Molecular characterization of G and P-types bovine rotavirus strains from Goiás, Brazil: high frequency of mixed P-type infections

In this study, 331 samples from calves less than one month old from a dairy herd in the district of Piracanjuba, state of Goiás, Brazil were tested for rotavirus. Thirty-three samples (9.9%) tested positive for rotavirus. Out of those, 31 were submitted to G and P characterization by reverse transcription followed by semi-nested polymerase chain reaction. Two samples were characterized as G6P[1], three as G10P[11] and five as G6P[11]. The majority of the samples (51.6%) displayed multiple P genotypes (P-genotype mixtures), including typical human genotypes P[4] and P[6M], suggesting the occurrence of co-infections and genetic reassortment. Also, the detection of human genotypes in bovine samples may be considered evidence of the zoonotic potential of rotaviruses. To our knowledge, this is the first report of such a high frequency of P genotype mixtures in bovine rotavirus samples. It also increases data on G and P rotavirus genotypes circulating in dairy herds in Brazil and can help in the development of more efficient immunization approaches, thereby controlling infection and reducing economical losses.

High-frequency linkage of co-expressing T-DNA in transgenic Arabidopsis thaliana transformed by vacuum-infiltration of Agrobacterium tumefaciens

The efficiency of co-expression and linkage of distinct T-DNAs present in separate Agrobacterium tumefaciens was analysed in Arabidopsis thaliana transformed by the vacuum infiltration method. Co-expression was monitored by the synthesis of three bacterial proteins involved in the production of polyhydroxybutyrate (PHB) in the plastids. Out of 80 kanamycin-resistant transgenic plants analysed, 13 plants were co-transformed with the two distinct T-DNAs and produced PHB. Of those, 7 lines had a kanamycin-resistance segregation ratio consistent with the presence of a single functional insert. Genetic linkage between the distinct T-DNAs was demonstrated for all 13 PHB-producing lines, while physical linkage between the distinct T-DNAs was shown for 12 out of 13 lines. T-DNAs were frequently linked in an inverted orientation about the left borders. Transformation of A. thaliana by the co-infiltration of two A. tumefaciens containing distinct T-DNAs is, thus, an efficient approach for the integration and expression of several transgenes at a single locus. This approach will facilitate the creation and study of novel metabolic pathways requiring the expression of numerous transgenes.

Wide-pulse-high-frequency neuromuscular stimulation of triceps surae induces greater muscle fatigue compared with conventional stimulation.

We compared the extent and origin of muscle fatigue induced by short-pulse-low-frequency [conventional (CONV)] and wide-pulse-high-frequency (WPHF) neuromuscular electrical stimulation. We expected CONV contractions to mainly originate from depolarization of axonal terminal branches (spatially determined muscle fiber recruitment) and WPHF contractions to be partly produced via a central pathway (motor unit recruitment according to size principle). Greater neuromuscular fatigue was, therefore, expected following CONV compared with WPHF. Fourteen healthy subjects underwent 20 WPHF (1 ms-100 Hz) and CONV (50 μs-25 Hz) evoked isometric triceps surae contractions (work/rest periods 20:40 s) at an initial target of 10% of maximal voluntary contraction (MVC) force. Force-time integral of the 20 evoked contractions (FTI) was used as main index of muscle fatigue; MVC force loss was also quantified. Central and peripheral fatigue were assessed by voluntary activation level and paired stimulation amplitudes, respectively. FTI in WPHF was significantly lower than in CONV (21,717 ± 11,541 vs. 37,958 ± 9,898 N·s P<0,001). The reductions in MVC force (WPHF: -7.0 ± 2.7%; CONV: -6.2 ± 2.5%; P < 0.01) and paired stimulation amplitude (WPHF: -8.0 ± 4.0%; CONV: -7.4 ± 6.1%; P < 0.001) were similar between conditions, whereas no change was observed for voluntary activation level (P > 0.05). Overall, our results showed a different motor unit recruitment pattern between the two neuromuscular electrical stimulation modalities with a lower FTI indicating greater muscle fatigue for WPHF, possibly limiting the presumed benefits for rehabilitation programs.

Use of high-frequency jet ventilation for percutaneous tumor ablation.

