Pub Date : 2024-07-16Epub Date: 2024-06-04DOI: 10.1128/jcm.00226-24
Sabine Pereyre, Nadège Hénin, Amandine Dolzy, Jennifer Guiraud, Cécile Laurier-Nadalié, Marie Gardette, Cécile Bébéar
Antimicrobial susceptibility testing (AST) of human mycoplasmas using microdilution is time-consuming. In this study, we compared the performance of MICRONAUT-S plates (Biocentric-Bruker) designed for AST of Ureaplasma parvum, Ureaplasma urealyticum, and Mycoplasma hominis with the results using the Clinical & Laboratory Standards Institute (CLSI) reference method. Then, we investigated the prevalence and mechanisms of resistance to tetracyclines, fluoroquinolones, and macrolides in France in 2020 and 2021. The two methods were compared using 60 strains. For the resistance prevalence study, U. parvum-, U. urealyticum-, and M. hominis-positive clinical specimens were collected for 1 month each year in 22 French diagnostic laboratories. MICs were determined using the MICRONAUT-S plates. The tet(M) gene was screened using PCR, and fluoroquinolone resistance-associated mutations were screened using PCR and Sanger sequencing. Comparing the methods, 99.5% (679/680) MICs obtained using the MICRONAUT-S plates concurred with those obtained using the CLSI reference method. For 90 M. hominis isolates, the tetracycline, levofloxacin, and moxifloxacin resistance rates were 11.1%, 2.2%, and 2.2%, respectively, with no clindamycin resistance. For 248 U. parvum isolates, the levofloxacin and moxifloxacin resistance rates were 5.2% and 0.8%, respectively; they were 2.9% and 1.5% in 68 U. urealyticum isolates. Tetracycline resistance in U. urealyticum (11.8%) was significantly (P < 0.001) higher than in U. parvum (1.2%). No macrolide resistance was observed. Overall, the customized MICRONAUT-S plates are a reliable, convenient tool for AST of human mycoplasmas. Tetracycline and fluoroquinolone resistance remain limited in France. However, the prevalence of levofloxacin and moxifloxacin resistance has increased significantly in Ureaplasma spp. from 2010 to 2015 and requires monitoring.
Importance: Antimicrobial susceptibility testing of human urogenital mycoplasmas using the CLSI reference broth microdilution method is time-consuming and requires the laborious preparation of antimicrobial stock solutions. Here, we validated the use of reliable, convenient plates designed for antimicrobial susceptibility testing that allows the simultaneous determination of the MICs of eight antibiotics of interest. We then investigated the prevalence and mechanisms of resistance of each of these bacteria to tetracyclines, fluoroquinolones, and macrolides in France in 2020 and 2021. We showed that the prevalence of levofloxacin and moxifloxacin resistance has increased significantly in Ureaplasma spp. from 2010 to 2015 and requires ongoing monitoring.
{"title":"Evaluation of commercial, customized microdilution plates for <i>Ureaplasma parvum</i>, <i>Ureaplasma urealyticum</i>, and <i>Mycoplasma hominis</i> antimicrobial susceptibility testing and determination of antimicrobial resistance prevalence in France.","authors":"Sabine Pereyre, Nadège Hénin, Amandine Dolzy, Jennifer Guiraud, Cécile Laurier-Nadalié, Marie Gardette, Cécile Bébéar","doi":"10.1128/jcm.00226-24","DOIUrl":"10.1128/jcm.00226-24","url":null,"abstract":"<p><p>Antimicrobial susceptibility testing (AST) of human mycoplasmas using microdilution is time-consuming. In this study, we compared the performance of MICRONAUT-S plates (Biocentric-Bruker) designed for AST of <i>Ureaplasma parvum</i>, <i>Ureaplasma urealyticum</i>, and <i>Mycoplasma hominis</i> with the results using the Clinical & Laboratory Standards Institute (CLSI) reference method. Then, we investigated the prevalence and mechanisms of resistance to tetracyclines, fluoroquinolones, and macrolides in France in 2020 and 2021. The two methods were compared using 60 strains. For the resistance prevalence study, <i>U. parvum</i>-, <i>U. urealyticum</i>-, and <i>M. hominis</i>-positive clinical specimens were collected for 1 month each year in 22 French diagnostic laboratories. MICs were determined using the MICRONAUT-S plates. The <i>tet</i>(M) gene was screened using PCR, and fluoroquinolone resistance-associated mutations were screened using PCR and Sanger sequencing. Comparing the methods, 99.5% (679/680) MICs obtained using the MICRONAUT-S plates concurred with those obtained using the CLSI reference method. For 90 <i>M</i>. <i>hominis</i> isolates, the tetracycline, levofloxacin, and moxifloxacin resistance rates were 11.1%, 2.2%, and 2.2%, respectively, with no clindamycin resistance. For 248 <i>U</i>. <i>parvum</i> isolates, the levofloxacin and moxifloxacin resistance rates were 5.2% and 0.8%, respectively; they were 2.9% and 1.5% in 68 <i>U</i>. <i>urealyticum</i> isolates. Tetracycline resistance in <i>U. urealyticum</i> (11.8%) was significantly (<i>P</i> < 0.001) higher than in <i>U. parvum</i> (1.2%). No macrolide resistance was observed. Overall, the customized MICRONAUT-S plates are a reliable, convenient tool for AST of human mycoplasmas. Tetracycline and fluoroquinolone resistance remain limited in France. However, the prevalence of levofloxacin and moxifloxacin resistance has increased significantly in <i>Ureaplasma</i> spp. from 2010 to 2015 and requires monitoring.