Genes encoding lipid II flippase MurJ and peptidoglycan hydrolases are required for chloroplast division in the moss Physcomitrella patens.

KEY MESSAGE: Homologous genes for the peptidoglycan precursor flippase MurJ, and peptidoglycan hydrolases: lytic transglycosylase MltB, and DD-carboxypeptidase VanY are required for chloroplast division in the moss Physcomitrella patens. The moss Physcomitrella patens is used as a model plant to study plastid peptidoglycan biosynthesis. In bacteria, MurJ flippase transports peptidoglycan precursors from the cytoplasm to the periplasm. In this study, we identified a MurJ homolog (PpMurJ) in the P. patens genome. Bacteria employ peptidoglycan degradation and recycling pathways for cell division. We also searched the P. patens genome for genes homologous to bacterial peptidoglycan hydrolases and identified genes homologous for the lytic transglycosylase mltB, N-acetylglucosaminidase nagZ, and LD-carboxypeptidase ldcA in addition to a putative DD-carboxypeptidase vanY reported previously. Moreover, we found a ß-lactamase-like gene (Pplactamase). GFP fusion proteins with either PpMltB or PpVanY were detected in the chloroplasts, whereas fusion proteins with PpNagZ, PpLdcA, or Pplactamase localized in the cytoplasm. Experiments seeking PpMurJ-GFP fusion proteins failed. PpMurJ gene disruption in P. patens resulted in the appearance of macrochloroplasts in protonemal cells. Compared with the numbers of chloroplasts in wild-type plants (38.9 ± 4.9), PpMltB knockout and PpVanY knockout had lower numbers of chloroplasts (14.3 ± 6.7 and 28.1 ± 5.9, respectively). No differences in chloroplast numbers were observed after PpNagZ, PpLdcA, or Pplactamase single-knockout. Chloroplast numbers in PpMltB/PpVanY double-knockout cells were similar to those in PpMltB single-knockout cells. Zymogram analysis of the recombinant PpMltB protein revealed its peptidoglycan hydrolase activity. Our results imply that PpMurJ, PpMltB and PpVanY play a critical role in chloroplast division in the moss P. patens.

Anesthetic management during tracheotomy in a child with respiratory distress caused by severe intubation-induced glottic stenosis.

We provided anesthetic management during a tracheotomy procedure for a child who demonstrated labored respiration during inspiration because of severe glottic stenosis and bilateral vocal cord paralysis caused by tracheal intubation. A 4-year-old boy developed acute respiratory depression associated with influenza pneumonia and had been under respiratory management with mechanical ventilation with tracheal intubation for 3 days. Following extubation, an upper-airway obstruction immediately appeared. The symptoms later worsened because of development of a common cold, and the patient underwent an emergency tracheotomy. For anesthetic management, we used a combination of ketamine with low-concentration sevoflurane inhalation. The tracheotomy was performed safely without respiratory complications by employing manual-assisted ventilation, while spontaneous breathing was preserved by use of a face mask.

Characterization of an intracellular cystatin homolog from the tick Haemaphysalis longicornis.

Cystatins are tight-binding inhibitors of papain-like cysteine proteases. On the basis of amino acid sequencing, cystatins can be subdivided into three closely related families, family 1, family 2 and family 3. Among them, only family 1 cystatins are intracellular. In this report, a gene encoding family 1 cystatin from tick Haemaphysalis longicornis (Hlcyst-1) has been identified. Its full-length cDNA is 437 bp, including an intact ORF encoding an expected protein with 98 amino acids. Sequence analysis demonstrated that it has significant homology with the known family 1 cystatin. The recombinant protein was expressed in a GST-fused soluble form in Escherichia coli, and its inhibitory activity against papain, cathepsin L, and cathepsin B was identified by fluorogenic substrate analysis. This is the first report of an intracellular cystatin homolog from the tick H. longicornis.

Perioperative management of a neonate with Cantrell syndrome.

Cantrell syndrome is a congenital malformation with a pentalogy characterized by defects involving the abdominal wall, lower sternum, anterior diaphragm, and diaphragmatic pericardium, as well as congenital cardiac anomalies. We recently managed anesthesia in a patient with this syndrome and herein report our experience. The patient was a 14-day-old male neonate, who had been diagnosed with Cantrell syndrome, including ventricular septal defect, left ventricular diverticulum, abdominal wall defect, omphalocele, and sternal hypoplasia. Surgical interventions to close the ventricular septal defect, resect the left ventricular diverticulum, and close the omphalocele were scheduled. After cardiac surgery, the hernial contents were returned to their original compartment and, subsequently, an attempt was made to suture the abdominal wall. However, blood pressure fell markedly and the attempt was discontinued. The chest was left open postoperatively and the patient was transferred to the intensive care unit (ICU), during which time circulatory and respiratory management was very complex. Issues requiring particular attention in the management of anesthesia for patients with this syndrome include complications of diverse cardiac malformations, pulmonary hypertension, pulmonary hypoplasia, and respiratory and circulatory failure associated with increased intraabdominal pressure due to primary closure of the omphalocele. Accordingly, extreme caution must be taken to restore respiratory and circulatory control.