PURPOSE: To report feasibility and potential benefits of high-frequency jet ventilation (HFJV) in tumor ablations techniques in liver, kidney, and lung lesions. METHODS: This prospective study included 51 patients (14 women, mean age 66 years) bearing 66 tumors (56 hepatic, 5 pulmonary, 5 renal tumors) with a median size of 16 ± 8.7 mm, referred for tumor ablation in an intention-to-treat fashion before preoperative anesthesiology visit. Cancellation and complications of HFJV were prospectively recorded. Anesthesia and procedure duration, as well as mean CO2 capnea, were recorded. When computed tomography guidance was used, 3D spacial coordinates of an anatomical target <2 mm in diameter on 8 slabs of 4 slices of 3.75-mm slice thickness were registered. RESULTS: HFJV was used in 41 of 51 patients. Of the ten patients who were not candidate for HFJV, two patients had contraindication to HFJV (severe COPD), three had lesions invisible under HFJV requiring deep inspiration apnea for tumor targeting, and five patients could not have HFJV because of unavailability of a trained anesthetic team. No specific complication or hypercapnia related to HFJV were observed despite a mean anesthetic duration of 2 h and ventilation performed in procubitus (n = 4) or lateral decubitus (n = 6). Measured internal target movement was 0.3 mm in x- and y-axis and below the slice thickness of 3.75 mm in the z-axis in 11 patients. CONCLUSIONS: HFJV is feasible in 80 % of patients allowing for near immobility of internal organs during liver, kidney, and lung tumor ablation.

High frequency of functionally active Melan-a-specific T cells in a patient with progressive immunoproteasome-deficient melanoma.

Tumor-reactive T cells play an important role in cancer immunosurveillance. Applying the multimer technology, we report here an unexpected high frequency of Melan-A-specific CTLs in a melanoma patient with progressive lymph node metastases, consisting of 18 and 12.8% of total peripheral blood and tumor-infiltrating CD8+ T cells, respectively. Melan-A-specific CTLs revealed a high cytolytic activity against allogeneic Melan-A-expressing target cells but failed to kill the autologous tumor cells. Loading of the tumor cells with Melan-A peptide reversed the resistance to killing, suggesting impaired function of the MHC class I antigen processing and presentation pathway. Mutations of the coding region of the HLA-A2 binding Melan-A26-35 peptide or down-regulation of the MHC class I heavy chain, the antigenic peptide TAP, and tapasin could be excluded. However, PCR and immunohistochemical analysis revealed a deficiency of the immunoproteasomes low molecular weight protein 2 and low molecular weight protein 7 in the primary tumor cells, which affects the quantity and quality of generated T-cell epitopes and might explain the resistance to killing. This is supported by our data, demonstrating that the resistance to killing can be partially reversed by pre-exposure of the tumor cells to IFN-gamma, which is known to induce the immunoproteasomes. Overall, this is the first report of an extremely high frequency of tumor-specific CTLs that exhibit competent T-cell-effector functions but fail to lyse the autologous tumor cells. Immunotherapeutic approaches should not only focus on the induction of a robust antitumor immune response, but should also have to target tumor immune escape mechanisms.

Reliable tidal volume estimates at the airway opening with an infant monitor during high-frequency oscillatory ventilation.

OBJECTIVE: To assess the suitability of a hot-wire anemometer infant monitoring system (Florian, Acutronic Medical Systems AG, Hirzel, Switzerland) for measuring flow and tidal volume (Vt) proximal to the endotracheal tube during high-frequency oscillatory ventilation. DESIGN: In vitro model study. SETTING: Respiratory research laboratory. SUBJECT: In vitro lung model simulating moderate to severe respiratory distress. INTERVENTION: The lung model was ventilated with a SensorMedics 3100A ventilator. Vt was recorded from the monitor display (Vt-disp) and compared with the gold standard (Vt-adiab), which was calculated using the adiabatic gas equation from pressure changes inside the model. MEASUREMENTS AND MAIN RESULTS: A range of Vt (1-10 mL), frequencies (5-15 Hz), pressure amplitudes (10-90 cm H2O), inspiratory times (30% to 50%), and Fio2 (0.21-1.0) was used. Accuracy was determined by using modified Bland-Altman plots (95% limits of agreement). An exponential decrease in Vt was observed with increasing oscillatory frequency. Mean DeltaVt-disp was 0.6 mL (limits of agreement, -1.0 to 2.1) with a linear frequency dependence. Mean DeltaVt-disp was -0.2 mL (limits of agreement, -0.5 to 0.1) with increasing pressure amplitude and -0.2 mL (limits of agreement, -0.3 to -0.1) with increasing inspiratory time. Humidity and heating did not affect error, whereas increasing Fio2 from 0.21 to 1.0 increased mean error by 6.3% (+/-2.5%). CONCLUSIONS: The Florian infant hot-wire flowmeter and monitoring system provides reliable measurements of Vt at the airway opening during high-frequency oscillatory ventilation when employed at frequencies of 8-13 Hz. The bedside application could improve monitoring of patients receiving high-frequency oscillatory ventilation, favor a better understanding of the physiologic consequences of different high-frequency oscillatory ventilation strategies, and therefore optimize treatment.

