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In Mycobacterium avium infections, macrophages play a critical role in the host defense response. Apoptosis inhibitor of macrophage (AIM), also known as CD5L, may represent a novel supportive therapy against various diseases, including metabolic syndrome and infectious diseases. The mechanisms of AIM include modulating lipid metabolism in macrophages and other host cells. We investigated the role of AIM in M. avium infections in vitro and in vivo. In a mouse model of M. avium pneumonia, foamy macrophages were induced 6 wk after infection. The bacteria localized in these macrophages. Flow cytometric analysis also confirmed that the percentage of CD11chighMHCclassIIhigh interstitial and alveolar macrophages, a cell surface marker defined as foamy macrophages, increased significantly after infection. AIM in alveolar lavage fluid and serum gradually increased after infection. Administration of recombinant AIM significantly increased the number of bacteria in the lungs of mice, accompanied by the induction of inflammatory cytokine and iNOS expression. In mouse bone marrow-derived macrophages, the mRNA expression of AIM after M. avium infection and the amount of AIM in the supernatant increased prior to the increase in intracellular bacteria. Infected cells treated with anti-AIM Abs had fewer bacteria and a higher percentage of apoptosis-positive cells than infected cells treated with isotype control Abs. Finally, AIM in the sera of patients with M. avium-pulmonary disease was measured and was significantly higher than in healthy volunteers. This suggests that AIM production is enhanced in M. avium-infected macrophages, increasing macrophage resistance to apoptosis and providing a possible site for bacterial growth.
Assuntos
Infecção por Mycobacterium avium-intracellulare , Mycobacterium avium , Camundongos , Animais , Macrófagos/fisiologia , Infecção por Mycobacterium avium-intracellulare/complicações , Infecção por Mycobacterium avium-intracellulare/microbiologia , Macrófagos Alveolares/microbiologia , ApoptoseRESUMO
L1-like metallo-ß-lactamases (MBLs) exhibit diversity and are highly conserved. Although the presence of the blaL1-like gene is known, the biochemical characteristics are unclear. This study aimed to characterize an L1-like MBL from Stenotrophomonas lactitubi. It showed 70.9-99.7% similarity to 50 L1-like amino acid sequences. The characteristic kinetic parameter was its high hydrolyzing efficiency for ampicillin and nitrocefin. Furthermore, L1-like from S. lactitubi was distinctly more susceptible to inhibition by EDTA than that to inhibition by 2,6-pyridinedicarboxylic acid.
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Antibacterianos , beta-Lactamases , Antibacterianos/farmacologia , Antibacterianos/química , beta-Lactamases/metabolismo , Stenotrophomonas/genética , Sequência de AminoácidosRESUMO
Escherichia coli ST131 is a multidrug-resistant lineage associated with the global spread of extended-spectrum ß-lactamase-producing organisms. Particularly, ST131 clade C1 is the most predominant clade in Japan, harboring blaCTX-M-14 at a high frequency. However, the process of resistance gene acquisition and spread remains unclear. Here, we performed whole-genome sequencing of 19 E. coli strains belonging to 12 STs and 12 fimH types collected between 1997 and 2016. Additionally, we analyzed the full-length genome sequences of 96 ST131-H30 clade C0 and C1 strains, including those obtained from this study and those registered in public databases, to understand how ST131 clade C1 acquired and spread blaCTX-M-14. We detected conjugative IncFII plasmids and IncB/O/K/Z plasmids carrying blaCTX-M-14 in diverse genetic lineages of E. coli strains from the 1990s to the 2010s, suggesting that these plasmids played an important role in the spread of blaCTX-M-14. Molecular phylogenetic and molecular clock analyses of the 96 ST131-H30 clade C0 and C1 strains identified 8 subclades. Strains harboring blaCTX-M-14 were clustered in subclades 4 and 5, and it was inferred that clade C1 acquired blaCTX-M-14 around 1993. All 34 strains belonging to subclade 5 possessed blaCTX-M-14 with ISEcp1 upstream at the same chromosomal position, indicating their common ancestor acquired blaCTX-M-14 in a single ISEcp1-mediated transposition event during the early formation of the subclade around 1999. Therefore, both the horizontal transfer of plasmids carrying blaCTX-M-14 to diverse genetic lineages and chromosomal integration in the predominant genetic lineage have contributed to the spread of blaCTX-M-14.
