Improved method for ex ovo-cultivation of developing chicken embryos for human stem cell xenografts

The characterization of human stem cells for the usability in regenerative medicine is particularly based on investigations regarding their differentiation potential in vivo. In this regard, the chicken embryo model represents an ideal model organism. However, the access to the chicken embryo is only achievable by windowing the eggshell resulting in limited visibility and accessibility in subsequent experiments. On the contrary, ex ovo-culture systems avoid such negative side effects. Here, we present an improved ex ovo-cultivation method enabling the embryos to survive 13 days in vitro. Optimized cultivation of chicken embryos resulted in a normal development regarding their size and weight. Our ex ovo-approach closely resembles the development of chicken embryos in ovo, as demonstrated by properly developed nervous system, bones, and cartilage at expected time points. Finally, we investigated the usability of our method for trans-species transplantation of adult stem cells by injecting human neural crest-derived stem cells into late Hamburger and Hamilton stages (HH26-HH28/E5-E6) of ex ovo-incubated embryos. We demonstrated the integration of human cells allowing experimentally easy investigation of the differentiation potential in the proper developmental context. Taken together, this ex ovo-method supports the prolonged cultivation of properly developing chicken embryos enabling integration studies of xenografted mammalian stem cells at late developmental stages.

Sensory enhancement, a pilot perceptual study of subdermal magnetic implants

Subdermal magnetic implants originated as an art form in the world of body modification. To date an in depth scientific analysis of the benefits of this implant has yet to be established. This research explores the concept of sensory extension of the tactile sense utilising this form of implantation. This relatively simple procedure enables the tactile sense to respond to static and alternating magnetic fields. This is not to say that the underlying biology of the system has changed; i.e. the concept does not increase our tactile frequency response range or sensitivity to pressure, but now does invoke a perceptual response to a stimulus that is not innately available to humans. Within this research two social surveys have been conducted in order to ascertain one, the social acceptance of the general notion of human enhancement, and two the perceptual experiences of individuals with the magnetic implants themselves. In terms of acceptance to the notion of sensory improvement (via implantation) ~39% of the general population questioned responded positively with a further ~25% of the respondents answering with the indecisive response. Thus with careful dissemination a large proportion of individuals may adopt this technology much like this if it were to become available for consumers. Interestingly of the responses collected from the magnetic implants survey ~60% of the respondents actually underwent the implant for magnetic vision purposes. The main contribution of this research however comes from a series of psychophysical testing. In which 7 subjects with subdermal magnetic implants, were cross compared with 7 subjects that had similar magnets superficially attached to their dermis. The experimentation examined multiple psychometric thresholds of the candidates including intensity, frequency and temporal. Whilst relatively simple, the experimental setup for the perceptual experimentation conducted was novel in that custom hardware and protocols were created in order to determine the subjective thresholds of the individuals. Abstract iv The overall purpose of this research is to utilise this concept in high stress scenarios, such as driving or piloting; whereby alerts and warnings could be relayed to an operator without intruding upon their other (typically overloaded) exterior senses (i.e. the auditory and visual senses). Hence each of the thresholding experiments were designed with the intention of utilising the results in the design of signals for information transfer. The findings from the study show that the implanted group of subjects significantly outperformed the superficial group in the absolute intensity threshold experiment, i.e. the implanted group required significantly less force than the superficial group in order to perceive the stimulus. The results for the frequency difference threshold showed no significant difference in the two groups tested. Interestingly however at low frequencies, i.e. 20 and 50 Hz, the ability of the subjects tested to discriminate frequencies significantly increased with more complex waveforms i.e. square and sawtooth, when compared against the typically used sinewave. Furthermore a novel protocol for establishing the temporal gap detection threshold during a temporal numerosity study has been established in this thesis. This experiment measured the subjects’ capability to correctly determine the number of concatenated signals presented to them whilst the time between the signals, referred to as pulses, tended to zero. A significant finding was that when altering the length of, the frequency of, and the number of cycles of the pulses, the time between pulses for correct recognition altered. This finding will ultimately aid in the design of the tactile alerts for this method of information transfer. Preliminary development work for the use of this method of input to the body, in an automotive scenario, is also presented within this thesis in the form of a driving simulation. The overall goal of which is to present warning alerts to a driver, such as rear-to-end collision, or excessive speeds on roads, in order to prevent incidents and penalties from occurring. Discussion on the broader utility of this implant has been presented, reflecting on its potential use as a basis for vibrotactile, and sensory substitution, devices. This discussion furthers with postulations on its use as a human machine interface, as well as how a similar implant could be used within the ear as a hearing aid device.

