Clinical impact of major pathogenic genotypes of Pseudomonas aeruginosa associated with refractory chronic suppurative otitis media.

PURPOSE: Chronic suppurative otitis media (CSOM) is characterized by persistent inflammation of the mucous membrane of the middle ear and mastoid. One of the primary causative agents of CSOM is P. aeruginosa, known for its production of virulent toxins and enzymes. Some cases of CSOM, improvement may not occur despite treatment lasting three weeks, leading to what is termed refractory CSOM. This research aims to characterize the P. aeruginosa strains isolated from patients with refractory CSOM in Gyeongsangnam-do, South Korea, providing insights into their pathogenic profiles. METHODS: We conducted a retrospective analysis of P. aeruginosa isolates from the otorrhea of patients diagnosed with CSOM at a tertiary hospital in Gyeongsangnam-do, over a period from January 2005 to August 2022. The strains were examined using multilocus sequence typing (MLST) and toxin gene assay to assess genetic diversity and virulence. RESULTS: 39 samples were obtained from 13 cases of refractory CSOM and 15 cases of non-refractory CSOM. The findings unveiled that the P. aeruginosa cultured from patients with refractory CSOM belonged to the P. aeruginosa sequence type 235 (ST235) strain, which harbors the exoU gene as a major virulence factor. CONCLUSION: The detection of ST235 in refractory CSOM signifies a challenging clinical scenario. Given the genotype's strong virulence and antibiotic resistance, identifying ST235 through MLST can guide effective management approaches, including potential surgical intervention. This study underscores the necessity of broader epidemiological investigations to understand ST235 behavior and advocates for patient education to mitigate the impacts of this formidable pathogen in CSOM.

Macrophage Depletion in CCR2-/- Mice Delays Bacterial Clearance and Enhances Neutrophil Infiltration in an Acute Otitis Media Model.

BACKGROUND: Otitis media (OM) is a common and potentially serious disease of childhood. Although OM is multifactorial on origin, bacterial infection is a unifying component. Many studies have established a critical role for innate immunity in bacterial clearance and OM resolution. A key component of innate immunity is the recruitment of immune and inflammatory cells, including macrophages. METHODS: To explore the role of macrophages in OM, we evaluated the expression of genes related to macrophage function during a complete episode of acute OM in the mouse caused by middle ear (ME) inoculation with Haemophilus influenzae. We also combined CCR2 deficiency with chlodronate liposome toxicity to deplete macrophages during OM. RESULTS: Macrophage genes were robustly regulated during OM. Moreover, macrophage depletion enhanced and prolonged the infiltration of neutrophils into the infected ME and increased the persistence of bacterial infection. CONCLUSIONS: The results illustrate the critical role played by macrophages in OM resolution.

Periostin induces kidney fibrosis after acute kidney injury via the p38 MAPK pathway.

Periostin plays a crucial role in fibrosis, and acute kidney injury results in a high risk of progression to chronic kidney disease. Therefore, we hypothesized that periostin was involved in the progression of acute kidney injury to kidney fibrosis. Unilateral ischemia-reperfusion injury (UIRI) was induced in 7- to 8-wk-old male wild-type and periostin-null mice, and the animals were observed for 6 wk. In vitro, human kidney-2 cells and primary-cultured human tubular epithelial cells were incubated under hypoxic conditions (5% O2, 5% CO2, and 90% N2) for 5 days. The cells were also cultured with recombinant periostin (rPeriostin) and a p38 mitogen-activated protein kinase (MAPK) inhibitor in a hypoxic incubator. At 6 wk after UIRI, interstitial fibrosis/tubular atrophy was significantly alleviated in periostin-null mice compared with wild-type controls. In addition, periostin-null mice had attenuated expression of fibrosis/apoptosis markers and phosphorylated-p38 MAPK compared with wild-type controls. In vitro, hypoxic injury increased the expression of fibrosis markers, periostin, and phosphorylated-p38 MAPK, which was comparable to or substantially greater than their expression levels following treatment with recombinant transforming growth factor-ß1 under normoxic conditions. Furthermore, rPeriostin treatment under hypoxic conditions enhanced fibrosis/apoptosis markers and phosphorylated-p38 MAPK. In contrast, p38 MAPK inhibition ameliorated hypoxia-induced fibrosis, and the addition of the p38 MAPK inhibitor to rPeriostin significantly ameliorated the changes induced by rPeriostin. In conclusion, periostin promotes kidney fibrosis via the p38 MAPK pathway following acute kidney injury triggered by a hypoxic or ischemic insult. Periostin ablation may protect against chronic kidney disease progression.

