Dilated cardiomyopathy in cats: survey of veterinary cardiologists and retrospective evaluation of a possible association with diet.

INTRODUCTION/OBJECTIVES: The objectives were to conduct a survey of cardiologists on their recent experiences with cats that have dilated cardiomyopathy (DCM) and to retrospectively review individual cases of feline DCM. ANIMALS, MATERIALS AND METHODS: Part one: A survey was distributed to cardiologists with questions regarding caseload and clinical management of cats with DCM diagnosed over the past two years. Part two: Cardiologists completing the survey were invited to submit data from cats recently diagnosed with DCM. Data on signalment, clinical signs, diet, echocardiographic measurements and outcome were recorded. RESULTS: Part one: From 52 completed surveys, many cardiologists responded that measuring and supplementing taurine and recommending a diet change in cats with DCM are common practices. Few (15%) cardiologists reported an increase in the number of feline DCM cases over the past two years, although some had cases that improved even if taurine deficiency was not present. Part two: Twenty of 37 (54%) cats ate low pea/lentil (low PL) diets, and 14/37 (38%) ate high PL diets at the time of diagnosis; three had incomplete diet information. Two of 13 cats (15%) in which taurine was measured had levels below the reference range. After adjusting for other variables, cats eating high PL diets that changed diets after diagnosis had a significantly longer survival time than that of cats eating high PL diets that did not change diets after diagnosis (P = 0.025). CONCLUSIONS: Additional research is warranted to determine whether there could be a possible association between diet and DCM in cats.

Multisession Anodal tDCS Protocol Improves Motor System Function in an Aging Population.

OBJECTIVES: The primary objective of this study was to investigate the effects of five consecutive, daily 20-minute sessions of M1 a-tDCS on motor learning in healthy, cognitively intact, aging adults. DESIGN: A total of 23 participants (51 to 69 years old) performed five consecutive, daily 20-minute sessions of a serial reaction time task (SRT task) concomitant with either anodal (n = 12) or sham (n = 11) M1 a-tDCS. RESULTS: We found a significant group × training sessions interaction, indicating that whereas aging adults in the sham group exhibited little-to-no sequence-specific learning improvements beyond the first day of training, reproducible improvements in the ability to learn new motor sequences over 5 consecutive sessions were the net result in age-equivalent participants from the M1 a-tDCS group. A significant main effect of group on sequence-specific learning revealed greater motor learning for the M1 a-tDCS group when the five learning sessions were averaged. CONCLUSION: These findings raise into prominence the utility of multisession anodal TDCS protocols in combination with motor training to help prevent/alleviate age-associated motor function decline.

Quebec Integrated Chronic Disease Surveillance System (QICDSS), an innovative approach.

INTRODUCTION: With the growing burden of chronic diseases, surveillance will play an essential role in improving their prevention and control. The Institut national de santé publique du Québec has developed an innovative chronic disease surveillance system, the Quebec Integrated Chronic Disease Surveillance System (QICDSS). We discuss the primary features, strengths and limitations of this system in this report. METHODS: The QICDSS was created by linking five health administrative databases. Updated annually, it currently covers the period from January 1, 1996, to March 31, 2012. The operational model comprises three steps: (1) extraction and linkage of health administrative data according to specific selection criteria; (2) analysis (validation of case definitions essentially) and production of surveillance measures; and (3) data interpretation, submission and dissemination of information. The QICDSS allows the surveillance of the following chronic diseases: diabetes, cardiovascular diseases, respiratory diseases, osteoporosis, osteoarticular diseases, mental disorders, Alzheimer's disease and related disorders. The system also lends itself to the analysis of multimorbidity and polypharmacy. RESULTS: For 2011-2012, the QICDSS contained information on 7 995 963 Quebecers with an average age of 40.8 years. Of these, 95.3% met at least one selection criterion allowing the application of case definitions for chronic disease surveillance. The actual proportion varied with age, from 90.1% for those aged 19 years or less to 99.3% for those aged 65 years or over. CONCLUSION: The QICDSS provides a way of producing population-based data on the chronic disease burden, health services and prescription drug uses. The system facilitates the integrated study of several diseases in combination, an approach rarely implemented until now in the context of population surveillance. The QICDSS possesses all the essential features of a surveillance system and supports the dissemination of information to public health decision-makers for future actions.