Effects of aphid infestation on Cd and Zn concentration in wheat

The effect of infestation by the aphid Metopolophium dirhodum on the concentration and mass partitioning of Cd and Zn was studied in wheat plants. Results show that infestation did not affect the concentration of either metal in the roots or shoots of wheat, but elevated concentrations in the ears. This appeared to result from the concentration of metal in the smaller ear mass of infested plants. Infestation did not significantly affect the mass partitioning of either metal in any part of the plant, some 10% of both metals being allocated to the roots. However, the two metals contrasted in their partitioning in the aerial parts of the plant, with ca. 60% of Cd mass partitioned in the shoots and ca. 67% of Zn mass partitioned in the ears. The possible effects of infestation on the transfer of Cd and Zn from the soil to cereal aphids are discussed.

Bmp4 regulates chick Ebf2 and Ebf3 gene expression in somite development

The chick Early B-cell Factor-2 and 3 (cEbf2 and cEbf3) genes are members of EBF family of helix loop helix transcription factors. The expression, regulation and importance of these genes have been extensively studied in lymphatic, nervous and muscular tissues. Recently, a new role for some members of EBF in bone development has been investigated. However, the expression profile and regulation in the axial skeleton precursor, the somite, have yet to be elucidated. Therefore, this study was aimed to investigate the expression and regulation of cEbf2 and cEbf3 genes in the developing chick embryo somite from HH4 to HH28. The spatiotemporal expression study revealed predominant localization of cEbf2 and cEbf3 in the lateral sclerotomal domains and later around vertebral cartilage anlagen of the arch and the proximal rib. Subsequently, microsurgeries, ectopic gene expression experiments were performed to analyze which tissues and factors regulate cEbf2 and cEbf3 expression. Lateral barriers experiments indicated the necessity for lateral signal(s) in the regulation of cEbf2 and cEbf3 genes. Results from tissue manipulations and ectopic gene expression experiments indicate that lateral plate-derived Bmp4 signals are necessary for the initiation and maintenance of cEbf2 and cEbf3 genes in somites. In conclusion, cEbf2 and cEbf3 genes are considered as lateral sclerotome markers which their expression is regulated by Bmp4 signals from the lateral plate mesoderm.

Paracrine effects of TLR4-polarised mesenchymal stromal cells are mediated by extracellular vesicles

Mesenchymal stromal cells (MSCs) are adult stem cells able to give rise to bone, cartilage and fat cells. In addition, they possess immunomodulatory and immunosuppressive properties that are mainly mediated through secretion of extracellular vesicles (EVs). In a previous issue of Journal of Translational Medicine, Ti and colleagues demonstrated that preconditioning of MSCs with bacterial lipopolysaccharides results in secretion of EVs that can polarise mac‑ rophages towards anti-inflammatory M2 phenotype. Moreover, the authors suggest that EVs of lipopolysaccharide (LPS)-treated MSCs are superior to EVs of untreated MSCs concerning their ability to support wound healing. Our commentary critically discusses parallel efforts of other laboratories to generate conditioned media from stem cells for therapeutic applications, and highlights impact and significance of the study of Ti et al. Finally, we summarise its limitations and spotlight areas that need to be addressed to better define the underlying molecular mechanisms.

Effects of rain shelter or simulated rain during grain filling and maturation on subsequent wheat grain quality in the UK

