Cryptococcus tetragattii Meningitis Associated with Travel, Taiwan.

Meningitis caused by Cryptococcus tetragattii fungus is rare and has been found in specific geographic regions. We report a case of meningitis caused by C. tetragattii (molecular type VGIV) in an immunocompetent patient in Taiwan. The patient had traveled to Egypt and was positive for granulocyte-macrophage colony-stimulating factor autoantibody.

Staphylococcus taiwanensis sp. nov., isolated from human blood.

A novel coagulase-negative Staphylococcus strain (NTUH-S172T) was isolated from human blood culture in Taiwan with preliminary identification of Staphylococcus saprophyticus. 16S rRNA gene analysis and multilocus sequence analysis (MLSA) showed that NTUH-S172T was most closely related to Staphylococcus haemolyticus. The average nucleotide identity and digital DNA-DNA hybridization values with the whole genome sequence were <95 % and<70 % when compared to the related species. Strain NTUH-S172T could be distinguished from S. haemolyticus by urease production and from Staphylococcus borealis by nitrate reduction. In addition, the matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) spectrum of NTHU-S172T was significantly different from that of S. haemolyticus, which could be used in clinical identification. In conclusion, it is proposed that this isolate represents a novel species, named Staphylococcus taiwanensis sp. nov., with type strain NTUH-S172T (=BCRC 81315T=JCM 34726T).

Emergence of aac(6')-Ie-aph(2'')-Ia-positive enterococci with non-high-level gentamicin resistance mediated by IS1216V: adaptation to decreased aminoglycoside usage in Taiwan.

OBJECTIVES: To explore the mechanisms mediating the different levels of gentamicin resistance in enterococci. METHODS: Susceptibility testing with gentamicin and PCR of resistance determinants were performed in 149 enterococcal isolates. Genetic relatedness was characterized by MLST and PFGE analysis. Sequences of the aac(6')-Ie-aph(2'')-Ia gene and its surrounding environment were determined by Illumina sequencing. Stability assays of gentamicin resistance were carried out to evaluate the probability of loss of the high-level gentamicin resistance (HLGR) phenotype. RESULTS: A total of 17 (11.4%) aac(6')-Ie-aph(2'')-Ia-positive enterococcal isolates (2 Enterococcus faecalis and 15 Enterococcus faecium) with non-HLGR phenotype were found. MLST analysis revealed that the 2 E. faecalis belonged to ST116 and ST618, while all the 15 E. faecium belonged to clonal complex 17. Sequence analysis demonstrated that IS1216V was inserted into the 5'-end of aac(6')-Ie-aph(2'')-Ia, leading to loss of HLGR phenotype. Three IS1216V insertion types were found, and type II and III were frequently found in E. faecium. Interestingly, a total of 38 aac(6')-Ie-aph(2'')-Ia-positive E. faecium with HLGR phenotype also had type II or type III IS1216V insertion. Sequencing of the aac(6')-Ie-aph(2'')-Ia-positive HLGR E. faecium E37 revealed that an intact aac(6')-Ie-aph(2'')-Ia was located adjacent to IS1216V-disrupted aac(6')-Ie-aph(2'')-Ia. In a non-antibiotic environment, E37 tended to lose HLGR phenotype with a probability of 1.57 × 10-4, which was largely attributed to homologous recombination between the intact and disrupted aac(6')-Ie-aph(2'')-Ia. CONCLUSIONS: This is first study to elucidate that the E. faecium is capable of changing its HLGR phenotype, which may contribute to adaptation to hospital environments with decreased usage of gentamicin.

Aeromonas Isolates from Fish and Patients in Tainan City, Taiwan: Genotypic and Phenotypic Characteristics.

