Polledness in Argentinean Creole cattle, five centuries surviving.

Polledness has been shown to have autosomal Mendelian inheritance, with the polled locus being dominant to the horned locus. This trait was mapped to the BTA1 centromeric end in several breeds. One of the distinctive attributes of Creole cattle, such as the Argentinean Creole, is the presence of long, lyre-shaped horns. However, polled native animals were reported before the introduction of modern selected European breeds. Here, we studied the origin of the polled mutation, either independent or introgressed, in a Creole line from the Creole cattle founder group at the IIACS-INTA Leales Experimental Station (northwest Argentina). The study sample (65 animals: 26 horned and 39 polled) was genotyped using high-density SNP microarrays and three previously reported genetic markers (P202 ID , P80kb ID and PG ). A genome-wide association study, selection signatures, linkage disequilibrium analysis and copy number variations were used to detect the responsible region and the segregating haplotypes/alleles. The interval mapped in the Leales herd (1.23-2.13 Mb) overlapped with the region previously reported in several European cattle breeds, suggesting that the same locus could be segregating in this population. The previously reported variants PF and PG were not detected, thus dismissing the Holstein-Friesian and Nellore origins of the polled phenotype in this native breed. Conversely, the presence of the Celtic variant PC suggests an almost complete co-segregation. The cluster analysis rejected the hypothesis of recent introgression, which is compatible with the historical record of polled Creole cattle in northwest Argentina.

Analysis of Doberman Pinscher and Toy Poodle samples with targeted next-generation sequencing.

Next-generation sequencing (NGS) technologies have enabled the identification of many causal variants of genetic disorders, the development of parentage tests and the analysis of multiple traits in domestic animals. In this study, we evaluated the performance of a Canine Targeted Genotyping-by-Sequencing (GBS) custom panel (Thermo Fisher Scientific, Waltham, Ma, USA) in a cohort of 95 dog DNA samples, comprising 76 Doberman Pinschers and 19 Toy Poodles from Argentina. The used panel included 383 targets (228 parentage SNVs, 137 genetic disorder markers and 18 trait markers). While paternity analysis showed correct duo (97.4%; LOD > 2.98E+13) and trio (100%; LOD > 2.20E+15) parentage assignment, the panel resulted still insufficient for excluding close relatives in inbred populations. In this sense, close relatives were wrongly assigned as parents in 12.6% of duos and 0.3% of trios. We detected 17 polymorphic markers (genetic disorders, n = 4; hair type, n = 3; coat color, n = 10) and estimated their allele frequencies in the studied breeds. The accuracy of targeted GBS results were evaluated for three markers that were associated with Progressive rod-cone degeneration, von Willebrand disease type 1 and dilated cardiomyopathy by pyrosequencing and Sanger sequencing genotyping, showing 94-100% concordance among assays. The targeted GBS custom panel resulted cost-effective strategy to study the prevalence of genetic disorders and traits in a large number of samples and to analyze genetic interactions between previously reported variants. Once assays based on AgriSeq technology were standardized, their uses are a good strategy for large-scale routine genetic evaluation of animal populations.

Out-of-sheath corticosteroid injections through the dorsal webspace for trigger finger and trigger thumb. A prospective cohort study. / Infiltración comisural dorsal en el tratamiento del pulgar y los dedos en resorte. Estudio de una cohorte prospectiva.

BACKGROUND AND OBJECTIVE: Steroid injections are effective in the treatment of trigger digits but the pain during the injection is an always-present accompanying effect. The aim of this study was to assess the effectiveness and perceived pain during an out-of-sheath corticosteroid injection through the dorsal webspace in the treatment of trigger digits. MATERIAL AND METHOD: A total of 126 consecutive patients were included. A subcutaneous (out-of-sheath) corticosteroid injection was performed through the dorsal webspace in all digits. In cases where signs or symptoms persisted, a second injection was offered. Visual analog scale for pain during the injection, DASH questionnaire, success rate and complications were collected. RESULTS: There were 86 women and 40 men with a mean age of 61 years. The mean visual analog scale for pain during the injection was 3.8. Twelve patients were lost to follow-up. The overall success was 68% and success after a single injection was 54%. The best result was achieved on the ring finger. Patients who were not previously operated on carpal tunnel syndrome responded better. No complications were noted. CONCLUSIONS: The extra-sheath corticosteroid injection through the dorsal webspace is effective and safe. It seems to be less painful than the reported scores for the palmar midline technique although it should be assessed in a comparative study.