Idiopathic benign paroxysmal positional vertigo in the elderly: a long-term follow-up study.

BACKGROUND: Benign paroxysmal positional vertigo (BPPV) is the most common cause of peripheral-type vertigo in the elderly. While some studies have investigated BPPV among the elderly, no study has focused on the isolated idiopathic BPPV (iBPPV) in the elderly. AIMS: To investigate the clinical features and recurrence rate in elderly patients diagnosed with iBPPV. METHODS: The authors prospectively reviewed the medical records of 627 patients diagnosed with BPPV, and a total of 370 patients fulfilled the inclusion criteria. The subjects were divided into two groups by age (experimental group ≥65 years and control group 17-64 years), and the gender, numbers of canalith reposition maneuver (CRM) for successful treatment, time elapsed from onset of symptoms to clinic visit, the affected semicircular canal, and the 1- and 5-year recurrence proportion were analyzed. RESULTS: The gender, numbers of CRM for successful treatment, affected semicircular canal, and the 1- and 5-year recurrence proportion showed no statistically significant difference between two groups (p > 0.05). The only exception was the experimental group which took a longer duration from manifestation of symptoms to hospital visit than control group (student's t test, p = 0.021). DISCUSSION: Several previous studies report that the elderly with iBPPV shows more protracted clinical course and much higher recurrence rate than younger adult. Unlike those reports, our study showed no significant differences about the clinical features and recurrence rate between age groups. CONCLUSION: The elderly with iBPPV could be treated as effective as general population.

Selective bio-oxidation of propane to acetone using methane-oxidizing Methylomonas sp. DH-1.

Propane is the major component of liquefied petroleum gas (LPG). Nowadays, the use of LPG is decreasing, and thus utilization of propane as a chemical feedstock is in need of development. An efficient biological conversion of propane to acetone using a methanotrophic whole cell as the biocatalyst was proposed and investigated. A bio-oxidation pathway of propane to acetone in Methylomonas sp. DH-1 was analyzed by gene expression profiling via RNA sequencing. Propane was oxidized to 2-propanol by particulate methane monooxygenase and subsequently to acetone by methanol dehydrogenases. Methylomonas sp. DH-1 was deficient in acetone-converting enzymes and thus accumulated acetone in the absence of any enzyme inhibition. The maximum accumulation, average productivity and specific productivity of acetone were 16.62 mM, 0.678 mM/h and 0.141 mmol/g cell/h, respectively, under the optimized conditions. Our study demonstrates a novel method for the bioconversion of propane to acetone using methanotrophs under mild reaction condition.

Intraoperative factors affecting conversion from cruciate retaining to cruciate substituting in total knee arthroplasty.

PURPOSE: The purpose of this study was to analyse the factors affecting the conversion from posterior cruciate ligament (PCL)-retaining (CR) to PCL-substituting (PS) prostheses during total knee arthroplasty (TKA). METHODS: A total of 920 TKAs, which had been preoperatively planned to undergo implantation of CR-type prostheses, were reviewed retrospectively. Of these, 83 knees (9.0 %) were converted intraoperatively to PS prostheses. The clinical and radiological factors of the non-converted (CR) and converted (PS) groups were compared. Clinically, age, gender, body mass index, angle of flexion contracture, size of the femoral component, and thickness of the polyethylene insert were compared between the CR and PS groups. Radiologically, the severity of the varus deformity and the posterior tibial slope angle (PSA) were compared between the CR and PS groups. RESULTS: No significant differences in age, gender, body mass index, range of motion, thickness of the polyethylene insert, or severity of varus deformity were identified. The average preoperative angle of flexion contracture was 5.9° ± 7.4° in the CR group and 8.1° ± 9.1° in the PS group (p = 0.002). The average preoperative PSA was 9.6° ± 4.0° in the CR group and 11.0° ± 5.0° in the PS group (p = 0.018). The conversion rates to a PS-type femoral component of size C, D, and E were 13.1, 7.0, and 6.3 %, respectively (p = 0.004). CONCLUSION: The conversion rate from CR- to PS-type prostheses was high in patients with severe flexion contracture, steep posterior slope, and a small femoral component size. These factors should be carefully considered for appropriate selection of prosthesis type. LEVEL OF EVIDENCE: IV.