</p><p><strong>Importance: </strong>Antimicrobial susceptibility testing of human urogenital mycoplasmas using the CLSI reference broth microdilution method is time-consuming and requires the laborious preparation of antimicrobial stock solutions. Here, we validated the use of reliable, convenient plates designed for antimicrobial susceptibility testing that allows the simultaneous determination of the MICs of eight antibiotics of interest. We then investigated the prevalence and mechanisms of resistance of each of these bacteria to tetracyclines, fluoroquinolones, and macrolides in France in 2020 and 2021. We showed that the prevalence of levofloxacin and moxifloxacin resistance has increased significantly in <i>Ureaplasma</i> spp. from 2010 to 2015 and requires ongoing monitoring.</p>","PeriodicalId":15511,"journal":{"name":"Journal of Clinical Microbiology","volume":" ","pages":"e0022624"},"PeriodicalIF":6.1,"publicationDate":"2024-07-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11324033/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141237116","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Candida auris is a multidrug-resistant fungal pathogen with a propensity to colonize humans and persist on environmental surfaces. C. auris invasive fungal disease is being increasingly identified in acute and long-term care settings. We have developed a prototype cartridge-based C. auris surveillance assay (CaurisSurV cartridge; "research use only") that includes integrated sample processing and nucleic acid amplification to detect C. auris from surveillance skin swabs in the GeneXpert instrument and is designed for point-of-care use. The assay limit of detection (LoD) in the skin swab matrix was 10.5 and 14.8 CFU/mL for non-aggregative (AR0388) and aggregative (AR0382) strains of C. auris, respectively. All five known clades of C. auris were detected at 2-3-5× (31.5-52.5 CFU/mL) the LoD. The assay was validated using a total of 85 clinical swab samples banked at two different institutions (University of California Los Angeles, CA and Wadsworth Center, NY). Compared to culture, sensitivity was 96.8% (30/31) and 100% (10/10) in the UCLA and Wadsworth cohorts, respectively, providing a combined sensitivity of 97.5% (40/41), and compared to PCR, the combined sensitivity was 92% (46/50). Specificity was 100% with both clinical (C. auris negative matrix, N = 31) and analytical (non-C. auris strains, N = 32) samples. An additional blinded study with N = 60 samples from Wadsworth Center, NY yielded 97% (29/30) sensitivity and 100% (28/28) specificity. We have developed a completely integrated, sensitive, specific, and 58-min prototype test, which can be used for routine surveillance of C. auris and might help prevent colonization and outbreaks in acute and chronic healthcare settings.
Importance: This study has the potential to offer a better solution to healthcare providers at hospitals and long-term care facilities in their ongoing efforts for effective and timely control of Candida auris infection and hence quicker response for any potential future outbreaks.
{"title":"A simple and sensitive test for <i>Candida auris</i> colonization, surveillance, and infection control suitable for near patient use.","authors":"Sukalyani Banik, Burcu Ozay, Marisol Trejo, YanChun Zhu, Charan Kanna, Cynthia Santellan, Bennett Shaw, Sukantha Chandrasekaran, Sudha Chaturvedi, Lindy Vejar, Soumitesh Chakravorty, David Alland, Padmapriya Banada","doi":"10.1128/jcm.00525-24","DOIUrl":"10.1128/jcm.00525-24","url":null,"abstract":"<p><p><i>Candida auris</i> is a multidrug-resistant fungal pathogen with a propensity to colonize humans and persist on environmental surfaces. <i>C. auris</i> invasive fungal disease is being increasingly identified in acute and long-term care settings. We have developed a prototype cartridge-based <i>C. auris</i> surveillance assay (CaurisSurV cartridge; \"research use only\") that includes integrated sample processing and nucleic acid amplification to detect <i>C. auris</i> from surveillance skin swabs in the GeneXpert instrument and is designed for point-of-care use. The assay limit of detection (LoD) in the skin swab matrix was 10.5 and 14.8 CFU/mL for non-aggregative (AR0388) and aggregative (AR0382) strains of <i>C. auris</i>, respectively. All five known clades of <i>C. auris</i> were detected at 2-3-5× (31.5-52.5 CFU/mL) the LoD. The assay was validated using a total of 85 clinical swab samples banked at two different institutions (University of California Los Angeles, CA and Wadsworth Center, NY). Compared to culture, sensitivity was 96.8% (30/31) and 100% (10/10) in the UCLA and Wadsworth cohorts, respectively, providing a combined sensitivity of 97.5% (40/41), and compared to PCR, the combined sensitivity was 92% (46/50). Specificity was 100% with both clinical (<i>C. auris</i> negative matrix, <i>N</i> = 31) and analytical (non-<i>C</i>. <i>auris</i> strains, <i>N</i> = 32) samples. An additional blinded study with <i>N</i> = 60 samples from Wadsworth Center, NY yielded 97% (29/30) sensitivity and 100% (28/28) specificity. We have developed a completely integrated, sensitive, specific, and 58-min prototype test, which can be used for routine surveillance of <i>C. auris</i> and might help prevent colonization and outbreaks in acute and chronic healthcare settings.</p><p><strong>Importance: </strong>This study has the potential to offer a better solution to healthcare providers at hospitals and long-term care facilities in their ongoing efforts for effective and timely control of <i>Candida auris</i> infection and hence quicker response for any potential future outbreaks.</p>","PeriodicalId":15511,"journal":{"name":"Journal of Clinical Microbiology","volume":" ","pages":"e0052524"},"PeriodicalIF":6.1,"publicationDate":"2024-07-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11250521/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141419320","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-07-16Epub Date: 2024-06-21DOI: 10.1128/jcm.01255-23
