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PloS One 2024Pleural empyema is a serious complication of pneumonia in children. Negative bacterial cultures commonly impede optimal antibiotic therapy. To improve bacterial... (Observational Study)
Observational Study
Pleural empyema is a serious complication of pneumonia in children. Negative bacterial cultures commonly impede optimal antibiotic therapy. To improve bacterial identification, we developed a molecular assay and evaluated its performance compared with bacterial culture. Our multiplex-quantitative PCR to detect Streptococcus pneumoniae, Streptococcus pyogenes, Staphylococcus aureus and Haemophilus influenzae was assessed using bacterial genomic DNA and laboratory-prepared samples (n = 267). To evaluate clinical performance, we conducted the Molecular Assessment of Thoracic Empyema (MATE) observational study, enrolling children hospitalised with empyema. Pleural fluids were tested by bacterial culture and multiplex-qPCR, and performance determined using a study gold standard. We determined clinical sensitivity and time-to-organism-identification to assess the potential of the multiplex-qPCR to reduce the duration of empiric untargeted antibiotic therapy. Using spiked samples, the multiplex-qPCR demonstrated 213/215 (99.1%) sensitivity and 52/52 (100%) specificity for all organisms. During May 2019-March 2023, 100 children were enrolled in the MATE study; median age was 3.9 years (IQR 2-5.6). A bacterial pathogen was identified in 90/100 (90%) specimens by multiplex-qPCR, and 24/100 (24%) by bacterial culture (P <0.001). Multiplex-qPCR identified a bacterial cause in 68/76 (90%) culture-negative specimens. S. pneumoniae was the most common pathogen, identified in 67/100 (67%) specimens. We estimate our multiplex-qPCR would have reduced the duration of untargeted antibiotic therapy in 61% of cases by a median 20 days (IQR 17.5-23, range 1-55). Multiplex-qPCR significantly increased pathogen detection compared with culture and may allow for reducing the duration of untargeted antibiotic therapy.
Topics: Humans; Child, Preschool; Empyema, Pleural; Male; Female; Multiplex Polymerase Chain Reaction; Child; Haemophilus influenzae; Staphylococcus aureus; Streptococcus pneumoniae; Streptococcus pyogenes; Infant; Hospitalization; Anti-Bacterial Agents; Sensitivity and Specificity; DNA, Bacterial
PubMed: 38917227
DOI: 10.1371/journal.pone.0304861 -
Microbiology Spectrum Jun 2024Bacteremia is associated with significant morbidity and mortality. The emergence of bacteria with antimicrobial resistance (AMR) has further exacerbated the poor...
UNLABELLED
Bacteremia is associated with significant morbidity and mortality. The emergence of bacteria with antimicrobial resistance (AMR) has further exacerbated the poor outcomes associated with bacteremia. The Taiwan Surveillance of Antimicrobial Resistance (TSAR) program was established in 1998 to monitor bacterial epidemiology and antimicrobial resistance trends across all patient types and age groups. Between 2002 and 2020, a total of 14,539 non-duplicate bacteremia isolates were collected biennially from 29 hospitals during the months of July-September as part of the TSAR program. The three most common bacteremia agents were (31%), (13.6%), and (12.7%) overall. However, there was a steady increase in the proportions of and isolated from bacteremia cases (both < 0.001), while the proportions of spp. decreased. Regarding antimicrobial resistance, there was a notable increase in rates of third-generation cephalosporin and fluoroquinolone non-susceptibility among and , while the rates of carbapenem non-susceptibility were elevated but remained milder in these two species, especially in . Of concern is the alarming increase in vancomycin resistance among , rising from 10.0% in 2004 to 47.7% in 2020. In contrast, the prevalence of methicillin-resistant has remained stable at 51.2% overall. In conclusion, , with increasing third-generation cephalosporin and fluoroquinolone resistance, is the predominant cause of bacteremia in Taiwan during the 18-year surveillance. The escalating proportion of in bacteremia, coupled with a concurrent upsurge in vancomycin resistance, presents a therapeutic challenge in the recent decade.
IMPORTANCE
AMR surveillance not only enables the identification of regional variations but also supports the development of coordinated efforts to combat AMR on a global scale. The TSAR has been a biennial, government-endorsed, multicenter study focusing on pathogens isolated from inpatients and outpatients in Taiwan hospitals since 1998. Our report presents an 18-year comprehensive analysis on blood isolates in the 2002-2020 TSAR program. The study highlights an alarming increase in the proportion of causing bacteremia accompanied by elevated vancomycin resistance. It is worth noting that this trend differs from the observations in the United States and China. Understanding the composition of bacteria causing bacteremia, along with their prevalence of antimicrobial resistance, holds significant importance in establishing healthcare and research priorities. Additionally, this knowledge serves as a critical factor in evaluating the effectiveness of preventive interventions.
