Identification of photoactivated adenylyl cyclases in Naegleria australiensis and BLUF-containing protein in Naegleria fowleri.

Complete genome sequencing of Naegleria gruberi has revealed that the organism encodes polypeptides similar to photoactivated adenylyl cyclases (PACs). Screening in the N. australiensis genome showed that the organism also encodes polypeptides similar to PACs. Each of the Naegleria proteins consists of a "sensors of blue-light using FAD" domain (BLUF domain) and an adenylyl cyclase domain (AC domain). PAC activity of the Naegleria proteins was assayed by comparing sensitivities of Escherichia coli cells heterologously expressing the proteins to antibiotics in a dark condition and a blue light-irradiated condition. Antibiotics used in the assays were fosfomycin and fosmidomycin. E. coli cells expressing the Naegleria proteins showed increased fosfomycin sensitivity and fosmidomycin sensitivity when incubated under blue light, indicating that the proteins functioned as PACs in the bacterial cells. Analysis of the N. fowleri genome revealed that the organism encodes a protein bearing an amino acid sequence similar to that of BLUF. A plasmid expressing a chimeric protein consisting of the BLUF-like sequence found in N. fowleri and the adenylyl cyclase domain of N. gruberi PAC was constructed to determine whether the BLUF-like sequence functioned as a sensor of blue light. E. coli cells expressing a chimeric protein showed increased fosfomycin sensitivity and fosmidomycin sensitivity when incubated under blue light. These experimental results indicated that the sequence similar to the BLUF domain found in N. fowleri functioned as a sensor of blue light.

Photomanipulation of antibiotic susceptibility and biofilm formation of Escherichia coli heterologously expressing photoactivated adenylyl cyclase.

A cyaA-deficient Escherichia coli strain was transformed by a plasmid carrying the gene for BsPAC, a photoactivated adenylyl cyclase identified from a Beggiatoa sp., and was subjected to an antibiotic susceptibility assay and biofilm formation assay under a light or dark condition. Cells expressing BsPAC that were incubated under blue light (470 nm) were more susceptible to fosfomycin, nalidixic acid and streptomycin than were cells incubated in the dark. Cells expressing BsPAC formed more biofilms when incubated under the light than did cells cultured in the dark. We concluded from these observations that it is possible to determine the importance of cAMP in antibiotic susceptibility and biofilm formation of E. coli by photomanipulating the cellular cAMP level by the use of BsPAC. A site-directed mutant of BsPAC in which Tyr7 was replaced by Phe functioned even in the dark, indicating that Tyr7 plays an important role in photoactivation of BsPAC. Results of mutational analysis of BsPAC should contribute to an understanding of the molecular basis for photoactivation of the protein.

Activity in the premotor area related to bite force control--a functional near-infrared spectroscopy study.

The purpose of this study was to elucidate the influence of bite force control on oxygenated hemoglobin (OxyHb) levels in regional cerebral blood flow as an indicator of brain activity in the premotor area. Healthy right-handed volunteers with no subjective or objective symptoms of problems of the stomatognathic system or cervicofacial region were included. Functional near-infrared spectroscopy (fNIRS) was used to determine OxyHb levels in the premotor area during bite force control. A bite block equipped with an occlusal force sensor was prepared to measure clenching at the position where the right upper and lower canine cusps come into contact. Intensity of clenching was shown on a display and feedback was provided to the subjects. Intensity was set at 20, 50 and 80% of maximum voluntary teeth clenching force. To minimize the effect of the temporal muscle on the working side of the jaw, the fNIRS probes were positioned contralaterally, in the left region. The findings of this study are: activation of the premotor area with bite force control was noted in all subjects, and in the group analysis OxyHb in the premotor cortex was significantly increased as the clenching strengthened at 20, 50 and 80% of maximum voluntary clenching force. These results suggest there is a possibility that the premotor area is involved in bite force control.

Functional near-infrared spectroscopy study on primary motor and somatosensory cortex response to biting and finger clenching.