Exploring the high frequency emission of radio loud X-ray binaries

We report millimetre-wave continuum observations of the X-ray binaries Cygnus X-3, SS 433, LSI+61 303, Cygnus X-1 and GRS 1915+105. The observations were carried out with the IRAM 30 m-antenna at 250 GHz (1.25 mm) from 1998 March 14 to March 20. These millimetre measurements are complemented with centimetre observations from the Ryle Telescope, at 15 GHz (2.0 cm) and from the Green Bank Interferometer at 2.25 and 8.3 GHz (13 and 3.6 cm). Both Cygnus X-3 and SS 433 underwent moderate flaring events during our observations, whose main spectral evolution properties are described and interpreted. A significant spectral steepening was observed in both sources during the flare decay, that is likely to be caused by adiabatic expansion, inverse Compton and synchrotron losses. Finally, we also report 250 GHz upper limits for three additional undetected X-ray binary stars: LSI+65 010, LSI+61 235 and X Per.

Firmground ichnofacies recording high-frequency marine flooding events (Langhian transgression, Vallè-Penedès Basin, Spain)

The decapod burrow Spongeliomorpha sudolica occurs associated with transgressive firmgrounds in the transition between Aragonian continental red beds and Langhian marine units in some of the inner sectors of the Vallès-Penedès Basin. This ichnospecies designates branching burrow systems with scratch marks in the walls produced by marine crustacean decapods. The occurrence of Spongeliomorpha represents an example of theGlossifungites ichnofacies. The several horizons where the traces are found are intercalated with continental red beds a few meters below the main transgressive surface, which is overlain by fossiliferous marine sandstones. The Spongeliomorpha-bioturbated layers record short, high frequency marine flooding surfaces that may be related either to actual sea-level changes or to variations in tectonic subsidence or sediment input. In any case, these flooding events punctuated the early phases of the Langhian transgression in the basin.

Mechanical ventilation in the newborn; a simplified approach. Part 2: High-frequency ventilation.

High frequency oscillatory ventilation (HFOV) is becoming an increasingly popular intervention in the neonatal intensive care unit. This article will attempt to explain the principles of HFOV. It is inherently more difficult to become skilled in this technique than in other forms of mechanical ventilation, so caution is warranted.

Degeneracy of antigen recognition as the molecular basis for the high frequency of naive A2/Melan-a peptide multimer(+) CD8(+) T cells in humans.

In contrast with the low frequency of most single epitope reactive T cells in the preimmune repertoire, up to 1 of 1,000 naive CD8(+) T cells from A2(+) individuals specifically bind fluorescent A2/peptide multimers incorporating the A27L analogue of the immunodominant 26-35 peptide from the melanocyte differentiation and melanoma associated antigen Melan-A. This represents the only naive antigen-specific T cell repertoire accessible to direct analysis in humans up to date. To get insight into the molecular basis for the selection and maintenance of such an abundant repertoire, we analyzed the functional diversity of T cells composing this repertoire ex vivo at the clonal level. Surprisingly, we found a significant proportion of multimer(+) clonotypes that failed to recognize both Melan-A analogue and parental peptides in a functional assay but efficiently recognized peptides from proteins of self- or pathogen origin selected for their potential functional cross-reactivity with Melan-A. Consistent with these data, multimers incorporating some of the most frequently recognized peptides specifically stained a proportion of naive CD8(+) T cells similar to that observed with Melan-A multimers. Altogether these results indicate that the high frequency of Melan-A multimer(+) T cells can be explained by the existence of largely cross-reactive subsets of naive CD8(+) T cells displaying multiple specificities.

High capacity audio watermarking using the high frequency band of the wavelet domain

This paper proposes a novel high capacity robust audio watermarking algorithm by using the high frequency band of the wavelet decomposition at which the human auditory system (HAS) is not very sensitive to alteration. The main idea is to divide the high frequency band into frames and, for embedding, to change the wavelet samples depending on the average of relevant frame¿s samples. The experimental results show that the method has a very high capacity (about 11,000 bps), without significant perceptual distortion (ODG in [¿1 ,0] and SNR about 30dB), and provides robustness against common audio signal processing such as additive noise, filtering, echo and MPEG compression (MP3).