Search

Assessing the numbers and distribution of at-risk megafauna such as the black rhino ( Diceros bicornis ) is key to effective conservation, yet such data are difficult to obtain. Many current monitoring technologies are invasive to the target animals and expensive. Satellite monitoring is emerging as a potential tool for very large animals ( e.g ., elephant) but detecting smaller species requires higher resolution imaging. Drones can deliver the required resolution and speed of monitoring, but challenges remain in delivering automated monitoring systems where internet connectivity is unreliable or absent. This study describes a model built to run on a drone to identify in situ images of megafauna. Compared with previously reported studies, this automated detection framework has a lower hardware cost and can function with a reduced internet bandwidth requirement for local network communication. It proposes the use of a Jetson Xavier NX, onboard a Parrot Anafi drone, connected to the internet throughout the flight to deliver a lightweight web-based notification system upon detection of the target species. The GPS location with the detected target species images is sent using MQ Telemetry Transport (MQTT), a lightweight messaging protocol using a publisher/subscriber architecture for IoT devices. It provides reliable message delivery when internet connection is sporadic. We used a YOLOv5l6 object detection architecture trained to identify a bounding box for one of five objects of interest in a frame of video. At an intersection over union (IoU) threshold of 0.5, our model achieved an average precision (AP) of 0.81 for black rhino (our primary target) and 0.83 for giraffe ( Giraffa giraffa) . The model was less successful at identifying the other smaller objects which were not our primary targets: 0.34, 0.25, and 0.42 for ostrich ( Struthio camelus australis ), springbok ( Antidorcas marsupialis ) and human respectively. We used several techniques to optimize performance and overcome the inherent challenge of small objects (animals) in the data. Although our primary focus for the development of the model was rhino, we included other species classes to emulate field conditions where many animal species are encountered, and thus reduce the false positive occurrence rate for rhino detections. To constrain model overfitting, we trained the model on a dataset with varied terrain, angle and lighting conditions and used data augmentation techniques ( i.e ., GANs). We used image tiling and a relatively larger ( i.e ., higher resolution) image input size to compensate for the difficulty faced in detecting small objects when using YOLO. In this study, we demonstrated the potential of a drone-based AI pipeline model to automate the detection of free-ranging megafauna detection in a remote setting and create alerts to a wildlife manager in a relatively poorly connected field environment., Competing Interests: Sky Alibhai and Zoe C. Jewell are employed by WildTrack Inc. Zoe Jewell and Sky Alibhai are Principal Research Associates JMP Division, SAS., (© 2022 Hua et al.)

Initiation is the rate-limiting step in translation. It is well-known that stable structure at a ribosome binding site (RBS) impedes initiation. The ribosome standby model of de Smit and van Duin, based on studies of the MS2 phage coat cistron, proposed how high translation rates can be reconciled with stable, inhibitory structures at an RBS. Here, we revisited the coat protein system and assessed the translation efficiency from its sequestered RBS by introducing standby mutations. Further experiments with gfp reporter constructs assessed the effects of 5'-tails-as standby sites-with respect to length and sequence contributions. In particular, combining in vivo and in vitro assays, we can show that tails of CA-dinucleotide repeats-and to a lesser extent, AU-repeats-dramatically increase translation rates. Tails of increasing length reach maximal rate-enhancing effects at 16-18 nucleotides. These standby tails are single-stranded and do not exert their effect by structure changes in the neighboring RBS stem-loop. In vitro translation and toeprinting assays furthermore demonstrate that standby effects are exerted at the level of translation initiation. Finally, as expected, destabilizing mutations within the coat RBS indicate an interplay with the effects of standby tails.

Increased numbers of CD8 + CD28 neg. T cells have been detected in the peripheral blood of patients with several types of malignancies. However, the role of these cells in anticancer immunity are not yet clear and CD8 + CD28 neg. T cells are a controversially discussed subpopulation reported both as immunosuppressive and cytotoxic. In this study, we examined the T cell receptor (TCR) repertoire and complementarity-determining region 3 sequences of CD28 neg. T cells in a melanoma patient with recurrent disease who achieved long-term disease-free status. As a result, the patient's oligoclonal CD8 + CD28 neg. T cell compartment holds TCRs that are public and specific for Melan-A as well as several public TCRs reported for common viral antigens. While over 80% of his CD8 + CD28 neg. T cells expressed a cytotoxicity marker, CD57, only 0.01% of CD8 + CD28 neg. T cells were positive for Foxp3. In conclusion, our results demonstrate that besides virus-specific also tumor-associated self-antigen targeting T cells accumulate in the CD28 neg. compartment of the immunological memory. Since the patient is in ongoing complete remission for more than 9 years, CD8 + CD28 neg. T cells with the Melan-A-specific TCR might contribute to antitumor immunity in this patient.

