A Case of Dysphagia and Dysarthria Improved by Flexible-palatal Lift/augmentation Combination Prosthesis.

Background: Palatal augmentation prosthesis (PAP) and palatal lift prosthesis (PLP) have been used to improve dysphagia and dysarthria. However, to date, there are few reports on their combined use. We report a quantitative evaluation of the effectiveness of a flexible-palatal lift/augmentation combination prosthesis (fPL/ACP) based on videofluoroscopic swallowing study (VFSS) and speech intelligibility testing. Case: An 83-year-old woman was admitted to our hospital with a hip fracture. She developed aspiration pneumonia at 1 month after partial hip replacement. Oral motor function tests revealed a motor deficit of the tongue and soft palate. VFSS showed delayed oral transit, nasopharyngeal reflux, and excessive pharyngeal residue. The cause of her dysphagia was assumed to be pre-existing diffuse large B-cell lymphoma and sarcopenia. To improve the dysphagia, an fPL/ACP was fabricated and applied. It improved the patient's oral and pharyngeal swallowing and speech intelligibility. In addition to prosthetic treatment, rehabilitation and nutritional support allowed her to be discharged. Discussion: The effects of fPL/ACP in the present case were similar to those of flexible-PLP and PAP. f-PLP assists in elevation of the soft palate and improved the nasopharyngeal reflux and hypernasal speech. PAP promotes tongue movement and results in improved oral transit and speech intelligibility. Therefore, fPL/ACP may be effective in patients with motor deficits in both the tongue and soft palate. To maximize the effect of the intraoral prosthesis, a transdisciplinary approach with concurrent swallowing rehabilitation, nutritional support, and physical and occupational therapy is necessary.

Effect of dentures on pharyngeal swallowing function in patients with dysphagia.

AIM: Dentures play an important role in improving masticatory and oropharyngeal swallowing functions in some edentulous patients without dysphagia. However, few studies have been conducted on patients with dysphagia. This study investigated the effect of dentures on pharyngeal swallowing function in patients with dysphagia. METHODS: Older inpatients with dysphagia who used well-fitting dentures were included in the study. Videofluoroscopic swallowing study findings with and without dentures were compared. Pharyngeal residue and area as spatial, the distance between the maxilla and mandible, hyoid bone/laryngeal displacement, and upper esophageal sphincter opening as kinematics, oral/pharyngeal transit time as temporal measurements, and patient-reported symptoms were evaluated. The primary outcome was the pharyngeal residue measured using the normalized residue ratio scale. Comparisons were made using the paired t-test, Wilcoxon signed-rank test and Fisher's exact test. RESULTS: The mean age of the 27 participants was 86.1 ± 6.8 years. The vallecular residue was more in those without dentures (with dentures: 0.01 [0-0.02], without dentures: 0.03 [0-0.08]; P = 0.003). The pyriform sinus residue showed no significant difference. Denture removal significantly increased the pharyngeal area. The distance between the maxilla and mandible decreased in the absence of dentures, and other kinematic measurements showed no significant differences. Oral/pharyngeal transit time was prolonged without dentures. CONCLUSIONS: Morphological changes caused by the removal of dentures led to pharyngeal expansion, which may result in increased vallecular residue. A treatment plan that considers the effect of dentures on pharyngeal swallowing function may provide rehabilitation that is more effective. Geriatr Gerontol Int 2021; 21: 907-912.

Seven years of progress in determining fungal diversity and characterization of fungi isolated from the Japanese Experiment Module KIBO, International Space Station.

The International Space Station (ISS) is a closed facility that orbits the earth carrying not only its crew but also microorganisms. We have participated in microbiota analysis projects for the Japanese Experiment Module KIBO (ISS; operations nomenclature: Microbe-I, II, III, and IV) and were in charge of fungal screening. The interior of KIBO was sampled using swabs and microbe detection sheets (MDSs) for fungal detection. The dominant genera obtained by culture were Aspergillus and Penicillium. DNA analyses of the fungal biota using a clone library showed that KIBO was dominated by Malassezia, a fungal inhabitant of human skin. Three fungal species, Aspergillus sydowii, Penicillium palitans, and Rhodotorula mucilaginosa, which grew under microgravity in KIBO were observed under a field emission-scanning electron microscope on the ground. No novel phenotypic characteristics were noted. The results of antifungal susceptibility testing of all isolates did not differ significantly from previous reports of corresponding fungi. In Microbe-I (August 2009), MDSs were culture negative, while in the next stages the CFU of MDSs were 10 for Microbe-II (February 2011), 24 for Microbe-III (October 2012), and 151 for Microbe-IV (February 2015). These results indicated that fungi inside KIBO are increasing and expanding over time, and therefore continuous surveillance is crucial.