Lucía Maccari, Paola Ceriana, Hugo Norberto Granchetti, Antonella Viviana Pezzaniti, Celeste Lucero, Melina Rapoport, Alejandra Menocal, Alejandra Corso, Fernando Pasteran
<p><p>Prompt and precise identification of carbapenemase-producing organisms is crucial for guiding clinical antibiotic treatments and limiting transmission. Here, we propose modifying the Blue Carba test (BCT) and Carba NP-direct (CNPd) to identify molecular carbapenemase classes, including dual carbapenemase strains, by adding specific Class A and Class B inhibitors. We tested 171 carbapenemase-producing Gram-negative bacilli strains-21 in Class A (KPC, NMC, SME), 58 in Class B (IMP, VIM, NDM, SPM), and 92 with dual carbapenemase production (KPC+NDM, KPC+IMP, KPC+VIM), all previously positive with BCT or CNPd. We also included 13 carbapenemase non-producers. β-lactamases were previously characterized by PCR. The improved BCT/CNPd methods detect imipenem hydrolysis from an imipenem-cilastatin solution, using pH indicators and Class A (avibactam) and/or Class B (EDTA) inhibitors. Results were interpreted visually based on color changes. CNPd achieved 99.4% sensitivity and 100% specificity in categorizing carbapenemases, while BCT had 91.8% sensitivity and 100% specificity. Performance varied by carbapenemase classes: both tests classified all Class A-producing strains. For Class B, the CNP test identified 57/58 strains (98.3%), whereas the BCT test, 45/58 strains (77.6%), with non-fermenters posing the greatest detection challenge. For Classes A plus B dual producers, both tests performed exceptionally well, with only one indeterminate strain for the BCT. The statistical comparison showed both methods had similar times to a positive result, with differences based on the carbapenemase class or bacterial group involved. This improved assay rapidly distinguishes major Class A or Class B carbapenemase producers among Gram-negative bacilli, including dual-class combinations, in less than 2 hours.</p><p><strong>Importance: </strong>Rapid and accurate identification of carbapenemase-producing organisms is of vital importance in guiding appropriate clinical antibiotic treatments and curbing their transmission. The emergence of negative bacilli carrying multiple carbapenemase combinations during and after the severe acute respiratory syndrome coronavirus 2 pandemic has posed a challenge to the conventional biochemical tests typically used to determine the specific carbapenemase type in the isolated strains. Several initiatives have aimed to enhance colorimetric methods, enabling them to independently identify the presence of Class A or Class B carbapenemases. Notably, no previous efforts have been made to distinguish both classes simultaneously. Additionally, these modifications have struggled to differentiate between carriers of multiple carbapenemases, a common occurrence in many Latin American countries. In this study, we introduced specific Class A and Class B carbapenemase inhibitors into the Blue Carba test (BCT) and Carba NP-direct (CNP) colorimetric assays to identify the type of carbapenemase, even in cases of multiple carbapenemase producers wit
及时、准确地识别产碳青霉烯酶生物对于指导临床抗生素治疗和限制传播至关重要。在此,我们建议修改蓝色卡巴试验(BCT)和卡巴NP-直接试验(CNPd),通过添加特异性A类和B类抑制剂来鉴定分子碳青霉烯酶类别,包括双重碳青霉烯酶菌株。我们检测了 171 株产碳青霉烯酶的革兰氏阴性杆菌,其中 21 株属于 A 类(KPC、NMC、SME),58 株属于 B 类(IMP、VIM、NDM、SPM),92 株产双重碳青霉烯酶(KPC+NDM、KPC+IMP、KPC+VIM),所有这些菌株之前都用 BCT 或 CNPd 检测呈阳性。我们还纳入了 13 种不产生碳青霉烯酶的细菌。β-内酰胺酶之前已通过 PCR 鉴定。改进的 BCT/CNPd 方法使用 pH 指示剂和 A 类(阿维巴坦)和/或 B 类(乙二胺四乙酸)抑制剂检测亚胺培南-西司他丁溶液中的亚胺培南水解作用。检测结果根据颜色变化直观判读。CNPd 对碳青霉烯酶分类的灵敏度为 99.4%,特异性为 100%,而 BCT 的灵敏度为 91.8%,特异性为 100%。碳青霉烯酶类别不同,检测结果也不同:两种检测方法都能对所有 A 类产菌菌株进行分类。对于 B 类,CNP 检测可识别 57/58 株菌株(98.3%),而 BCT 检测可识别 45/58 株菌株(77.6%),其中非发酵菌株的检测难度最大。对于 A 类和 B 类双重生产者,两种检测方法的表现都非常出色,BCT 检测方法仅检测出一株不确定菌株。统计比较显示,两种方法得出阳性结果的时间相近,但因涉及的碳青霉烯酶类别或细菌群而有所不同。这种改进后的检测方法能在不到 2 小时的时间内快速区分革兰氏阴性杆菌中主要的 A 类或 B 类碳青霉烯酶生产者,包括双类组合:重要意义:快速、准确地鉴定碳青霉烯酶产生菌对指导适当的临床抗生素治疗和遏制其传播至关重要。在严重急性呼吸系统综合征冠状病毒 2 大流行期间和之后,出现了携带多种碳青霉烯酶组合的阴性杆菌,这给通常用于确定分离菌株中特定碳青霉烯酶类型的传统生化检验带来了挑战。有几项计划旨在改进比色法,使其能够独立识别 A 类或 B 类碳青霉烯酶的存在。值得注意的是,此前还没有人尝试过同时区分这两种类型。此外,这些改良方法也很难区分多种碳青霉烯酶的携带者,而这在许多拉美国家都很常见。在这项研究中,我们将特定的 A 类和 B 类碳青霉烯酶抑制剂引入蓝色卡巴试验(BCT)和卡巴 NP-直接(CNP)比色测定法中,以确定碳青霉烯酶的类型,即使在这些类别中存在多种碳青霉烯酶生产者的情况下也是如此。这些更新后的检测方法具有极高的灵敏度和特异性(≥ 90%),而且所有检测都能在 2 小时内快速完成,通常只需 45 分钟即可完成。这些对 BCT 和 CNP 检测方法的内部改进为确定主要碳青霉烯酶类别提供了一种快速、直接和经济有效的方法。它们可以作为分子生物学或免疫层析技术的可行替代方法,在整个过程中充当初始诊断步骤。