The present study aimed to isolate Aeromonas from fish sold in the markets as well as in sushi and seafood shops and compare their virulence factors and antimicrobial characteristics with those of clinical isolates. Among the 128 fish isolates and 47 clinical isolates, Aeromonas caviae, A. dhakensis, and A. veronii were the principal species. A. dhakensis isolates carried at least 5 virulence genes, more than other Aeromonas species. The predominant genotype of virulence genes was hlyA lip alt col ela in both A. dhakensis and A. hydrophila isolates, alt col ela in A. caviae isolates, and act in A. veronii isolates. A. dhakensis, A. hydrophila, and A. veronii isolates more often exhibited hemolytic and proteolytic activity and showed greater virulence than A. caviae isolates in Caenorhabditis elegans and the C2C12 cell line. However, the link between the genotypes and phenotypes of the studied virulence genes in Aeromonas species was not evident. Among the four major clinical Aeromonas species, nearly all (99.0%) A. dhakensis, A. hydrophila, and A. veronii isolates harbored blaCphA, which encodes a carbapenemase, but only a minority (6.7%, 7/104) were nonsusceptible to carbapenem. Regarding AmpC ß-lactamase genes, blaAQU-1 was exclusively found in A. dhakensis isolates, and blaMOX3 was found only in A. caviae isolates, but only 7.6% (n = 6) of the 79 Aeromonas isolates carrying blaAQU-1 or blaMOX3 exhibited a cefotaxime resistance phenotype. In conclusion, fish Aeromonas isolates carry a variety of combinations of virulence and ß-lactamase resistance genes and exhibit virulence phenotypes and antimicrobial resistance profiles similar to those of clinical isolates.IMPORTANCEAeromonas species can cause severe infections in immunocompromised individuals upon exposure to virulent pathogens in the environment, but the characteristics of environmental Aeromonas species remain unclear. Our study showed that several pathogenic Aeromonas species possessing virulence traits and antimicrobial resistance similar to those of Aeromonas isolates causing clinical diseases were present in fish intended for human consumption in Tainan City, Taiwan.

Distribution of Staphylococcal Cassette Chromosome (SCC) mec Element Types in Fusidic Acid-Resistant Staphylococcus epidermidis and Identification of a Novel SCC7684 Element.

We analyzed the staphylococcal cassette chromosome mec (SCCmec) types of 143 fusidic acid- and methicillin-resistant Staphylococcus epidermidis isolates. The most frequent SCCmec type was SCCmec III/SCCHg (53%), followed by SCCmec IV (29%). Clonal spreading of SCCmec III/SCCHg strains contributed to the increased prevalence of SCCmec III. A novel non-mec SCC structure, SCC7684, adjacent to SCCmec III, which carries a new ccrC allotype (ccrC3 allele 1) and contains heavy metal resistance genes, was identified in 14 isolates.

Novel Structure of Enterococcus faecium-Originated ermB-Positive Tn1546-Like Element in Staphylococcus aureus.

We determined the resistance determinants in 274 erythromycin-resistant methicillin-susceptible Staphylococcus aureus (MSSA) isolates during a 13-year period, 2000 to 2012. The resistance phenotypes, inducible macrolide-lincosamide-streptogramin (iMLS), constitutive MLS (cMLS), and macrolide-streptogramin (MS) resistance phenotypes, were examined by a double-disk diffusion D test. The ermB gene was more frequent (35%; 97/274) than ermC (27%; 75/274) or ermA (21%; 58/274). All 97 ermB-positive isolates harbored Tn551 and IS1216V The majority (89/97) of ermB-positive isolates displayed the cMLS phenotype and carried mobile element structure (MES)-like structures, which has been previously reported in sequence type 59 (ST59) methicillin-resistant S. aureus (MRSA). The remaining 8 ermB-carrying isolates, belonging to ST7 (n = 4), ST5 (n = 3), and ST59 (n = 1), were sasK intact and did not carry MES-like structures. Unlike a MES-like structure that was located on the chromosome, the ermB elements on sasK-intact isolates were located on plasmids by S1 nuclease pulsed-field gel electrophoresis (PFGE) analysis and conjugation tests. Sequence data for the ermB-containing region (14,566 bp) from ST59 NTUH_3874 revealed that the best match was a Tn1546-like element in plasmid pMCCL2 DNA (GenBank accession number AP009486) of Macrococcus caseolyticus Tn1546 is recognized as an enterococcal transposon and was known from the vancomycin resistance gene cluster in vancomycin-resistant Enterococcus (VRE). So far, acquisitions of Tn1546 in S. aureus have occurred in clonal complex 5 (CC5) MRSA, but not in MSSA. This is the first report that MSSA harbors an Enterococcus faecium-originated ermB-positive Tn1546-like element located on a plasmid.