PubMed: 38916365
DOI: 10.1128/spectrum.00608-24 -
Microbiology Spectrum Jun 2024Methicillin-resistant (MRSA) bacteremia is a serious clinical challenge with high mortality rates. Antibiotic combination therapy is currently used in cases of...
Methicillin-resistant (MRSA) bacteremia is a serious clinical challenge with high mortality rates. Antibiotic combination therapy is currently used in cases of persistent infection; however, the limited development of new antibiotics will likely increase the need for combination therapy, and better methods are needed for identifying effective combinations for treating persistent bacteremia. To identify pairwise combinations with the most consistent potential for benefit compared to monotherapy with a primary anti-MRSA agent, we conducted a systematic study with an high-throughput methodology. We tested daptomycin and vancomycin each in combination with gentamicin, rifampicin, cefazolin, and oxacillin, and ceftaroline with daptomycin, gentamicin, and rifampicin. Combining cefazolin with daptomycin lowered the daptomycin concentration required to reach 95% growth inhibition (IC) for all isolates tested and lowered daptomycin IC below the sensitivity breakpoint for five out of six isolates that had daptomycin minimum inhibitory concentrations at or above the sensitivity breakpoint. Similarly, vancomycin ICs were decreased when vancomycin was combined with cefazolin for 86.7% of the isolates tested. This was a higher percentage than was achieved by adding any other secondary antibiotic to vancomycin. Adding rifampicin to daptomycin or vancomycin did not always reduce ICs and failed to produce synergistic interaction in any of the isolates tested; the addition of rifampicin to ceftaroline was frequently synergistic and always lowered the amount of ceftaroline required to reach the IC. These analyses rationalize further evaluation of three drug pairs for MRSA bacteremia: daptomycin+cefazolin, vancomycin+cefazolin, and ceftaroline+rifampicin.IMPORTANCEBloodstream infections caused by methicillin-resistant (MRSA) have a high mortality rate despite the availability of vancomycin, daptomycin, and newer antibiotics including ceftaroline. With the slow output of the antibiotic pipeline and the serious clinical challenge posed by persistent MRSA infections, better strategies for utilizing combination therapy are becoming increasingly necessary. We demonstrated the value of a systematic high-throughput approach, adapted from prior work testing antibiotic combinations against tuberculosis and other mycobacteria, by using this approach to test antibiotic pairs against a panel of MRSA isolates with diverse patterns of antibiotic susceptibility. We identified three antibiotic pairs-daptomycin+cefazolin, vancomycin+cefazolin, and ceftaroline+rifampicin-where the addition of the second antibiotic improved the potency of the first antibiotic across all or most isolates tested. Our results indicate that these pairs warrant further evaluation in the clinical setting.
PubMed: 38916355
DOI: 10.1128/spectrum.00976-24 -
Microbiology Spectrum Jun 2024Elexacaftor/tezacaftor/ivacaftor (ETI) therapy has revolutionized the treatment of cystic fibrosis (CF) for most affected individuals but the effects of treatment on...
Elexacaftor/tezacaftor/ivacaftor (ETI) therapy has revolutionized the treatment of cystic fibrosis (CF) for most affected individuals but the effects of treatment on sinus microbiota are still unknown. Changes to the airway microbiota in CF are associated with disease state and alterations to the bacterial community after ETI initiation may require changes to clinical management regimens. We collected sinus swab samples from the middle meatus in an observational study of 38 adults with CF and chronic rhinosinusitis (CRS) from 2017 to 2021 and captured the initiation of ETI therapy. We performed 16S and custom amplicon sequencing to characterize the sinus microbiota pre- and post-ETI. Real-time quantitative PCR (RT-qPCR) was performed to estimate total bacterial abundance. Sinus samples from people with CF (pwCF) clustered into three community types, dependent on the dominant bacterial organism: a -dominant, -dominant, and mixed dominance cluster. Shannon's diversity index was low and not significantly altered post-ETI. Total bacterial load was not significantly lowered post-ETI. spp. abundance was significantly reduced post-ETI, but eradication was not observed. spp. became the dominant organism in most individuals post-ETI and we showed the presence of methicillin-resistant (MRSA) in the sinus both pre- and post-ETI. We also demonstrated that the sinus microbiome is predictive of the presence of spp., spp., and spp. in the sputum. spp. and spp., including MRSA, persist in the sinuses of pwCF after ETI therapy, indicating that these pathogens will continue to be important in CF airway disease management in the era of highly effective modulator therapies (HEMT).IMPORTANCEHighly effective modulator therapies (HEMT), such as elexacaftor/tezacaftor/ivacaftor (ETI), for cystic fibrosis (CF) have revolutionized patient care and quality of life for most affected individuals. The effects of these therapies on the microbiota of the airways are still unclear, though work has already been published on changes to microbiota in the sputum. Our study presents evidence for reduced relative abundance of spp. in the sinuses following ETI therapy. We also show that spp. becomes the dominant organism in the sinus communities of most individuals in this cohort after ETI therapy. We identified methicillin-resistant (MRSA) in the sinus microbiota both pre- and post-therapy. These findings demonstrate that pathogen monitoring and treatment will remain a vital part of airway disease management for people with cystic fibrosis (pwCF) in the era of HEMT.