{"title":"Improved Blue Carba test and Carba NP test for detection and classification of major Class A and B carbapenemases, including dual producers, among Gram-negative bacilli.","authors":"Lucía Maccari, Paola Ceriana, Hugo Norberto Granchetti, Antonella Viviana Pezzaniti, Celeste Lucero, Melina Rapoport, Alejandra Menocal, Alejandra Corso, Fernando Pasteran","doi":"10.1128/jcm.01255-23","DOIUrl":"10.1128/jcm.01255-23","url":null,"abstract":"<p><p>Prompt and precise identification of carbapenemase-producing organisms is crucial for guiding clinical antibiotic treatments and limiting transmission. Here, we propose modifying the Blue Carba test (BCT) and Carba NP-direct (CNPd) to identify molecular carbapenemase classes, including dual carbapenemase strains, by adding specific Class A and Class B inhibitors. We tested 171 carbapenemase-producing Gram-negative bacilli strains-21 in Class A (KPC, NMC, SME), 58 in Class B (IMP, VIM, NDM, SPM), and 92 with dual carbapenemase production (KPC+NDM, KPC+IMP, KPC+VIM), all previously positive with BCT or CNPd. We also included 13 carbapenemase non-producers. β-lactamases were previously characterized by PCR. The improved BCT/CNPd methods detect imipenem hydrolysis from an imipenem-cilastatin solution, using pH indicators and Class A (avibactam) and/or Class B (EDTA) inhibitors. Results were interpreted visually based on color changes. CNPd achieved 99.4% sensitivity and 100% specificity in categorizing carbapenemases, while BCT had 91.8% sensitivity and 100% specificity. Performance varied by carbapenemase classes: both tests classified all Class A-producing strains. For Class B, the CNP test identified 57/58 strains (98.3%), whereas the BCT test, 45/58 strains (77.6%), with non-fermenters posing the greatest detection challenge. For Classes A plus B dual producers, both tests performed exceptionally well, with only one indeterminate strain for the BCT. The statistical comparison showed both methods had similar times to a positive result, with differences based on the carbapenemase class or bacterial group involved. This improved assay rapidly distinguishes major Class A or Class B carbapenemase producers among Gram-negative bacilli, including dual-class combinations, in less than 2 hours.</p><p><strong>Importance: </strong>Rapid and accurate identification of carbapenemase-producing organisms is of vital importance in guiding appropriate clinical antibiotic treatments and curbing their transmission. The emergence of negative bacilli carrying multiple carbapenemase combinations during and after the severe acute respiratory syndrome coronavirus 2 pandemic has posed a challenge to the conventional biochemical tests typically used to determine the specific carbapenemase type in the isolated strains. Several initiatives have aimed to enhance colorimetric methods, enabling them to independently identify the presence of Class A or Class B carbapenemases. Notably, no previous efforts have been made to distinguish both classes simultaneously. Additionally, these modifications have struggled to differentiate between carriers of multiple carbapenemases, a common occurrence in many Latin American countries. In this study, we introduced specific Class A and Class B carbapenemase inhibitors into the Blue Carba test (BCT) and Carba NP-direct (CNP) colorimetric assays to identify the type of carbapenemase, even in cases of multiple carbapenemase producers wit","PeriodicalId":15511,"journal":{"name":"Journal of Clinical Microbiology","volume":" ","pages":"e0125523"},"PeriodicalIF":6.1,"publicationDate":"2024-07-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11250402/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141432002","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-07-16Epub Date: 2024-06-18DOI: 10.1128/jcm.00207-24
Shao-Hua Yu, Keun-Ju Kim, Chien-Chang Lee, Yanely Pineiro Puebla, Gelza Mae A Zabat, Hong-Mo Shih, Po-Ren Hsueh
The Panbio COVID-19/Flu A&B Panel (Abbott) is an in vitro diagnostic rapid test designed for the qualitative detection of nucleocapsid proteins SARS-CoV-2 and nucleoprotein influenza A and B antigens in nasal mid-turbinate (NMT) swab specimens from symptomatic individuals meeting COVID-19 and influenza clinical and/or epidemiological criteria. This study, the largest global one to date using fresh samples, aimed to assess the diagnostic sensitivity and specificity of the Panbio COVID-19/Flu A&B Panel in freshly collected NMT swab specimens from individuals suspected of respiratory viral infection consistent with COVID-19 and/or influenza within the first 5 days of symptom onset compared with results obtained with the cobas SARS-CoV-2 and influenza A/B qualitative assay (cobas 6800/8800 systems), which were tested using nasopharyngeal swab samples. A total of 512 evaluable subjects were enrolled in the COVID-19 cohort across 18 sites, and 1,148 evaluable subjects were enrolled in the influenza cohort across 22 sites in the Asia-Pacific, Europe, and the USA. The Panbio COVID-19/Flu A&B Panel demonstrated a sensitivity of 80.4% and a specificity of 99.7% for COVID-19. For influenza A, the sensitivity and specificity rates were 80.6% and 99.3%, respectively. Likewise, for influenza B, the sensitivity and specificity rates were 80.8% and 99.4%, respectively. In conclusion, the Panbio COVID-19/Flu A&B Panel emerges as a suitable rapid test for detecting COVID-19 and influenza in symptomatic subjects across diverse global populations, exhibiting high sensitivity. The assay achieved a sensitivity of 94.4% in samples with Ct ≤24 for COVID-19 and 92.6% in samples with Ct ≤30 for influenza A and B.
Importance: The Panbio COVID-19/Flu A&B Panel is a suitable rapid test for detecting COVID-19 and influenza in symptomatic subjects across diverse global populations, exhibiting high sensitivity. The assay achieved a sensitivity of 94.0% in samples with Ct ≤24 for COVID-19 and 92.6% in samples with Ct ≤30 for influenza A and B.