Standalone Effects of a Cognitive Behavioral Intervention Using a Mobile Phone App on Psychological Distress and Alcohol Consumption Among Japanese Workers: Pilot Nonrandomized Controlled Trial.

BACKGROUND: Research that investigates standalone effects of a mobile phone-based cognitive behavioral therapy without any human contact for reducing both psychological distress and risky drinking has been advancing; however, the number of studies is still limited. A mobile phone app called Self Record that facilitates cognitive restructuring through self-monitoring of daily thoughts and activities was developed in Japan. OBJECTIVE: This study conducted a nonrandomized controlled pilot trial of the Self Record app to investigate standalone effects of the intervention on psychological distress and alcohol consumption among Japanese workers. Additionally, we examined moderating effects of negative mood regulation expectancies, which are beliefs about one's ability to control one's negative mood. METHODS: A quasi-experimental design with a 1-month follow-up was conducted online in Japan from February 2016 to March 2016. A research marketing company recruited participants. The selection criteria were being a Japanese full-time worker (age 20-59 years), experiencing mild to moderate psychological distress, and having some interest in self-record apps. Assignment to group was based on participants' willingness to use the app in the study. All participants completed outcome measures of negative mood regulation expectancies, positive well-being, general distress, depression, anxiety, and typical/most weekly alcohol consumption. RESULTS: From the recruitment, 15.65% (1083/6921) of participants met the inclusion criteria. Of these, 51.43% (557/1083) enrolled in the study: 54.9% (306/557) in the intervention group and 45.1% (251/557) in the control group. At the 1-month follow-up, 15.3% (85/557) of participants had dropped out. Intention-to-treat analyses revealed that participants in the intervention group reported increased typical drinking (η2=.009) and heavy drinking (η2=.001). Adherence to using the app was low; 64.8% (199/306) of participants in the intervention group discontinued using the app on the first day. Additionally, 65.7% (366/557) of the total sample did not correctly answer the validity checks in the outcome measures (eg, "Please select 'mildly agree' for this item"). Therefore, per-protocol analyses were conducted after removing these participants. Results showed that continuing app users (42/127) in the intervention group reported increases in anxiety (η2=.006), typical drinking (η2=.005), and heavy drinking (η2=.007) compared to those in the control group (85/127). Negative mood regulation expectancies moderated the effects of the intervention for general distress (beta=.39). CONCLUSIONS: Results were contrary to our hypotheses. Self-recording methods of standalone mobile phone interventions may heighten individuals' awareness of their pathological thought and drinking behavior, but may be insufficient to decrease them unless combined with a more intense or face-to-face intervention. Limitations include high attrition in this study; measures to improve the response rate are discussed.

Sequence characterization and expression patterns of two defensin-like antimicrobial peptides from the tick Haemaphysalis longicornis.

Two cDNAs encoding defensin-like antimicrobial peptides were cloned and sequenced from the tick Haemaphysalis longicornis. The full-length cDNA of Hlgut-defensin (H. longicornis midgut defensin) is 333bp, encoding an expected protein with 73 amino acids. The full-length cDNA of Hlsal-defensin (H. longicornis salivary gland defensin) is 382bp, encoding an expected protein with 81 amino acids. The antibacterial activities of the synthetic peptides based on the Hlgut-defensin and Hlsal-defensin sequences were tested against a variety of Gram-positive and Gram-negative bacteria. Using real-time PCR, the tissue-specific expression of two defensin-like peptides were determined and it was also found that the gene transcripts of Hlgut-defensin and Hlsal-defensin were significantly induced by a lipopolysaccharide (LPS) injection.

A secreted cystatin from the tick Haemaphysalis longicornis and its distinct expression patterns in relation to innate immunity.