Chronic kidney disease after orthotopic liver transplantation in recipients receiving tacrolimus.

INTRODUCTION: Chronic kidney disease (CKD) is common in liver transplant recipients receiving calcineurin inhibitors. METHOD AND POPULATION: The goals of this case-control study were to identify risk factors associated with CKD and its effect on mortality in 294 liver transplant recipients receiving calcineurin inhibition with tacrolimus. RESULTS: Hepatitis C virus (HCV) was the most common indication (42%) for transplantation. CKD 4 and 5 (estimated glomerular filtration rate (eGFR) of <=29 ml/min/1.73 m2) developed in 10.8% of recipients during a mean follow-up of 52 months. The incidence density of CKD was 2.56 per 100 patient-years. End-stage renal disease developed in 2.7%. By univariate analysis, CKD patients were older (mean±sd, 57±10 vs. 51±11, p<0.05) with hypertension (56 vs. 32%, p<0.05), had lower preoperative hematocrit (31±6 vs. 34±5, p<0.05), alanine aminotransferase (median (95% confidence limit) 46 (34­80) vs. 68 (56­77), <0.05) and eGFR (56±28 vs. 91±35 ml/ min/1.73 m2, p<0.05), had higher preoperative prothrombin time (16.1 (14.6­17.2) vs. 14.8 (14.5­15.1) seconds, p<0.05), and required more perioperative renal replacement therapy (RRT) (41% vs. 6.5%, p<0.05) compared to controls. Perioperative need for RRT (hazard ratio (95% CI) 2.72 (1.05­7.03)) and lower preoperative eGFR: 60­89 (4.08 (1.23­13.5)), 30­59 (4.26 (1.18­15.36)), and<=29 (5.91 ((1.28­27.19)) vs. eGFR>=90 ml/min/1.73 m2 were independently associated with development of CKD adjusting for important covariates. The development of CKD (2.36 (1.22­4.59)) was independently associated with late mortality with an attributable risk of 12.8%. CONCLUSION: Data demonstrate that CKD is an important clinical event associated with increased risk for death after primary liver transplantation.

Dorsal web injection technique in the treatment of trigger finger and trigger thumb. Anatomical study. / Infiltración comisural dorsal en el tratamiento del pulgar y los dedos en resorte. Estudio anatómico.

BACKGROUND AND OBJECTIVE: Corticosteroid injection is an effective treatment for trigger digits but the pain during the injection is an ever-present side effect. Since the palmar skin has a high density of sensitive receptors, injecting through dorsal skin could be less painful. Our aim was to assess whether a dorsal technique through the dorsal web is safe for extra-sheath injection of trigger fingers and thumb. MATERIAL AND METHOD: This is an anatomical study in sixteen cadaveric hands. An injection through the dorsal web was performed on each digit. After careful resection of the palmar skin, the distance between the needle and the main anatomical structures was measured. The risk of major injury was considered high when the mean distance from the needle to the neurovascular bundle was below 1mm. RESULTS: The mean distance from the needle to the neurovascular bundle was 1.77mm. Two neurovascular injuries in 112 injections were observed, one nerve and one artery. The safest digit was the thumb while the most dangerous was the index finger. At the ring finger, the technique was safer when it was carried out from the dorso-radial. CONCLUSIONS: A subcutaneous injection near the flexor tendon sheath can be carried out through the dorsal web with a low, but present, risk of neurovascular injury. It could be useful for injection in the treatment of trigger finger and trigger thumb but it should be assessed in a clinical study.