The purpose of this study was to compare the influence of biting and finger clenching intensity on the concentration of oxygenated hemoglobin (OxyHb) in regional cerebral blood flow (rCBF) as an indicator of brain activity in the primary motor (MI) and somatosensory (SI) cortices. Functional near-infrared spectroscopy (fNIRS) was used in 8 healthy subjects. Subjects were required to do biting (bite) and finger clenching (fclench) at 20, 50 and 80% of maximum force. To minimize the effect of temporal muscle activity on the working side of the jaw, the fNIRS probes were positioned contralaterally, in the left temporal region. Activation of MI and SI cortices with bite and fclench was noted in all subjects, irrespective of the intensity of bite and fclench. A significant increase was observed in OxyHb in MI and SI between 20% and both 50 and 80% intensity. In MI cortex, OxyHb showed a significant increase between 80% and both 20 and 50% fclench intensity. The results suggest that intensity of bite and fclench influences activation levels in MI and SI. Further, an activation was more obvious with bite than fclench.

Functional near-infrared spectroscopy study on primary motor and sensory cortex response to clenching.

The purpose of this study was to elucidate the influence of clenching and clenching intensity on oxygenated hemoglobin (OxyHb) levels in regional cerebral blood flow as an indicator of brain activity in the primary motor and sensory cortices. Functional near-infrared spectroscopy (fNIRS) was used to minimize the effect of clenching-associated muscle activity in eight healthy subjects. Subjects were required to clench at 20%, 50% and 80% of maximum clenching force. To minimize the effect of temporal muscle activity on the working side of the jaw, the fNIRS probes were positioned contralaterally, in the left temporal region. Activation of the primary motor and sensory cortices with clenching was noted in all subjects, irrespective of intensity of clenching. A significant increase was observed in OxyHb in the primary motor cortex between at 80% and both 20% and 50% clenching intensity. In the primary sensory cortex, OxyHb showed a significant increase between all levels of clenching intensity. The results suggest that clenching elicits activation of both the primary motor and sensory cortices, and that intensity of clenching influences activation levels in the brain.

In search of necessary mouthguard thickness. Part 1: From the viewpoint of shock absorption ability.

PURPOSE: To date, the minimum thickness required for a mouthguard has been assumed to be around 2 mm to 4 mm. However, this figure is based mostly on experience and is yet to be standardized. The purpose of this study is to determine the minimum thickness required to obtain sufficient energy absorption. METHODS: The thicknesses of the tested ethylene vinyl acetate) samples were 1, 2, 3, 4, 5, and 6 mm. The pendulum- type testing equipment used in the present study was also used in a series of earlier studies. Three types of sensors (strain gauge, accelerator, and load cell) and two different impact objects (a steel ball and baseball) were used. RESULTS: The results showed that all the above-mentioned mouthguard thicknesses reduced shocks for all the three types of sensors and both types of impact objects; little difference was observed between sensors and clear results were obtained for the steel ball. An improvement in the energy absorption was observed with an initial increase in the thickness. However, a further increase in the thickness from 4 mm to 5 mm and 6 mm tended to yield a smaller improvement in energy absorption. CONCLUSION: Within the limitations of this study, from the viewpoint of energy absorption ability, the minimum thickness required for a mouthguard is 4 mm, which is generally too large from the viewpoint of player comfort. This finding indicates the necessity of improving the impact absorption ability of mouthguards by considering new designs and developing new materials.

A vacuum technique to increase anterior thickness of athletic mouthguards to achieve a full-balanced occlusion.

A full-balanced occlusion is essential for mouthguards. It has been reported that a balanced occlusion for upper and lower anterior teeth is essential for prevention of injuries occurring to the maxillary anterior teeth and alveolar bone caused by horizontal direct impact. The support of the mandibular teeth through the mouthguard is critical to prevent maxillary front tooth injury from a direct impact force. However, some vacuum mouthguard designs may not achieve a full-balanced occlusion. For example, when a player has a malocclusion, an elongated molar or premolar tooth, an open bite, a large over jet or a maxillary protrusion. An improved vacuum fabrication method is necessary to obtain full balanced occlusion in these cases as opposed to conventional vacuum type single-layer mouthguard technique.