PubMed: 38916354
DOI: 10.1128/spectrum.00787-24 -
Microbiology Spectrum Jun 2024The incidence of heterogeneous vancomycin-intermediate (hVISA) infection is increasing and is associated with vancomycin treatment failures. However, studies...
The incidence of heterogeneous vancomycin-intermediate (hVISA) infection is increasing and is associated with vancomycin treatment failures. However, studies investigating the risk factors for treatment failure in hVISA infection are limited. Patients with hVISA bacteremia treated with vancomycin over 7 days between August 2008 and June 2020 were enrolled in this study. Clinical and microbiological characteristics were compared between vancomycin treatment failure and success groups to identify the risk factors for vancomycin treatment failure. Among the 180 patients with hVISA bacteremia, 102 patients treated with vancomycin over 7 days were included. Vancomycin treatment failed in 80 (78%) patients. Patients in the vancomycin treatment failure group were older ( < 0.001) and more frequently had solid cancer ( = 0.04) than those in the vancomycin treatment success group. Solid organ transplantation (SOT) was more frequent ( < 0.001) in the vancomycin treatment success group. The Charlson comorbidity index ( = 0.01) and Acute Physiology and Chronic Health Evaluation II scores ( < 0.001) were higher in the vancomycin treatment failure group. In multivariate analysis, independent risk factors for vancomycin treatment failure were old age and severity of bacteremia. SOT and vancomycin minimal inhibitory concentration (MIC) ≤ 1.0 mg/L using the broth microdilution (BMD) method were associated with successful vancomycin treatment. Old age and infection severity were independent risk factors for vancomycin treatment failure. Vancomycin MIC using the BMD method is an important risk factor for vancomycin treatment failure, and its use should be considered in hVISA bacteremia.IMPORTANCEIn this study, we assessed the clinical and microbiological characteristics of heterogeneous vancomycin-intermediated (hVISA) bacteremia and identified risk factors for vancomycin treatment failure. We found that advanced age and severity of infection were independent risk factors for vancomycin treatment failure. On the other hand, solid organ transplantation and a low vancomycin minimal inhibitory concentration were associated with successful vancomycin treatment. This study highlights the importance of vancomycin minimal inhibitory concentration in hVISA bacteremia.
PubMed: 38916352
DOI: 10.1128/spectrum.00333-24 -
Microbiology Spectrum Jun 2024Sonicating explanted prosthetic implants to physically remove biofilms is a recognized method for improving the microbiological diagnosis of prosthetic joint infection...
UNLABELLED
Sonicating explanted prosthetic implants to physically remove biofilms is a recognized method for improving the microbiological diagnosis of prosthetic joint infection (PJI); however, chemical and enzymatic treatments have been investigated as alternative biofilm removal methods. We compared the biofilm dislodging efficacy of sonication followed by the addition of enzyme cocktails with different activity spectra in the diagnosis of PJI with that of the sonication of fluid cultures alone. Consecutive patients who underwent prosthesis explantation due to infection at our institution were prospectively enrolled for 1 year. The diagnostic procedure included the collection of five intraoperative tissue cultures, sonication of the removed devices, and conventional culture of the sonication fluid. The resulting sonication fluid was also treated with an enzyme cocktail consisting of homemade dispersin B (0.04 µg/mL) and proteinase K (Sigma; 100 µg/mL) for 45 minutes at 37°C. The resulting sonication (S) and sonication with subsequent enzymatic treatment (SE) fluids were plated for aerobic and anaerobic culture broth for 7 days (aerobic) or 14 days (anaerobic). Identification was performed by matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (Bruker). We included 107 patients from whom a prosthetic implant had been removed, among which PJI was diagnosed in 36 (34%). The sensitivity of S alone was significantly greater than that of SE alone (82% vs 71%; < 0.05). Four patients with PJI were positive after sonication alone but negative after sonication plus enzymatic treatment. The four microorganisms missed after the addition of the enzyme cocktail were , two coagulase-negative and . In conclusion, sonication alone was more sensitive than sonication followed by enzymatic treatment. The combination of these two methods had no synergistic effect; in contrast, the results suggest that the combination of both dislodgment methods affects the viability of gram-positive microorganisms.