External Marginal Gap Evaluation of Different Resin-filling Techniques for Class II Restorations-A Micro-CT and SEM Analysis.

CLINICAL RELEVANCE: Secondary caries are the main reason for the failure of restorations, class II being the most affected. Techniques that promote less gap percentage are important. Flowable bulk fill composites used at such locations have been shown to decrease gap formation while being a faster procedure than an incremental technique.

A new device for the fixation of anterior cruciate ligament tendon grafts. Design and experimental study.

In this paper the design and experimental analysis is presented of a new fixation device of anterior cruciate ligament (ACL) grafts of the knee. This device is inserted into the bone tunnel, after the graft, in the same way as an interference screw. However, the fixation device described in this paper has been designed in such a way that, after the insertion of a threaded element in its interior, some of its components expand in a radial direction, pressing against the walls of the bone tunnel and thereby increasing the fixation of the graft. This expansion device can be used in both the femur and the tibia. The device proposed in this paper was compared with an interference screw for load failure and fixation stiffness in experiments performed using porcine bones. The failure load was significantly higher in the new expansion device group (633+/-202 N) than in the interference screw group (471+/-179 N). The stiffness obtained when the new device was used (59+/-20 N/mm) was also significantly higher than that obtained using the interference screw (37+/-19 N/mm) (t-test, P<0.05). According to these results, this new device could be considered a good alternative to improve fixation of anterior cruciate ligament grafts.

Quantification of Cryptosporidium parvum in anaerobic digesters treating manure by (reverse-transcription) quantitative real-time PCR, infectivity and excystation tests.

The survival of Cryptosporidium parvum oocysts in anaerobic digesters treating manure was investigated for mesophilic, thermophilic, and a combined treatment (mesophilic-thermophilic-mesophilic) under different retention times of oocysts in the reactors. C. parvum DNA was extracted with an optimised protocol, and its amount determined by quantitative real-time PCR (qPCR). Results indicated noteworthy differences in DNA content after the different treatments. DNA was not degraded during the process. However, excystation and infectivity tests showed a reduction of viable oocyst numbers of > or = 2 and > or = 5 log units after the thermophilic treatment in two different experiments. Thus qPCR-targeting DNA can overestimate the number of oocysts that survive and remain viable after anaerobic digestion. However, targeting DNA is suitable to indicate the presence or absence of oocysts. Reverse transcription qPCR (RT-qPCR) targeting C. parvum hsp70 mRNA successfully indicated the presence of viable fraction of oocysts.

Mesophilic-thermophilic-mesophilic anaerobic digestion of liquid dairy cattle manure.

The potential of a mesophilic-thermophilic-mesophilic anaerobic digestion system was investigated with respect to improvement of both digestion and sanitation efficiencies during treatment of liquid cattle manure. The pilot plant produced a high methane yield from liquid dairy cattle manure of 0.24 m3 (kg VSfed)(-1) Considering the low system loading rate of 1.4-1.5 kg VS (m3 d)(-1), digestion efficiency compared to conventional processes did not appear improved. The minimum guaranteed retention time in the tubular thermophilic reactor was increased compared to a continuously stirred tank reactor. Levels of intestinal enterococci in raw liquid manure as determined with cultivation methods were reduced by 2.5 -3 log units to a level of around 10(2) cfu/mL. This sanitizing effect was achieved both during mesophilic-thermophilic-mesophilic and thermophilic-mesophilic treatment, provided the thermophilic digester was operated at 53-55 degrees C. A change in feeding interval from 1 h to 4 h did not significantly alter methane yield and sanitation efficiency. It was proposed that a two-stage, thermophilic-mesophilic anaerobic digestion system would be able to achieve the same sanitizing effect and equal or better digestion efficiency at lower costs.

Hygienization by anaerobic digestion: comparison between evaluation by cultivation and quantitative real-time PCR.