Emergence of a small colony variant of vancomycin-intermediate Staphylococcus aureus in a patient with septic arthritis during long-term treatment with daptomycin.

OBJECTIVES: Small colony variants (SCVs) of Staphylococcus aureus are associated with persistent and drug-resistant infections. We demonstrated for the first time the emergence of SCVs in a patient with vancomycin-intermediate S. aureus (VISA) infection during long-term treatment with daptomycin. METHODS: A 73-year-old man with septic arthritis was infected with VISA. The patient was treated with daptomycin; however, the patient remained infected with VISA, with continuous isolation of VISA from his blood during long-term treatment. Five VISA isolates were characterized by: PFGE; genotyping including staphylococcal cassette chromosome mec (SCCmec), spa and MLST; antimicrobial susceptibility testing; and scanning and transmission electron microscopy. WGS and fatty acid analysis were also performed. RESULTS: The five VISA isolates were from a single clone of ST239/spa3(t037) and, of these, the first three were SCCmecIII positive and daptomycin susceptible, whereas the last two were SCCmecIII negative and daptomycin resistant and exhibited the characteristics of SCVs. The first and last isolates showed 13 remarkable genetic differences in SCCmec and the mprF, cls2, clpX and fabF genes. Of these, mutation of fabF (encoding the fatty acid synthase) seemed to be partially responsible for the slow growth and ultrastructural features, including an abnormal intercellular substance, and for the daptomycin resistance of SCVs. CONCLUSIONS: For the first time, we identified SCVs of VISA in a patient with septic arthritis during long-term treatment with daptomycin. Daptomycin-resistant SCVs of VISA were evolved in a stepwise manner and the mutation of fabF is likely responsible for the physical and ultrastructural characteristics and daptomycin resistance.

A novel fusidic acid resistance determinant, fusF, in Staphylococcus cohnii.

OBJECTIVES: To determine MICs of fusidic acid for and identify genetic determinants of resistance in Staphylococcus cohnii isolates. METHODS: Susceptibility to fusidic acid was determined by the standard agar dilution method in 24 S. cohnii subsp. urealyticus clinical isolates, 7 S. cohnii subsp. cohnii clinical isolates and 2 reference strains. Sequencing of a novel resistance determinant, fusF, and its flanking regions was performed by long and accurate PCR and inverse PCR. To evaluate the function of fusF, the MIC of fusidic acid was determined for recombinant Staphylococcus aureus carrying a plasmid expressing fusF. RESULTS: A total of 25 S. cohnii subsp. urealyticus (24 clinical isolates and 1 reference strain) and 2 S. cohnii subsp. cohnii displayed low-level resistance to fusidic acid (MICs 2-16 mg/L). Sequencing of a 4259 bp fragment from S. cohnii subsp. urealyticus ATCC 49330 revealed a novel resistance gene, designated fusF, which displayed 70.5% nucleotide and 67.3% amino acid identity to fusD. Expression of fusF in S. aureus confers resistance to fusidic acid. CONCLUSIONS: A novel FusB-family gene, fusF, was identified as a major resistance determinant in S. cohnii clinical isolates resistant to fusidic acid.

A novel staphylococcal cassette chromosomal element, SCCfusC, carrying fusC and speG in fusidic acid-resistant methicillin-resistant Staphylococcus aureus.

A high prevalence of fusC (16/46, 59%) was found in fusidic acid-resistant methicillin-resistant Staphylococcus aureus isolates collected from 2008 to 2010. Nucleotide sequencing of fusC and flanking regions revealed a novel staphylococcal cassette chromosome (SCC) structure, SCCfusC, which was integrated into rlmH and located upstream from SCCmec. The SCCfusC element contained speG, which may contribute to the polyamine resistance.

Comparison of ERIC carbapenem-resistant Enterobacteriaceae test, BD Phoenix CPO detect panel, and NG-test CARBA 5 for the detection of main carbapenemase types of carbapenem-resistant Enterobacterales.