IMPORTANCE
While the potential of sonication and enzymes as biofilm dispersal agents has been previously described, the originality of our work resides in the combination of both methods, which is hypothesized to enhance the ability to remove biofilm and, therefore, improve the microbiological diagnosis of PJI.
PubMed: 38916322
DOI: 10.1128/spectrum.00020-24 -
Microbiology Spectrum Jun 2024is a leading cause of healthcare-associated infections globally. Vancomycin-resistant (VRSA), those with high-level resistance [minimum inhibitory concentration (MIC)...
UNLABELLED
is a leading cause of healthcare-associated infections globally. Vancomycin-resistant (VRSA), those with high-level resistance [minimum inhibitory concentration (MIC) of 16-32 µg/mL vancomycin], are uncommon, whereas vancomycin-intermediate (VISA; MIC of 4-8 µg/mL), are isolated more frequently and develop during long-term and/or repeated use of the antibiotic. VISA can be difficult to eradicate and infections may persist. Our knowledge of mechanisms that underlie the development of VISA is incomplete. We used a genomics approach to investigate the VISA phenotype in three prominent lineages. All VISA clinical isolates tested had increased cell wall thickness compared with vancomycin-susceptible strains. Growth rates of clonal complex (CC) 5, CC8, and CC45 clinical isolates were reduced in 2 µg/mL vancomycin compared to media alone. Culture in 2 and 4 µg/mL vancomycin sequentially for two weeks reduced susceptibility to daptomycin, televancin, tigecycline, and vancomycin in a majority of CC5, CC8, and CC45 isolates tested. We identified alleles reported previously to contribute to the VISA phenotype, but unexpectedly, these alleles were unique to each CC. A subtherapeutic concentration of vancomycin elicited changes in the VISA transcriptome-common and unique-among the three CCs tested. Multiple genes, including those encoding a glycerate kinase, an M50 family metallopeptidase, and an uncharacterized membrane protein, were upregulated among all three lineages and not reported previously as associated with VISA. Although there are lineage-specific changes in DNA sequence, our findings suggest changes in the VISA transcriptome constitute a general response to stress that confers reduced susceptibility to multiple antibiotics.
IMPORTANCE
Our understanding of the mechanisms that underlie the development of vancomycin-intermediate (VISA) is incomplete. To provide a more comprehensive view of this process, we compared genome sequences of clonal complex (CC) 5, CC8, and CC45 VISA clinical isolates and measured changes in the transcriptomes of these isolates during culture with a subtherapeutic concentration of vancomycin. Notably, we identified differentially expressed genes that were lineage-specific or common to the lineages tested, including genes that have not been previously reported to contribute to a VISA phenotype. Changes in gene expression were accompanied by reduced growth rate, increased cell wall thickness, and reduced susceptibility to daptomycin, televancin, tigecycline, and vancomycin. Our results provide support to the idea that changes in gene expression contribute to the development of VISA among three CCs that are a prominent cause of human infections.
PubMed: 38916317
DOI: 10.1128/spectrum.00486-24 -
Frontiers in Cellular and Infection... 2024is a major causative pathogen of osteomyelitis. Intracellular infections of resident bone cells including osteocytes can persist despite gold-standard clinical...