The burden of readmissions after rib fractures among older adults.

BACKGROUND: The index hospitalization morbidity and mortality of rib fractures among older adults (aged ≥65 years) is well-known, yet the burden and risks for readmissions after rib fractures in this vulnerable population remain understudied. We aimed to characterize the burdens and etiologies associated with 3-month readmissions among older adults who suffer rib fractures. We hypothesized that readmissions would be common and associated with modifiable etiologies. METHODS: This survey-weighted retrospective study using the 2017 and 2019 National Readmissions Database evaluated adults aged ≥65 years hospitalized with multiple rib fractures and without major extrathoracic injuries. The main outcome was the proportion of patients experiencing all-cause 3-month readmissions. We assessed the 5 leading principal readmission diagnoses overall and delineated them by index hospitalization discharge disposition (home or facility). Sensitivity analysis using clinical classification categories characterized readmissions that could reasonably represent rib fracture-related sequelae. RESULTS: In 2017, 25,092 patients met the inclusion criteria, with 20% (N = 4,894) experiencing 3-month readmissions. Six percent of patients did not survive their readmission. The 5 leading principal readmission diagnoses were sepsis (many associated with secondary diagnoses of pneumonia [41%] or urinary tract infections [41%]), hypertensive heart/kidney disease, hemothorax, pneumonia, and respiratory failure. In 2019, a comparable 3-month readmission rate of 23% and identical 5 leading diagnoses were found. Principal readmission diagnosis of hemothorax was associated with the shortest time to readmission (median [interquartile range]:9 [5-23] days). Among patients discharged home after index hospitalization, pleural effusion-possibly representing mischaracterized hemothorax-was among the leading principal readmission diagnoses. Some patients readmitted with a principal diagnosis of hemothorax or pleural effusion had these diagnoses at index hospitalization; a lower proportion of these patients underwent pleural fluid intervention during index hospitalization compared with readmission. On sensitivity analysis, 30% of 3-month readmissions were associated with principal diagnoses suggesting rib fracture-related sequelae. CONCLUSION: Readmissions are not infrequent among older adults who suffer rib fractures, even in the absence of major extrathoracic injuries. Future studies should better characterize how specific complications associated with readmissions, such as pneumonia, urinary tract infections, and delayed hemothoraces, could be mitigated.

Engineering of Pseudomonas putida to produce medium-chain-length polyhydroxyalkanoate from crude glycerol.

The development of biodegradable polymers is crucial for addressing environmental issues and waste management challenges, and a medium-chain-length polyhydroxyalkanoate(MCL-PHA) exhibits significant application potential in diverse industrial and environmental contexts owing to its versatility and biodegradability. Here, Pseudomonas putida was metabolically engineered to produce MCL-PHA from crude glycerol. To increase the precursor pool, we first deleted the phaC1ZC2 operon and introduced a plasmid-based overexpression of phaC2 and phaG, and the MCL-PHA content derived from glycerol increased to 18.27 wt% at 60 h. Subsequently, by optimizing the acoA expression through promoter selection and UTR design, the MCL-PHA content further increased to 19.93 wt% at 72 h. Additionally, a notable increase in MCL-PHA production was achieved using PhaC2 designed to have no substrate-trapping effect (PhaC2A477A478). This improvement was guided by filling structural data gaps using Alphafold2 and docking simulations that revealed the substrate-trapping phenomenon. High-level production of MCL-PHA was achieved through fed-batch fermentation using the final engineered P. putida from refined glycerol, which yielded 34.9 g/L of MCL-PHA with 44.64 wt% at 180 h. Furthermore, using crude glycerol as the sole carbon source enabled the production of 49.5 g/L of MCL-PHA with 45.41 wt% at 180 h in fed-batch culture.

Effect of high dose intravenous vitamin C on idiopathic sudden sensorineural hearing loss: a prospective single-blind randomized controlled trial.