In order to assess hygienization by anaerobic digestion, a comparison between evaluation by cultivation and quantitative real-time PCR (qPCR) including optimized DNA extraction and quantification was carried out for samples from a full-scale fermenter cascade (F1, mesophilic; F2, thermophilic; F3, mesophilic). The system was highly effective in inactivating (pathogenic) viable microorganisms, except for spore-formers. Conventionally performed cultivation underestimated viable organisms particularly in F2 and F3 by a factor of at least 10 as shown by data from extended incubation times, probably due to the rise of sublethally injured (active but not cultivable) cells. Incubation should hence be extended adequately in incubation-based hygiene monitoring of stressed samples, in order to minimize contamination risks. Although results from qPCR and cultivation agreed for the equilibrated compartments, considerably higher qPCR values were obtained for the fermenters. The difference probably corresponded to DNA copies from decayed cells that had not yet been degraded by the residual microbial activity. An extrapolation from qPCR determination to the quantity of viable organisms is hence not justified for samples that had been exposed to lethal stress.

Physical interaction between Langerhans cells and T-lymphocytes during antigen presentation in vitro.

Physical interaction between Langerhans cells and T cells is an essential requirement for antigen presentation. In this study we report the ultrastructural characteristics of the antigen-specific physical interactions that occur in vitro between murine Langerhans cells and T cells. Epidermal Langerhans cells enriched to 78% purity by a panning method were pulsed with 2,4-dinitrophenyl-Limulus polyphemus hemocyanin and co-incubated with syngeneic T lymphocytes primed in vivo with the same antigen. A substantial number of conjugates constituted by Langerhans cells surrounded by three or more lymphocytes were obtained after 60 min of incubation at 4 degrees C. Electron microscopy of the conjugates revealed that Langerhans cells and T lymphocytes interacted by two type of contacts. Type I was characterized by glycocalyx-glycocalyx interaction that occurred in relation to protrusions or microvilli of both cells. Type II was characterized by wide and tight areas of close apposition between Langerhans cells and T-lymphocyte plasma membranes. In these areas there were zones with intercellular bridges and small septilaminar junctions highly suggestive of gap junctions. An electron-immunogold procedure demonstrated the presence of DNP-LPH antigen on the type I contact. Our findings suggest that type I contact may represent the locus for antigen presentation whereas the type II contact may be involved in keeping adhesiveness between Langerhans cells and lymphocytes during antigen presentation.

Pituitary response to luteinizing hormone-releasing hormone analog at sea level and high altitudes.

Two groups of subjects living at different altitudes were stimulated with synthetic luteinizing hormone-releasing hormone (LH-RH) or with an LH-RH analog. One hundred micrograms LH-RH was given subcutaneously to 19 men and 19 women, all normal, of reproductive age, and living at sea level, and to 19 men and 20 women living at a high altitude (2940 m). In neither sex, however, were the differences in peak responses for both gonadotropins significantly different between the sea level and high-altitude groups. Plasma levels of LH and follicle-stimulating hormone (FSH) were also measured after administration of [D-Leu6, des Gly-NH2(10)]-LH-RH ethylamide (25 micrograms subcutaneously) in 32 men and 37 women living at sea level and at high altitude. The analog induced a powerful and protracted stimulation of the release of LH and FSH irrespective of altitude and sex. The responses of men and women to the LH-RH analog, both for LH as well as for FSH, were significantly greater at sea level than at high altitudes.

Cells of the carotid body express connexin43 which is up-regulated by cAMP.

We identified a gap junction protein subunit, connexin43 (Cx43) by immunofluorescence and immunoblotting, in cultured rat carotid body cells and in whole organs. In 1-week-old cultures, all cells were flat but after 3 h exposure to 8Br-cAMP (1 mM), tyrosine hydroxylase (TH) positive cells (chemoreceptors), but not TH negative cells, adopted a round body with multiple thin arborization processes. The incidence of dye coupling between cultured cells of the same type increased from 26% in controls to 73% after treatment with 8Br-cAMP. In control cultures, Cx43 immunoreactivity showed a diffuse perinuclear distribution and after 8Br-cAMP treatment, it was also found at cell-cell contacts. Both 8Br-cAMP-induced dye coupling and cellular redistribution of Cx43 were blocked by pretreatment with actinomycin D (5 microM), a mRNA transcription blocker. Moreover, 3 h exposure to 8Br-cAMP increased the levels of Cx43 in entire organs. We suggest that conditions that promote a sustained increase in cytosolic cAMP up-regulate coupling between carotid body cells in a transcription-dependent manner. The possible functional significance of these findings is discussed.