{"title":"Performance evaluation of the Panbio COVID-19/Flu A&B Panel for detection of SARS-CoV-2, influenza A, and influenza B antigens using mid-turbinate nasal swabs.","authors":"Shao-Hua Yu, Keun-Ju Kim, Chien-Chang Lee, Yanely Pineiro Puebla, Gelza Mae A Zabat, Hong-Mo Shih, Po-Ren Hsueh","doi":"10.1128/jcm.00207-24","DOIUrl":"10.1128/jcm.00207-24","url":null,"abstract":"<p><p>The Panbio COVID-19/Flu A&B Panel (Abbott) is an <i>in vitro</i> diagnostic rapid test designed for the qualitative detection of nucleocapsid proteins SARS-CoV-2 and nucleoprotein influenza A and B antigens in nasal mid-turbinate (NMT) swab specimens from symptomatic individuals meeting COVID-19 and influenza clinical and/or epidemiological criteria. This study, the largest global one to date using fresh samples, aimed to assess the diagnostic sensitivity and specificity of the Panbio COVID-19/Flu A&B Panel in freshly collected NMT swab specimens from individuals suspected of respiratory viral infection consistent with COVID-19 and/or influenza within the first 5 days of symptom onset compared with results obtained with the cobas SARS-CoV-2 and influenza A/B qualitative assay (cobas 6800/8800 systems), which were tested using nasopharyngeal swab samples. A total of 512 evaluable subjects were enrolled in the COVID-19 cohort across 18 sites, and 1,148 evaluable subjects were enrolled in the influenza cohort across 22 sites in the Asia-Pacific, Europe, and the USA. The Panbio COVID-19/Flu A&B Panel demonstrated a sensitivity of 80.4% and a specificity of 99.7% for COVID-19. For influenza A, the sensitivity and specificity rates were 80.6% and 99.3%, respectively. Likewise, for influenza B, the sensitivity and specificity rates were 80.8% and 99.4%, respectively. In conclusion, the Panbio COVID-19/Flu A&B Panel emerges as a suitable rapid test for detecting COVID-19 and influenza in symptomatic subjects across diverse global populations, exhibiting high sensitivity. The assay achieved a sensitivity of 94.4% in samples with Ct ≤24 for COVID-19 and 92.6% in samples with Ct ≤30 for influenza A and B.</p><p><strong>Importance: </strong>The Panbio COVID-19/Flu A&B Panel is a suitable rapid test for detecting COVID-19 and influenza in symptomatic subjects across diverse global populations, exhibiting high sensitivity. The assay achieved a sensitivity of 94.0% in samples with Ct ≤24 for COVID-19 and 92.6% in samples with Ct ≤30 for influenza A and B.</p>","PeriodicalId":15511,"journal":{"name":"Journal of Clinical Microbiology","volume":" ","pages":"e0020724"},"PeriodicalIF":6.1,"publicationDate":"2024-07-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11250729/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141419322","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
In this study, we investigated the genomic changes in a major methicillin-resistant Staphylococcus aureus (MRSA) clone following a significant outbreak at a hospital. Whole-genome sequencing of MRSA isolates was utilized to explore the genomic evolution of post-outbreak MRSA strains. The epidemicity of the clone declined over time, coinciding with the introduction of multimodal infection control measures. A genome-wide association study (GWAS) identified multiple genes significantly associated with either high or low epidemic success, indicating alterations in mobilome, virulence, and defense mechanisms. Random Forest models pinpointed a gene related to fibrinogen binding as the most influential predictor of epidemicity. The decline of the MRSA clone may be attributed to various factors, including the implementation of new infection control measures, single nucleotide polymorphisms accumulation, and the genetic drift of a given clone. This research underscores the complex dynamics of MRSA clones, emphasizing the multifactorial nature of their evolution. The decline in epidemicity seems linked to alterations in the clone's genetic profile, with a probable shift towards decreased virulence and adaptation to long-term carriage. Understanding the genomic basis for the decline of epidemic clones is crucial to develop effective strategies for their surveillance and management, as well as to gain insights into the evolutionary dynamics of pathogen genomes.