The aim of this prospective single-blind randomized controlled study was to evaluate the therapeutic efficacy of high dose intravenous vitamin C (HDVC) added to systemic steroid in patients with idiopathic sudden sensorineural hearing loss (ISSNHL). Between August 2010 and August 2011, 72 ISSNHL patients who participated in this study were randomly allocated to two groups: 36 to a control group, members of which were given systemic steroid treatment for 15 days, and 36 to a HDVC group, members of which were given HDVC (200 mg/kg/day) for 10 days in addition to steroid therapy followed by oral vitamin C (2,000 mg) for 30 days after discharge. Finally, we analyzed each group: 35 as a control group and 32 as a HDVC group. Auditory evaluations were performed by pure tone audiometry (PTA) before and ~1 month after treatment using Siegel's criteria. HDVC group showed significantly greater complete and partial recovery improvement (p = 0.035). In addition, the complete recovery rate in the HDVC group was more than twice that of the control group (p = 0.031). In the HDVC group, PTA improved from 67.6 ± 19.8 dB HL before treatment to 37.1 ± 28.8 dB HL at 1 month after treatment, whereas in the control group, PTA improved from 70.3 ± 12.4 to 47.6 ± 25.2 dB HL, which represented a significant intergroup difference (p = 0.030). In conclusion, HDVC may enhance hearing recovery in ISSNHL patients, which suggests that HDVC reduces levels of reactive oxygen metabolites produced by inner ear ischemia or inflammation, and that HDVC could be considered for the treatment of ISSNHL.

Outcomes of Endoscopic Congenital Cholesteatoma Removal in South Korea.

Importance: Transcanal endoscopic ear surgery (TEES) provides minimally invasive transcanal access to the middle ear and improves middle ear visibility during cholesteatoma resection. However, the literature on outcomes following TEES alone for the removal of congenital cholesteatoma (CC) is lacking and limited to small series. Objective: To assess outcomes of TEES for CC limited to the middle ear and/or mastoid antrum and to explore the risk factors associated with recidivism (ie, recurrent and/or residual cholesteatoma). Design, Setting, and Participants: This cohort study evaluated retrospective, multicenter data for 271 children with CC who underwent TEES at 9 tertiary referral hospitals in South Korea between January 1, 2013, and December 31, 2021, and had a follow-up of at least 6 months after surgery. Main Outcomes and Measures: Outcomes included the incidence of residual cholesteatoma and audiometric data after TEES. A multivariable analysis using Cox proportional hazards regression models was used to assess associations between cholesteatoma characteristics and recidivism, with hazard ratios (HRs) and 95% CIs reported. Results: Of the 271 patients (mean [SD] age, 3.5 [2.9] years; 194 [71.6%] boys, 77 [28.4%] girls), 190 had Potsic stage I CC (70.1%), 21 (7.7%) had stage II, 57 (21.0%) had stage III, and 3 (1.1%) had stage IV. Thirty-six patients (13.3%) with residual cholesteatoma were found, including 15 (7.9%) with Potsic stage I, 3 (14.3%) with stage II, and 18 (31.6%) with stage III. In the multivariable analysis, invasion of the malleus (HR, 2.257; 95% CI, 1.074-4.743) and posterosuperior quadrant location (HR, 3.078; 95% CI, 1.540-6.151) were associated with the incidence of recidivism. Overall, hearing loss (>25 dB on auditory behavioral test or >30 dB of auditory evoked responses) decreased from 24.4% to 17.7% after TEES. Conclusions and Relevance: This cohort study involved the largest known population to date of CC removed by TEES. The findings suggest that TEES may be feasible and effective for the removal of CC limited to the middle ear and/or mastoid antrum in children.

Outcomes of Endoscopic Tympanoplasty for Large Perforations: A Multicenter Retrospective Study in South Korea.