is a major causative pathogen of osteomyelitis. Intracellular infections of resident bone cells including osteocytes can persist despite gold-standard clinical intervention. The mechanisms by which intracellular evades antibiotic therapy are unknown. In this study, we utilised an infection model of human osteocytes to investigate whether antibiotic-mediated dysregulation of autophagy contributes to this phenomenon. Infected or non-infected osteocyte-like cells were exposed to combinations of rifampicin, vancomycin, and modulators of autophagy. Intracellular bacterial growth characteristics were assessed using colonyforming unit (CFU) analysis, viable bacterial DNA abundance, and the rate of escape into antibiotic-free medium, together with measures of autophagic flux. Rifampicin, alone or in combination with vancomycin, caused a rapid decrease in the culturability of intracellular bacteria, concomitant with stable or increased absolute bacterial DNA levels. Both antibiotics significantly inhibited autophagic flux. However, modulation of autophagic flux did not affect viable bacterial DNA levels. In summary, autophagy was shown to be a factor in the host-pathogen relationship in this model, as its modulation affected the growth state of intracellular with respect to both their culturability and propensity to escape the intracellular niche. While rifampicin and vancomycin treatments moderately suppressed autophagic flux acutely, this did not explain the paradoxical response of antibiotic treatment in decreasing culturability whilst failing to clear bacterial DNA and hence intracellular bacterial load. Thus, off-target effects of rifampicin and vancomycin on autophagic flux in osteocyte-like cells could not explain the persistent infection in these cells.
Topics: Autophagy; Staphylococcus aureus; Osteocytes; Anti-Bacterial Agents; Humans; Vancomycin; Rifampin; Staphylococcal Infections; Host-Pathogen Interactions; DNA, Bacterial
PubMed: 38915921
DOI: 10.3389/fcimb.2024.1403289 -
BioRxiv : the Preprint Server For... Jun 2024Microbes must adapt to diverse biotic and abiotic factors encountered in host environments. Polyamines are an abundant class of aliphatic molecules that play essential...
Microbes must adapt to diverse biotic and abiotic factors encountered in host environments. Polyamines are an abundant class of aliphatic molecules that play essential roles in fundamental cellular processes across the tree of life. Surprisingly, the bacterial pathogen is highly sensitive to polyamines encountered during infection, and acquisition of a polyamine resistance locus has been implicated in spread of the prominent USA300 methicillin-resistant lineage. At present, alternative pathways of polyamine resistance in staphylococci are largely unknown. Here we applied experimental evolution to identify novel mechanisms and consequences of adaption when exposed to increasing concentrations of the polyamine spermine. Evolved populations of exhibited striking evidence of parallel adaptation, accumulating independent mutations in the potassium transporter genes and . Mutations in either or are sufficient to confer polyamine resistance and function in an additive manner. Moreover, we find that ktr mutations provide increased resistance to multiple classes of unrelated cationic antibiotics, suggesting a common mechanism of resistance. Consistent with this hypothesis, ktr mutants exhibit alterations in cell surface charge indicative of reduced affinity and uptake of cationic molecules. Finally, we observe that laboratory-evolved ktr mutations are also present in diverse natural isolates, suggesting these mutations may contribute to antimicrobial resistance during human infections. Collectively this study identifies a new role for potassium transport in polyamine resistance with consequences for susceptibility to both host-derived and clinically-used antimicrobials.
PubMed: 38915543
DOI: 10.1101/2024.06.15.599172 -
RSC Advances Jun 2024Multidrug-resistant bacteria resulting from the abuse and overuse of antibiotics have become a huge crisis in global public health security. Therefore, it is urgently...
Multidrug-resistant bacteria resulting from the abuse and overuse of antibiotics have become a huge crisis in global public health security. Therefore, it is urgently needed to develop new antibacterial drugs with unique mechanisms of action. As a versatile moiety, morpholine has been widely employed to enhance the potency of numerous bioactive molecules. In this study, a series of ruthenium-based antibacterial agents modified with the morpholine moiety were designed and characterized, aiming to obtain a promising metalloantibiotic with a multitarget mechanism. Antibacterial activity screening demonstrated that the most active complex Ru(ii)-3 exhibited the strongest potency against () with an MIC value of only 0.78 μg mL, which is better than most clinically used antibiotics. Notably, Ru(ii)-3 not only possessed excellent bactericidal efficacy, but could also overcome bacterial resistance. Importantly, Ru(ii)-3 very efficiently removed biofilms produced by bacteria, inhibited the secretion of bacterial exotoxins, and enhanced the activity of many existing antibiotics. The results of mechanism studies confirmed that Ru(ii)-3 could destroy the bacterial membrane and induce ROS production in bacteria. Furthermore, animal infection models confirmed that Ru(ii)-3 showed significant anti-infective activity . Overall, this work demonstrated that a morpholine-modified ruthenium-based agent is a promising antibiotic candidate in tackling the crisis of drug-resistant bacteria.
PubMed: 38915333
DOI: 10.1039/d4ra02667e