TITRE: Le Système intégré de surveillance des maladies chroniques du Québec (SISMACQ), une approche novatrice. INTRODUCTION: Avec l'accroissement du fardeau des maladies chroniques, la surveillance est fondamentale pour améliorer la prévention et la prise en charge de ces dernières. L'Institut national de santé publique du Québec a donc développé un système novateur de surveillance des maladies chroniques, le Système intégré de surveillance des maladies chroniques du Québec (SISMACQ), dont les principales caractéristiques, les forces et les limites sont présentées ici. MÉTHODOLOGIE: Le SISMACQ est le résultat du jumelage de cinq fichiers médicoadministratifs. Mises à jour annuellement, ses données couvrent actuellement la période du 1er janvier 1996 au 31 mars 2012. Trois étapes en caractérisent le modèle opérationnel : 1) l'extraction et le jumelage des données médico-administratives grâce à divers critères de sélection; 2) les analyses (principalement la validation des définitions) et la production des mesures de surveillance et 3) l'interprétation, le dépôt et la diffusion de l'information. Le SISMACQ permet actuellement l'étude des maladies chroniques suivantes : diabète, maladies cardiovasculaires, maladies respiratoires, ostéoporose, maladies ostéoarticulaires, troubles mentaux et Alzheimer et maladies apparentées. Il permet également l'analyse de la multimorbidité et de la polypharmacie. RÉSULTATS: Pour 2011-2012, le SISMACQ contenait des données sur 7 995 963 Québécois, et leur moyenne d'âge était de 40,8 ans. Parmi eux, 95,3 % répondaient à au moins un critère de sélection permettant l'application de définitions de cas pour la surveillance des maladies chroniques. Cette proportion variait avec l'âge : de 90,1 % chez les Québécois de 19 ans et moins à 99,3 % chez ceux de 65 ans et plus. CONCLUSION: Le SISMACQ permet la production de données, à l'échelle de la population, sur le fardeau de plusieurs maladies chroniques, sur l'utilisation des services de santé et sur la consommation de médicaments. Il rend possible l'étude intégrée de la combinaison de plusieurs maladies, une approche jusqu'à présent rarement mise en oeuvre dans un contexte de surveillance populationnelle. Le SISMACQ répond aux attributs essentiels d'un système de surveillance et aide à la diffusion de l'information auprès des décideurs en vue d'actions en santé publique.

Agreement between survey data and Régie de l'assurance maladie du Québec (RAMQ) data with respect to the diagnosis of asthma and medical services use for asthma in children.

INTRODUCTION: The goal of this study was to assess the agreement between the results of a respiratory health survey conducted in Montréal on children aged 6 months to 12 years and the Régie de l'assurance maladie du Québec (RAMQ, Quebec health insurance board) database in terms of the diagnosis of asthma and medical services use. A secondary aim was to evaluate the effect of the survey method used (Internet-based survey or telephone survey). METHODS: We assessed whether a diagnosis of asthma was made for 7922 children. In addition, we compared the use of medical services for asthma (emergency department visits and hospitalizations) in the 12 months preceding the survey for the 402 children considered to have asthma, using 2 groups of respiratory diagnoses and 2 data linkage periods. The agreement between the 2 data sources was evaluated using the kappa statistic (κ) and sensitivity and specificity, as well as percentages of agreement, overreporting and under-reporting with respect to health services use. RESULTS: Moderate agreement was found between the 2 data sources (survey and RAMQ data) in terms of the diagnosis of asthma (κ = 0.54 and κ = 0.60 depending on the definition used). Specificity was high (93% and 96%), but sensitivity varied (50% and 65%). Respondents over-reported health services use, resulting in moderate kappa values (0.49 for emergency department visits and 0.48 for hospitalizations). However, when more diagnoses were included in the definition and when the linkage period was extended (15 rather than 12 months), the kappa values increased (0.59 for emergency department visits and 0.64 for hospitalizations) and sensitivity and specificity were high. Slightly higher agreement was obtained for the Internet-based survey relative to the telephone survey. CONCLUSION: The findings validate the use of survey data with respect to the diagnosis of pediatric asthma and major health services use for this disease.