OBJECTIVES: Endoscopic tympanoplasty (ET) provides minimally invasive transcanal access to the middle ear and improves middle ear visibility for the treatment of tympanic membrane (TM) perforations. However, the literature on surgical outcomes for large TM perforations is lacking and limited to small series. This study aimed to evaluate the clinical benefits of ET for large TM perforations. METHODS: This retrospective cohort study was conducted at nine tertiary referral hospitals in South Korea, where 252 patients who underwent ET as primary surgery from September 2019 to August 2021 were included. The outcome measures included the graft success rate and pre- and postoperative audiometric data. RESULTS: In 239 patients, the graft success rate of ET for large or subtotal perforations was 86.2% (206 patients), while the graft failure rate was 13.8% (33 patients). The graft failure rate was directly correlated with surgical techniques, including overlay and medial or lateral underlay tympanoplasty (P=0.027). Lateral underlay tympanoplasty showed the most favorable. RESULTS: Sex, laterality, etiology, site and size of perforation, operation time, and graft materials did not vary significantly between the graft success and failure groups (P>0.05). The mean air-bone gap (ABG) improved significantly in both groups (graft success group: 10.0±0.6 dB and graft failure group: 7.7±0.3 dB; P<0.001). However, the ABG improvement did not significantly differ between the groups. Analysis of covariance revealed that the postoperative 500-Hz bone conduction threshold improved after successful ET (adjusted coefficient, -11.351; 95% confidence interval, -21.491 to -1.212; P=0.028). CONCLUSION: This study involved the largest population to date of large TM perforations treated by ET. The study findings suggest that ET is feasible and effective in treating large TM perforations.

Sensation recovery of auricle following chronic ear surgery by retroauricular incision.

The aim of this prospective analysis was to objectify and quantify the sensory loss in the auricular area that occurs following surgery for chronic otitis media or cholesteatoma, and to assess the exact recovery time of the auricular sensation. Till now, no study has yet been conducted on the sensory loss that is developed after retroauricular incision is performed for chronic ear surgery. Forty-eight patients underwent surgery via retroauricular incision for chronic otitis media or for chronic otitis media or cholesteatoma between March 2009 and January 2010. The skin around the auricle was divided into six areas. The sensation of each area was assessed before the surgery and 3 days, 7 days, 1 month, 3 months, and 6 months after the surgery, using the Semmes-Weinstein monofilaments. The patients were also asked to record the sensation that they felt using the visual analog scale (VAS). In the objective sensory assessment, only area 5, which corresponds to the retroauricular area, showed significant sensory loss. Three months after the surgery, the sensation was recovered to a level comparable to that before the surgery in most of the patients. The mean subjective VAS score was 10 prior to the surgery, 8.56 (± 1.08) 3 months after the surgery, and 9.32 (± 0.74) 6 months after the surgery, respectively. In conclusion, following chronic ear surgery, the sensation of the auricle was recovered to the previous level within 3 months. Therefore, patients who are to undergo retroauricular incision should be informed that they will experience temporary sensory loss for approximately 3 months after the surgery.

The ECRG4 cleavage product augurin binds the endotoxin receptor and influences the innate immune response during otitis media.

Otitis media (OM), the most common disease of childhood, is typically characterized by bacterial infection of the middle ear (ME). Prominent features of OM include hyperplasia of the ME mucosa, which transforms from a monolayer of simple squamous epithelium with minimal stroma into a full-thickness respiratory epithelium in 2-3 days after infection. Analysis of the murine ME transcriptome during OM showed down-regulation of the tumor suppressor gene Ecrg4 that was temporally related to mucosal hyperplasia and identified stromal cells as the primary ECRG4 source. The reduction in Ecrg4 gene expression coincided with the cleavage of ECRG4 protein to release an extracellular fragment, augurin. The duration of mucosal hyperplasia during OM was greater in Ecrg4 -/- mice, the number of infiltrating macrophages was enhanced, and ME infection cleared more rapidly. ECRG4-null macrophages showed increased bacterial phagocytosis. Co-immunoprecipitation identified an association of augurin with TLR4, CD14 and MD2, the components of the lipopolysaccharide (LPS) receptor. The results suggest that full-length ECRG4 is a sentinel molecule that potentially inhibits growth of the ME stroma. Processing of ECRG4 protein during inflammation, coupled with a decline in Ecrg4 gene expression, also influences the behavior of cells that do not express the gene, limiting the production of growth factors by epithelial and endothelial cells, as well as the activity of macrophages.

Clinical characteristics and treatment of benign paroxysmal positional vertigo after traumatic brain injury.