{"title":"Genomic evolution of ST228 SCCmec-I MRSA 10 years after a major nosocomial outbreak.","authors":"Florian Mauffrey, Claire Bertelli, Gilbert Greub, Laurence Senn, Dominique S Blanc","doi":"10.1128/jcm.00203-24","DOIUrl":"10.1128/jcm.00203-24","url":null,"abstract":"<p><p>In this study, we investigated the genomic changes in a major methicillin-resistant <i>Staphylococcus aureus</i> (MRSA) clone following a significant outbreak at a hospital. Whole-genome sequencing of MRSA isolates was utilized to explore the genomic evolution of post-outbreak MRSA strains. The epidemicity of the clone declined over time, coinciding with the introduction of multimodal infection control measures. A genome-wide association study (GWAS) identified multiple genes significantly associated with either high or low epidemic success, indicating alterations in mobilome, virulence, and defense mechanisms. Random Forest models pinpointed a gene related to fibrinogen binding as the most influential predictor of epidemicity. The decline of the MRSA clone may be attributed to various factors, including the implementation of new infection control measures, single nucleotide polymorphisms accumulation, and the genetic drift of a given clone. This research underscores the complex dynamics of MRSA clones, emphasizing the multifactorial nature of their evolution. The decline in epidemicity seems linked to alterations in the clone's genetic profile, with a probable shift towards decreased virulence and adaptation to long-term carriage. Understanding the genomic basis for the decline of epidemic clones is crucial to develop effective strategies for their surveillance and management, as well as to gain insights into the evolutionary dynamics of pathogen genomes.</p>","PeriodicalId":15511,"journal":{"name":"Journal of Clinical Microbiology","volume":" ","pages":"e0020324"},"PeriodicalIF":6.1,"publicationDate":"2024-07-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11250417/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141457174","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-07-16Epub Date: 2024-05-29DOI: 10.1128/jcm.00154-24
Jinghao Zhang, Jieli Xu, Siquan Shen, Li Ding, Weiwei Yang, Chengkang Tang, Qingyu Shi, Hu Zhao, Yan Guo, Renru Han, Fupin Hu
The increasing use of ceftazidime-avibactam has led to the emergence of a wide range of ceftazidime-avibactam-resistant blaKPC-2 variants. Particularly, the conventional carbapenemase phenotypic assay exhibited a high false-negative rate for KPC-2 variants. In this study, three colloidal gold immunoassays, including the Gold Mountainriver CGI test, Dynamiker CGI test and NG-Test CARBA5, and GeneXpert Carba-R, were used to detect the presence of KPC-2 carbapenemase and its various variants in 42 Klebsiella pneumoniae strains. These strains covered blaKPC-2 (13/42) and 16 other blaKPC-2 variants including blaKPC-12 (1/42), blaKPC-23 (1/42), blaKPC-25 (1/42), blaKPC-33 (6/42), blaKPC-35 (1/42), blaKPC-44 (1/42), blaKPC-71 (1/42), blaKPC-76 (8/42), blaKPC-78 (1/42), blaKPC-79 (1/42), blaKPC-100 (1/42), blaKPC-127 (1/42), blaKPC-128 (1/42), blaKPC-144 (1/42), blaKPC-157 (2/42), and blaKPC-180 (1/42). For KPC-2 strains, all four assays showed 100% negative percentage agreement (NPA) and 100% positive percentage agreement (PPA) with sequencing results. For all 16 KPC-2 variants, GeneXpert Carba-R showed 100% NPA and 100% PPA, and the three colloidal gold immunoassays showed 100% NPA, while the PPAs of the Gold Mountainriver CGI test, Dynamiker CGI test, and NG-Test CARBA5 were 87.5%, 87.5%, and 68.8%, respectively. We also found a correlation between the mutation site in the amino acid of the variants and false-negative results by colloidal gold immunoassays. In conclusion, the GeneXpert Carba-R has been proven to be a reliable method in detecting KPC-2 and its variants, and the colloidal gold immunoassay tests offer a practical and cost-effective approach for their detection. For the sample with a negative result by a colloidal gold immunoassay test but not matching the drug-resistant phenotype, it is recommended to retest using another type of kit or the GeneXpert Carba-R assay, which can significantly improve the accuracy of detection.
{"title":"Comparison of three colloidal gold immunoassays and GeneXpert Carba-R for the detection of <i>Klebsiella pneumoniae bla</i><sub>KPC-2</sub> variants.","authors":"Jinghao Zhang, Jieli Xu, Siquan Shen, Li Ding, Weiwei Yang, Chengkang Tang, Qingyu Shi, Hu Zhao, Yan Guo, Renru Han, Fupin Hu","doi":"10.1128/jcm.00154-24","DOIUrl":"10.1128/jcm.00154-24","url":null,"abstract":"<p><p>The increasing use of ceftazidime-avibactam has led to the emergence of a wide range of ceftazidime-avibactam-resistant <i>bla</i><sub>KPC-2</sub> variants. Particularly, the conventional carbapenemase phenotypic assay exhibited a high false-negative rate for KPC-2 variants. In this study, three colloidal gold immunoassays, including the Gold Mountainriver CGI test, Dynamiker CGI test and NG-Test CARBA5, and GeneXpert Carba-R, were used to detect the presence of KPC-2 carbapenemase and its various variants in 42 <i>Klebsiella pneumoniae</i> strains. These strains covered <i>bla</i><sub>KPC-2</sub> (13/42) and 16 other <i>bla</i><sub>KPC-2</sub> variants including <i>bla</i><sub>KPC-12</sub> (1/42), <i>bla</i><sub>KPC-23</sub> (1/42), <i>bla</i><sub>KPC-25</sub> (1/42), <i>bla</i><sub>KPC-33</sub> (6/42), <i>bla</i><sub>KPC-35</sub> (1/42), <i>bla</i><sub>KPC-44</sub> (1/42), <i>bla</i><sub>KPC-71</sub> (1/42), <i>bla</i><sub>KPC-76</sub> (8/42), <i>bla</i><sub>KPC-78</sub> (1/42), <i>bla</i><sub>KPC-79</sub> (1/42), <i>bla</i><sub>KPC-100</sub> (1/42), <i>bla</i><sub>KPC-127</sub> (1/42), <i>bla</i><sub>KPC-128</sub> (1/42), <i>bla</i><sub>KPC-144</sub> (1/42), <i>bla</i><sub>KPC-157</sub> (2/42), and <i>bla</i><sub>KPC-180</sub> (1/42). For KPC-2 strains, all four assays showed 100% negative percentage agreement (NPA) and 100% positive percentage agreement (PPA) with sequencing results. For all 16 KPC-2 variants, GeneXpert Carba-R showed 100% NPA and 100% PPA, and the three colloidal gold immunoassays showed 100% NPA, while the PPAs of the Gold Mountainriver CGI test, Dynamiker CGI test, and NG-Test CARBA5 were 87.5%, 87.5%, and 68.8%, respectively. We also found a correlation between the mutation site in the amino acid of the variants and false-negative results by colloidal gold immunoassays. In conclusion, the GeneXpert Carba-R has been proven to be a reliable method in detecting KPC-2 and its variants, and the colloidal gold immunoassay tests offer a practical and cost-effective approach for their detection. For the sample with a negative result by a colloidal gold immunoassay test but not matching the drug-resistant phenotype, it is recommended to retest using another type of kit or the GeneXpert Carba-R assay, which can significantly improve the accuracy of detection.</p>","PeriodicalId":15511,"journal":{"name":"Journal of Clinical Microbiology","volume":" ","pages":"e0015424"},"PeriodicalIF":6.1,"publicationDate":"2024-07-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11250111/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141161178","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-07-16Epub Date: 2024-06-10DOI: 10.1128/jcm.00479-24