Characterization of the translation elongation factor 1-α gene in a wide range of pathogenic Aspergillus species.

PURPOSE: We aimed to evaluate the resolving power of the translation elongation factor (TEF)-1α gene for phylogenetic analysis of Aspergillus species. METHODOLOGY: Sequences of 526 bp representing the coding region of the TEF-1α gene were used for the assessment of levels of intra- and inter-specific nucleotide polymorphism in 33 species of Aspergillus, including 57 reference, clinical and environmental strains. RESULTS: Analysis of TEF-1α sequences indicated a mean similarity of 92.6 % between the species, with inter-species diversity ranging from 0 to 70 nucleotides. The species with the closest resemblance were A. candidus/A. carneus, and A. flavus/A. oryzae/A. ochraceus, with 100 and 99.8 % identification, respectively. These species are phylogenetically very close and the TEF-1α gene appears not to have sufficient discriminatory power to differentiate them. Meanwhile, intra-species differences were found within strains of A. clavatus, A. clavatonanicus, A. candidus, A. fumigatus, A. terreus, A. alliaceus, A. flavus, Eurotium amstelodami and E. chevalieri. The tree topology with strongly supported clades (≥70 % bootstrap values) was almost compatible with the phylogeny inferred from analysis of the DNA sequences of the beta tubulin gene (BT2). However, the backbone of the tree exhibited low bootstrap values, and inter-species correlations were not obvious in some clades; for example, tree topologies based on BT2 and TEF-1α genes were incompatible for some species, such as A. deflectus, A. janus and A. penicillioides. CONCLUSION: The gene was not phylogenetically more informative than other known molecular markers. It will be necessary to test other genes or larger genomic regions to better understand the taxonomy of this important group of fungi.

Characterization of fungi isolated from the equipment used in the International Space Station or Space Shuttle.

As a part of a series of studies regarding the microbial biota in manned space environments, fungi were isolated from six pieces of equipment recovered from the Japanese Experimental Module "KIBO" of the International Space Station and from a space shuttle. Thirty-seven strains of fungi were isolated, identified and investigated with regard to morphological phenotypes and antifungal susceptibilities. The variety of fungi isolated in this study was similar to that of several previous reports. The dominant species belonged to the genera Penicillium, Aspergillus and Cladosporium, which are potential causative agents of allergy and opportunistic infections. The morphological phenotypes and antifungal susceptibilities of the strains isolated from space environments were not significantly different from those of reference strains on Earth.

Cross-reactivity in Cryptococcus antigen latex agglutination test in two commercial kits.

This article presents an examination of the cross-reactivity of pathogenic fungi with Cryptococcus neoformans in two commercial Cryptococcus antigen latex agglutination tests performed across 39 fungal strains. Some fungi were newly indicated as Cryptococcus cross-reactive, and the two kits showed differences in cross-reactive fungi.

Translation elongation factor 1-α gene as a potential taxonomic and identification marker in dermatophytes.

Intra- and interspecies variations of the translation elongation factor 1-α (Tef-1α) gene were evaluated as a new identification marker in a wide range of dermatophytes, which included 167 strains of 30 species. An optimized pan-dermatophyte primer pair was designed, and the target was sequenced. Consensus sequences were used for multiple alignment and phylogenetic tree analysis and the levels of intra- and interspecific nucleotide polymorphism were assessed. Between species, the analyzed part of the Tef-1α gene varied in length from 709 to 769 nucleotides. Significant numbers of species including Trichophyton rubrum, T. tonsurans, T. schoenleinii, T. concentricum, T. violaceum, Epidermophyton floccosum, Microsporum ferrugineum, M. canis, M. audouinii, T. equinum, T. eriotrephon, and T. erinacei were invariant in Tef-1α and had sufficient barcoding distance with neighboring species. Although overall consistency was found between ITS phylogeny as the current molecular marker of dermatophytes and Tef-1α, a higher discriminatory power of Tef-1α appeared particularly useful in some clades of closely related species such as the A. vanbreuseghemii, T. rubrum, A. benhamiae, and A. otae complexes. Nevertheless, we stress that a single gene can not specify species borderlines among dermatophytes and multiple lines of evidence based on a multilocus inquiry may ascertain an incontrovertible evaluation of kinship.