BACKGROUND: Traumatic brain injury (TBI) has been reported to be a common cause of benign paroxysmal positional vertigo (BPPV). However, only a few studies have investigated BPPV after TBI. The aim of this study was to identify the clinical characteristics of BPPV after TBI and to determine whether there are clinical differences between BPPV after TBI and idiopathic BPPV. METHODS: The authors reviewed the medical records of 192 consecutive patients with positional vertigo after head injury during the period 2003 to 2009 and investigated 112 patients with idiopathic BPPV treated over the same period. The clinical characteristics of BPPV after TBI and the clinical differences between the traumatic BPPV and idiopathic BPPV groups were investigated. RESULTS: A total of 32 patients with BPPV after TBI fulfilled the inclusion criteria. Twenty-four patients in the traumatic BPPV group had posterior semicircular canal-BPPV and 11 patients lateral semicircular canal-BPPV. A total of 58 repositioning maneuver sessions were performed in these 32 patients. Members of the traumatic BPPV group required more treatment sessions than members of the idiopathic group (p<0.05), but no tendency to recur was observed in the traumatic group (p>0.05). Recurrence rates in the traumatic and idiopathic BPPV groups were 15.6% and 18.8%, respectively (p>0.05). CONCLUSIONS: It is likely that BPPV after TBI is more difficult to treat than idiopathic BPPV, but no tendency to recur was observed in patients who developed BPPV after TBI compared with idiopathic BPPV. Further prospective clinical meta-analytic studies are needed to investigate the outcome of BPPV after TBI.

Prolonged Positional Downbeat Nystagmus in Benign Paroxysmal Positional Vertigo: A Case Report and Literature Review.

Purpose This study aimed to report an unusual case of benign paroxysmal positional vertigo (BPPV), who showed prolonged positional downbeat nystagmus without latency and was diagnosed with cupulolithiasis of the anterior canal (AC). We compared this case with one of typical AC-BPPV, and possible mechanisms underlying the atypical characteristics were discussed. Method Two patients diagnosed with AC-BPPV were reported. Positional testing using video-oculography goggles was performed, and outcomes were measured via medical records and analysis of videos of the nystagmus. Results Downbeat nystagmus was observed in the contralateral Dix-Hallpike test in both cases. The torsional component was subtle or absent, but motion was induced toward the affected ear. The two cases differed in latency and duration of vertigo, as well as habituation. The patient with atypical nystagmus showed little or no latency and longer duration. Moreover, there was no habituation on repeated tests. The nystagmus showed several differences from that of typical AC-BPPV. Conclusions Based on our case, AC-BPPV may induce various unusual clinical manifestations of nystagmus. Accurate diagnosis requires careful consideration of the patient's symptoms and the characteristics of the nystagmus. Supplemental Material https://doi.org/10.23641/asha.14265356.

IgG4-related disease presenting as otogenic skull base osteomyelitis.

IgG4-related disease (IgG4-RD) is an emerging clinical disease entity characterized by tumefactive lesions at multiple sites with a dense lymphoplasmacytic infiltrate rich in IgG4+ plasma cells. Although almost any organ can be affected, IgG4-RD is most likely to involve the submandibular, lacrimal, or parotid glands in the head and neck region. However, skull base involvement presenting as otogenic skull base osteomyelitis (SBO) is rare. We encountered a 70-year-old male with IgG4-RD presenting primarily with severe otalgia and otorrhea. He had uncontrolled diabetes mellitus and showed clinical manifestations of otogenic SBO. Tissue immunostaining revealed typical features of increased IgG4-positive plasma cells, and hematological examination showed elevated serum IgG4 concentrations. Treatment with corticosteroids significantly improved well-being and partially resolved the lesion based on computed tomography (CT) scan.

Pseudo-spontaneous nystagmus in lateral semicircular canal benign paroxysmal positional vertigo: Correlation with bow and lean test in a pitch plane.