Hansheng Wang, Dan Yu, Xiao Chen, Yanhui Zhou, Xin Qian, Dan Liu, Lei Wang, Yijun Tang, Meifang Wang
The diagnosis of invasive pulmonary fungal disease depends on histopathology and mycological culture; there are few studies on touch imprints of bronchoscopic biopsies or lung tissue biopsies for the diagnosis of pulmonary filamentous fungi infections. The purpose of the present study was to explore the detection accuracy of rapid on-site evaluation of touch imprints of bronchoscopic biopsies or lung tissue biopsies for the filamentous fungi, and it aims to provide a basis for initiating antifungal therapy before obtaining microbiological evidence. We retrospectively analyzed the diagnosis and treatment of 44 non-neutropenic patients with invasive pulmonary filamentous fungi confirmed by glactomannan assay, histopathology, and culture from February 2017 to December 2023. The diagnostic positive rate and sensitivity of rapid on-site evaluation for these filamentous fungi identification, including diagnostic turnaround time, were calculated. Compared with the final diagnosis, the sensitivity of rapid on-site evaluation was 81.8%, and the sensitivity of histopathology, culture of bronchoalveolar lavage fluid, and glactomannan assay of bronchoalveolar lavage fluid was 86.4%, 52.3%, and 68.2%, respectively. The average turnaround time of detecting filamentous fungi by rapid on-site evaluation was 0.17 ± 0.03 hours, which was significantly faster than histopathology, glactomannan assay, and mycological culture. A total of 29 (76.3%) patients received earlier antifungal therapy based on ROSE diagnosis and demonstrated clinical improvement. Rapid on-site evaluation showed good sensitivity and accuracy that can be comparable to histopathology in identification of pulmonary filamentous fungi. Importantly, it contributed to the triage of biopsies for further microbial culture or molecular detection based on the preliminary diagnosis, and the decision on early antifungal therapy before microbiological evidence is available.
{"title":"Performance of rapid on-site evaluation of touch imprints of bronchoscopic biopsies or lung tissue biopsies for the diagnosis of invasive pulmonary filamentous fungi infections in non-neutropenic patients.","authors":"Hansheng Wang, Dan Yu, Xiao Chen, Yanhui Zhou, Xin Qian, Dan Liu, Lei Wang, Yijun Tang, Meifang Wang","doi":"10.1128/jcm.00479-24","DOIUrl":"10.1128/jcm.00479-24","url":null,"abstract":"<p><p>The diagnosis of invasive pulmonary fungal disease depends on histopathology and mycological culture; there are few studies on touch imprints of bronchoscopic biopsies or lung tissue biopsies for the diagnosis of pulmonary filamentous fungi infections. The purpose of the present study was to explore the detection accuracy of rapid on-site evaluation of touch imprints of bronchoscopic biopsies or lung tissue biopsies for the filamentous fungi, and it aims to provide a basis for initiating antifungal therapy before obtaining microbiological evidence. We retrospectively analyzed the diagnosis and treatment of 44 non-neutropenic patients with invasive pulmonary filamentous fungi confirmed by glactomannan assay, histopathology, and culture from February 2017 to December 2023. The diagnostic positive rate and sensitivity of rapid on-site evaluation for these filamentous fungi identification, including diagnostic turnaround time, were calculated. Compared with the final diagnosis, the sensitivity of rapid on-site evaluation was 81.8%, and the sensitivity of histopathology, culture of bronchoalveolar lavage fluid, and glactomannan assay of bronchoalveolar lavage fluid was 86.4%, 52.3%, and 68.2%, respectively. The average turnaround time of detecting filamentous fungi by rapid on-site evaluation was 0.17 ± 0.03 hours, which was significantly faster than histopathology, glactomannan assay, and mycological culture. A total of 29 (76.3%) patients received earlier antifungal therapy based on ROSE diagnosis and demonstrated clinical improvement. Rapid on-site evaluation showed good sensitivity and accuracy that can be comparable to histopathology in identification of pulmonary filamentous fungi. Importantly, it contributed to the triage of biopsies for further microbial culture or molecular detection based on the preliminary diagnosis, and the decision on early antifungal therapy before microbiological evidence is available.</p>","PeriodicalId":15511,"journal":{"name":"Journal of Clinical Microbiology","volume":" ","pages":"e0047924"},"PeriodicalIF":6.1,"publicationDate":"2024-07-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11250116/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141296150","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
The rpoB gene has been proposed as a promising phylogenetic marker for bacterial identification, providing theoretically improved species-level resolution compared to the 16S rRNA gene for a range of clinically important taxa. However, its utility in diagnostic microbiology has been limited by the lack of broad-range primers allowing for its amplification from most species with a single PCR assay. Here, we present an assay for broad-range partial amplification and Sanger sequencing of the rpoB gene. To reduce cross-reactivity and allow for rpoB amplification directly from patient samples, primers were based on the dual priming oligonucleotide principle. The resulting amplicon is ~550 base pairs in length and appropriate for species-level identification. Systematic in silico evaluation of a wide selection of taxa demonstrated improved resolution within multiple important genera, including Enterococcus, Fusobacterium, Mycobacterium, Streptococcus, and Staphylococcus species and several genera within the Enterobacteriaceae family. Broad-range rpoB amplification and Sanger sequencing of 115 bacterial isolates provided unambiguous species-level identification for 97 (84%) isolates, as compared to 57 (50%) using a clinical 16S rRNA gene assay. Several unresolved taxonomic matters disguised by the low resolution of the 16S rRNA gene were revealed using the rpoB gene. Using a collection of 33 clinical specimens harboring bacteria and assumed to contain high concentrations of human DNA, the rpoB assay identified the pathogen in 29 specimens (88%). Broad-range rpoB amplification and sequencing provides a promising tool for bacterial identification, improving discrimination between closely related species and making it amenable for use in culture-based and culture-independent diagnostic approaches.