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Phylogenetic analysis based on single-nucleotide polymorphism (SNP)-based through whole-genome sequencing is recognized as the standard method for probing nosocomial transmission. However, the application of WGS is constrained by the high cost of equipment and the need for diverse analysis tools, which limits its widespread use in clinical laboratory settings. In Japan, the prevalent use of PCR-based open reading frame typing (POT) for tracing methicillin-resistant Staphylococcus aureus (MRSA) transmission routes is attributed to its simplicity and ease of use. Although POT's discriminatory power is considered insufficient for nosocomial transmission analysis, conclusive data supporting this notion is lacking. This study assessed the discriminatory capabilities of SNP analysis and POT across 64 clinical MRSA strains. All 21 MRSA strains of ST5/SCCmec IIa, having more than 16 SNPs, demonstrated distinct clones. Conversely, two strains shared the same POT number and were identified as group A. Among the 12 MRSA strains of ST8/SCCmec IVl with over nine SNPs, five fell into POT group B, and five into POT group C. All four MRSA strains of ST8/SCCmec IVa were classified into POT group D, although they included strains with more than 30 SNPs. Among the 27 MRSA strains of ST1/SCCmec IVa, 14 were classified into POT group E. However, except for two clusters (each comprising two or three strains), all had SNP counts >10 (Fig. 1-D). SNP analysis of MRSA in CC1/SCCmec IV showed that several strains had the same number of SNPs in POT number (106-183-37), even among bacteria with >100 SNPs, indicating POT's limited use in detailed nosocomial transmission analysis.
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Infecção Hospitalar , Staphylococcus aureus Resistente à Meticilina , Reação em Cadeia da Polimerase , Polimorfismo de Nucleotídeo Único , Infecções Estafilocócicas , Sequenciamento Completo do Genoma , Staphylococcus aureus Resistente à Meticilina/genética , Staphylococcus aureus Resistente à Meticilina/isolamento & purificação , Polimorfismo de Nucleotídeo Único/genética , Humanos , Infecção Hospitalar/transmissão , Infecção Hospitalar/microbiologia , Infecções Estafilocócicas/transmissão , Infecções Estafilocócicas/microbiologia , Infecções Estafilocócicas/epidemiologia , Sequenciamento Completo do Genoma/métodos , Reação em Cadeia da Polimerase/métodos , Fases de Leitura Aberta/genética , Filogenia , Japão , Genoma Bacteriano/genéticaRESUMO
BACKGROUND: Mycobacterium avium is associated with pulmonary disease in otherwise healthy adults. Several clarithromycin-refractory cases have been reported, including some cases caused by clarithromycin-susceptible strains. OBJECTIVES: To characterize the reason for the discrepancy between clinical response and antibiotic susceptibility results. METHODS: We conducted population analysis of clarithromycin-tolerant and heteroresistant subpopulations of M. avium cultured in vitro and in homogenates of infected lungs of mice. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were determined for 28 M. avium and two M. kansasii strains. Mice were intranasally infected with M. avium and treated with or without clarithromycin (100 mg/kg) thrice weekly. They were sacrificed on day 35 and the bacteria in lung homogenates were tested for clarithromycin resistance. Population analysis assays were performed based on colony growth on plates containing two-fold dilutions of clarithromycin. RESULTS: The MBC/MIC ratios were ≥8 in all 28 strains of M. avium tested. In the population analysis assay, several colonies were observed on the plates containing clarithromycin concentrations above the MIC (2-64 mg/L). No growth of M. kansasii colonies was observed on the plates containing clarithromycin concentrations ≥2 mg/L. M. avium in the homogenates of infected lungs showed clearer clarithromycin-resistant subpopulations than in vitro, regardless of clarithromycin exposure. CONCLUSION: M. avium shows intrinsic heterogeneous resistance (heteroresistance) to clarithromycin. This may explain the observed discrepancies between clarithromycin susceptibility testing results and clinical response to clarithromycin treatment. Further studies are needed to confirm a link between heteroresistance and clinical outcomes.