TITRE: Concordance entre les résultats d'une enquête et les données de la Régie de l'assurance maladie du Québec (RAMQ) pour le diagnostic d'asthme et pour l'utilisation des services médicaux pour asthme chez les enfants. INTRODUCTION: L'objectif est d'évaluer la concordance, quant au diagnostic de l'asthme et à l'utilisation des services médicaux, entre les résultats d'une enquête réalisée à Montréal sur la santé respiratoire des enfants de 6 mois à 12 ans et la base de données de la Régie de l'assurance maladie du Québec (RAMQ). Nous avons voulu également évaluer l'effet du mode d'enquête (Internet ou téléphone). MÉTHODOLOGIE: On a mesuré l'existence d'un diagnostic d'asthme pour 7 922 enfants. On a comparé également l'utilisation des services médicaux pour asthme (visites à l'urgence et hospitalisations) dans les douze derniers mois précédant l'enquête pour les 402 enfants considérés comme asthmatiques, à l'aide de deux groupes de diagnostics respiratoires et deux périodes de couplage. La concordance entre les deux sources a été évaluée à l'aide du coefficient kappa de Cohen (κ), de la sensibilité etde la spécificité ainsi qu'enmesurant, pour l'utilisation des services, les proportions d'accord, de sur-déclaration et de sous-déclaration. RÉSULTATS: La concordance entre les deux sources (enquête et RAMQ) est modérée pour le diagnostic d'asthme (κ = 0,54 et κ = 0,60 selon la définition utilisée). La spécificité est élevée (93 % et 96 %) et la sensibilité variable (50 % et 65 %). Il y a une surdéclaration d'utilisation des services par les répondants, avec des coefficients kappa modérés (0,49 pour les visites aux urgences et 0,48 pour les hospitalisations), mais ces derniers augmentent lorsque l'on inclut davantage de diagnostics dans la définition et que l'on allonge (à 15 mois plutôt que 12) la période de couplage (0,59 pour les visites et 0,64 pour les hospitalisations). La sensibilité et la spécificité sont élevées. La concordance est légèrement plus élevée pour l'enquête par Internet que par téléphone. CONCLUSION: Ces résultats valident l'utilisation des données d'enquête concernant l'asthme pédiatrique et l'utilisation des services principaux en relation avec cette maladie.

Nontransmission of porcine circovirus 2 (PCV2) by embryo transfer.

Two experiments were conducted to determine the association of porcine circovirus type 2 (PCV2) with embryos and the risk of viral transmission by embryo transfer. In the first experiment, 240 embryos from uninfected donors were exposed to PCV2a 10(4)TCID50/mL in vitro before transfer to seronegative recipients; in the second experiment, 384 embryos recovered from infected donors, 10 days after donor inoculation with PCV2, were transferred to seronegative recipients. In total, 1120 embryos and/or ova were collected from 37 viral-free donors (experiment 1) and 1019 from 59 PCV2-infected donors (experiment 2) (P < 0.01). The washing and/or disinfection procedure recommended by the International Embryo Transfer Society was applied to embryos in both experiments. Transfer of embryos experimentally exposed in vitro to high titers of virus caused seroconversion of recipients (58%; N = 7/12) and their piglets (81%; N = 13/16). Postmortem, PCV2 DNA was detected in various organs of embryo transfer recipients and their embryo transfer-derived piglets. In contrast, the transfer of embryos recovered from infectious PCV2 donors did not result in the seroconversion of embryo recipients (N = 24) or their embryo transfer-derived piglets (N = 76). Neither PCV2 DNA nor infectious virus was detected in the tissues of either recipients or embryo transfer-derived piglets collected postmortem in the second experiment. The results obtained in this study indicate that the transmission of PCV2 from infected donors by embryo transfer is unlikely if the sanitary recommendations of the International Embryo Transfer Society are followed. In practical terms, this means that embryo transfer can be successfully used for the intentional elimination of PCV2 and to create virus-free offspring for the safe exchange of swine genetic materials.

