Emergence and Persistent Dominance of SARS-CoV-2 Omicron BA.2.3.7 Variant, Taiwan.

Since April 2022, waves of SARS-CoV-2 Omicron variant cases have surfaced in Taiwan and spread throughout the island. Using high-throughput sequencing of the SARS-CoV-2 genome, we analyzed 2,405 PCR-positive swab samples from 2,339 persons and identified the Omicron BA.2.3.7 variant as a major lineage within recent community outbreaks in Taiwan.

Voluntary attendance of small-group brainstorming tutoring courses intensify new clerk's "excellence in clinical care": a pilot study.

BACKGROUND: Clerkship provides a unique way of transferring the knowledge and skills gathered during medical school's curriculum into real-ward clinical care environment. The annual program evaluation has indicated that the training of clerks in diagnostic and clinical reasoning skills needed to be enhanced. Recently, "clinical excellence" program have been promoted in our institution to augment the excellence in clinical care of new clerks. Current study aims to evaluate whether this pilot program improve the "clinical excellence" of new clerks. METHODS: In a pilot study, groups of new clerks in years 2013 and 2014 voluntarily attended either a small-group brainstorming course or a didactic classroom tutoring courses as part of their 3-month internal medicine clinical rotation block. A third group of new clerks did not join either of the above courses and this group served as the control group. Pre-block/post-block self-assessment and post-block 5-station mini-Objective Subjective Clinical Examinations (OSCEs) were used to evaluate the effectiveness of these two additional courses that trained diagnostic and clinical reasoning skills. RESULTS: Overtime, the percentages of new clerks that attended voluntarily either the small-group brainstorming or classroom tutoring courses were increased. Higher post-block self-assessed diagnostic and clinical reasoning skill scores were found among individuals who attended the small-group brainstorming courses compared to either the didactic group or the control group. In a corresponding manner, the small-group brainstorming group obtained higher summary OSCEdiag and OSCEreason scores than either the didactic group or control group. For all basic images/laboratory OSCE stations, the individual diagnostic skill (OSCEdiag) scores of the small-group brainstorming group were higher than those of the didactic group. By way of contrast, only the clinical reasoning skill (OSCEreason) scores of the basic electrocardiogram and complete blood count + biochemistry OSCE station of thesmall-group brainstorming group were higher than those of the didactic group. Among the small-group brainstorming group, clerks with higher cumulative learning hours (>30-h) had significant higher OSCEdiag and OSCEreason scores (>400) than those with less cumulative learning hours. CONCLUSION: Our pilot study provides a successful example of the use of a small-group tutoring courses for augmenting the diagnostic and clinical reasoning skills of new clerks. The positive results obtained during the initial 2-year long pilot "clinical excellence" program have encouraged the formal implementation of this course as part of the clerkship curriculum.

A persistent level of Cisd2 extends healthy lifespan and delays aging in mice.

The CISD2 gene, which is an evolutionarily conserved novel gene, encodes a transmembrane protein primarily associated with the mitochondrial outer membrane. Significantly, the CISD2 gene is located within the candidate region on chromosome 4q where a genetic component for human longevity has been mapped. Previously, we have shown that Cisd2 deficiency shortens lifespan resulting in premature aging in mice. Additionally, an age-dependent decrease in Cisd2 expression has been detected during normal aging. In this study, we demonstrate that a persistent level of Cisd2 achieved by transgenic expression in mice extends their median and maximum lifespan without any apparent deleterious side effects. Cisd2 also ameliorates age-associated degeneration of the skin, skeletal muscles and neurons. Moreover, Cisd2 protects mitochondria from age-associated damage and functional decline as well as attenuating the age-associated reduction in whole-body energy metabolism. These results suggest that Cisd2 is a fundamentally important regulator of lifespan and provide an experimental basis for exploring the candidacy of CISD2 in human longevity.

The quantitative assessment of the role played by basic amino acid clusters in the nuclear uptake of human ribosomal protein L7.

In this study, we used a multiple copy (EGFP)(3) reporter system to establish a numeric nuclear index system to assess the degree of nuclear import. The system was first validated by a FRAP assay, and then was applied to evaluate the essential and multifaceted nature of basic amino acid clusters during the nuclear import of ribosomal protein L7. The results indicate that the sequence context of the basic cluster determines the degree of nuclear import, and that the number of basic residues in the cluster is irrelevant; rather the position of the pertinent basic residues is crucial. Moreover, it also found that the type of carrier protein used by basic cluster has a great impact on the degree of nuclear import. In case of L7, importin ß2 or importin ß3 are preferentially used by clusters with a high import efficiency, notwithstanding that other importins are also used by clusters with a weaker level of nuclear import. Such a preferential usage of multiple basic clusters and importins to gain nuclear entry would seem to be a common practice among ribosomal proteins in order to ensure their full participation in high rate ribosome synthesis.