A novel family of repetitive DNA sequences amplified site-specifically on the W chromosomes in Neognathous birds.

A novel family of repetitive DNA sequences was molecularly cloned from ApaI-digested genomic DNA of two Galliformes species, Japanese quail (Coturnix japonica) and guinea fowl (Numida meleagris), and characterized by chromosome in-situ hybridization and filter hybridization. Both the repeated sequence elements produced intensely painted signals on the W chromosomes, whereas they weakly hybridized to whole chromosomal regions as interspersed-type repetitive sequences. The repeated elements of the two species had high similarity of nucleotide sequences, and cross-hybridized to chromosomes of two other Galliformes species, chicken (Gallus gallus) and blue-breasted quail (Coturnix chinensis). The nucleotide sequences were conserved in three other orders of Neognathous birds, the Strigiformes, Gruiformes and Falconiformes, but not in Palaeognathous birds, the Struthioniformes and Tinamiformes, indicating that the repeated sequence elements were amplified on the W chromosomes in the lineage of Neognathous birds after the common ancestor diverged into the Palaeognathae and Neognathae. They are components of the W heterochromatin in Neognathous birds, and a good molecular cytogenetic marker for estimating the phylogenetic relationships and for clarifying the origin of the sex chromosome heterochromatin and the process of sex chromosome differentiation in birds.

Does hard insertion and space improve shock absorption ability of mouthguard?

Mouthguards are expected to reduce sports-related orofacial injuries. Numerous studies have been conduced to improve the shock absorption ability of mouthguards using air cells, sorbothane, metal wire, or hard material insertion. Most of these were shown to be effective; however, the result of each study has not been applied to clinical use. The aim of this study was to develop mouthguards that have sufficient prevention ability and ease of clinical application with focus on a hard insertion and space. Ethylene vinyl acetate (EVA) mouthguard blank used was Drufosoft and the acrylic resin was Biolon (Dreve-Dentamid GMBH, Unna, Germany). Three types of mouthguard samples tested were constructed by means of a Dreve Drufomat (Type SO, Dreve-Dentamid) air pressure machine: the first was a conventional laminated type of EVA mouthguard material; the second was a three layer type with acrylic resin inner layer (hard-insertion); the third was the same as the second but with space that does not come into contact with tooth surfaces (hard + space). As a control, without any mouthguard condition (NOMG) was measured. A pendulum type impact testing machine with interchangeable impact object (steel ball and baseball) and dental study model (D17FE-NC.7PS, Nissin, Tokyo, Japan) with the strain gages (KFG-1-120-D171-11N30C2: Kyowa, Tokyo, Japan) applied to teeth and the accelerometer to the dentition (AS-A YG-2768 100G, Kyowa) were used to measure transmitted forces. Statistical analysis (anova, P < 0.01) showed significant differences among four conditions of NOMG and three different mouthguards in both objects and sensor. About acceleration: in a steel ball which was a harder impact object, shock absorption ability of about 40% was shown with conventional EVA and hard-insertion and about 50% with hard + space. In a baseball that was softer compared with steel ball, a decrease rate is smaller, reduction (EVA = approximately 4%, hard-insertion = approximately 12%, hard + space = approximately 25%) was admitted in the similar order. A significant difference was found with all the combinations except for between EVA and hard-insertion with steel ball (Tukey test). About distortion: both buccal and lingual, distortions had become small in order of EVA, hard-insertion, and hard + space, too. The decrease rate is larger than acceleration, EVA = approximately 47%, hard-insertion = 80% or more, and hard +space = approximately 98%, in steel ball. EVA = approximately 30%, hard-insertion = approximately 75%, and hard + space = approximately 98% in baseball. And a significant difference was found with all the combinations (Tukey test). Especially, hard + space has decreased the distortion of teeth up to several percentages. Acceleration of the maxilla and distortions of the tooth became significantly smaller when wearing any type of mouthguard, in both impact objects. But the effect of mouthguard was clearer in the distortion of the tooth and with steel ball. Considering the differences of mouthguards, the hard-insertion and the hard + space had significantly greater buffer capacity than conventional EVA. Furthermore, hard + space shows quite high shock absorption ability in the tooth distortion. Namely, hard + space has decreased the distortion of teeth up to several percentages in both impact objects.