OBJECTIVES: We investigated the incidence and characteristics of pseudo-spontaneous nystagmus (PSN) in benign paroxysmal positional vertigo involving the lateral semicircular canal (LC-BPPV) and evaluated the correlation between PSN and the bow and lean test. METHODS: We examined nystagmus in the sitting position using video-oculography goggles in 131 LC-BPPV patients. The positioning test and bow and lean test were also performed. Patients were divided into canalolithiasis and cupulolithiasis groups according to the character of nystagmus. In each group, the incidence and direction of PSN, correlation with the bow and lean test, and treatment outcome were analyzed. RESULTS: PSN was observed in 25 cases (19.1%) in LC-BPPV patients, 7 of which were canalolithiasis and 18 of which were cupulolithiasis (p = 0.098). Of the 25 patients with PSN, 21 (84%) exhibited nystagmus consistent with the lean test whereas 4 (16%) exhibited nystagmus consistent with the bow test. In patients with PSN, nystagmus was observed in the bow and lean test in all cases (23/23), but in patients without PSN, no nystagmus was observed in 13 cases (13/87) in the bow and lean test (p = 0.048). The number of barbecue maneuvers performed until the end of treatment was 1.4 ± 0.7 in patients with PSN and 1.4 ± 0.9 in those without PSN (p = 0.976). CONCLUSION: We identified PSN in patients with LC-BPPV irrelevant of subtype. Moreover, all patients with PSN showed nystagmus in the bow and lean test. The direction of PSN was mostly consistent with that of the lean test (21/25, 84%). The presence of PSN was not related to the treatment outcome in this study.

Enhanced Production of Bacterial Cellulose in Komagataeibacter xylinus Via Tuning of Biosynthesis Genes with Synthetic RBS.

Bacterial cellulose (BC) has outstanding physical and chemical properties, including high crystallinity, moisture retention, and tensile strength. Currently, the major producer of BC is Komagataeibacter xylinus. However, due to limited tools of expression, this host is difficult to engineer metabolically to improve BC productivity. In this study, a regulated expression system for K. xylinus with synthetic ribosome binding site (RBS) was developed and used to engineer a BC biosynthesis pathway. A synthetic RBS library was constructed using green fluorescent protein (GFP) as a reporter, and three synthetic RBSs (R4, R15, and R6) with different strengths were successfully isolated by fluorescence-activated cell sorting (FACS). Using synthetic RBS, we optimized the expression of three homologous genes responsible for BC production, pgm, galU, and ndp, and thereby greatly increased it under both static and shaking culture conditions. The final titer of BC under static and shaking conditions was 5.28 and 3.67 g/l, respectively. Our findings demonstrate that reinforced metabolic flux towards BC through quantitative gene expression represents a practical strategy for the improvement of BC productivity.

Development and characterization of a Nannochloropsis mutant with simultaneously enhanced growth and lipid production.

BACKGROUND: The necessity to develop high lipid-producing microalgae is emphasized for the commercialization of microalgal biomass, which is environmentally friendly and sustainable. Nannochloropsis are one of the best industrial microalgae and have been widely studied for their lipids, including high-value polyunsaturated fatty acids (PUFAs). Many reports on the genetic and biological engineering of Nannochloropsis to improve their growth and lipid contents have been published. RESULTS: We performed insertional mutagenesis in Nannochloropsis salina, and screened mutants with high lipid contents using fluorescence-activated cell sorting (FACS). We isolated a mutant, Mut68, which showed improved growth and a concomitant increase in lipid contents. Mut68 exhibited 53% faster growth rate and 34% higher fatty acid methyl ester (FAME) contents after incubation for 8 days, resulting in a 75% increase in FAME productivity compared to that in the wild type (WT). By sequencing the whole genome, we identified the disrupted gene in Mut68 that encoded trehalose-6-phosphate (T6P) synthase (TPS). TPS is composed of two domains: TPS domain and T6P phosphatase (TPP) domain, which catalyze the initial formation of T6P and dephosphorylation to trehalose, respectively. Mut68 was disrupted at the TPP domain in the C-terminal half, which was confirmed by metabolic analyses revealing a great reduction in the trehalose content in Mut68. Consistent with the unaffected N-terminal TPS domain, Mut68 showed moderate increase in T6P that is known for regulation of sugar metabolism, growth, and lipid biosynthesis. Interestingly, the metabolic analyses also revealed a significant increase in stress-related amino acids, including proline and glutamine, which may further contribute to the Mut68 phenotypes. CONCLUSION: We have successfully isolated an insertional mutant showing improved growth and lipid production. Moreover, we identified the disrupted gene encoding TPS. Consistent with the disrupted TPP domain, metabolic analyses revealed a moderate increase in T6P and greatly reduced trehalose. Herein, we provide an excellent proof of concept that the selection of insertional mutations via FACS can be employed for the isolation of mutants with improved growth and lipid production. In addition, trehalose and genes encoding TPS will provide novel targets for chemical and genetic engineering, in other microalgae and organisms as well as Nannochloropsis.