Late-assembly of human ribosomal protein S20 in the cytoplasm is essential for the functioning of the small subunit ribosome.

Using immuno-fluorescent probing and Western blotting analysis, we reveal the exclusive cytoplasm nature of the small subunit ribosomal protein S20. To illustrate the importance of the cellular compartmentation of S20 to the function of small subunit 40S, we created a nuclear resident S20NLS mutant gene and examined polysome profile of cells that had been transfected with the S20NLS gene. As a result, we observed the formation of recombinant 40S carried S20NLS but this recombinant 40S was never found in the polysome, suggesting such a recombinant 40S was translation incompetent. Moreover, by the tactic of the energy depletion and restoration, we were able to restrain the nuclear-resided S20NLS in the cytoplasm. Yet, along a progressive energy restoration, we observed the presence of recombinant 40S subunits carrying the S20NLS in the polysome. This proves that S20 needs to be cytoplasmic in order to make a functional 40S subunit. Furthermore, it also implies that the assembly order of ribosomal protein in eukaryote is orderly regulated.

Draft genome sequence of the human pathogen Streptomyces somaliensis, a significant cause of actinomycetoma.

We report the draft genome sequence of the human pathogen Streptomyces somaliensis (DSM 40738), a pathogen within a genus of largely saprophytic organisms. S. somaliensis causes severe and debilitating deep tissue and bone infections. The genome sequence is deposited in DDBJ/EMBL/GenBank with the accession number AJJM01000000.

Analysis of developmental gene conservation in the Actinomycetales using DNA/DNA microarray comparisons.

Based on available genome sequences, Actinomycetales show significant gene synteny across a wide range of species and genera. In addition, many genera show varying degrees of complex morphological development. Using the presence of gene synteny as a basis, it is clear that an analysis of gene conservation across the Streptomyces and various other Actinomycetales will provide information on both the importance of genes and gene clusters and the evolution of morphogenesis in these bacteria. Genome sequencing, although becoming cheaper, is still relatively expensive for comparing large numbers of strains. Thus, a heterologous DNA/DNA microarray hybridization dataset based on a Streptomyces coelicolor microarray allows a cheaper and greater depth of analysis of gene conservation. This study, using both bioinformatical and microarray approaches, was able to classify genes previously identified as involved in morphogenesis in Streptomyces into various subgroups in terms of conservation across species and genera. This will allow the targeting of genes for further study based on their importance at the species level and at higher evolutionary levels.

An Actinobacterial Isolate, Streptomyces sp. YX44, Produces Broad-Spectrum Antibiotics That Strongly Inhibit Staphylococcus aureus.

The need for new antibiotics is increasing due to their overuse, and antibiotic resistance has become one of the major threats worldwide to public health, food safety, and clinical treatment. In this study, we describe an actinobacterial isolate, YX44, which belongs to the genus Streptomyces. This Streptomyces was isolated from a drinking pipe located in Osaka, Japan, and has the ability to inhibit Gram-positive bacteria, Gram-negative bacteria, and various fungi. YX44 fermentation broth shows strong activity against Escherichia coli and Staphylococcus aureus, as well as also inhibiting clinical isolates of multidrug-resistant Staphylococcus aureus. The YX44 antibacterial substances in the broth are relatively heat-stable, show high stability from the pH range 1 to 11, and have good solubility in both organic and non-organic solvents. Size-exclusion chromatography revealed that the YX44 antibacterial compounds are less than 1000 Da in size. LC-MS was able to identify three possible candidate molecules with molecular weights of 308, 365, 460, and 653 g/mol; none of these sizes correspond to any well-known antibiotics. Our results show that Streptomyces sp. YX44 seems to produce a number of novel antibiotics with high pH stability and good solubility that have significant activity against S. aureus, including multidrug-resistant strains.

Diagnosis of Arboleda-Tham syndrome by whole genome sequencing in an Asian boy with severe developmental delay.