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Claritromicina , Farmacorresistência Bacteriana , Testes de Sensibilidade Microbiana , Mycobacterium avium , Claritromicina/farmacologia , Claritromicina/uso terapêutico , Animais , Camundongos , Mycobacterium avium/efeitos dos fármacos , Pulmão/microbiologia , Feminino , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , HumanosRESUMO
L1 metallo-ß-lactamases produced by Stenotrophomonas maltophilia exhibit high diversity. Here, we characterized the genomes of Stenotrophomonas species harboring blaL1-like genes using publicly available genome sequences. Our findings provide evidence that Stenotrophomonas species with blaL1-like genes constitute a complex comprising many species with high genetic diversity, and similarities between blaL1-like genes are lower than those of the genome. This suggests that the diversity of blaL1-like is attributable to species diversity in Stenotrophomonas species harboring blaL1-like and the rapid evolutionary changes in blaL1-like genes.
Assuntos
Stenotrophomonas maltophilia , Stenotrophomonas , Stenotrophomonas/genética , beta-Lactamases/genética , Stenotrophomonas maltophilia/genéticaRESUMO
BACKGROUND: Pseudomonas otitidis belongs to the genus Pseudomonas and causes various infections, including ear, skin, and soft tissue infections. P. otitidis has a unique susceptibility profile, being susceptible to penicillins and cephalosporins but resistant to carbapenems, due to the production of the metallo-ß-lactamase called POM-1. This revealed genetic similarities with Pseudomonas aeruginosa, which can sometimes lead to misidentification. CASE PRESENTATION: We report the case of a 70-year-old Japanese male who developed cellulitis and bacteremia during chemotherapy for multiple myeloma. He was initially treated with meropenem, but blood culture later revealed gram-negative bacilli identified as P. otitidis using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Carbapenem resistance was predicted from previous reports; therefore, we switched to dual therapy with levofloxacin and cefepime, and favorable treatment results were obtained. CONCLUSION: This is the first reported case of P. otitidis cellulitis and bacteremia in an immunocompromised patient. Carbapenems are typically used in immunocompromised patients and P. otitidis is often resistant to it. However, its biochemical properties are similar to those of Pseudomonas aeruginosa; therefore, its accurate identification is critical. In the present study, we rapidly identified P. otitidis using MALDI-TOF MS and switched from carbapenems to an appropriate antimicrobial therapy, resulting in a successful outcome.
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Bacteriemia , Infecções por Pseudomonas , Humanos , Masculino , Idoso , Antibacterianos/uso terapêutico , Celulite (Flegmão)/diagnóstico , Celulite (Flegmão)/tratamento farmacológico , Pseudomonas , Carbapenêmicos/uso terapêutico , Infecções por Pseudomonas/diagnóstico , Infecções por Pseudomonas/tratamento farmacológico , Pseudomonas aeruginosa , Bacteriemia/diagnóstico , Bacteriemia/tratamento farmacológico , Hospedeiro Imunocomprometido , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodosRESUMO
We investigated the epidemiology and antimicrobial susceptibility profiles of seven major Gram-negative bacilli (Escherichia coli, Klebsiella spp., Enterobacter spp., Pseudomonas aeruginosa, Acinetobacter spp., Stenotrophomonas maltophilia, and Bacteroides spp.) that caused bacteremia in Japan. We collected clinical information and isolates from patients aged 20 years or older who developed bacteremia during a year at three Japanese university hospitals and performed microbiological examination. In total, 628 cases were included, half of which were caused by E. coli (315 isolates). P. aeruginosa (56 isolates) was isolated most frequently among non-fermenting bacteria and 33 Bacteroides spp. were isolated. Mortality rates were the highest for P. aeruginosa (7-day, 16.1%; 30-day, 26.8%). The 7- and 30-day mortality rates ranged 3.8-9.0% and 8.3-17.6%, respectively, for Enterobacterales, and they were 15.2% each for Bacteroides spp. Regarding antimicrobial resistance, Enterobacterales and Acinetobacter spp. showed susceptibility to carbapenems and amikacin (98.0-100.0%). The susceptibility rates to ceftolozane/tazobactam ranged 82.4-99.0% for Enterobacterales and 92.9% for P. aeruginosa. More than 30.0% of E. coli isolates were resistant to fluoroquinolone. Extended-spectrum ß-lactamase (ESBL) producers were found in 21.0% of E. coli and approximately 80% of those were resistant to fluoroquinolones. The susceptibility of the 33 Bacteroides spp. to carbapenems, ampicillin/sulbactam, and piperacillin/tazobactam was 100.0%. Among the ESBL producers, blaCTX-M group 9 was the major subgroup in E. coli (77.3%), and blaCTX-M group 1 was detected in 18.2% of E. coli and 50.0% of Klebsiella spp. Continuous surveillance is needed to understand the epidemiology and consider appropriate therapeutic strategies.