Stimulating the immune system to attack cancer is a promising approach, even for the control of advanced cancers. Several cytokines that promote interferon-γ-dominated immune responses show antitumor activity, with interleukin 12 (IL-12) being of major importance. Here, we used an antibody-IL-12 fusion protein (NHS-IL12) that binds histones of necrotic cells to treat human sarcoma in humanized mice. Following sarcoma engraftment, NHS-IL12 therapy was combined with either engineered IL-7 (FcIL-7) or IL-2 (IL-2MAB602) for continuous cytokine bioavailability. NHS-IL12 strongly induced innate and adaptive antitumor immunity when combined with IL-7 or IL-2. NHS-IL12 therapy significantly improved survival of sarcoma-bearing mice and caused long-term remissions when combined with IL-2. NHS-IL12 induced pronounced cancer cell senescence, as documented by strong expression of senescence-associated p16 INK4a and nuclear translocation of p-HP1γ, and permanent arrest of cancer cell proliferation. In addition, this cancer immunotherapy initiated the induction of myogenic differentiation, further promoting the hypothesis that efficient antitumor immunity includes mechanisms different from cytotoxicity for efficient cancer control in vivo .

The lifelong generation of αβT cells enables us to continuously build immunity against pathogens and malignancies despite the loss of thymic function with age. Homeostatic proliferation of post-thymic naïve and memory T cells and their transition into effector and long-lived memory cells balance the decreasing output of naïve T cells, and recent research suggests that also αβT-cell development independent from the thymus may occur. However, the sites and mechanisms of extrathymic T-cell development are not yet understood in detail. γδT cells represent a small fraction of the overall T-cell pool, and are endowed with tremendous phenotypic and functional plasticity. γδT cells that express the Vδ1 gene segment are a minor population in human peripheral blood but predominate in epithelial (and inflamed) tissues. Here, we characterize a CD4(+) peripheral Vδ1(+) γδT-cell subpopulation that expresses stem-cell and progenitor markers and is able to develop into functional αβT cells ex vivo in a simple culture system and in vivo. The route taken by this process resembles thymic T-cell development. However, it involves the re-organization of the Vδ1(+) γδTCR into the αβTCR as a consequence of TCR-γ chain downregulation and the expression of surface Vδ1(+)Vβ(+) TCR components, which we believe function as surrogate pre-TCR. This transdifferentiation process is readily detectable in vivo in inflamed tissue. Our study provides a conceptual framework for extrathymic T-cell development and opens up a new vista in immunology that requires adaptive immune responses in infection, autoimmunity, and cancer to be reconsidered.

Escherichia coli produces proteinaceous surface structures called curli that are involved in adhesion and biofilm formation. CsgD is the transcriptional activator of curli genes. We show here that csgD expression is, in part, controlled post-transcriptionally by two redundant small RNAs (sRNAs), OmrA and OmrB. Their overexpression results in curli deficiency, in accordance with the inhibition of chromosomally encoded, FLAG-tagged CsgD. Downregulation of csgD occurs by a direct antisense interaction within the csgD 5'-UTR, far upstream of the ribosome-binding site (RBS). OmrA/B downregulate plasmid-borne csgD-gfp fusions in vivo, and inhibit CsgD translation in vitro. The RNA chaperone Hfq is required for normal csgD mRNA and OmrA/B levels in the cell, and enhances sRNA-dependent inhibition of csgD translation in vitro. Translational inhibition involves two phylogenetically conserved secondary structure modules that are supported by chemical and enzymatic probing. The 5'-most element is necessary and sufficient for regulation, the one downstream comprises the RBS and affects translational efficiency. OmrA/B are two antisense RNAs that regulate a transcription factor to alter a morphotype and group behaviour.