Diagnosis of a 9-month-old boy brought to our genetics clinic with chief complaints of developmental delay (DD), failure to thrive, microcephaly, trunk hypotonia and hypertonia of the extremities. Multiple congenital defects but no significant syndromes or diseases were impressed. The chromosomal analysis and array comparative genomic hybridization (aCGH) revealed no significant pathogenic changes. Whole Genome Sequencing (WGS) identified a p.Glu1139fs de novo mutation of the KAT6A gene. The patient's phenotype was consistent clinically with Arboleda-Tham syndrome (ARTHS). Reviewing the literature showed that this is the first patient in Taiwan detected by WGS and that it involves a novel mutation. Comparing the highly variable clinical presentations of this syndrome with our patient, this boy's features and severe developmental defects seem to be due to a late-truncating mutation at the carboxyl end of the KAT6A protein. Our study demonstrates the power of WGS to confirm a diagnosis within 4 weeks for this rare condition.

Genome sequencing and comparative analysis of Klebsiella pneumoniae NTUH-K2044, a strain causing liver abscess and meningitis.

Nosocomial infections caused by antibiotic-resistant Klebsiella pneumoniae are emerging as a major health problem worldwide, while community-acquired K. pneumoniae infections present with a range of diverse clinical pictures in different geographic areas. In particular, an invasive form of K. pneumoniae that causes liver abscesses was first observed in Asia and then was found worldwide. We are interested in how differences in gene content of the same species result in different diseases. Thus, we sequenced the whole genome of K. pneumoniae NTUH-K2044, which was isolated from a patient with liver abscess and meningitis, and analyzed differences compared to strain MGH 78578, which was isolated from a patient with pneumonia. Six major types of differences were found in gene clusters that included an integrative and conjugative element, clusters involved in citrate fermentation, lipopolysaccharide synthesis, and capsular polysaccharide synthesis, phage-related insertions, and a cluster containing fimbria-related genes. We also conducted comparative genomic hybridization with 15 K. pneumoniae isolates obtained from community-acquired or nosocomial infections using tiling probes for the NTUH-K2044 genome. Hierarchical clustering revealed three major groups of genomic insertion-deletion patterns that correlate with the strains' clinical features, antimicrobial susceptibilities, and virulence phenotypes with mice. Here we report the whole-genome sequence of K. pneumoniae NTUH-K2044 and describe evidence showing significant genomic diversity and sequence acquisition among K. pneumoniae pathogenic strains. Our findings support the hypothesis that these factors are responsible for the changes that have occurred in the disease profile over time.

Genetic diversity of capsular polysaccharide biosynthesis in Klebsiella pneumoniae clinical isolates.

Klebsiella pneumoniae is an enteric pathogen causing community-acquired and hospital-acquired infections in humans. Epidemiological studies have revealed significant diversity in capsular polysaccharide (CPS) type and clinical manifestation of K. pneumoniae infection in different geographical areas of the world. We have sequenced the capsular polysaccharide synthesis (cps) region of seven clinical isolates and compared the sequences with the publicly available cps sequence data of five strains: NTUH-K2044 (K1 serotype), Chedid (K2 serotype), MGH78578 (K52 serotype), A1142 (K57 serotype) and A1517. Among all strains, six genes at the 5' end of the cps clusters that encode proteins for CPS transportation and processing at the bacterial surface are highly similar to each other. The central region of the cps gene clusters, which encodes proteins for polymerization and assembly of the CPS subunits, is highly divergent. Based on the collected sequence, we found that either the wbaP gene or the wcaJ gene exists in a given K. pneumoniae strain, suggesting that there is a major difference in the CPS biosynthesis pathway and that the K. pneumoniae strains can be classified into at least two distinct groups. All isolates contain gnd, encoding gluconate-6-phosphate dehydrogenase, at the 3' end of the cps gene clusters. The rmlBADC genes were found in CPS K9-positive, K14-positive and K52-positive strains, while manC and manB were found in K1, K2, K5, K14, K62 and two undefined strains. Our data indicate that, while overall genomic organization is similar between different pathogenic K. pneumoniae strains, the genetic variation of the sugar moiety and polysaccharide linkage generate the diversity in CPS molecules that could help evade host immune attack.

Two-component signal transduction systems in Streptomyces and related organisms studied using DNA comparative microarray analysis.