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Staphylococcus aureus bacteremia results in substantial mortality. Rapid identification and the determination of methicillin susceptibility are crucial for immediate treatment with appropriate antibiotics. In the present study, we aimed to evaluate the basic assay performance of GeneSoC®, a novel rapid quantitative polymerase chain reaction (qPCR) method, for the detection of methicillin-susceptible (MS) or -resistant (MR) S. aureus in blood culture (BC) bottles. qPCR pimers and probes were desinged for femA and mecA genes to diagnose S. aureus and its methicilline-resistance status. GeneSoC® system can detect target genes within 12 min per sample using microfludic thermal cycling. A total of 100 BC-positive samples, showing clusters of gram-positive cocci using microscopy, were tested. The analytical sensitivity was demonstrated for the target sequence of femA and mecA genes at 10 copies/µL, respectively. The detection limit of the MRSA bacterial burden using this system was 104 and 103 CFU/mL for femA and mecA, respectively. Compared with culture-based identification and susceptibility testing, the sensitivity and specificity for the detection of femA (+)/mecA (+) MRSA using GeneSoC® were 90.9 and 98.9%, respectively, whereas the sensitivity and specificity for detection of femA (+)/mecA (-) MSSA were 96.2% and 97.3%, respectively. In conclusion, although this was a small sample and pilot study, the GeneSoC® system is beneficial for rapid, reliable, and highly sensitive real-time testing of MRSA and MSSA in BC bottles.
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Staphylococcus aureus Resistente à Meticilina , Infecções Estafilocócicas , Humanos , Staphylococcus aureus/genética , Resistência a Meticilina/genética , Reação em Cadeia da Polimerase em Tempo Real , Meticilina/farmacologia , Meticilina/uso terapêutico , Hemocultura , Projetos Piloto , Infecções Estafilocócicas/tratamento farmacológico , Staphylococcus aureus Resistente à Meticilina/genética , Proteínas de Bactérias/genéticaRESUMO
OBJECTIVES: Isavuconazole is a convenient triazole antifungal agent with a broad antifungal spectrum. A randomized, open-label study (ClinicalTrials.gov, NCT03471988) was conducted to evaluate the efficacy and safety of isavuconazole in Japanese patients with deep-seated mycoses. PATIENTS AND METHODS: In Cohort A, patients with aspergillosis (chronic pulmonary aspergillosis and invasive aspergillosis) were randomized in a 2:1 ratio to isavuconazole or voriconazole, and in Cohort B, patients with cryptococcosis and mucormycosis were assigned to isavuconazole for up to 84 days of treatment. The overall outcome was evaluated according to the clinical, radiological, and mycological responses at Days 42 and 84 and at the end of treatment (EOT). RESULTS: A total of 103 participants were enrolled and received the study drug. The overall response rate of patients with chronic pulmonary aspergillosis in the isavuconazole (52 patients) and voriconazole (27 patients) groups was 82.7% and 77.8% at EOT, respectively. The response rate in patients with cryptococcosis (10 patients, isavuconazole group only) was 90.0%. One of three participants with invasive aspergillosis and one of three participants with mucormycosis responded in the isavuconazole group. In the safety evaluation, the incidence of adverse events in participants with chronic pulmonary aspergillosis was similar in both groups. Adverse drug reactions were reported in 32 (61.5%) patients receiving isavuconazole and 23 (85.2%) patients receiving voriconazole. CONCLUSIONS: Isavuconazole showed efficacy and safety in Japanese patients with chronic pulmonary aspergillosis and cryptococcosis, for which the drug is not currently indicated.