[Mouthguard effect of tooth distortion during clenching].

PURPOSE: A mouthguard can protect stomatognathic systems from traumatic damage. However, severe occlusal wear of teeth and loss of teeth have often been found in players clinically. These problems might originate in strong clenching during sports. Although it is thought that a mouthguard may be effective for these types of clenching, the relation between mouthguards and clenching has not been sufficiently examined. In this study, the effect of a mouthguard (Drufosoft 3mm, EVA) on tooth distortion caused by clenching was measured and examined at three different clenching strengths. METHODS: As a test tooth, a lower first molar was selected. A strain gauge applied to the outer surface of the buccal cusp was used to measure the distortion. A muscle balance monitor (GC) was used to regulate clenching strengths (10, 50, and 100%). The maximum-effort clenching without a mouthguard was assumed to be the 100% clenching strength. Measurements were conducted with or without mouthguard. A maximum value during clenching was assumed to be date of distortion by using analytical software AcquKnowledge (BIOPAC System Inc.). Statistical analysis software SPSS (SPSS Japan Inc.) was used for the Mann-Whitney test. RESULTS: 1. The tooth distortion by clenching, regardless of the presence of the mouthguard, increased as clenching power strengthened, from 10, 50 to 100%. 2. The tooth distortion, regardless of strength of clenching, was decreased by wearing the mouthguard in all subjects. At 50 and 100% clenching, it was decreased significantly by the mouthguard in all subjects. CONCLUSIONS: Mouthguards decreaseed the tooth distortion caused by clenching. Therefore, a mouthguard may prevent not only traumatic injuries in contact sports but also damage to teeth and periodontal tissues and so on, which occur due to frequent strong clenching in many sports.

Are all mouthguards the same and safe to use? The influence of occlusal supporting mouthguards in decreasing bone distortion and fractures.

The safety benefits of mouthguards have been demonstrated in many studies, with many authors and sports dentists strongly recommending the wearing of mouthguards. However, wearing a mouthguard with incorrect occlusion might cause a variety of problems. It comes as no surprise that a traumatic blow to the chin, while wearing an insufficient mouthguard lacking anterior contact, can result in severe distortions to the mandibular bone, and bone fractures. The aim of this study was to clarify how ineffective insufficient occlusal supporting mouthguards are and how dangerous they can be to use. Consequently, in this study, occlusal supportive areas were varied and accelerations of head and distortions of the mandible were measured using an artificial skull model and a pendulum impact device. As a result, the distortions of the mandible tended to increase as the supported area decreased. On the contrary, accelerations of the head decreased as the occlusion part decreased. Thus, a lot of impact energy was consumed in the distortion of the mandible; accordingly, it seemed that only a little destructive energy was transferred to the head. From this study, it would seem that wearing a mouthguard, which is insufficient in the occlusion, has the potential of causing a bone fracture of the mandible. Consequently, mouthguards should have proper occlusion.

The Bh (black at hatch) gene that causes abnormal feather pigmentation maps to chromosome 1 of the Japanese quail.

Japanese quail embryos normally have longitudinal black and brown stripes formed by colored feather buds on their back whereas an autosomal dominant mutation, black at hatch (Bh), disrupts this pigmentation pattern by causing overall black and brown coating in heterozygotes and homozygotes, respectively. These phenotypes of the Bh mutant embryos suggest that the Bh locus plays an important role in the pigment pattern formation of plumage, but its genetic origin, including cloning of the responsible gene, has been insufficiently studied. In this study, we adapted genetically directed representational difference analysis with elimination of excessive clones (GDRDA-WEEC) to Bh quails and isolated two genetic markers linked to the Bh locus as DNA fragments. Cytogenetic study by fluorescence in situ hybridization (FISH) of the DNA fragments used as probes demonstrated that the marker loci were located in the same region on the long arm of chromosome 1. Close genetic linkage between the Bh and the marker loci, and the chromosomal location of the latter suggested that the Bh locus is located on the long-arm of chromosome 1 of the Japanese quail.