Two component sensor-response regulator systems (TCSs) are very common in the genomes of the Streptomyces species that have been fully sequenced to date. It has been suggested that this large number is an evolutionary response to the variable environment that Streptomyces encounter in soil. Notwithstanding this, TCSs are also more common in the sequenced genomes of other Actinomycetales when these are compared to the genomes of most other eubacteria. In this study, we have used DNA/DNA genome microarray analysis to compare 14 Streptomyces species and one closely related genus to Streptomyces coelicolor in order to identify a core group of such systems. This core group is compared to the syntenous and non-syntenous TCSs present in the genome sequences of other Actinomycetales in order to separate the systems into those present in Actinomycetales in general, the Streptomyces specific systems and the species specific systems. Horizontal transfer does not seem to play a very important role in the evolution of the TCS complement analyzed in this study. However, cognate pairs do not necessarily seem to evolve at the same pace, which may indicate the evolutionary responses to environmental variation may be reflected differently in sequence changes within the two components of the TCSs. The overall analysis allowed subclassification of the orphan TCSs and the TCS cognate pairs and identification of possible targets for further study using gene knockouts, gene overexpression, reporter genes and yeast two hybrid analysis.

Once the circle has been broken: dynamics and evolution of Streptomyces chromosomes.

Chromosomal instability has been a hallmark of Streptomyces genetics. Deletions and circularization often occur in the less-conserved terminal sequences of the linear chromosomes, which contain swarms of transposable elements and other horizontally transferred elements. Intermolecular recombination involving these regions also generates gross exchanges, resulting in terminal inverted repeats of heterogeneous size and context. The structural instability is evidently related to evolution of the Streptomyces chromosomes, which is postulated to involve linearization of hypothetical circular progenitors via integration of a linear plasmid. This scenario is supported by several bioinformatic analyses.

Unintentional ingestion of Cordyceps fungus-infected cicada nymphs causing ibotenic acid poisoning in Southern Vietnam.

BACKGROUND: Cordyceps fungus found in infected cicada nymphs ("cicada flowers") is utilized in traditional Chinese medicine. Cordyceps fungus toxicity in humans has not been previously reported. We report 60 cases of apparent Cordyceps poisoning in Southern Vietnam. METHODS: We retrospectively collected demographic and clinical data from the medical records (21 cases) and by telephone interview (39 cases) of patients admitted to seven hospitals in Southern Vietnam following ingestion of cicada flowers between 2008 and 2015. We also determined the species of Cordyceps present in the cicada flowers and performed a partial chemical analysis of the fungus. RESULTS: Sixty cases of toxic effects following ingestion of cicada flowers were documented. Symptom onset occurred within 60 minutes following ingestion. Symptoms included dizziness, vomiting, salivation, mydriasis, jaw stiffness, urinary retention, seizures, agitated delirium, hallucinations, somnolence and coma. None of the patients suffered liver or kidney injury. There was one fatality. The Cordyceps fungus involved in these poisoning was identified as Ophiocordyceps heteropoda. The presence of ibotenic acid was confirmed, but musimol and muscarine were absent. CONCLUSIONS: Cicada infected with Ophiocordyceps heteropoda in Vietnam contain ibotenic acid and are associated with a clinical syndrome consistent with its effects.

Effects of a new parallel primary healthcare centre and on-campus training programme on history taking, physical examination skills and medical students' preparedness: a prospective comparative study in Taiwan.

OBJECTIVES: The primary healthcarecentre (PHCC) is the first place that medical students experience patient contact. Usually, medical students are frustrated by a lack of proper skills training for on-campus history taking (HT), physical examination (PE) and self-directed learning (SDL) to prepare for their PHCC and inhospital patient contact. For pre-clerks, this study aims to compare the effectiveness of PHCC training and PHCC training in combination with on-campus HT and PE training modules (PHCC+on-campus) on their clerkship preparedness. DESIGN: This comparative study utilised prospective, consecutive, end of pre-clerkship group objective structured clinical examination (GOSCE), beginning of clerkship OSCE and self-administered Preparation for Hospital Practice Questionnaire (PHPQ). SETTING/PARTICIPANTS: 128 pre-clinical clerk volunteers (64 each year) receiving PHCC training (7 week PHCCtraining in addition to 7 week assignment based group learning, academic year 2014, controls) and PHCC training in combination with on-campus module training (academic year 2015, 7 week PHCCtraining in addition to 7 week on-campus sessions) were sequentially assessed before the module (week 1), at the end of the module (week 14) and at the beginning of clerkship (week 25). RESULTS: For overall HT and PE skills, both PHCC and PHCC+on-campus module trained pre-clerks performed better on OSCE than GOSCE. Additionally, the improvement was accompanied by higher self-reported PHPQ scores in 'confidence/coping' and 'SDL' domains. At the end of the pre-clerkship and the beginning of the clerkship stages, the degree of improvement in preparedness in 'confidence/coping' and 'SDL' domains was higher for those in the PHCC+on-campus group than for those in the PHCC group. Among the PHCC+on-campus module participants, a positive association was observed between high mean PHPQ-SDL scores and high OSCE scores. CONCLUSIONS: Our study suggests that the PHCC+on-campus module, which is paired faculty led and pre-trained dyad student assisted, is effective in developing a preclinical clerk's HT and PE skills and intensifying SDL/patient management abilities to prepare for hospital practice in clerkship.