The effects of simulated additional rain (ear wetting, 25 mm) or of rain shelter imposed at different periods after anthesis on grain quality at maturity and the dynamics of grain filling and desiccation were investigated in UK field-grown crops of wheat (Triticum aestivum L., cvar Tybalt) in 2011 and in 2012 when June–August rainfall was 255.0 and 214.6 mm, respectively, and above the decadal mean (157.4 mm). Grain filling and desiccation were quantified well by broken-stick regressions and Gompertz curves, respectively. Rain shelter for 56 (2011) or 70 d (2012) after anthesis, and to a lesser extent during late maturation only, resulted in more rapid desiccation and hence progress to harvest maturity whereas ear wetting had negligible effects, even when applied four times. Grain-filling duration was also affected as above in 2011, but with no significant effect in 2012. In both years, there were strong positive associations between final grain dry weight and duration of filling. The treatments affected all grain quality traits in 2011: nitrogen (N) and sulphur (S) concentrations, N:S ratio, sodium dodecyl sulphate (SDS) sedimentation volume, Hagberg Falling Number (HFN), and the incidence of blackpoint. Only N concentration and blackpoint were affected significantly by treatments in 2012. Rain shelter throughout grain filling reduced N concentration, whereas rain shelter reduced the incidence of blackpoint and ear wetting increased it. In 2011, rain shelter throughout reduced S concentration, increased N:S ratio and reduced SDS. Treatment effects on HFN were not consistent within or between years. Nevertheless, a comparison between the extreme treatment means in 2012 indicated damage from late rain combined with ear wetting resulted in a reduction of c. 0.7 s in HFN/mm August rainfall, whilst that between samples taken immediately after ear wetting at harvest maturity or 7 d later suggested recovery from damage to HFN upon re-drying in planta. Hence, the incidence of blackpoint was the only grain quality trait affected consistently by the diverse treatments. The remaining aspects of grain quality were comparatively resilient to rain incident upon developing and maturing ears of cvar Tybalt. No consistent temporal patterns of sensitivity to shelter or ear wetting were detected for any aspect of grain quality.

Evaluation of directional microphone drift in digital hearing aids

The occurrence of directional microphone drift following hearing aid use has been infrequently examined. This study uses the front-to-side ratio to evaluate changes in directional microphone output from new behind-the-ear hearing aids and following approximately three months of hearing aid use. Results indicate no overall significant differences in the front-to-side ratio between initial and follow-up measurements.

Vitamin A deficiency in preschool children of Recife, Northeast of Brazil

Vitamin A deficiency in preschool children of Recife, Northeast of Brazil. The purpose of the study was to evaluate the extent of vitamin A deficiency (VAD) among preschool children in the city of Recife, Northeast Brazil. The sample comprised 344 children of both sexes, 24 to 60 months old, in 18 public day care centres in the city of Recife, in 2007. The nutritional status of vitamin A was assessed by biochemical (serum retinol) and dietetic (vitamin A rich-food consumption) indicators and the pondo-stature status through anthropometric indicators weight-for-age, height-for-age and weight-for-height. The prevalence of hyporetinolemia (<0.70 mu mol / L) was 7.7% (IC95% 4.88 - 11.81), which characterizes the VAD as a light-type public health problem, according to World Health Organization criteria. On the other hand, 29.6% (IC95% 24.22 - 35.63) of children had acceptable or marginal levels (0.70 to 1.04 mu mol/L) of retinol. Regarding the vitamin A rich-food intake, values below the EAR (Estimated Average Requirement) - 210 mu g/day for children of 1 to 3 years old and 275 mu g/day for children of 4 to 8 years old - were 8.1% and 21.3% respectively. The prevalence of anthropometrical deficits (<-2 scores -Z) in preschool children were 2.5% for the indicator weight-for-age, 8.6% for height-for-age and 1.5% for weight-for-height. The research findings point out to the importance of institutionalization for the appropriate nutritional status of children and maintenance of adequate reserves of vitamin A. However, more studies are needed focusing on non-institutionalized preschool, or children living outside the privileged environment of public day care centres.

Chronic Suppurative Otitis Media in Cleft Palate: Microorganism Etiology and Susceptibilities

Objective: To investigate the microbial etiology of suppurative chronic otitis media (SCOM) in patients with complete cleft lip and palate and isolated cleft palate and to determine the sensitivity of isolated microorganisms to antibiotics by drug diffusion from impregnated discs in agar and the minimum inhibitory concentration of each drug to these microorganisms by drug dilution in agar. Design/Patients: Effusion samples of SCOM obtained from 40 patients with cleft lip and palate registered at the Hospital for Rehabilitation of Craniofacial Anomalies, University of Sao Paulo, at Bauru, Brazil, were bacteriologically analyzed by cultures. The isolated bacteria were submitted to an in vitro susceptibility test to clinically used drugs. Results: Positive cultures were obtained in 100% of studied cases. Among the 57 strains observed, the most frequent were Pseudomonas aeruginosa (35%), Staphylococcus aureus (15.5%), Enterococcus faecalis (14%), and Proteus mirabilis (12%). The frequency of Gram-negative bacilli (enterobacteriaceae and nonfermentative bacilli) was 67%. Pseudomonas aeruginosa presented the highest sensitivity to ciprofloxacin, and enterobacteriaceae exhibited the highest sensitivity to gentamicin. The strains of S. aureus and E. faecalis presented the highest sensitivity to imipenem and sulfamethoxazole/trimethoprim, respectively. Conclusion: Patients with cleft lip and palate presenting with SCOM exhibited 100% positive cultures, with the highest frequency of Pseudomonas and enterobacteriaceae. With regard to the action of antibiotics, imipenem was effective against the four species of isolated microorganisms, followed by ciprofloxacin, which was effective against 75% of isolated species.