Regulation of glycogen content in rat pineal gland by norepinephrine.

In the rat pineal gland the glycogen stores were cytochemically localized in astrocytes and pinealocytes. Moreover, it was found that norepinephrine (NE) induced a time- and concentration-dependent reduction in pineal glycogen content and yielded lactic acid. The NE effect was prevented by blocking alpha1- but not alpha2 or beta-adrenoceptors. Activation of alpha2-adrenoceptors induced a small decrease in glycogen levels that could have pre- and postsynaptic components. Activation of beta-adrenoceptors with 10(-12)-10(-3) M isoproterenol (ISO) induced a bell shape concentration-response curve, presumably due to desensitization, since the response induced by 10(-4) M ISO was greater with shorter period of stimulation. On the other hand, activation of alpha1-adrenoceptors with 10(-12)-10(-3) M phenylephrine (PHN) induced a hyperbolic concentration-response curve with a maximum at concentrations above 10(-8) M. Moreover, treatment with ISO drastically reduced the response induced by PHN concentrations lower but not higher than 10(-6) M, favoring a concentration-dependent response between 10(-6) and 10(-4) M PHN, similar to that induced by equimolar NE concentrations. Thus, the NE-induced reduction in glycogen content of the rat pineal gland is mainly mediated by alpha1-adrenoceptors and modulated by intracellular mechanisms activated by beta-adrenoceptors.

The ependymocytes of the bovine subcommissural organ are functionally coupled through gap junctions.

The subcommissural organ (SCO) is a circunventricular organ secreting glycoproteins into the ventricle. It is richly innervated by (1) serotonergic fibers originated in raphe nuclei, that would exert an inhibitory control, and (2) peptidergic fibers of unknown function. Due to the scarce number of the latter, their functional significance might largely depends on whether the cells of the SCO are functionally coupled through gap junctions. This investigation was designed to answer this question. The bovine SCO, either freshly isolated or maintained in organ culture, was processed for immunoblot and immunocytochemistry, using an anti-connexin43 antibody, and dye coupling studies. It was found that the cells of the SCO in situ are functionally coupled through gap junctions made at least of connexin43, but in cultured explants are not. The possibility that coupling of the SCO may be controlled by the neural input and undergoes circadian variations is discussed.

Inter-arm asymmetry in bone mineral content and bone area in postmenopausal recreational tennis players.

OBJECTIVE: To determine whether long-term recreational tennis participation is associated with increased bone and muscle mass in the dominant compared to the non-dominant arm, in postmenopausal tennis players. METHODS: Dual-energy X-ray whole body scans were performed in 10 postmenopausal tennis players (mean +/- S.D.; 59.7 +/- 4.8 years) and 12 postmenopausal non-active women (62.8 +/- 7.2 years) matched for age and height. Tennis players started in this sport at 31.4 +/- 8.8 years, and had been playing for 26.8 +/- 6.8 years, at least 3 h per week. In addition, shoulder isokinetic muscular strength was also evaluated. RESULTS: Tennis participation resulted in a lower whole mass and fat mass in both arms (P < 0.05). Dominant arm muscle mass and shoulder joint isokinetic strength were essentially similar in both groups, however, the tennis players showed 8% greater bone mineral content and 7% greater osseous area in the dominant than in the non-dominant arm (P < 0.05). No inter-arm differences were found in bone mineral density. A relationship was observed between the length of tennis participation and the degree of inter-arm asymmetry in bone mineral content (r = 0.81, P < 0.01) and bone area (r = 0.78, P < 0.01). However, the BMD of the arms was essentially similar in both groups. No differences were observed in femoral and lumbar bone mass and density between groups. CONCLUSIONS: Long-term tennis participation is associated with increased bone mass, but not BMD, in the dominant arm of postmenopausal tennis players. The magnitude of the inter-arm asymmetry of postmenopausal tennis players is proportional to the length of tennis participation.