Position paper: improving governance for effective veterinary services in developing countries--a priority for donor funding.

Livestock contributes significantly to the world economy. However, animal diseases and food safety are still major constraints on livestock-sector productivity, economic growth, the reduction of poverty and food security. Efficient and effective governance of Veterinary Services throughout the world is a fundamental requirement for addressing the global animal health and related public health threats. Recent work by the World Organisation for Animal Health (OIE) through the application of the Tool for the Evaluation of Performance of Veterinary Services (PVS Tool) and related Gap Analysis (both of which form part of the PVS Pathway) has indicated that a significant proportion of the national Veterinary Services worldwide do not meet the essential requirements for good governance. This shortcoming poses a significant risk for many developing countries and their trading partners when considered in the context of the growing trade in animal-source foods, and the burgeoning global livestock population. Well-managed, transparent and credible Veterinary Services, in both the public and private sector, are essential for mitigating animal disease risks and ensuring sustainable incomes for vulnerable producers. They are also vital for limiting the public health risks posed by zoonotic diseases. This paper is intended to highlight the impact of governance on the delivery of veterinary services in a development context and the benefits generated by improving veterinary governance. It recognises 'global public good' elements embedded in the good governance of Veterinary Services, and it could also provide an operational development investment roadmap that builds on the OIE PVS Pathway, and innovative financing options based on government commitments supported by donor programmes.

Variation of daily warm season mortality as a function of micro-urban heat islands.

BACKGROUND: Little attention has been paid to how heat-related health effects vary with the micro-urban variation of outdoor temperatures. This study explored whether people located in micro-urban heat islands are at higher risk of mortality during hot summer days. METHODS: Data used included (1) daily mortality for Montreal (Canada) for June-August 1990-2003, (2) daily mean ambient outdoor temperatures at the local international airport and (3) two thermal surface images (Landsat satellites, infrared wavelengths). A city-wide temperature versus daily mortality function was established on the basis of a case-crossover design; this function was stratified according to the surface temperature at decedents' place of death. RESULTS: The risk of death on warm summer days in areas with higher surface temperatures was greater than in areas with lower surface temperatures. CONCLUSIONS: This study suggests that measures aimed at reducing the temperature in micro-urban heat islands (eg, urban greening activities) may reduce the health impact of hot temperatures. Further studies are needed to document the variation of heat-related risks within cities and to evaluate the health benefits of measures aimed at reducing the temperature in micro-urban heat islands.

Biotic disturbance, recolonization, and early succession of bacterial assemblages in intertidal sediments.

The role of disturbance in structuring natural microbial communities has been largely unexplored. Disturbance associated with invertebrate ingestion can reduce bacterial biomass and alter metabolic activities and compositions of bacterial assemblages in marine sediments. The primary objectives of the research presented here were to test whether ingestion by a taxonomically diverse group of deposit feeders constituted a disturbance, and to determine the mechanisms by which bacterial assemblages recover following deposit-feeder ingestion. To test the question of disturbance, we compared fresh egesta vs surficial sediments with respect to bacterial assemblage structure. In emersed intertidal sediments, microbial recovery could be due to regrowth of bacterial populations surviving gut passage or to immigration from adjacent sediments. To differentiate between these modes of recolonization we used field manipulative experiments to exclude migration by isolating freshly extruded fecal coils of three deposit-feeding species from surrounding sediments. We then followed the quantitative and qualitative recovery in egesta and sediments through time using epifluorescence microscopy and PCR-DGGE analysis of 16S rDNA. Our findings indicate that (1) the degree and nature of the disturbance to bacterial assemblages from deposit feeding varies among invertebrate taxa, (2) recovery was significant but incomplete over 3 h, and (3) recolonization of biotically disturbed sediments is dominated by immigration.