BACKGROUND: This study aimed to assess the performance of three commercial panels, the ERIC Carbapenem-Resistant Enterobacteriaceae Test (ERIC CRE test), the NG-Test CARBA 5 (NG CARBA 5), and the BD Phoenix CPO Detect Panel (CPO panel), for the detection of main types of carbapenemases among carbapenem-resistant Enterobacterales (CRE). METHODS: We collected 502 isolates of carbapenem-resistant Enterobacterales (CRE) demonstrating intermediate or resistant profiles to at least one carbapenem antibiotic (ertapenem, imipenem, meropenem, or doripenem). Carbapenemase genes and their specific types were identified through multiplex PCR and sequencing methods. Subsequently, the ERIC CRE test, CPO panel, and NG CARBA 5 assay were conducted on these isolates, and the results were compared with those obtained from multiplex PCR. RESULTS: The results indicated that the ERIC CRE test exhibited an overall sensitivity and specificity of 98.1% and 93.6%, respectively, which were comparable to 99.1% and 90.6% for the NG CARBA 5. However, the CPO panel demonstrated a sensitivity of only 56.2% in identifying Ambler classes, exhibiting the poorest sensitivity for class A. Moreover, while the ERIC CRE test outperformed the NG CARBA 5 in identifying multi-gene isolates with multiple carbapenemase-encoding genes, the CPO panel failed to accurately classify these isolates. CONCLUSIONS: Our findings support the utilization of the ERIC CRE test as one of the methods for detecting carbapenemases in clinical laboratories. Nonetheless, further optimization is imperative for the CPO panel to enhance its accuracy in determining carbapenemase classification and address limitations in detecting multi-gene isolates.

Staphylococcus hsinchuensis sp. nov., Isolated from Soymilk.

A novel coagulase-negative Staphylococcus strain (H164T) was isolated from soymilk in Taiwan. Comparative sequence analysis of the 16S rRNA gene revealed that the H164T strain is a member of the genus Staphylococcus. We used multilocus sequence analysis (MLSA) and phylogenomic analyses to demonstrate that the novel strain was closely related to Staphylococcus gallinarum, Staphylococcus nepalensis, Staphylococcus cohnii, and Staphylococcus urealyuticus. The average nucleotide identity and digital DNA-DNA hybridization values between H164T and its closest relatives were <95% and <70%, respectively. The H164T strain could also be distinguished from its closest relatives by the fermentation of d-fructose, d-maltose, d-trehalose, and d-mannitol, as well as by the activities of α-glucosidase and alkaline phosphatase. The major cellular fatty acids were C15:0 iso and C15:0 anteiso, and the predominant menaquinones were MK-7 and MK-8, respectively. The major cellular fatty acids and predominant menaquinones were C15:0 iso and C15:0 anteiso and MK-7 and MK-8, respectively. In conclusion, this strain represents a novel species, named Staphylococcus hsinchuensis sp. nov., with the type strain H164T (=BCRC 81404T = NBRC 116174T).

New structure of phage-related islands carrying fusB and a virulence gene in fusidic acid-resistant Staphylococcus epidermidis.

Nucleotide sequencing of the fusB-flanking regions in two fusidic acid-resistant Staphylococcus epidermidis isolates with the type IV aj1-leader peptide (LP)-fusB structure (lacking aj1) revealed that their fusB gene was located on novel phage-related islands inserted downstream of smpB and are here referred to as SeRIfusB-3692 and SePIfusB-857. The novel SePIfusB-857 structure was followed by SeCI857, forming a composite pathogenicity island which contained a putative virulence gene, vapE. The linkage of fusB and vapE may contribute to bacterial adaption.

Potentials of organic tellurium-containing compound AS101 to overcome carbapenemase-producing Escherichia coli.