Perthes' disease. A study of constitutional aspects in adulthood.

We studied 16 patients suffering from osteoarthritis of the hip who had had Perthes' disease during childhood. They were compared clinically and radiologically with a control group who had not had Perthes' disease, in order to assess whether a generalised, pre-existing constitutional disorder was present. Nine patients with a previous history of Perthes' disease had some other skeletal abnormality, but only three presented with clinical symptoms. Only one patient in the control group was found to have an abnormality but was symptom-free. Our findings provide further evidence that patients with Perthes' disease may have a generalised abnormality related to chondrogenesis which can produce other skeletal anomalies that persist into adult life.

Longitudinal bone growth after sciatic denervation in rats.

The right sciatic nerve of 50 one-month-old male rats was cut under general anaesthesia. Groups of animals were sacrificed at intervals of up to 12 weeks after operation and the length of the femora, tibiae and first and fifth metatarsals were measured with a caliper accurate to 0.05 mm. From the first week, both metatarsals were between 3% and 5% shorter on the denervated side, but there was no further increase of the discrepancy. The femora were less than 1% longer in the denervated limb at the second and eighth week. No difference was found between the lengths of the tibiae. The various factors which could possibly be responsible for these findings are discussed.

Evaluating real-time PCR for the quantification of distinct pathogens and indicator organisms in environmental samples.

We evaluated quantitative real-time PCR (qPCR) and RTqPCR (for RNA species) for their ability to quantify microorganisms and viruses in problematic environmental samples such as cattle manure, digester material, wastewater and soil. Important developments included a standard spiking approach which compensated for methodological bias and allowed sample-to-sample comparison and reliable quantification. Programme CeTe was developed to calculate endogenous concentrations of target organisms (nucleic acid copies) for each sample separately from the generated standard curves. The approach also permitted assessment of the detection limit of the complete method, including extraction. It varied from sample to sample, due to different extraction efficiencies and variable co-extraction of PCR inhibitors. False negative results were thereby avoided. By using this approach we were able to optimise a DNA extraction protocol from the different tested sample types. Protocols for the extraction of RNA species from environmental samples were also optimised. DNA was (almost) not degraded after lethal shock (autoclaving) in the sterile environment. In contrast, the parallel selective cultivation and qPCR results for various microbial parameters from an anaerobic digester chain suggested that DNA from decaying organisms was readily recycled in metabolically active environments. It may, therefore, be used to determine viable organisms in samples exhibiting substantial metabolic turnover. It is proposed that our standard spiking approach, including data evaluation by the program CeTe, should be considered in future standardisation and norms for the quantification of nucleic acid containing organisms in environmental and product samples.

[Anesthesia for vertebral osteoclasis in a patient with severe predominantly cervical spondylitis ankylopoietica]. / Anestesia para la intervención de osteoclasia vertebral en un paciente con espondilitis anquilopoyética grave de predominio cervical.

We present a patient suffering severe, mainly cervical, ankylopoietic spondylitis who was scheduled for posterior vertebral osteoclasis under local anesthesia. We describe handling of anesthesia, particularly airway control by flexible bronchoscopy in an awake, locally anesthetized patient sedated with benzodiazepines and inhalational anesthetics as compared with other alternatives: an awake patient who is not intubated, under local anesthesia and sedated with narcotics and gaseous anesthetics, or a patient under general anesthesia with placement of a flexible bronchoscope and neurological monitoring by evoked potentials. The surgical technique was Simmons' posterior vertebral osteoclasis, with the column extended and immobilized with wire.