Local topical cooling with gel-packs or cryo-cuffs is often used after surgery or after knee injuries. Mostly beneficial, but some deleterious effects have been reported in literature. The spatial distribution of heat/cold through different tissues of the knee in time may be responsible for different effects of cooling on the traumatized or inflamed tissue and have yet to be studied. Parallel computer simulation was used to study, non-invasively, temperature changes in the knee during cooling. A 3-D computer model of the knee, with a spatial resolution of 1mm, was derived from cross sections available in the visible human dataset. Heat transfer in a non-homogenous knee tissue was modeled with a diffusion equation, which was solved by the explicit finite difference method. The heat transfer between blood and tissue and tissue metabolism have also been modeled and simulated. Two different simulations were performed: cooling with a liquid at constant temperature (cryo-cuff) and topical cooling with frozen gel-packs. The simulated results were analyzed and compared with relevant measurements.

Giardia lamblia is a common intestinal-dwelling protozoan and causes diarrhoea in humans and animals worldwide. For several years, a small number of drugs such as the 5-nitroimidazole metronidazole (MET) or the thiazolide nitazoxanide (NTZ) have been used for chemotherapy against giardiasis. However, various pre-clinical and clinical investigations revealed that antigiardial chemotherapy may be complicated by emergence of giardial resistance to these drugs. The present study addressed the question if isoflavones with antigiardial activity, such as daidzein (DAI) or formononetin (FOR), may serve as alternative compounds for treatment of giardiasis. For this purpose, the potential of G. lamblia clone WB C6 to form resistance to FOR and related isoflavones was tested in vitro. In the line of these experiments, a clone (C3) resistant to isoflavones, but sensitive to MET and NTZ, was generated. Affinity chromatography on DAI-agarose using cell-free extracts of G. lamblia trophozoites resulted in the isolation of a polypeptide of approximately 40 kDa, which was identified by mass spectrometry as a nucleoside hydrolase (NH) homologue (EAA37551.1). In a nucleoside hydrolase assay, recombinant NH hydrolysed all nucleosides with a preference for purine nucleosides and was inhibited by isoflavones. Using quantitative RT-PCR, the expression of genes that are potentially involved in resistance formation was analysed, namely NH and genes encoding variant surface proteins (VSPs, TSA417). The transcript level of the potential target NH was found to be significantly reduced in C3. Moreover, drastic changes were observed in VSP gene expression. This may indicate that resistance formation in Giardia against isoflavones is linked to, and possibly mediated by, altered gene expression. Taken together, our results suggest FOR or related isoflavones as an alternative antigiardial agent to overcome potential problems of resistance to drugs like MET or NTZ. However, the capacity of Giardia to develop resistance to isoflavones can potentially interfere with this alternative treatment of the disease.

Objectives: The characterization of Giardia lamblia WB C6 strains resistant to metronidazole and to the nitro-thiazole nitazoxanide [2-acetolyloxy-N-(5-nitro 2-thiazolyl) benzamide] as the parent compound of thiazolides, a novel class of anti-infective drugs with a broad spectrum of activities against a wide variety of helminths, protozoa and enteric bacteria., Methods: Issuing from G. lamblia WB C6, we have generated two strains exhibiting resistance to nitazoxanide (strain C4) and to metronidazole (strain C5) and determined their susceptibilities to both drugs. Using quantitative RT-PCR, we have analysed the expression of genes that are potentially involved in resistance formation, namely genes encoding pyruvate oxidoreductases (POR1 and POR2), nitroreductase (NR), protein disulphide isomerases (PDI2 and PDI4) and variant surface proteins (VSPs; TSA417). We have cloned and expressed PDI2 and PDI4 in Escherichia coli. Using an enzyme assay based on the polymerization of insulin, we have determined the activities of both enzymes in the presence and absence of nitazoxanide., Results: Whereas C4 was cross-resistant to nitazoxanide and to metronidazole, C5 was resistant only to metronidazole. Transcript levels of the potential targets for nitro-drugs POR1, POR2 and NR were only slightly modified, PDI2 transcript levels were increased in both resistant strains and PDI4 levels in C4. This correlated with the findings that the functional activities of recombinant PDI2 and PDI4 were inhibited by nitazoxanide. Moreover, drastic changes were observed in VSP gene expression., Conclusions: These results suggest that resistance formation in Giardia against nitazoxanide and metronidazole is linked, and possibly mediated by, altered gene expression in drug-resistant strains compared with non-resistant strains of Giardia.