The prognosis of dysphagia patients over 100 years old.

Several reports have recently been published regarding dysphagia in very elderly patients, and centenarian dysphagia patients have become more common in Japan. The aim of this study was to assess the prognosis of dysphagia in very elderly patients. Participants were 24 centenarian dysphagia patients. For each patient, we collected information on age, care level, past medical history, and changes in oral intake according to the Functional Oral Intake Scale (FOIS). Patients were divided into two groups based on the mode of food intake at the time of transfer or discharge: the per oral-only group (the PO-only group, i.e., oral intake alone) and the tube feeding-dependent group (the TF-dependent group, i.e., combination of oral intake and tube feeding, or tube feeding alone). In both groups, the FOIS score decreased significantly from pre-hospitalization to the time of transfer or discharge (p=0.006 for both). The FOIS score at initial assessment was higher in the PO-only group with the TF-dependent group (p=0.0004). Furthermore, the frequency of a FOIS score of 4 at initial assessment was significantly higher in the PO-only group, and the frequency of a FOIS score of 1 was significantly higher in the TF-dependent group (p=0.0006). These findings collectively suggest that oral intake can be recovered if the FOIS score is ≥ 4 at initial assessment, is difficult if the score is 1, and may be possible with a FOIS score of 2.

Utilization of matrix-assisted laser desorption and ionization time-of-flight mass spectrometry for identification of infantile seborrheic dermatitis-causing Malassezia and incidence of culture-based cutaneous Malassezia microbiota of 1-month-old infants.

Matrix-assisted laser desorption and ionization time-of-flight mass spectrometry (MALDI-TOF-MS) has been utilized for identification of various microorganisms. Malassezia species, including Malassezia restricta, which is associated with seborrheic dermatitis, has been difficult to identify by traditional means. This study was performed to develop a system for identification of Malassezia species with MALDI-TOF-MS and to investigate the incidence and variety of cutaneous Malassezia microbiota of 1-month-old infants using this technique. A Malassezia species-specific MALDI-TOF-MS database was developed from eight standard strains, and the availability of this system was assessed using 54 clinical strains isolated from the skin of 1-month-old infants. Clinical isolates were cultured initially on CHROMagar Malassezia growth medium, and the 28S ribosomal DNA (D1/D2) sequence was analyzed for confirmatory identification. Using this database, we detected and analyzed Malassezia species in 68% and 44% of infants with and without infantile seborrheic dermatitis, respectively. The results of MALDI-TOF-MS analysis were consistent with those of rDNA sequencing identification (100% accuracy rate). To our knowledge, this is the first report of a MALDI-TOF-MS database for major skin pathogenic Malassezia species. This system is an easy, rapid and reliable method for identification of Malassezia.

Usefulness of a handheld nebulizer in cough test to screen for silent aspiration.

Cough test to screen for silent aspiration (SA) was reported, and the effectiveness was excellent. However, the device was rather large so that the portability was poor. So, the purpose of this study is to investigate the usefulness of a handheld nebulizer for the test and verify the reproducibility of the method. The subjects were 160 patients who were suspected of having dysphagia and underwent videofluorography (VF) or videoendoscopy (VE). They inhaled 1.0 % citric acid-physiologic saline orally for 1 min using a handheld nebulizer, and the examiner observed the number of coughs: more than five coughs was considered as negative (normal), while less than four coughs was regarded as positive. Among the subjects, 70 patients administered the cough test and VF or VE twice or more at some intervals. The k coefficient was calculated in reproducibility. Using the results of the VF or VE examination as the standards, for SA detection, the sensitivity was 0.86, specificity was 0.71, positive predictive value was 0.53, and negative predictive value was 0.93. The k coefficient was 0.79. In conclusion, the handheld nebulizer was useful in the cough test to screen for SA. Furthermore, satisfactory reproducibility was shown.

Cryptococcus lacticolor sp. nov. and Rhodotorula oligophaga sp. nov., novel yeasts isolated from the nasal smear microbiota of Queensland koalas kept in Japanese zoological parks.

