We analyzed the occurrence and mechanisms of fusidic acid resistance present in staphylococci isolated from 59 healthy volunteers. The fingers of the volunteers were screened for the presence of ...staphylococci, and the collected isolates were tested for resistance to fusidic acid. A total of 34 fusidic acid resistant staphylococcal strains (all were coagulase-negative) were isolated from 22 individuals (22/59, 37.3%). Examination of the resistance genes revealed that acquired fusB or fusC was present in Staphylococcus epidermidis, Staphylococcus capitis subsp. urealyticus, Staphylococcus hominis subsp. hominis, Staphylococcus warneri and Staphylococcus haemolyticus. Resistance islands (RIs) carrying fusB were found in S. epidermidis and S. capitis subsp. urealyticus, while staphylococcal chromosome cassette (SCC)-related structures harboring fusC were found in S. hominis subsp. hominis. Genotypic analysis of S. epidermidis and S. hominis subsp. hominis indicated that the fus elements were disseminated in diverse genetic strain backgrounds. The fusC elements in S. hominis subsp. hominis strains were highly homologous to SCCfusC in the epidemic sequence type (ST) 239/SCCmecIII methicillin-resistant S. aureus (MRSA) or the pseudo SCCmec in ST779 MRSA. The presence of acquired fusidic acid resistance genes and their genetic environment in commensal staphylococci suggested that the skin commensal staphylococci may act as reservoir for fusidic acid resistance genes.
Chronic periodontitis is characterized by inflammation of periodontal tissues, leading to bone resorption and tooth loss. The goal of treatment is to regenerate periodontal tissues including bone and ...cementum lost as a consequence of disease. The local delivery of tetracycline was proven to be effective in controlling localized periodontal infection without apparent side effects. Previous studies suggested that lovastatin has a significant role in new bone formation; however, the local delivery of lovastatin might enhance its therapeutic effects. A number of local delivery devices have been developed recently, including poly(D,L-lactide-co-glycolide acid) (PLGA) nanoparticles. The aim of this study was to develop a local delivery device, PLGA-lovastatin-chitosan-tetracycline nanoparticles, which allows the sequential release of tetracycline and lovastatin to effectively control local infection and promote bone regeneration in periodontitis. The size and microstructure of nanoparticles were examined by transmission electron microscopy, Nanoparticle Size Analyzer, and Fourier transform infrared spectroscopy. The release of tetracycline and lovastatin was quantified using a UV-Vis spectrophotometer. Furthermore, the cytotoxic effect and alkaline phosphatase activity of the nanoparticles in osteoblast cell cultures as well as antibacterial activity against periodontal pathogens were investigated. Finally, the bone regeneration potential of PLGA nanoparticles in three-walled defects in beagle dogs was investigated. The results indicated that PLGA-lovastatin-chitosan-tetracycline nanoparticles showed good biocompatibility, antibacterial activity, and increased alkaline phosphatase activity. The volumetric analysis from micro-CT revealed significantly increased new bone formation in defects filled with nanoparticles in dogs. This novel local delivery device might be useful as an adjunctive treatment in periodontal regenerative therapy.
Phage-derived bacteriocins (tailocins) are ribosomally synthesized structures produced by bacteria in order to provide advantages against competing strains under natural conditions. Tailocins are ...highly specific in their target range and have proven to be effective for the prevention and/or treatment of bacterial diseases under clinical and agricultural settings. We describe the discovery and characterization of a new tailocin locus encoded within genomes of
and
subsp.
, which may enable the development of tailocins as preventative treatments against phytopathogenic infection by these species.
To evaluate the initial cutoff values, rates of screen positives, and genotypes for the large-scale newborn screening program for multiple mucopolysaccharidoses (MPS) in Taiwan.
More than 100 000 ...dried blood spots were collected consecutively as part of the national Taiwan newborn screening programs. Enzyme activities were measured by tandem mass spectrometry from dried blood spot punches. Genotypes were obtained when a second newborn screening specimen again had a decreased enzyme activity. Additional clinical evaluation was then initiated based on enzyme activity and/or genotype.
Molecular genetic analysis for cases with low enzyme activity revealed 5 newborns with pathogenic alpha-L-iduronidase mutations, 3 newborns with pathogenic iduronate-2-sulfatase mutations, and 1 newborn was a carrier of an arylsulfatase B mutation. Several variants of unknown pathogenic significance were also identified, most likely causing pseudodeficiency.
The highly robust tandem mass spectrometry-based enzyme assays for MPS-I, MPS-II, and MPS-VI allow for high-throughput newborn screening for these lysosomal storage disorders. Optimized cutoff values combined with second tier testing could largely eliminate false-positive results. Accordingly, newborn screening for these lysosomal storage disorders is possible.