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Aspergilose , Criptococose , Infecções Fúngicas Invasivas , Mucormicose , Aspergilose Pulmonar , Humanos , Voriconazol/efeitos adversos , Mucormicose/tratamento farmacológico , Japão , Triazóis/efeitos adversos , Antifúngicos/efeitos adversos , Aspergilose/tratamento farmacológico , Infecções Fúngicas Invasivas/tratamento farmacológico , Aspergilose Pulmonar/tratamento farmacológico , Criptococose/tratamento farmacológicoRESUMO
The trends and prevalence of antimicrobial susceptibility of pathogens vary by country, region, and time. Long-term regular surveillance is required to investigate trends in the antimicrobial resistance of various isolated bacterial pathogens. We report the results of a nationwide surveillance on the antimicrobial susceptibility of bacterial respiratory pathogens in Japan conducted by the Japanese Society of Chemotherapy, the Japanese Association for Infectious Diseases, and the Japanese Society for Clinical Microbiology. The isolates were collected from clinical specimens obtained from adult patients who visited a collaborating medical facility between June 2019 and December 2020 and were diagnosed with respiratory tract infections by a physician. Antimicrobial susceptibility testing was performed in a centralized laboratory according to the methods recommended by the Clinical and Laboratory Standards Institute. Susceptibility testing was performed for 932 strains (201 Staphylococcus aureus, 158 Streptococcus pneumoniae, 6 S. pyogenes, 136 Haemophilus influenzae, 127 Moraxella catarrhalis, 141 Klebsiella pneumoniae, and 163 Pseudomonas aeruginosa) collected from 32 facilities in Japan. The proportions of methicillin-resistant S. aureus and penicillin-resistant S. pneumoniae were 35.3% and 0%, respectively. In H. influenzae, 16.2% and 16.9% were ß-lactamase-producing ampicillin resistant and ß-lactamase-negative ampicillin resistant, respectively. Extended-spectrum ß-lactamase-producing K. pneumoniae accounted for 5.0% of all K. pneumoniae infections. Carbapenemase-producing K. pneumoniae and multi-drug-resistant P. aeruginosa with metallo-ß-lactamase were not detected in this study. This surveillance will be a useful reference for treating respiratory infections in Japan and will provide evidence to enhance the appropriate use of antimicrobial agents.
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Doenças Transmissíveis , Staphylococcus aureus Resistente à Meticilina , Infecções Respiratórias , Adulto , Humanos , Ampicilina , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Bactérias , beta-Lactamases , Doenças Transmissíveis/tratamento farmacológico , Farmacorresistência Bacteriana , Haemophilus influenzae , Testes de Sensibilidade Microbiana , Infecções Respiratórias/tratamento farmacológico , Infecções Respiratórias/epidemiologia , Infecções Respiratórias/microbiologia , JapãoRESUMO
BACKGROUND: Intrawound vancomycin powder is effective in preventing surgical site infection after spine surgery. In a previous study, vancomycin-induced cytotoxicity in osteoblasts was investigated in vitro, and vitamin D3 was verified to be a candidate drug aiding recovery from vancomycin-induced cytotoxicity. The treatment practices involving osteogenesis-promoting drugs vary widely. Teriparatide, an anabolic agent, highly promotes bone formation by inducing osteoblast activation, increasing bone formation and mineral density, and preventing vertebral fractures. Hence, teriparatide may be administered in combination with vancomycin. METHODS: MC3T3-E1 cells were cultured in minimum essential medium supplemented with 10% fetal bovine serum at 37 °C in a humidified incubator containing 5% CO2. The experimental concentrations of vancomycin (2500, 5000, and 7500 µg/mL) were determined based on previous reports and our preliminary experiments. Teriparatide (100 ng/mL) was administered concomitantly to prevent cytotoxicity in osteoblasts, using pulsed vancomycin for 24 h (measured at 1, 3, and 7 days). Cell numbers and morphological changes in cells treated with vancomycin or vancomycin plus 100 ng/mL teriparatide were measured. Osteoblast differentiation was assessed using alkaline phosphatase staining, alkaline phosphatase activity, and alizarin red S staining. RESULTS: Teriparatide showed a recovery effect when vancomycin (7500 µg/mL) was administered only for 24 h. Microscopic examination revealed that teriparatide had a protective effect on osteoblasts exposed to 7500 µg/mL vancomycin. Addition of teriparatide led to the recovery of alkaline phosphatase staining and alizarin red staining. CONCLUSION: Vancomycin-induced cytotoxicity in osteoblasts could be inhibited by administering teriparatide concomitantly with vancomycin.