A total of 515 yeast strains were isolated from the nasal smears of Queensland koalas and their breeding environments in Japanese zoological parks between 2005 and 2012. The most frequent species in the basidiomycetous yeast biota isolated from koala nasal passages was Cryptococcus neoformans, followed by Rhodotorula minuta. R. minuta was the most frequent species in the breeding environments, while C. neoformans was rare. Seven strains representing two novel yeast species were identified. Analyses of the 26S rDNA (LSU) D1/D2 domain and nuclear ribosomal DNA internal transcribed spacer region sequences indicated that these strains represent new species with close phylogenetic relationships to Cryptococcus and Rhodotorula. A sexual state was not found for either of these two novel yeasts. Key phenotypic characters confirmed that these strains could be placed in Cryptococcus and Rhodotorula. The names Cryptococcus lacticolor sp. nov. (type strain TIMM 10013(T) = JCM 15449(T) = CBS 10915(T) = DSM 21093(T), DDBJ/EMBL/Genbank Accession No.; AB375774 (ITS) and AB375775 (26S rDNA D1/D2 region), MycoBank ID; MB 802688, Fungal Barcoding Database ID; 3174), and Rhodotorula oligophaga sp. nov. (type strain TIMM 10017(T) = JCM 18398(T) = CBS 12623(T) = DSM 25814(T), DDBJ/EMBL/Genbank Accession No.; AB702967 (ITS) and AB702967 (26S rDNA D1/D2 region), MycoBank ID; MB 802689, Fungal Barcoding Database ID; 3175) are proposed for these new species.

Detection and identification of probable endemic fungal pathogen, Cryptococcus gattii, and worldwide pathogen, Cryptococcus neoformans, by real-time PCR.

A real-time PCR method for detection and identification of Cryptococcus neoformans and Cryptococcus gattii was developed and evaluated using DNA from single-colony or koala nasal smears. Two TaqMan minor groove binder probes that distinguished between these species were designed corresponding to the internal sequences of the CAP59 gene for both species. The real-time PCR assay had 100% specificity, as assessed using 13 reference strains and 300 environmental strains. Twelve smear samples from healthy koalas were analyzed by direct real-time PCR. This method successfully detected C. gattii and C. neoformans in one and three koalas, respectively.

Effect of the reclining position in patients after oral tumor surgery.

OBJECTIVES: This study investigated the effects of reclining position on swallowing functions in the patients who had undergone surgery for oral tumors. SUBJECTS AND METHODS: 53 Patients after oral tumor surgery who had undergone the videofluoroscopic swallowing study in the sitting position and reclining position were sampled. We measured Oral transit time (OTT), Pharyngeal transit time (PTT), Penetration-Aspiration scale, Locating of the leading edge of bolus at swallow onset, Route of bolus transit and Transit velocity. RESULTS: Laryngeal penetration and aspiration were less likely to occur in the reclining position. The mean OTT was significantly shorter in the reclining position than in the sitting position. The leading edge of bolus at swallow onset was significantly lower in the reclining position than in the sitting position. The bolus went along the posterior wall of the pharynx in more patients in the reclining position. The mean PTT and the transit velocity did not significantly differ between the reclining position and the sitting position. CONCLUSION: This study suggested that, in patients after surgery on oral tumors, the reclining position is a useful posture to aid oral transit and to readily prevent aspiration and laryngeal penetration.

Human external ear canal as the specific reservoir of Malassezia slooffiae.

The incidence of Malassezia species recovered from the external ear canal was characterized using culture medium optimized for Malassezia spp., CHROMagar Malassezia. The results of this study indicated that in healthy individuals M. slooffiae was the dominant Malassezia species followed by M. restricta.

Quantitation of fungal DNA contamination in commercial zymolyase and lyticase used in the preparation of fungi.

Small amounts of contaminants may lead to false-positive results in sensitive polymerase chain reaction (PCR) detection systems. To analyze contaminants and understand the usability of beta-glucanases in fungal preparations, we estimated the ribosomal DNA (rDNA) contamination in Zymolyase-100T and Lyticase by quantitative PCR. The amount of rDNA contamination determined by real-time PCR was 9210 copies/unit for Zymolyase-100T and 0.0323 copies/unit for Lyticase. The observations regarding these enzyme products indicate that careful consideration of contaminating DNA included in the reagents used for molecular diagnostics is necessary.

Enhanced gene replacements in Ku80 disruption mutants of the dermatophyte, Trichophyton mentagrophytes.