Pigs expressing salivary phytase produce low-phosphorus manure.

To address the problem of manure-based environmental pollution in the pork industry, we have developed the phytase transgenic pig. The saliva of these pigs contains the enzyme phytase, which allows the pigs to digest the phosphorus in phytate, the most abundant source of phosphorus in the pig diet. Without this enzyme, phytate phosphorus passes undigested into manure to become the single most important manure pollutant of pork production. We show here that salivary phytase provides essentially complete digestion of dietary phytate phosphorus, relieves the requirement for inorganic phosphate supplements, and reduces fecal phosphorus output by up to 75%. These pigs offer a unique biological approach to the management of phosphorus nutrition and environmental pollution in the pork industry.

Soluble, template-dependent extracts from Nicotiana benthamiana plants infected with potato virus X transcribe both plus- and minus-strand RNA templates.

We have developed a method to convert membrane-bound replication complexes isolated from Nicotiana benthamiana plants infected with potato virus X (PVX) to a soluble, template-dependent system for analysis of RNA synthesis. Analysis of RNA-dependent RNA polymerase activity in the membrane-bound, endogenous template extracts indicated three major products, which corresponded to double-stranded versions of PVX genomic RNA and the two predominant subgenomic RNAs. The endogenous templates were removed from the membrane-bound complex by treatment with BAL 31 nuclease in the presence of Nonidet P-40 (NP-40). Upon the addition of full-length plus- or minus- strand PVX transcripts, the corresponding-size products were detected. Synthesis was not observed when red clover necrotic mosaic dianthovirus (RCNMV) RNA 2 templates were added, indicating template specificity for PVX transcripts. Plus-strand PVX templates lacking the 3' terminal region were not copied, suggesting that elements in the 3' region were required for initiation of RNA synthesis. Extracts that supported RNA synthesis from endogenous templates could also be solublized using sodium taurodeoxycholate and then rendered template-dependent by BAL 31 nuclease/NP-40 treatment. The solubilized preparations copied both plus- and minus-strand PVX transcripts, but did not support synthesis from RCNMV RNA 2. These membrane-bound and soluble template-dependent systems will facilitate analyses of viral and host components required for PVX RNA synthesis.

1971 Medicare amendment: reflections on the passage of the end-stage renal disease Medicare program.

In this article I reflect on events between 1968 and 1972 that influenced the enactment of the End-Stage Renal Disease (ESRD) Amendment to Public Law 92-603.

Assessment of stallion spermatozoa viability by flow cytometry and light microscope analysis.

Viability of spermatozoa can be assessed by numerous methods, but many are slow and poorly repeatable, and subjectively assess only 100 to 200 spermatozoa per ejaculate. We collected two ejaculates from each of 4 stallions, and extended them to 50x10(6) sperm/mL in a nonfat dried milk solids glucose extender (EZ Mixin). Half the ejaculate was freeze-killed by immersing in liquid nitrogen for 10 min. Aliquots using appropriate volumes of live and freeze-killed spermatozoa provided the following ratios of live:dead spermatozoa: 100:0, 75:25, 50:50, 25:75, 0:100. We determined the viability of each aliquot by 1) motility; 2) eosin-nigrosin staining; and 3) dual fluorescent staining. For the latter, aliquots incubated with SYBR-14 and propidium iodide had live and dead spermatozoa quantitated by fluorescent microscope (2 x 100 sperm/sample) and flow cytometry (10,000 sperm/sample). We found a linear relationship between the ratio of live:dead spermatozoa and the percentage of spermatozoa counted as live (P<0.0001). For fresh spermatozoa, correlation coefficients of the known live:dead ratio were high for all methods (eosin-nigrosin, r>0.75; fluorescent microscope, r>0.76; flow cytometry, r>0.75; motility, r>0.76). To determine viability of cryopreserved equine spermatozoa, we froze 17 fresh ejaculates from 6 stallions in a glycine extender. Each sample was thawed, extended 1:1 with EZ Mixin and evaluated as above. Cryopreserved spermatozoa assessed by flow cytometry tended to be less well correlated (r<0.68) with the other methods, and estimates were significantly higher with eosin-nigrosin staining (P<0.001). This study shows that different methods may equally estimate viability of fresh equine spermatozoa. However, evaluation by flow cytometry appears to be less precise with cryopreserved spermatozoa.