Neuron-specific enolase (NSE) represents the gamma gamma- and alpha gamma- isoforms of the dimeric glycolytic enzyme enolase. NSE is predominantly found in neurons and neuroendocrine cells and has proven to be a marker for tumors derived from these cells. It is widely accepted in the monitoring of patients with small cell lung cancer and is also of value as an aid in diagnosis. Recently it has become of interest in the monitoring of brain damage. Monoclonal antibodies against gamma-enolase were raised in mice and selected for optimal performance on the Cobas Core enzyme immunoassay system. The antibody combination of choice was MAb 18E5 for capturing and MAb 84B10 for detection which is accomplished by using a horseradish peroxidase conjugate and the substrate 3,3',5,5'-tetramethylbenzidine. The resulting assay is a one-step enzyme immunoassay of the sandwich type. It is performed on the fully automated Cobas Core immunoassay analyzer with a total assay time of 45 min. The sample volume is 10 microliters. Calibration is done by a 1-point recalibration using a lot-specific master calibration curve provided with the kit. The dynamic range is 0-200 ng/ml. The analytical detection limit (standard 0 + 2SD) of the Cobas Core NSE EIA II was 0.1 ng/ml. Intra- and interassay coefficients of variation were < 5% and < 6%, respectively. A Hook Effect was not observed up to a concentration of 20'000 ng/ml. Test results correlated closely with the well established polyclonal Cobas Core NSE EIA (r = 0.99). In summary, the Cobas Core NSE EIA II is a rapid, reliable and convenient test for measuring NSE in human serum.

Mitochondrial aspartate aminotransferase is a homodimeric protein with 2 x 402 amino acid residues. The enzyme in solution undergoes ligand-induced and syncatalytic conformational changes which appear to correspond to shifts in the equilibrium between the crystallographically defined open and closed conformation. In the closed conformation, the small domain of each subunit has rotated as a rigid body by 13 degrees and 14 degrees towards the large coenzyme-binding domain and has closed the active-site pocket. Molecular dynamics simulations at 300 K of 120-ps duration were started from the crystal structures of the unliganded pyridoxal form (open form) of the dimeric enzyme and the 2-methylaspartate-liganded closed form in which the 2-methyl group had been removed. Both structures contained the crystal water molecules and were placed in a 5-A shell of water. The rms fluctuations of the individual C alpha atoms during the simulations agreed well with the corresponding B factors of the crystal structures. Superposition of the initial structures and the average structures of the last 20 ps showed in both simulations extensive C alpha deviations in the case of the whole subunit but much smaller changes in the individual large and small domains, indicating a movement of the two domains relative to each other. In the simulation of the open form, superposition of the large domains made evident a displacement of the small domain towards its position in the closed crystal structure, which can be described by a rotation of the small domain by about 13 degrees around the twofold symmetry (z) axis. A significantly less extensive rearrangement of parts of the small domain, i.e. a rotation of about 5 degrees around the z axis, was observed in the simulation of the substrate-liganded enzyme (closed form) which, in contrast to the open form, showed only small conformational changes around the active site. In both simulations an additional rotation of the small domain by 9 degrees around the x axis occurred. The actual domain movement is estimated to occur in a time range at least two orders of magnitude larger than the simulation time of 120 ps. Apparently, the surface tension of the unrestrained nonspherical shell of water accelerates the simulated conformational change which, however, quite closely imitates the geometric features of the extensive movement of the small domains (each approximately 130 residues).

Three medical/surgical units in a Midwestern medical center introduced a two-way wireless communication system to test its effect on environmental noise, staff communication, timeliness of response to patient requests, nurse fatigue and job satisfaction. Data were collected through focus groups, surveys, pedometer studies and work sampling. Results provide for nurse managers the first objective evaluation of the potential of this new device and a framework for designing other nursing evaluations of the effects of a new technology.