Proteins capable of selective and specific inhibition of cysteine protease have been identified as cystatins and are isolated from a variety of microbes and tissues of animals and plants. The physiological function of these proteins has been proposed to be the regulation of protein turnover and defense against pathogens as well as the balance of the host-parasite immune relationship. Genes encoding cystatins have been found in several species of ticks, but the function of cystatin in ticks is not understood. We cloned a gene encoding cystatin from tick H. longicornis and designated it as Hlcyst-2 (H. longicornis cystatin-2). Its full-length cDNA is 569 bp, and it encodes a putative 133 amino acid protein with an obvious signal peptide. Sequence analysis demonstrated that it has significant homology with the known cystatin. The recombinant protein was expressed in a GST-fused soluble form in Escherichia coli and purified by affinity chromatography. The inhibitory activity of the recombinant protein against papain, cathepsin L, and cathepsin B was identified by fluorogenic substrate analysis. Cystatin was mostly expressed in the tick midgut and hemocyte. Blood feeding induced significantly increased expression in the midgut. Real-time PCR confirmed that LPS-injected adult ticks expressed Hlcyst-2 1.6 more times than the PBS-injected control; Babesia gibsoni-infected larvae ticks expressed Hlcyst-2 1.8 more times than normal larvae ticks. The recombinant protein also showed a significant growth-inhibitory effect on Babesia bovis cultured in vitro. These results indicated this cystatin Hlcyst-2 is involved in tick innate immunity.

The management of difficult intubation in infants: a retrospective review of anesthesia record database.

We retrospectively reviewed the anesthesia records of infants < 1 year of age for elucidating the incidence of difficult intubation and airway management in a single general hospital. The electronic data records from a total of 753 consecutive anesthesiological procedures in 513 different infants were analyzed. After excluding data with a lack of records of laryngoscopic findings, a total of 497 procedures (389 different infants) with either remarks of difficult intubation (requiring > 10 min for tracheal intubation) or records of Cormack-Lehane grade were included. Demographic data are median age 5 (range, 0-11) months, height 61 (33-84) cm, body weight 6.0 (1.1 - 11.8) kg. The number of cases with ASA physical status I, II, III and IV was 182 (36.6 %), 135 (27.3 %), 177 (35.5 %) and 3 (0.6 %), respectively. Cormack-Lehane grade 1, 2, 3 and 4 was seen in 450 (90.5 %), 32 (6.4 %), 6 (1.2 %) and 6 (1.2 %) cases, respectively. Document of difficult intubation was found in 12 cases (2.4 %, 10 different infants) with a lack of record of Cormack-Lehane grade in 3 cases. Of these 10 infants, nine had multiple congenital anomalies including heart diseases and cleft palate. Without premedication, general anesthesia was induced with intravenous midazolam or sevoflurane in the 12 cases. Tracheal intubation was performed after disappearance of spontaneous respiration except three cases who were intubated in the awake state or under sedation. Elapsed time from induction of anesthesia to intubation was 17 (14-29) min. Although mask ventilation was adequate in all cases, two cases (one infant) developed hypoxia and bradycardia during tracheal intubation. No remarkable decrease of SpO2 or bradycardia less than 100 bpm was detected in other cases. In conclusion, we found difficult intubation in 2.4 % of infants undergoing general anesthesia. Although muscle relaxants are useful for facilitating tracheal intubation, it should be carefully used with the preparation of other airway devices in infants with predicted difficult intubation.

A case of Wegener's granulomatosis associated with progressive dysphagia owing to esophageal involvement.

A 52-year-old woman was admitted to our hospital with progressive dysphagia. Upper gastrointestinal endoscopy revealed esophageal stenosis and computed tomographic scan revealed symmetrical wall thickness of the thoracic esophagus. Biopsies findings from a lesion were unremarkable. However, a definitive diagnosis of Wegener's granulomatosis was based on positive anti-neutrophil cytoplasmic antibodies directed against proteinase 3 and otorhinolaryngological manifestations. Esophageal complications are rarely reported in Wegener's granulomatosis; however, clinicians should be aware of the possibility of esophageal involvement.

Phosphaturic mesenchymal tumor with symptoms related to osteomalacia that appeared one year after tumorectomy.

A 45-year-old man was admitted to our hospital because of bone pain and hypophosphatemia. He had undergone surgery 2 years previously for a "benign unclassified mesenchymal tumor" in the skull, but there were no clinical symptoms related to osteomalacia. His laboratory examination revealed low serum phosphate, high alkaline phosphatase, and normal calcium levels. The diagnosis of tumor-induced osteomalacia due to phosphaturic mesenchymal tumor mixed connective tissue variant (PMTMCT) was made by re-examining the pathologic specimens. Oral supplementation with phosphate and 1-25-dihydroxyvitamin D relieved his clinical symptoms and laboratory values returned to normal. However, subcutaneous administration of octreotide had no clinical effect. Clinicians and pathologists should be aware of the existence of PMTMCT especially nonphosphaturic or asymptomatic variants of this disorder.