A model of four hierarchical levels to train Chinese residents' teaching skills for "practice-based learning and improvement" competency.

OBJECTIVES: The current study focused on validating a protocol for training and auditing the resident's practice-based learning and improvement (PBLI) and quality improvement (QI) competencies for primary care. METHODS: Twelve second-year (R2), 12 first-year (R1) and 12 postgraduate year-1 residents were enrolled into group A, B and C, respectively, as trainees. After three training protocols had been completed, a writing test, self-assessed questionnaire and mini-OSTE and end-of-rotation assessment were used in auditing the PBLI competency, performance and teaching ability of trainees. RESULTS: Baseline expert-assessed PBLI and QI knowledge application tool writing scores were low for the R1 and R2 residents. After three training protocols, PBLI and QI proficiencies, performance and teaching abilities were improved to similar levels cross the three training levels of residents based on the expert-assessed writing test-audited assessments and on the faculty and standardized clerk-assessed end-of-rotation-/mini-OSTE-audited assessments. CONCLUSION: The different four-level hierarchical protocols used to teach group A, B and C were equally beneficial and fitted their needs; namely the different levels of the trainees. Specifically, each level was able to augment their PBLI and QI proficiency. This educational intervention helps medical institutions to train residents as PBLI instructors.

Draft Genome Sequence of the Oxytetracycline-Producing Bacterium Streptomyces rimosus ATCC 10970.

We report the draft genome of Streptomyces rimosus (ATCC 10970), a soil isolate that produces oxytetracycline, a commercially important and clinically useful antibiotic.

Copy Number Change of the NDM-1 sequence in a multidrug-resistant Klebsiella pneumoniae clinical isolate.

The genetic features of the antimicrobial resistance of a multidrug resistant Klebsiella pneumoniae strain harboring bla NDM-1 were investigated to increase our understanding of the evolution of NDM-1. The strain, KPX, came from a Taiwanese patient with a hospitalization history in New Delhi. Complete DNA sequencing was performed; and the genes responsible for antimicrobial resistance were systematically examined and isolated by library screening. KPX harbored two resistance plasmids, pKPX-1 and pKPX-2, which are 250-kb and 141-kb in size, respectively, with bla NDM-1 present on pKPX-1. The plasmid pKPX-1 contained genes associated with the IncR and IncF groups, while pKPX-2 belonged to the IncF family. Each plasmid carried multiple antimicrobial resistance genetic determinants. The gene responsible for resistance to carbapenems was found on pKPX-1 and that for resistance to aztreonam was found on pKPX-2. To our surprise, we discovered that bla NDM-1 exists on pKPX-1 as multiple copies in the form of tandem repeats. Amplification of bla NDM-1 was found to occur by duplication of an 8.6-kb unit, with the copy number of the repeat varying from colony to colony. This repeat sequence is identical to that of the pNDM-MAR except for two base substitutions. The copy number of bla NDM-1 of colonies under different conditions was assessed by Southern blotting and quantitative PCR. The bla NDM-1 sequence was maintained in the presence of the antimicrobial selection; however, removal of antimicrobial selection led to the emergence of susceptible bacterial populations with a reduced copy number or even the complete loss of the bla NDM-1 sequence. The dynamic nature of the NDM-1 sequence provides a strong argument for judicious use of the broad-spectrum antimicrobials in order to reduce the development and spread of antimicrobial resistance among pathogens.

Chromosome diversity and similarity within the Actinomycetales.

Many chromosomes from Actinomycetales, an order within the Actinobacteria, have been sequenced over the last 10 years and the pace is increasing. This group of Gram-positive and high G+C% bacteria is economically and medically important. However, this group of organisms also is just about the only order in the kingdom Bacteria to have a relatively high proportion of linear chromosomes. Chromosome topology varies within the order according to the genera. Streptomyces, Kitasatospora and Rhodococcus, at least as chromosome sequencing stands at present, have a very high proportion of linear chromosomes, whereas most other genera seem to have circular chromosomes. This review examines chromosome topology across the Actinomycetales and how this affects our concepts of chromosome evolution.