The frequency of targeted gene disruption via homologous recombination is low in the clinically important dermatophyte, Trichophyton mentagrophytes. The Ku genes, Ku70 and Ku80, encode key components of the nonhomologous end-joining pathway involved in DNA double-strand break repair. Their deletion increases the homologous recombination frequency, facilitating targeted gene disruption. To improve the homologous recombination frequency in T. mentagrophytes, the Ku80 ortholog was inactivated. The nucleotide sequence of the Ku80 locus containing a 2788-bp ORF encoding a predicted product of 728 amino acids was identified, and designated as TmKu80. The predicted TmKu80 product showed a high degree of amino acid sequence similarity to known fungal Ku80 proteins. Ku80 disruption mutant strains of T. mentagrophytes were constructed by Agrobacterium tumefaciens-mediated genetic transformation. The average homologous recombination frequency was 73.3 +/- 25.2% for the areA/nit-2-like nitrogen regulatory gene (tnr) in Ku80(-) mutants, about 33-fold higher than that in wild-type controls. A high frequency (c. 67%) was also obtained for the Tri m4 gene encoding a putative serine protease. Ku80(-) mutant strains will be useful for large-scale reverse genetics studies of dermatophytes, including T. mentagrophytes, providing valuable information on the basic mechanisms of host invasion.

Agrobacterium tumefaciens-mediated transformation of the dermatophyte, Trichophyton mentagrophytes: an efficient tool for gene transfer.

Agrobacterium tumefaciens-mediated transformation (ATMT) was used to facilitate gene transfer into the clinically important dermatophyte, Trichophyton mentagrophytes (teleomorph: Arthroderma vanbreuseghemii). A binary vector containing a hygromycin B resistance cassette was introduced into A. tumefaciens, and the resultant strain was co-cultivated with fungal small conidia. Transformation yielded a large number of hygromycin B-resistant transformants. Hybridization analysis showed that most of the transformants harboured a single copy of T-DNA randomly integrated into the genome. Transformation frequency was increased to more than 200 per 10(7) conidia by optimizing the co-cultivation time and temperature. ATMT was then used for targeted gene disruption mediated by homologous recombination. Using a PCR-based strategy, we isolated the areA/nit-2-like nitrogen regulatory gene (tnr:Trichophytonnitrogen regulator) from T. mentagrophytes. A binary vector containing two regions of the tnr locus flanking the hygromycin B resistance cassette was constructed and introduced into T. mentagrophytesvia ATMT. Transformants with disruption of the areA/nit-2-like gene (tnr) were obtained in three of four independent disruption experiments, most of which showed homologous recombination via double crossover without additional ectopic integration of the disruption construct.

Genetic transformation of the dermatophyte, Trichophyton mentagrophytes, based on the use of G418 resistance as a dominant selectable marker.

BACKGROUND: Dermatophytes are closely related keratinophilic fungal pathogens and are the causative agents of a superficial cutaneous infection called dermatophytosis (ringworm). A lack of gene manipulation techniques has prevented detailed analyses of the mechanisms of host invasion by dermatophytes. We have introduced the tetracycline-regulatable (TR) gene expression system into dermatophytes to facilitate functional analyses of genes essential for growth and virulence. As the TR gene expression system consists of two plasmid vector components, two dominant selectable markers are required for genetic transformation. In dermatophytes, only the hygromycin B phosphotransferase gene (hph) is available as a selectable marker. OBJECTIVE: We investigated the possibility of G418 resistance as a secondary selectable marker for genetic transformation in dermatophytes. METHODS: A series of plasmid vectors carrying the neomycin phosphotransferase gene (nptII) were introduced into the protoplasts of Trichophyton mentagrophytes, one of the most clinically important dermatophyte species, by polyethylene glycol (PEG)-mediated transformation. Transformants were selected on selective medium containing G418 at 300-500 microg/ml. RESULTS: Molecular biological analyses indicated that colonies appearing on the selective medium harbored nptII in their chromosomes. Colonies produced from protoplasts transformed with the enhanced green fluorescent protein (eGFP) gene-T. mentagrophytes cyclophilin cDNA (TmcypB) fusion vector also exhibited GFP fluorescence throughout their mycelia, but accumulation of the GFP-TmCYPB fusion protein in specific intracellular compartments was not observed. CONCLUSIONS: This study has provided a new selectable marker for genetic transformation in dermatophytes.