Moho map of South America from receiver functions and surface waves

We estimate crustal structure and thickness of South America north of roughly 40 degrees S. To this end, we analyzed receiver functions from 20 relatively new temporary broadband seismic stations deployed across eastern Brazil. In the analysis we include teleseismic and some regional events, particularly for stations that recorded few suitable earthquakes. We first estimate crustal thickness and average Poisson`s ratio using two different stacking methods. We then combine the new crustal constraints with results from previous receiver function studies. To interpolate the crustal thickness between the station locations, we jointly invert these Moho point constraints, Rayleigh wave group velocities, and regional S and Rayleigh waveforms for a continuous map of Moho depth. The new tomographic Moho map suggests that Moho depth and Moho relief vary slightly with age within the Precambrian crust. Whether or not a positive correlation between crustal thickness and geologic age is derived from the pre-interpolation point constraints depends strongly on the selected subset of receiver functions. This implies that using only pre-interpolation point constraints (receiver functions) inadequately samples the spatial variation in geologic age. The new Moho map also reveals an anomalously deep Moho beneath the oldest core of the Amazonian Craton.

New Species of Earless Lizard Genus Heterodactylus (Squamata: Gymnophthalmidae) from the Highlands of Chapada Diamantina, State of Bahia, Brazil

A new species of Heterodactylus is described based on two specimens obtained in the highlands of Chapada Diamantina, state of Bahia, Brazil. The new lizard is characterized by very elongate body and tail, absence of external ear opening, presence of moveable eyelids, absence of prefrontals and frontoparietals, a vestigial interparietal, 37-39 dorsal and 27-29 ventral transverse scale rows, 23-25 scales around midbody, six gular scale rows, and 10-11 and 14-15 fourth finger and fourth toe infradigital lamellae, respectively. The new species is most similar to Heterodactylus lundii from which it differs by the absence of contact between frontal and interparietal, by having wider than long parietals, smooth posterior dorsal scales, posterior ventral scales almost twice longer than wide, a lower number of scales around midbody, last supralabial in contact with the granules of the ear depression, and a more elongate body. The new species occurs about 1,100 km north of the northernmost known record of H. lundii. Species of Heterodactylus seem to be restricted to areas of cold climates associated with high latitudes and mountainous areas of eastern Brasil.

Potamotrygon tigrina, a new species of freshwater stingray from the upper Amazon basin, closely related to Potamotrygon schroederi Fernandez-Yepez, 1958 (Chondrichthyes: Potamotrygonidae)

A new species of Neotropical freshwater stingray, family Potamotrygonidae, is described from the Rio Nanay in the upper Rio Amazonas basin of Peru. Potamotrygon tigrina, n. sp., is easily distinguished from all congeners by its conspicuous dorsal disc coloration, composed of bright yellow to orange vermiculations strongly interwoven with a dark-brown to deep-black background. Additional features that in combination diagnose P. tigrina, n. sp., include the presence of a single angular cartilage, low and not closely grouped dorsal tail spines, and coloration of tail composed of relatively wide and alternating bands of creamy white and dark brown to black. Potamotrygon tigrina is closely related to Potamotrygon schroederi Fernandez-Yepez, 1958, which occurs in the Rio Negro (Brazil) and Rio Orinoco (Venezuela, Colombia). Both species are very similar in proportions and counts, and share features hypothesized to be derived within Potamotrygonidae, related to their specific angular cartilage morphology, distal tail color, dorsal tail-spine pattern, and ventral lateral-line system. To further substantiate the description of P. tigrina, n. sp., we provide a redescription of P. schroederi based on material from the Rio Negro (Brazil) and Rio Orinoco (Venezuela). Specimens from the two basins differ in number of vertebral centra and slightly in size and frequency of rosettes on dorsal disc, distinctions that presently do not warrant their specific separation. Potamotrygon tigrina is frequently commercialized in the international aquarium trade but virtually nothing is known of its biology or conservation status.