Stem-loop structure in the 5' region of potato virus X genome required for plus-strand RNA accumulation.

Computer-generated thermodynamic predictions and solution structure probing indicated two stem-loop structures, stem-loop 1 (SL1; nt 32-106) and stem-loop 2 (SL2; nt 143-183), within the 5' 230 nt of potato virus X (PVX) RNA. Because the existence of SL1 was further supported by covariation analysis of several PVX strains, the functional significance of this structure was investigated by site-directed mutational analysis in a tobacco protoplast system. In general, mutations that reduced genomic plus-strand RNA accumulation similarly affected coat protein accumulation, indicating that subgenomic plus-strand RNA was also affected. In contrast, minus-strand RNA levels remained relatively unchanged. Mutational analysis of the stem C (SC) region of SL1 indicated that pairing was more important than sequence, which was consistent with the covariation analysis. Alterations that increased length and stability of either SC or stem D (SD) were deleterious to plus-strand RNA accumulation. The formation of internal loop C between SC and SD, as well as specific nucleotides within this loop, were also required. Several modifications were made to the terminal GAAA tetraloop, a motif known for enhanced RNA stability. Both GANA and GAAG motifs resulted in wild-type levels of RNA accumulation. However, a UUCG tetraloop was detrimental, indicating that the sequence of this element was important beyond just providing stabilization of the structure. These data indicate that multiple features of SL1 are critical for accumulation of PVX plus-strand RNA.

Effect of ambient temperatures during oocyte recovery on in vitro production of bovine embryos.

Recovery of oocytes from ovaries collected at slaughter was carried out at three ambient temperatures (25 degrees, 30 degrees and 35 degrees C) to assess the effect on subsequent embryonic production in vitro. Oocytes recovered at each temperature were thereafter maintained at temperatures > or =35 degrees C as they were subjected to in vitro maturation, fertilization and culture (IVM/IVF/IVC). The oocytes and resulting embryos within each temperature group were subsequently evaluated for their rates of fertilization, cleavage and development to blastocysts, as well as for the number of cells/blastocyst. The results demonstrate that exposure of cumulus-ocyte-complexes (COCs) to temperatures below 35 degrees C during oocyte recovery is detrimental to optimal embryo production. Although the fertilization and cleavage rates of oocytes recovered at temperatures below 35 degrees C were not significantly lower than that of the controls, the percentage of oocytes recovered at 35 degrees C that developed to the blastocyst stage following fertilization and culture (33.7%) was significantly greater than those from oocytes recovered at either 25 degrees C (22.4%) or 30 degrees C (19.5%). The mean numbers of blastomeres/embryo were significantly lower in embryos derived from oocytes collected at either 25 degrees or 30 degrees compared with those collected at 35 degrees C. The results of this study suggest that exposure of COCs to temperatures below 35 degrees C during oocyte recovery may significantly decrease both the quantity and quality of embryos produced by in vitro methods.

The Kinetics of Bacteriolysis in the Gut of the Deposit Feeder Arenicola marina.

