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Circumstance Document: Pathogenic MYH9 h.5797delC Mutation in a Patient Along with Evident Thrombocytopenia and Nephropathy.
New blood culture instrumentation and medium formulations have led to improved time to positivity (TTP) for positive blood cultures. Data regarding the necessity of pediatric blood culture bottles with contemporary blood culture systems are sparse. We compared performance of three commercial blood culture systems, evaluating impact of blood volumes in standard and pediatric blood culture media across systems. Simulated blood cultures with packed red blood cells (PRBCs) and three Gram-positive, four Gram-negative, and one anaerobic organism (final concentrations ranging from 0.5 to 19 CFU/ml blood) on the Virtuo, VersaTrek, and Bactec FX instruments were evaluated with FAN Plus, Redox, and Bactec Plus media, respectively. For each medium/instrument/organism combination, 1-, 3-, 5-, and 10-ml blood volumes were evaluated in triplicate. Detection rate was not affected by blood volume. Aerobic organisms that demonstrated variable rates of detection were Kingella kingae, Haemophilus influenzae, and Neisseria meningitidis. Bacteroides fragilis was detected in 83%, 100%, and 100% of Virtuo, VersaTrek, and Bactec anaerobic bottles, respectively. The average TTP of standard medium for aerobic organisms detected on Virtuo was decreased compared to those for VersaTrek (-2.3 h) and Bactec (-4.9 h). Compared to standard medium, detection rate and TTP were unchanged on Virtuo, while TTP was reduced with pediatric medium for 2/8 organisms tested on Bactec and 7/8 organisms on VersaTrek, illustrating the potential benefit of pediatric medium on VersaTrek or Bactec when low blood volumes ( less then 5 ml) are collected. These results demonstrate that TTP is decreased on the Virtuo compared to VersaTrek and Bactec for many microorganisms associated with bloodstream infection (BSI) but may have species-specific limitations.In vivo diagnostic imaging of bacterial infections is currently reliant on targeting their metabolic pathways, an ineffective method to identify microbial species with low metabolic activity. Here, we establish HS-198 as a small-molecule fluorescent conjugate that selectively targets the highly conserved bacterial protein HtpG (high-temperature protein G), within Borrelia burgdorferi, the bacterium responsible for Lyme disease. We describe the use of HS-198 to target morphologic forms of B. burgdorferi in both the logarithmic growth phase and the metabolically dormant stationary phase as well as in inactivated spirochetes. Furthermore, in a murine infection model, systemically injected HS-198 identified B. burgdorferi as revealed by imaging in postnecropsy tissue sections. These findings demonstrate how small-molecule probes directed at conserved bacterial protein targets can function to identify the microbe using noninvasive imaging and potentially as scaffolds to deliver antimicrobial agents to the pathogen.New and rapid diagnostic methods are needed for the detection of antimicrobial resistance to aid in curbing drug-resistant infections. Targeted liquid chromatography-tandem mass spectrometry (LC-MS/MS) is a method that could serve this purpose, as it can detect specific peptides of antimicrobial resistance mechanisms with high accuracy. In the current study, we developed an accurate and rapid targeted LC-MS/MS assay based on parallel reaction monitoring for detection of the most prevalent aminoglycoside-modifying enzymes and 16S rRNA methyltransferases in Escherichia coli and Klebsiella pneumoniae that confer resistance to aminoglycosides. Specific tryptic peptides needed for detection were selected and validated for AAC(3)-Ia, AAC(3)-II, AAC(3)-IV, AAC(3)-VI, AAC(6')-Ib, AAC(6')-Ib-cr, ANT(2″)-I, APH(3')-VI, ArmA, RmtB, RmtC, and RmtF. In total, 205 isolates containing different aminoglycoside resistance mechanisms that consisted mostly of E. coli and K. pneumoniae were selected for assay development and evaluation. Mass spectrometry results were automatically analyzed and were compared to whole-genome sequencing results. Of the 2,460 isolate and resistance mechanism combinations tested, 2,416 combinations matched. Discrepancies were further analyzed by repeating LC-MS/MS analysis and performing additional PCRs. Mass spectrometry results were also used to predict resistance and susceptibility to gentamicin, tobramycin, and amikacin in only the E. coli and K. pneumoniae isolates (n = 191). The category interpretations were correctly predicted for gentamicin in 97.4% of the isolates, for tobramycin in 97.4% of the isolates, and for amikacin in 82.7% of the isolates. Targeted LC-MS/MS can be applied for accurate and rapid detection of aminoglycoside resistance mechanisms.The recent increase in macrolide-resistant Mycoplasma pneumoniae in Asia has become a continuing problem. A point-of-care testing method that can quickly detect M. pneumoniae and macrolide-resistant mutations (MR mutations) is critical for proper antimicrobial use. Smart Gene (Mizuho Medy Co., Ltd., Tosu City, Saga, Japan) is a compact and inexpensive fully automatic gene analyzer that combines amplification with PCR and the quenching probe method to specify the gene and MR mutations simultaneously. We performed a clinical evaluation of this device and its reagents on pediatric patients with suspected M. pneumoniae respiratory infections and evaluated the impact of the assay on antimicrobial selection. Using real-time PCR as a comparison control, the sensitivity of Smart Gene was 97.8% (44/45), its specificity was 93.3% (98/105), and its overall concordance rate was 94.7% (142/150). The overall concordance rate of Smart Gene diagnosis of MR mutations in comparison with sequence analysis was 100% (48/48). The ratio of MR mutations was significantly higher at high-level medical institutions than at a primary medical clinic (P = 0.023), and changes in antibiotic therapy to drugs other than macrolides were significantly more common in patients with MR mutations (P = 0.00024). see more Smart Gene demonstrated excellent utility in the diagnosis of M. pneumoniae and the selection of appropriate antimicrobials for MR mutations at primary medical institutions, which play a central role in community-acquired pneumonia care. The use of this device may reduce referrals to high-level medical institutions for respiratory infections, thereby reducing the medical and economic burdens on patients.
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