A new microteiid lizard of the genus Acratosaura (Squamata: Gymnophthalmidae) from Serra do Sincora, State of Bahia, Brazil

A new species of the genus Acratosaura is described on the basis of two specimens obtained at the ""campos rupestres"" (rocky meadows) near Mucuge municipality, state of Bahia, Brazil. Acratosaura spinosa sp. nov., has ear openings and eyelids, pentadactyl limbs lacking a claw on first toe, a single frontonasal, prefrontals, frontoparietals, parietals, interparietals, occipitals, no collar fold, three pairs of genials, three supraoculars and three superciliaries and is further characterized by presenting 28-31 rows of strongly keeled, lanceolate and imbricate dorsal scales, sides of the neck with keeled scales, striate temporal scales, four longitudinal and 17-18 transverse rows of smooth ventral scales, 28-30 scales around the body, 14 and 19-20 infradigital lamellae under finger IV and toe IV, respectively, 13 total preanal and femoral pores in male, absent in female. The new species differs strikingly in color pattern from its only congener A. mentalis. The two species occur sympatricaly in the high altitude open habitats near the type locality.

Prevalence of GJB2 (Connexin-26) and GJB6 (Connexin-30) Mutations in a Cohort of 300 Brazilian Hearing-Impaired Individuals: Implications for Diagnosis and Genetic Counseling

Objective: Hereditary nonsyndromic deafness is an autosomal recessive condition in about 80% of cases, and point mutations in the GJB2 gene (connexin 26) and two deletions in the GJB6 gene (connexin 30), del(GJB6-D13S1830) and del(GJB6-D13S1854), are reported to account for 50% of recessive deafness, Aiming at establishing the frequencies of GJB2 mutations and GJB6 deletions in the Brazilian population, we screened 300 unrelated individuals with hearing impairment, who were not affected by known deafness related syndromes. Methods: We firstly screened the most frequently reported mutations, c.35delG and c.167delT in the GJB2 gene, and del(GJB6-D13S1830) and del(GJB6-D13S1854) in the GJB6 gene, through specific techniques. The detected c.35delG and c.167delT mutations were validated by sequencing. Other mutations in the GJB2 gene were screened by single-strand conformation polymorphism and the coding region was sequenced when abnormal patterns were found. Results: Pathogenic mutations in GJB2 and GJB6 genes were detected in 41 individuals (13.7%), and 80.5% (33/41) presented these mutations in homozygosis or compound heterozygosis, thus explaining their hearing defect. The c.35delG in the GJB2 gene was the most frequent mutation (37/300; 12.4%), detected in 23% familial and 6.2% the sporadic cases. The second most frequent mutation (1%; 3/300) was the del(GJB6- D13S1830), always found associated with the c.35delG mutation. Nineteen different sequence variations were found in the GJB2 gene. In addition to the c.35delG mutation, nine known pathogenic alterations were detected 0 67delT, p.Trp24X, p.Val37lle, c.176_191del16, c.235delC, p.Leu90Pro, p.Arg127His, c.509insA, and p.Arg184Pro, Five substitutions had been previously considered benign polymorphisms: c.-15C>T, p.Val27lle, p.Met34hr, p.Ala40Ala, and p.Gly160Ser. Two previously reported Mutations of unknown pathogenicity were found (p.Lys168Arg, and c.684C>A), and two novel substitutions, p.Leu81Val (c.G241C) and p.Met195Val (c.A583G), both in heterozygosis without an accompanying mutation in the other allele. None of these latter four variants of undefined status was present in a sample of 100 hearing controls. Conclusions: The present study demonstrates that Mutations in the GJB2 gene and del(GJB6 D13S1830) are important causes of hearing impairment in Brazil, thus justifying their screening in a routine basis. The diversity of variants in our sample reflects the ethnic heterogeneity of the Brazilian population.

Revision of the Asian spider genus Pandava Lehtinen (Araneae: Titanoecidae): description of five new species and first record of Titanoecidae from Africa

The spider genus Pandava Lehtinen previously included two species, P. laminata (Thorell) and P. hunanensis Yin and Bao, known from Asia and the Pacific Islands. The genus is diagnosed by the reduced tegular process on the male palp and the anterior position of the copulatory openings on the female epigynum. In this paper, we present updated descriptions for the known species of Pandava and we describe five new species: Pandava shiva sp. nov. from Pakistan, Pandava sarasvati sp. nov. from Myanmar; Pandava ganesha sp. nov., Pandava kama sp. nov. and Pandava ganga sp. nov., all from India. We also update the generic distribution including the first records of Titanoecidae from Africa.