A turbidimetric technique was used to measure the bacteriolytic activities of digestive fluids collected seasonally from the deposit-feeding polychaete Arenicola marina. Lysis of cultured sedimentary bacteria was monitored spectrophotometrically. The kinetics of the lytic reaction were characterized with respect to temperature, pH, reactant concentration, and time. Bacteriolysis generally followed saturation kinetics behavior and was apparently a first-order process with respect to the substrate (bacteria). Deviation from a simple Michaelis-Menten relationship, however, was observed at high substrate levels, at which inhibition of lysis was evident. The lytic rate decreased rapidly and sharply through time as the substrate was depleted. Lytic factors in the gut fluid acted as catalysts; rates increased with concentration of gut fluid, but no depletion in lytic capacity through time was observed. The lytic rate increased exponentially for all seasonal samples over the range of environmental temperatures (4 to 31(deg)C) but showed little relationship with pH under conditions likely to be experienced in situ. The lytic rate also varied among bacterial strains and with culture age.

Comparison of development of pig zygotes and embryos in simple and complex culture media.

The development of pig zygotes and embryos in three culture media was investigated. Zygotes and morulae, recovered from superovulated prepubertal gilts after slaughter, were randomly allocated to culture in one of three simple media: (i) Chatot, Ziomek, Bavister medium (CZB) in 5% CO2:95% air; (ii) Hepes-buffered CZB medium (H-CZB) in 100% air; or in a complex solution enriched with vitamins and amino acids; or (iii) modified Eagle's minimal essential medium (Pig-MEM) in 5% CO2:95% air. The relative effectiveness of the various media and gas combinations was assessed by determining the percentage of zygotes and morulae that developed into blastocysts and hatched from their zonae, as well as by the mean number of cells per embryo. Of zygotes cultured either in CZB in CO2:air, or in H-CZB in air alone, 65% and 57% developed into blastocysts, respectively; no zygotes cultured in Pig-MEM in CO2:air developed beyond the four-cell stage. After 115 h of culture, embryos resulting from zygotes cultured in either CZB or H-CZB contained approximately 50 cells per embryo. Morulae differed from zygotes in their developmental potential in the three media in that virtually all morulae cultured in CZB, H-CZB or Pig-MEM developed to the expanded blastocyst stage in vitro. However, of blastocysts resulting from morulae cultured in Pig-MEM, 97% hatched from their zonae, whereas only 8% and 5%, respectively, of those resulting from morulae cultured in CZB or H-CZB did so.(ABSTRACT TRUNCATED AT 250 WORDS)

In vitro and in vivo survival of cryopreserved sheep embryos.

Experiments were conducted to devise an efficient method to cryopreserve ovine embryos for field application. Embryos were surgically collected from superovulated ewes on Day 6 after natural breeding; oocytes were collected from ovaries obtained at the abattoir. Osmotic behavior of oocytes and embryos was determined by measuring their responses to hypertonic solutions of CsCl or sucrose. Embryos and oocytes contracted osmotically by decreasing their volumes proportionally to the reciprocal of the solution's osmolality. The respective nonosmotic volumes of embryos in CsCl and sucrose were 13.8 and 13.5% of their isotonic volume, and those of oocytes were 18.5 and 19.6%. Tests of the permeability of morulae to commonly used cryoprotectants, ethylene glycol (EG), propylene glycol (PG), dimethyl sulfoxide (DMSO), and glycerol (Glyc), showed that the order of permeability was EG > PG > DMSO approximately Glyc. Comparison of the efficacy of cryoprotective agents indicated that the respective survivals of embryos frozen in EG, PG, and DMSO were 76.9, 62.5, and 55.6%, based on their development into hatched blastocysts in vitro. Therefore, EG appeared to be superior to the other two cryoprotectants for freezing sheep embryos. To determine the functional survival of embryos in vivo, 67 embryos frozen in EG were thawed and directly diluted with phosphate-buffered saline; 47 of these (70%) appeared morphologically normal and were transferred into 14 recipients. Five of these recipients, which had received a total of 16 embryos, became pregnant. Ten lambs were born, showing that the method employed in this study for cryopreservation of sheep embryos followed by their direct dilution out of EG has potential application for practical field use.