BACKGROUND: The issue of carbapenem-resistant Escherichia coli was aggravated yearly. The previous studies reported the varied but critical epidemiology of carbapenem-resistant E. coli among which the carbapenemase-producing strains were regarded as one of the most notorious issues. AS101, an organic tellurium-containing compound undergoing clinical trials, was revealed with antibacterial activities. However, little is known about the antibacterial effect of AS101 against carbapenemase-producing E. coli (CPEC). MATERIALS AND METHODS: The minimum inhibitory concentration (MIC) of AS101 against the 15 isolates was examined using a broth microdilution method. The scanning electron microscopy, pharmaceutical manipulations, reactive oxygen species level, and DNA fragmentation assay were carried out to investigate the antibacterial mechanism. The sepsis mouse model was employed to assess the in vivo treatment effect. RESULTS: The blaNDM (33.3%) was revealed as the dominant carbapenemase gene among the 15 CPEC isolates, followed by the blaKPC gene (26.7%). The MICs of AS101 against the 15 isolates ranged from 0.5 to 32 µg/ml, and 99.9% of bacterial eradication was observed at 8 h, 4 h, and 2 h for 1×, 2×, and 4 × MIC, respectively. The mechanistic investigations suggest that AS101 would enter the bacterial cell, and induce ROS generation, leading to DNA fragmentation. The in vivo study exhibited that AS101 possessed a steady treatment effect in a sepsis mouse model, with an up to 83.3% of survival rate. CONCLUSION: The in vitro activities, mechanisms, and in vivo study of AS101 against CPEC were unveiled. Our finding provided further evidence for the antibiotic development of AS101.

Prediction of methicillin-resistant Staphylococcus aureus and carbapenem-resistant Klebsiella pneumoniae from flagged blood cultures by combining rapid Sepsityper MALDI-TOF mass spectrometry with machine learning.

This study investigated combination of the Rapid Sepsityper Kit and a machine learning (ML)-based matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) approach for rapid prediction of methicillin-resistant Staphylococcus aureus (MRSA) and carbapenem-resistant Klebsiella pneumoniae (CRKP) from positive blood culture bottles. The study involved 461 patients with monomicrobial bloodstream infections. Species identification was performed using the conventional MALDI-TOF MS Biotyper system and the Rapid Sepsityper protocol. The data underwent preprocessing steps, and ML models were trained using preprocessed MALDI-TOF data and corresponding labels. The interpretability of the model was enhanced using SHapely Additive exPlanations values to identify significant features. In total, 44 S. aureus isolates comprising 406 MALDI-TOF MS files and 126 K. pneumoniae isolates comprising 1249 MALDI-TOF MS files were evaluated. This study demonstrated the feasibility of predicting MRSA among S. aureus and CRKP among K. pneumoniae isolates using MALDI-TOF MS and Sepsityper. Accuracy, area under the receiver operating characteristic curve, and F1 score for MRSA/methicillin-susceptible S. aureus were 0.875, 0.898 and 0.904, respectively; for CRKP/carbapenem-susceptible K. pneumoniae, these values were 0.766, 0.828 and 0.795, respectively. In conclusion, the novel ML-based MALDI-TOF MS approach enables rapid identification of MRSA and CRKP from flagged blood cultures within 1 h. This enables earlier initiation of targeted antimicrobial therapy, reducing deaths due to sepsis. The favourable performance and reduced turnaround time of this method suggest its potential as a rapid detection strategy in clinical microbiology laboratories, ultimately improving patient outcomes.

Direct prediction of carbapenem-resistant, carbapenemase-producing, and colistin-resistant Klebsiella pneumoniae isolates from routine MALDI-TOF mass spectra using machine learning and outcome evaluation.

The objective of this study was to develop a rapid prediction method for carbapenem-resistant Klebsiella pneumoniae (CRKP) and colistin-resistant K. pneumoniae (ColRKP) based on routine MALDI-TOF mass spectrometry (MS) results in order to formulate a suitable and rapid treatment strategy. A total of 830 CRKP and 1462 carbapenem-susceptible K. pneumoniae (CSKP) isolates were collected; 54 ColRKP isolates and 1592 colistin-intermediate K. pneumoniae (ColIKP) isolates were also included. Routine MALDI-TOF MS, antimicrobial susceptibility testing, NG-Test CARBA 5, and resistance gene detection were followed by machine learning (ML). Using the ML model, the accuracy and area under the curve for differentiating CRKP and CSKP were 0.8869 and 0.9551, respectively, and those for ColRKP and ColIKP were 0.8361 and 0.8447, respectively. The most important MS features of CRKP and ColRKP were m/z 4520-4529 and m/z 4170-4179, respectively. Of the CRKP isolates, MS m/z 4520-4529 was a potential biomarker for distinguishing KPC from OXA, NDM, IMP, and VIM. Of the 34 patients who received preliminary CRKP ML prediction results (by texting), 24 (70.6%) were confirmed to have CRKP infection. The mortality rate was lower in patients who received antibiotic regimen adjustment based on the preliminary ML prediction (4/14, 28.6%). In conclusion, the proposed model can provide rapid results for differentiating CRKP and CSKP, as well as ColRKP and ColIKP. The combination of ML-based CRKP with preliminary reporting of results can help physicians alter the regimen approximately 24 h earlier, resulting in improved survival of patients with timely antibiotic intervention.