Mitochondrial aspartate aminotransferase was selectively labeled with various maleimide-linked nitroxide spin labels at the conformationally sensitive Cys166. The mobility of the spin group was found to increase with increasing length of the spacer between the nitroxide and maleimide moiety. The label with the ethylcarbamoyl group, a spacer of intermediate length, responded sensitively to conformational changes of aspartate aminotransferase. The modification with this label decreased the enzymic activity to 30% of its initial value and increased the affinity for various substrates and inhibitors 5-10-fold. Identical ESR spectra were obtained for the pyridoxal and pyridoxamine form of the enzyme. These spectra are complex, consisting of an isotropic and at least two anisotropic components. The spectral complexity is attributed to different modes of interaction of the spin label with its local protein environment giving rise to different motional states. The same changes in the ESR spectra have been observed upon formation of the adsorption complex of the pyridoxal form with a competitive inhibitor and on formation of covalent intermediates of the transamination reaction. Essentially, the isotropic component is converted to a new anisotropic one as the local environment changes due to a conformational adaptation of aspartate aminotransferase. The ESR data are consistent with an equilibrium between two conformational states of the enzyme but inconsistent with individual protein conformations of the various intermediates of the transamination reaction. The two conformational states may be assigned to the open and closed conformations as defined by X-ray crystallography. In the adsorption complex of the pyridoxal enzyme, and in the covalent intermediates, the two-state equilibrium appears to be shifted towards the closed conformation in which the spin label is more rigidly bound, as also suggested by molecular dynamic simulations of the label modelled into aspartate aminotransferase. In contrast the formation of adsorption complexes between the pyridoxamine form and aspartate or maleate was not accompanied by the same shift of the conformational equilibrium.

The motives for the selection of primary patients in a neonatal intensive care unit were identified by 34 participants in a series of staff development programs on primary nursing. Motives were identified and ranked in order of importance. These included: (1) medical problems; (2) continuity of care; (3) impression of parents; (4) impression of the infant; (5) altruism; (6) nurses' self-esteem; and (7) impression of other staff. This paper considers the potential impact of these motives on the ability of the nurse to fulfill the expectations of primary nursing practice.

Absorption and circular dichroism spectra of stable enzyme-substrate intermediates of aspartate aminotransferase were recorded at subzero temperatures (down to -65 degrees C) in the cryosolvent water/methanol. The intermediates were formed either between the pyridoxal form of the enzyme and its amino acid substrates, or between the pyridoxamine form and its oxo acid substrates. Kd values determined by spectroscopic titration were very close to the Km values reported for the different substrates. The adsorption complex of the pyridoxal form was probably obtained on addition of cysteine sulfinate. This complex is characterized by an increased absorption at 430 nm together with a positive Cotton effect, as also observed in the case of the complex with the competitive inhibitor maleate indicating protonation of the internal aldimine. Addition of the substrates aspartate or glutamate to the pyridoxal form seemed to result in the direct accumulation of the external aldimine which showed a slight decrease in both the absorbance and the Cotton effect at 360 nm. Additionally, a bathochromic shift of 5 nm was observed in the case of glutamate. At 430 nm, only a minor increase in absorbance, but not in circular dichroism, was observed with aspartate, and no changes were found with glutamate and the substrate analog 2-methylaspartate, indicating a deprotonated external aldimine. Presumably, the ketimine intermediate was obtained on addition of the oxo acids 2-oxoglutarate or oxalacetate to the pyridoxamine form. The intermediate showed a slight bathochromic shift (2 nm) of the absorption band and decreased circular dichroism. On formation of the ketimine, a tyrosine residue, probably active-site Tyr225, becomes partly ionized. The finding that the external aldimine can probably be accumulated in the conversion of the pyridoxal to the pyridoxamine form with the natural substrates would confirm the proton abstraction at C alpha to be the rate-limiting step in the tautomerization, although with cysteine sulfinate, the formation of the external aldimine might contribute to the rate limitation. Accumulation of the ketimine in the reverse direction would indicate that the proton abstraction at C4' is rate-limiting in this half-reaction. The results demonstrate the feasibility of further structural investigations of true enzyme-substrate intermediates.

The challenge in meeting the caloric and metabolic demands of infants who are critically ill, debilitated, or neurophysically immature is discussed. Advances in nutrition that have greatly improved neonatal survival are presented. Intravenous nutrition is presented as an appropriate mode of therapy to provide or supplement nutritional support in neonates who cannot derive all the requirements from oral feedings.