Assuntos
Teriparatida , Vancomicina , Humanos , Vancomicina/toxicidade , Teriparatida/farmacologia , Teriparatida/uso terapêutico , Fosfatase Alcalina , Diferenciação Celular , Osteogênese , OsteoblastosRESUMO
A plethora of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) diagnostic tests are available, each with different performance specifications, detection methods, and targets. This narrative review aims to summarize the diagnostic technologies available and how they are best selected to tackle SARS-CoV-2 infection as the pandemic evolves. Seven key settings have been identified where diagnostic tests are being deployed: symptomatic individuals presenting for diagnostic testing and/or treatment of COVID-19 symptoms; asymptomatic individuals accessing healthcare for planned non-COVID-19-related reasons; patients needing to access emergency care (symptom status unknown); patients being discharged from healthcare following hospitalization for COVID-19; healthy individuals in both single event settings (e.g. airports, restaurants, hotels, concerts, and sporting events) and repeat access settings (e.g. workplaces, schools, and universities); and vaccinated individuals. While molecular diagnostics remain central to SARS-CoV-2 testing strategies, we have offered some discussion on the considerations for when other tools and technologies may be useful, when centralized/point-of-care testing is appropriate, and how the various additional diagnostics can be deployed in differently resourced settings. As the pandemic evolves, molecular testing remains important for definitive diagnosis, but increasingly widespread point-of-care testing is essential to the re-opening of society.
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COVID-19 , SARS-CoV-2 , Humanos , Teste para COVID-19 , COVID-19/diagnóstico , Pandemias , Testes Imediatos , Sensibilidade e EspecificidadeRESUMO
Biochemical properties of the novel subclass B3 metallo-ß-lactamase (MBL) PJM-1 expressed in Pseudoxanthomonas japonensis, which is often isolated from the environment, were determined. The 906-bp blaPJM-1 gene in P. japonensis is a species-specific MBL gene, and PJM, with 301 predicted amino acids, has 81.8% amino acid identity with AIM-1. In this study, PJM-1 was recombinantly expressed and purified. PJM-1 showed a low catalytic activity against ceftazidime and cefepime, and it was strongly inhibited by EDTA.
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Ceftazidima , beta-Lactamases , Aminoácidos , Antibacterianos/farmacologia , Proteínas de Bactérias/metabolismo , Cefepima , Ceftazidima/farmacologia , Ácido Edético , Xanthomonadaceae , beta-Lactamases/metabolismoRESUMO
OBJECTIVES: Molecular analysis of meropenem-resistant mechanisms in mutants emerging from long-term in vitro meropenem exposure to borderline meropenem-susceptible carbapenemase-producing Enterobacterales (CPE) and non-CPE. METHODS: Escherichia coli TUM13867 harbouring both blaIMP-6- and blaCTX-M-2-carrying IncN plasmid and Citrobacter koseri TUM13189 with blaCTX-M-2-carrying chromosome were used. Meropenem MIC was 1â mg/L against both strains. Each strain was cultured in the hollow-fibre infection model (HFIM) to approximately 1â×â106 colony formation unit (cfu)/mL, and meropenem 1â g q8h treatment was initiated. Then, changes in total and meropenem-resistant populations were observed for 124â h. Meropenem resistance mechanisms were analysed using full-length whole-genome sequencing (WGS), reverse-transcription quantitative PCR and digital PCR. RESULTS: Meropenem reduced TUM13867 and TUM13189 to approximately 5 and 2 log10â cfu/mL, respectively, at 2â h after initiation, but regrowth was observed at 24â h. The meropenem-resistant mutant emergence frequency at 120 and 124â h was 4.4â×â10-4 for TUM13867 and 7.6â×â10-1 for TUM13189. Meropenem MIC of the mutants derived from TUM13867 (TUM20902) and TUM13189 (TUM20903) increased 4- and 16-fold, respectively. TUM20902, which harboured pMTY20902_IncN plasmid with a 27 505-bp deletion that included blaCTX-M-2, and blaIMP-6 showed 4.21-fold higher levels of transcription than the parental strain. TUM20903 had a 49 316-bp deletion that included ompC and a replicative increase of blaCTX-M-2 to three copies. CONCLUSIONS: Molecular analysis including full-length WGS revealed that the resistance mechanisms of meropenem-resistant mutants that emerged during long-term in vitro meropenem exposure were increased blaIMP-6 transcripts in CPE and increased blaCTX-M-2 transcripts due to gene triplication and OmpC loss resulting from ompC deletion in non-CPE.