Cleavage and 3H-uridine incorporation in bovine embryos of high in vitro developmental potential.

The timing of genome activation in bovine embryos is still not well defined. The objective of this study was therefore to investigate transcription in bovine embryos with a high potential to develop in culture after in vitro fertilization, by examining, autoradiographically, their incorporation of 3H-uridine. Initial experiments determined that developmental potential in vitro could be related to the time of first division of the zygote. Embryos that completed their first cleavage within 30 hours of exposure to sperm were more likely to develop into blastocysts (65.7%) and to hatch (50.9%). Using such embryos, it was found that 10 of 12 8-cell and all 11 4-cell stage embryos were labeled after a 2-4-hr exposure to 3H-Uridine. Among 2-cell stage embryos, 0 of 23, 3 of 17, 8 of 15, and 3 of 4 were labeled after exposure to 3H-uridine of 2, 4, 7, and 10 hr, respectively. Treatment with alpha-amanatin (10-100 micrograms/ml) blocked 3H-uridine incorporation but did not inhibit cleavage during the first 4 cell cycles. It was concluded that transcription occurs as early as the 2-cell stage in bovine embryos in vitro but is not critical to the first four cell cycles.

Effects of Ureaplasma diversum on bovine oviductal explants: quantitative measurement using a calmodulin assay.

Calmodulin (CAM) acts as an intracellular regulator of calcium, an important mediator of many cell processes. We used the CAM assay and electron microscopy to investigate the effects of Ureaplasma diversum on bovine oviductal explants obtained aseptically from slaughtered cows. A stock suspension of U. diversum (treated specimens) and sterile broth (controls) was added to replicates of cultured explants and incubated at 38 degrees C in an atmosphere of 5.5% CO2 for 48 hours. Explants were examined for ciliary activity, extracellular CAM loss, and for histological and ultrastructural changes. Explants and their culture media were examined for changes in CAM concentration. All experiments were replicated three times. In addition, U. diversum, medium and broth were assayed for CAM content. The concentrations of CAM in explants and media changed significantly (p < 0.05) in samples which were inoculated with U. diversum when compared to controls. The controls and infected specimens did not differ histologically or ultrastructurally, but U. diversum was seen to be closely associated with infected explant tissue. In view of this close affinity it is assumed the loss of CAM from the oviductal cells was causally related, but this was not proven. The failure to show cell membrane injury on light and electron microscopic examination was probably related to the short duration of the experiment and may only point out the sensitivity of the CAM assay in detecting early cell membrane injury. Compromise in characteristics of the medium to support both, the viability of oviductal cells and U. diversum limited the experimental time to 48 hours.(ABSTRACT TRUNCATED AT 250 WORDS)

Immunofluorescence assay for effects on field abundance of a naturally occurring pseudomonad during passage through the gut of a marine deposit feeder, Abarenicola pacifica.

In a seasonal study we used immunofluorescence to follow a specific bacterial population, as well as total numbers, through the fore-, mid-, and hindgut of a deposit feeder, Abarenicola pacifica. We chose a pseudomonad because of its high ambient abundance. On five dates, we collected A. pacifica gut contents, with concurrent measurements of sedimentary food quality (chlorophyll a, protein, bacterial abundance), animal egestion rates (inversely proportional to gut residence time), and temperature. Increasing bacterial numbers from ingested sediment to foregut contents, and decreases from foregut to midgut indicate significant selection and digestion, respectively, of both the pseudomonad and the total bacterial community. Inverse correlations between egestion rate and digestive removal of bacteria offer some support for the prediction that digestion of bacteria is proportional to time spent exposed to digestive enzymes, although the significance of the associated statistical tests is marginal. No hindgut growth of the pseudomonad was observed, likely due to the short gut residence time of A. pacifica. The pseudomonad showed variation of less than a factor of 3 in its ambient sedimentary abundance over the year.