSD) growth as a function of gestational age (GA), allowing an estimate of GA during the first month of pregnancy. We found that pregnancy diagnosis may be carried out as early as 9 days after mating in alpacas and 7 days in llamas. This diagnosis was found to be accurate at 23 days in alpacas and 34 days in llamas. The empirical relations that best describe the relationship between GSD and GA were GA = logGSD + 1.2339/0.0585 r = 0.85; P < 0.001 in alpacas, and GA = logGSD + 1.2649/0.0546 r = 0.77, P < 0.001 in llamas, where GA is measured in days and GSD in centimeters. Our results also indicate that ultrasonography is a reliable technique for early pregnancy diagnosis. Furthermore, the empirical formulae reliably make it possible to estimate GA from GSD during the first month of pregnancy and their use might improve the efficiency of camelid breeders.

A comparative study of salivary secretion by parotid and mandibular glands of anaesthetized Capra hircus: effect of pilocarpine.

A study was made of basal secretion and the effect of the infusion of pilocarpine on the flow and composition of saliva in the parotid and mandibular glands of the anaesthetized lactating goat. In the parotid gland there was a basal flow (1.6 +/- 0.29 microliter/min) which was not present in the mandibular gland. There is a statistically significant dose-effect relationship between pilocarpine and salivary flow in both glands. Salival composition and its variation with respect to the flow of saliva did not conform to either of the two glands to an exclusive monogastric or ruminant model.

Biliary secretion in the conscious preruminant goat: use of a re-entrant cannula.

A double re-entrant bile cannula is described for use in conscious goats which permits study of bile flow and composition.

Estimulacion sensorial programada en lactantes de bajo nivel socioeconomico. / Programed sensorial stimulation in infants of low socio-economical back-ground

Se estudiaron dos grupos de ninos sanos de bajo nivel socio-economico, beneficiarios del Servicio de Salud de Valdivia, desde su nacimiento hasta el primer ano de vida con el fin de probar la eficacia de la aplicacion de un programa de estimulacion temprana. Para ello un grupo permanecio en sus condiciones ambientales habituales y el otro recibio el programa de estimulacion. Al comparar los coeficientes de desarrollo de ambos grupos aplicando un analisis de regresion minimo cuadratica se encontraron diferencias significativas entre los valores de ambos grupos siendo mayores los indices de coeficiente de desarrollo en el grupo de estudio, sin embargo el incremento del coeficiente de desarrollo no fue significativamente distinto entre los dos grupos. Con respecto a variables de crecimiento como peso, talla, perimetro craneano no hubo diferencias en el incremento en los ninos estudiados

Identification of seven isolants of infectious bronchitis virus by interference with the B-1 isolant of Newcastle disease virus.

Seven isolants of infectious bronchitis virus (IBV) interfered with the development of hemagglutinins (HA) by the B-1 isolant of Newcaslte disease virus (NDV) in eggs previously inoculated with these IBV isolants. The I50 [defined as the minimum amount of IBV needed to produce, upon inoculation in embryonating chicken eggs (ECE), HA interference in 50% of these ECE reioculated 10 hr later with 10(2,41 EID50 (HA) of the B-1 strain of NDV] was less than 1 EID50. The interference was considered specific for IBV because if could be eliminated by addition of homologous anti-IBV serum. Furthermore, three other common poultry viruses (laryngotracheitis, fowlpox, and CELO viruses) did not interfere with NDV. In view of its sensitivity, specificity, and simplicity of performance, the IBV-NDV interference test appears to have considerable potential as a diagnostic test for IBV under field conditions.

Two new isolants of infectious bronchitis virus with polyvalent immunogenicity.

The immunizing capacity of two new field isolants of avian infectious bronchitis virus (IBV) was tested in vivo. Chickens vaccinated with either isolant resisted challenge with 6 serotypes of IBV, including some of the types most commonly isolated from chickens. The two new isolants, identified herein as California "G" and California "S," appear to be immunologically different from each other and from the other 6 types tested. The polyvalent immunogenicity of both G and S is noteworthy, and potentially a very promising characteristic for the development of effective vaccines. The immunogenicity is slightly better for G than for S, but both are still pathogenic for chickens. Therefore, they should be attenuated before being used as vaccines.

Identification of infectious bronchitis virus by interference with the B-1 isolant of Newcastle disease virus. Waxing and waning of interference.

The Massachusetts and the Connecticut types of infectious bronchitis virus (IBV) were identified by interference in embryonating chicken eggs (ECE) with the production of hemagglutinin by the B-1 isolant of Newcastle disease virus (NDV). This interference test appears to be specific because the above interference was eliminated by adding type-specific anti-IBV serum to the IBV-NDV system; however, interference was not detectable when fowlpox virus (FPV) and infectious laryngotracheitis virus (LTV) were substituted for IBV. Specificity of the interference test was dependent upon a system involving IBV, NDV, FPV, and LTV. The test can be done in 3 days and requires minimum laboratory facilities. Most of the experiments were done with the Massachusetts type of IBV. Only a few were with the Connecticut type. The interfering action of the above two types of IBV over the B-1 isolant of NDV waxed between the 24th and 54th hr after inoculation of NDV; it was waning at the 54th-60th hr postinoculation and was undetectable by the 66th-72nd hr.