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Antibacterianos , Proteínas de Bactérias , Meropeném/farmacologia , Antibacterianos/farmacologia , Testes de Sensibilidade Microbiana , Proteínas de Bactérias/genética , beta-Lactamases/genética , Escherichia coli/genética , PlasmídeosRESUMO
BACKGROUND: There is no comprehensive study on PAM-like MBLs. OBJECTIVES: Our aim was to characterize novel B3 MBL variants, PAM-2 and PAM-3, from Pseudomonas tohonis clinical isolates. METHODS: We evaluated the antimicrobial susceptibility and the MBL gene composition of three novel P. tohonis clinical isolates identified at a Japanese hospital, using the broth microdilution method and WGS, respectively. We characterized the PAM-2 and PAM-3 proteins using recombinant protein expression and biochemical evaluations. RESULTS: Low carbapenem MICs (meropenem MICâ=â0.125-1â mg/L) were observed for all three P. tohonis isolates; however, the isolates produced MBLs. We identified blaPAM-2 and blaPAM-3 as potential genes, belonging to a novel subclass of B3 MBLs. Their genomic sequence was similar to that of blaPAM-1 from Pseudomonas alcaligenes. PAM-2 and PAM-3 comprised 287 amino acids and exhibited 90% amino acid identity with PAM-1, 73% identity with POM-1 from Pseudomonas otitidis and 61% identity with L1 from Stenotrophomonas maltophilia. Biochemical evaluations of recombinant PAM-2 and PAM-3 revealed similar kcat/Km ratios and demonstrated catalytic activity against all the tested ß-lactams, except for aztreonam. In addition, the kcat/Km ratio for imipenem was 40-fold lower than that for meropenem. CONCLUSIONS: P. tohonis harbours a species-specific PAM-family MBL gene. This enzyme has higher hydrolytic activity against meropenem compared with that against imipenem.
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Infecções por Pseudomonas , beta-Lactamases , Antibacterianos/farmacologia , Humanos , Imipenem/farmacologia , Meropeném/farmacologia , Testes de Sensibilidade Microbiana , Pseudomonas/genética , Pseudomonas aeruginosa/genética , beta-Lactamases/metabolismoRESUMO
BACKGROUND: The bioactivities of commensal duodenal microbiota greatly influence the biofunction of hosts. We investigated the role of Helicobacter pylori infection in extra-gastroduodenal diseases by determining the impact of H. pylori infection on the duodenal microbiota. We sequenced 16 S rRNA genes in samples aspirated from the descending duodenum of 47 (male, 20; female, 27) individuals who were screened for gastric cancer. Samples were analysed using 16 S rRNA gene amplicon sequencing, and the LEFSe and Kyoto Encyclopaedia of Genes and Genomes methods were used to determine whether the duodenal microflora and microbial biofunctions were affected using H. pylori infection. RESULTS: Thirteen and 34 participants tested positive and negative for H. pylori, respectively. We identified 1,404 bacterial operational taxonomic units from 23 phyla and 253 genera. H. pylori infection changed the relative mean abundance of three phyla (Proteobacteria, Actinobacteria, and TM7) and ten genera (Neisseria, Rothia, TM7-3, Leptotrichia, Lachnospiraceae, Megasphaera, F16, Moryella, Filifactor, and Paludibacter). Microbiota features were significantly influenced in H. pylori-positive participants by 12 taxa mostly classified as Gammaproteobacteria. Microbial functional annotation revealed that H. pylori significantly affected 12 microbial metabolic pathways. CONCLUSIONS: H. pylori disrupted normal bacterial communities in the duodenum and changed the biofunctions of commensal microbiota primarily by upregulating specific metabolic pathways. Such upregulation may be involved in the onset of diseases associated with H. pylori infection.
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Duodeno/microbiologia , Microbioma Gastrointestinal/genética , Infecções por Helicobacter/complicações , Helicobacter pylori/patogenicidade , Redes e Vias Metabólicas/genética , Microbiota/genética , Idoso , Bacteroidetes/genética , Duodeno/patologia , Disbiose/microbiologia , Feminino , Mucosa Gástrica/microbiologia , Helicobacter pylori/genética , Humanos , Masculino , Pessoa de Meia-Idade , Proteobactérias/genética , RNA Ribossômico 16S/genéticaRESUMO
INTRODUCTION: Bacterial culture remains the gold standard for the diagnosis of legionellosis. However, past reports indicate that most physicians use the urinary antigen test (UAT) alone. Combining it with other tests is important, especially in patients with negative UAT results. The aim of this study was to investigate the current situation of legionellosis diagnostics and clarify the issues that need to be addressed. METHODS: Between March 1, 2021 and April 30, 2021, a questionnaire survey was conducted in an anonymous manner among physicians working in Japan. Questionnaires were generated on a website and asked questions in a multiple-choice format. RESULTS: Valid responses were received from 309 physicians during the study period. Most (92.9%) physicians reported using UAT as the initial test for patients suspected of having legionellosis, and <10% reported using other tests (e.g., culture, nucleic acid amplification test [NAAT], Gimenez staining, and serum antibody titer measurement [ATM]). When the initial test result was negative, 63% of physicians reported not conducting additional tests. Even when they chose to run additional tests, at most 27.8%, 23.6%, 12.3%, and 10.4% of all physicians used NAAT, culture, Gimenez staining, and serum ATM, respectively. The major reasons for not using tests other than UAT were "unavailability in the medical facility," "long turn-around time," and "difficult to collect sputum." CONCLUSIONS: The present survey revealed that most physicians in Japan used UAT alone for diagnosing legionellosis. Eliminating barriers to creating a reasonable environment and edification of physicians are needed to improve the current situation.