Banana (Musa spp.) is one of the world's most important staple and cash crops. Despite accumulating genetic and transcriptomic data, low transformation efficiency in agronomically important Musa spp. ...render translational researches in banana difficult by using conventional knockout approaches. To develop tools for translational research in bananas, we developed a virus induced-gene silencing (VIGS) system based on a banana-infecting cucumber mosaic virus (CMV) isolate, CMV 20. CMV 20 genomic RNA 1, 2, and 3, were separately cloned in Agrobacterium pJL89 binary vectors, and a cloning site was introduced on RNA 2 immediately after the 2a open reading frame to insert the gene targeted for silencing. An efficient Agrobacterium inoculation method was developed for banana, which enabled the CMV 20 VIGS vector infection rate to reach 95% in our experiments. CMV 20-based silencing of Musa acuminata cv. Cavendish (AAA group) glutamate 1-semialdehyde aminotransferase (MaGSA) produced a typical chlorotic phenotype and silencing of M. acuminata phytoene desaturase (MaPDS) produced a photobleachnig phenotype. We show this approach efficiently reduced GSA and PDS transcripts to 10% and 18% of the control, respectively. The high infection rate and extended silencing of this VIGS system will provide an invaluable tool to accelerate functional genomic studies in banana.
High-end cutoffs of thyroid-stimulating hormone (TSH) have been emphasized for hypothyroidism therapy in extremely preterm infants, but the significance of low TSH levels remains unknown. This study ...hypothesized that the spectrum of TSH levels by newborn screening after birth signifies specific morbidities in extremely preterm neonates.
The multicenter population cohort analyzed 434 extremely preterm neonates receiving TSH screening at 24-96 h of age in 2008-2019. Neonates were categorized by blood TSH levels into group 1: TSH <0.5 µU/mL, group 2: 0.5 ≤ TSH <2 µU/mL, group 3: 2 ≤ TSH <4 µU/mL, and group 4: TSH ≥4 µU/mL. Neonatal morbidities were categorized using the modified Neonatal Therapeutic Intervention Scoring System.
The four groups differed in gestational age, birth weight, and the postnatal age at blood sampling so did the proportions of mechanical ventilation usage (p = 0.01), hypoxic respiratory failure (p = 0.005), high-grade intraventricular hemorrhage (p = 0.007), and periventricular leukomalacia (p = 0.048). Group 1 had higher severity scores for respiratory distress syndrome (RDS; effect size 0.39 95% confidence interval CI: 0.18-0.59) and brain injury (0.36 0.15-0.57) than group 2, which remained significant after adjusting for gestational age, birth weight, dopamine usage, and the postnatal age at TSH screening (RDS: mean + 0.45 points 95% CI: 0.11-0.79; brain injury: +0.32 0.11-0.54).
Low TSH levels in extremely preterm neonates are associated with severe RDS and brain injuries. Studies recruiting more neonates with complete thyroid function data are necessary to understand central-peripheral interactions of the hypothalamic-pituitary-thyroid axis.
Tandem mass spectrometry (MS/MS) is the most universal platform currently available for the analysis of enzymatic activities and biomarkers in dried blood spots (DBS) for applications in newborn ...screening (NBS). Among the MS/MS applications in NBS, the most common is flow-injection analysis (FIA-) MS/MS, where the sample is introduced as a bolus injection into the mass spectrometer without the prior fractionation of analytes. Liquid chromatography combined with MS/MS (LC-MS/MS) has been employed for second-tier tests to reduce the false-positive rate associated with several nonspecific screening markers, beginning two decades ago. More recently, LC-MS/MS has been applied to primary screening for new conditions for which FIA-MS/MS or other methods, including genomic screening, are not yet adequate. In addition to providing a list of the currently used LC-MS/MS-based assays for NBS, the authors share their experience regarding the maintenance requirements of LC-MS/MS vs. FIA-MS/MS systems. The consensus is that the maintenance of LC-MS/MS and FIA-MS/MS instrumentation is similar, and LC-MS/MS has the advantage of allowing for a larger number of diseases to be screened for in a multiplex, cost-effective fashion with a high throughput and an adequate turnaround time.
Patients with glucose-6-phosphate dehydrogenase deficiency might develop acute hemolytic anemia, chronic hemolytic anemia, and neonatal hyperbilirubinemia when exposed to high levels of oxidative ...stress. Severe hemolysis may occur in not only patients but also female carriers under certain conditions. However, 80%–85% of female carriers were undetected in an existing newborn screening program because of their wide-ranging levels of enzyme activity.
We developed a cost- and time-efficient multiplex SNaPshot assay using dried blood spots.
By detecting 21 common mutations in Taiwan and Southeast Asia, the assay could determine 98.2% of the mutant alleles in our cohort of Taiwanese newborns. The 9 undetermined mutant alleles were consequently detected by Sanger sequencing, of which 5 unpublished variations—c.187G > A (Pingtung), c.585G > C (Tainan), c.586A > T (Changhua), c.743G > A (Chiayi), and c.1330G > A (Tainan-2)—were detected. Furthermore, 13% of mild mutations were missed in male infants whose enzyme levels at 6.1–7.0 U/gHb in the newborn screening program when set the cutoff value at 6.0 U/gHb. We therefore suggest increasing the cutoff value and applying the multiplex SNaPshot assay as the second tier for neonatal screening.
Our approach could significantly increase the detection rate of male patients and female carriers with a reasonable cost and a reasonable number of clinic referrals.
•We propose a cost- and time-efficient multiplex SNaPshot assay using dried blood spots.•With the 21 G6PD mutations selected, the assay could determine 98.2% of mutant alleles in Taiwan.•We strongly suggest applying the proposed assay to the current newborn screening program.•The proposed assay could increase detection rates and control clinic referrals.
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.
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.
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.
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.
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