Haplotype distribution of SARS-CoV-2 variants in low and high vaccination rate countries during ongoing global COVID-19 pandemic in early 2021.

The widespread severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continuously impacts our economic and public health. The potential of emerging variants to increase transmissibility and evade vaccine-induced immunity lets us put more effort to research on viral mutations and explore the pathogenic haplotypes. In this study, we characterized the haplotype and sub-haplotype diversity of SARS-CoV-2 global variants in January-March and the areas with low and high COVID19 vaccination rates in May 2021 by analyzing viral proteome of complete genome sequences published. Phylogenetic tree analysis of the proteomes of SARS-CoV-2 variants with Neighbor-Joining and Maximum Parsimony methods indicated that haplotype 2 variant with nsp12 P323L and Spike D614G was dominant (98.81%), including new sub-haplotypes 2A_1 to 2A_3, 2B_1 to 2B_3, and 2C_1 to 2C_2 emerged post-one-year COVID-19 outbreak. In addition, the profiling of sub-haplotypes indicated that sub-haplotype 2A_1 with the mutations at N501Y, A570D, D614G, P681H, T716I, S982A, and D118H in Spike was over 58% in May 2021 in the high partly vaccinated rate group (US, Canada, and Germany). Meanwhile, the new haplotype 2C_3 bearing the mutations at EFR156-158del, T19R, A222V, L452R, T478K, and D614G in Spike occupied over 54.8% in May 2021 in the low partly vaccinated rate group (India, Malaysia, Taiwan, and Vietnam). Sub-haplotypes 2A_1 and 2C_3 had a meaningful alternation of ACE2-specific recognition site, neutralization epitopes, and furin cleavage site in SARS-CoV-2 Spike protein. The results discovered the haplotype diversity and new sub-haplotypes of SARS-CoV-2 variants post one-year pandemic in January-March 2021, showing the profiles of sub-haplotypes in the groups with low and high partly vaccinated rates in May 2021. The study reports the emergence of new SARS-CoV-2 sub-haplotypes during ongoing pandemic and vaccination in early 2021, which might help inform the response to vaccination strategies.

Evaluation of the Rapid Sepsityper protocol and specific MBT-Sepsityper module for the identification of bacteremia and fungemia using Bruker Biotyper MALDI-TOF MS.

The rapid identification method, the Rapid Sepsityper protocol with a specific MBT-Sepsityper module (Bruker Daltonics), based on the MALDI Biotyper platform, accurately identified 93.5% (116/124) of microorganisms at the species level in the 124 flagged blood culture samples from patients with monomicrobial bloodstream infections. Gram-negative bacilli (95.6%, 43/45) had a higher identification rate than Gram-positive cocci (93.3%, 70/75) and yeasts (75%, 3/4). The Rapid Sepsityper protocol displayed poor identification performance for polymicrobial samples.

Potentially conjugative plasmids harboring Tn6636, a multidrug-resistant and composite mobile element, in Staphylococcus aureus.

OBJECTIVES: This study aimed to provide detailed genetic characterization of Tn6636, a multidrug-resistant and composite mobile element, in clinical isolates of Staphylococcus aureus. METHODS: A total of 112 ermB-positive methicillin-susceptible S. aureus (MSSA) and 224 ermB-positive methicillin-resistant S. aureus (MRSA) isolates collected from 2000 to 2015 were tested for the presence of Tn6636. Detection of the plasmids harboring Tn6636 was performed by S1 nuclease digestion pulsed-field gel electrophoresis (PFGE) analysis, conjugation test, and whole genome sequencing (WGS). RESULTS: Prevalence of Tn6636 in MSSA is higher than that in MRSA. Ten MSSA isolates and 10 MRSA isolates carried Tn6636. The 10 MSSA isolates belonged to three sequence types (ST), including ST7 (n = 6), ST5 (n = 3), and ST59 (n = 1). The 10 MRSA isolates belonged to ST188 (n = 8) and ST965 (n = 2). Analysis of plasmid sequences revealed that Tn6636 was harbored by six different mosaic plasmids. In addition to resistance genes, some plasmids also harbored toxin genes. CONCLUSION: The presence of multi-resistant Tn6636 in plasmids of both MSSA and MRSA with various STs suggests its broad dissemination. Results indicate that Tn6636 has existed for at least 16 years in Taiwan. The mosaic plasmids harboring Tn6636 can be transferred by conjugation. Ongoing surveillance of Tn6636 is essential to avoid continued spreading of resistant plasmids.