Assuntos
Legionella pneumophila , Legionelose , Médicos , Testes Diagnósticos de Rotina , Humanos , Japão , Legionelose/diagnósticoRESUMO
Legionella pneumophila is a major causative pathogen of community-acquired pneumonia (CAP), but recently the novel coronavirus disease 2019 (COVID-19) became the most common causative pathogen of CAP. Because L. pneumophila CAP is clinically distinct from bacterial CAPs, the Japan Society for Chemotherapy (JSC) developed a simple scoring system, the Legionella Score, using six parameters for the presumptive diagnosis of L. pneumophila pneumonia. We investigated the clinical and laboratory differences of L. pneumophila CAP and COVID-19 CAP and validated the Legionella Score in both CAP groups. We analyzed 102 patients with L. pneumophila CAP and 956 patients with COVID-19 CAP. Dyspnea and psychiatric symptoms were more frequently observed and cough was less frequently observed in patients with L. pneumophila CAP than those with COVID-19 CAP. Loss of taste and anosmia were observed in patients with COVID-19 CAP but not observed in those with L. pneumophila CAP. C-reactive protein and lactate dehydrogenase levels in L. pneumophila CAP group were significantly higher than in the COVID-19 CAP group. In contrast, sodium level in the L. pneumophila CAP group was significantly lower than in the COVID-19 CAP group. The median Legionella Score was significantly higher in the L. pneumophila CAP group than the COVID-19 CAP group (score 4 vs 2, p < 0.001). Our results demonstrated that the JSC Legionella Score had good diagnostic ability during the COVID-19 pandemic. However, physicians should consider COVID-19 CAP when loss of taste and/or anosmia are observed regardless of the Legionella Score.
Assuntos
Ageusia , COVID-19 , Infecções Comunitárias Adquiridas , Legionella pneumophila , Legionella , Doença dos Legionários , Pneumonia , Anosmia , COVID-19/diagnóstico , Infecções Comunitárias Adquiridas/tratamento farmacológico , Humanos , Doença dos Legionários/microbiologia , Pandemias , Pneumonia/microbiologiaRESUMO
INTRODUCTION: Influenza remains a clinically heavy burden worldwide. It is well known that some populations are at high risk of complications from influenza, whereas, even previously healthy people might suffer from severe influenza. The objective of this study was to clarify clinical manifestations of hospitalized patients without risk factors infected with influenza. METHODS: The clinical data for patients who were severely ill with influenza, and required hospitalization were gathered and analyzed between November 2014 and August 2020 (6 influenza seasons) using an internet-surveillance system. Among them, the patients who had no risk factors of complications from influenza were extracted. RESULTS: Finally, a total of 91 patients (9.0% of all influenza-related hospitalizations) without risk factors were analyzed. The no risk group was younger than the risk group, though other significant differences of clinical characteristics were not recognized between the groups. Pneumonia was the most common cause of hospitalization in the no risk group, and primary influenza viral pneumonia was the most common pneumonia. Antiviral drugs were administered in 96.7% of the no-risk group, and artificial ventilation was performed in 18.7%. In-hospital death was recorded for 3 patients without risk factors. CONCLUSIONS: Severe complications of influenza which required hospitalization may occur in a certain degree of patients with no risk factors. Efforts are needed to diagnose and treat influenza appropriately even in previously healthy younger patients. Continuous nationwide surveillance will be required to clarify risk factors for severe influenza even in previously healthy younger patients. (UMIN000015989).