Bacteremia due to extended-spectrum-ß-lactamase-producing Aeromonas spp. at a medical center in Southern Taiwan.

Although extended-spectrum-ß-lactamase (ESBL)-producing aeromonads have been increasingly reported in recent years, most of them were isolates from case reports or environmental isolates. To investigate the prevalence of ESBL producers among Aeromonas blood isolates and the genes encoding ESBLs, consecutive nonduplicate Aeromonas blood isolates collected at a medical center in southern Taiwan from March 2004 to December 2008 were studied. The ESBL phenotypes were examined by clavulanate combination disk test and the cefepime-clavulanate ESBL Etest. The presence of ESBL-encoding genes, including bla(TEM), bla(PER), bla(CTX-M), and bla(SHV) genes, was evaluated by PCR and sequence analysis. The results showed that 4 (2.6%) of 156 Aeromonas blood isolates, 1 Aeromonas hydrophila isolate and 3 Aeromonas caviae isolates, expressed an ESBL-producing phenotype. The ESBL gene in two A. caviae isolates was bla(PER-3), which was located in both chromosomes and plasmids, as demonstrated by Southern hybridization. Of four patients with ESBL-producing Aeromonas bacteremia, two presented with catheter-related phlebitis and the other two with primary bacteremia. Three patients had been treated with initial noncarbapenem ß-lactams for 5 to 10 days, and all survived. In conclusion, ESBL producers exist among Aeromonas blood isolates, and clinical suspicion of ESBL production should be raised in treating infections due to cefotaxime-resistant Aeromonas isolates.

The extent of molecular variation in novel SARS-CoV-2 after the six-month global spread.

The pandemic spread of Coronavirus Disease 2019 (COVID-19) is still ongoing since severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is identified as the etiologic pathogen late December 2019. After over six-month spread of COVID-19, SARS-CoV-2 causes critical threats to global public health and economy. The investigations on evolution and genotyping on genetic variations are of great importance, therefore, the present study characterized the molecular variation of SARS-CoV-2 by analyzing 4230 complete genome sequences from the worldwide samples collected during the first 6-month pandemic. Phylogenetic tree analysis with Neighbor-Joining and Maximum-Parsimony methods indicated that the haplotypes of SARS-CoV-2 genome sequences were classified into four clades with the unique nucleotide and amino acid changes: T27879C (ORF8 L84S) in clade 1 (25.34%), A23138G (spike D614G) in clade 2 (63.54%), G10818T (nsp6 L37F), C14540T (nsp12 T442I), and G25879T (ORF3a V251F) in clade 3 (2.58%), and miscellaneous changes in clade 4 (8.54%). Interestingly, subclade 2B with the amino acid changes at nsp2 T85I, Spike D614G, and ORF3a Q57H was firstly reported on March 4, 2020 in United States of America, becoming the most frequent sub-haplogroup in the world (36.21%) and America (45.81%). Subclade 1C with the amino acid changes at nsp13 P504L and ORF8 L84S was becoming the second most frequent sub-haplogroup in the world (19.91%) and America (26.29%). Subclade 2A with the amino acid changes in Spike D614G and Nucleocapsid R203K and G204R was highly prevalent in Asia (18.82%) and Europe (29.72%). The study highlights the notable clades and sub-clades with unique mutations, revealing the genetic and geographical relevant post the six-month outbreak of COVID-19. This study thoroughly observed the genetic feature of SARS-CoV-2 haplotyping, providing an epidemiological trend of COVID-19.