{"title":"Broad-range amplification and sequencing of the <i>rpoB</i> gene: a novel assay for bacterial identification in clinical microbiology.","authors":"Joanna Małgorzata Bivand, Ruben Dyrhovden, Audun Sivertsen, Marit Gjerde Tellevik, Robin Patel, Øyvind Kommedal","doi":"10.1128/jcm.00266-24","DOIUrl":"10.1128/jcm.00266-24","url":null,"abstract":"<p><p>The <i>rpoB</i> gene has been proposed as a promising phylogenetic marker for bacterial identification, providing theoretically improved species-level resolution compared to the 16S rRNA gene for a range of clinically important taxa. However, its utility in diagnostic microbiology has been limited by the lack of broad-range primers allowing for its amplification from most species with a single PCR assay. Here, we present an assay for broad-range partial amplification and Sanger sequencing of the <i>rpoB</i> gene. To reduce cross-reactivity and allow for <i>rpoB</i> amplification directly from patient samples, primers were based on the dual priming oligonucleotide principle. The resulting amplicon is ~550 base pairs in length and appropriate for species-level identification. Systematic <i>in silico</i> evaluation of a wide selection of taxa demonstrated improved resolution within multiple important genera, including <i>Enterococcus, Fusobacterium</i>, <i>Mycobacterium</i>, <i>Streptococcus</i>, and <i>Staphylococcus</i> species and several genera within the <i>Enterobacteriaceae</i> family. Broad-range <i>rpoB</i> amplification and Sanger sequencing of 115 bacterial isolates provided unambiguous species-level identification for 97 (84%) isolates, as compared to 57 (50%) using a clinical 16S rRNA gene assay. Several unresolved taxonomic matters disguised by the low resolution of the 16S rRNA gene were revealed using the <i>rpoB</i> gene. Using a collection of 33 clinical specimens harboring bacteria and assumed to contain high concentrations of human DNA, the <i>rpoB</i> assay identified the pathogen in 29 specimens (88%). Broad-range <i>rpoB</i> amplification and sequencing provides a promising tool for bacterial identification, improving discrimination between closely related species and making it amenable for use in culture-based and culture-independent diagnostic approaches.</p>","PeriodicalId":15511,"journal":{"name":"Journal of Clinical Microbiology","volume":" ","pages":"e0026624"},"PeriodicalIF":6.1,"publicationDate":"2024-07-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11324016/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141331069","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-07-16Epub Date: 2024-06-06DOI: 10.1128/jcm.00416-24
Jung-Ho Youn, Lorenzo Walker, Seth Carlson, Craig Soutar, Karen Frank, Adrian Zelazny, Sanchita Das
{"title":"Mitigation of errors on an FDA-approved platform for cytomegalovirus viral load assay.","authors":"Jung-Ho Youn, Lorenzo Walker, Seth Carlson, Craig Soutar, Karen Frank, Adrian Zelazny, Sanchita Das","doi":"10.1128/jcm.00416-24","DOIUrl":"10.1128/jcm.00416-24","url":null,"abstract":"","PeriodicalId":15511,"journal":{"name":"Journal of Clinical Microbiology","volume":" ","pages":"e0041624"},"PeriodicalIF":6.1,"publicationDate":"2024-07-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11250505/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141261670","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-07-16Epub Date: 2024-05-31DOI: 10.1128/jcm.00369-24
Julia Serrano-Lobo, Elena Reigadas, Patricia Muñoz, Pilar Escribano, Jesús Guinea
Azole resistance screening in Aspergillus fumigatus sensu stricto can be routinely carried out by using azole-containing agar plates (E.Def 10.2 procedure); however, conidial suspension filtering and inoculum adjustment before inoculum preparation are time-consuming. We evaluated whether skipping the filtration and inoculum adjustment steps negatively influenced the performance of the E.Def 10.2 procedure. A. fumigatus sensu stricto isolates (n = 98), previously classified as azole susceptible or azole resistant (E.Def 9.4 method), were studied. Azole-resistant isolates had either the wild-type cyp51A gene sequence (n = 1) or the following cyp51A gene substitutions: TR34-L98H (n = 41), G54R (n = 5), TR46-Y121F-T289A (n = 1), or G448S (n = 1). In-house azole-containing agar plates were prepared according to the EUCAST E.Def 10.2 procedure. Conidial suspensions obtained by adding distilled water (Tween 20 0.1%) were either filtered and the inocula adjusted to 0.5 McFarland or left unfiltered and unadjusted. Agreements between the agar screening methods using inocula prepared by each procedure were high for itraconazole (99%), voriconazole (100%), and posaconazole (94.9%). Sensitivity and specificity (considering the susceptibility category as per the microdilution E.Def 9.4 method as the gold standard) of E.Def 10.2 were 100% to rule in or rule out resistance when unfiltered and unadjusted suspensions were used; the resistance phenotype of isolates harboring the TR34-L98H, G54R, or TR46-Y121F-T289A substitutions was correctly detected. Unfiltered and unadjusted conidial suspensions do not negatively influence the performance of the E.Def 10.2 method when screening for azole resistance in A. fumigatus sensu stricto.
Importance: Azole resistance screening in Aspergillus fumigatus sensu stricto can be routinely carried out by using azole-containing plates (E.Def 10.2 procedure); however, conidial suspension filtering and inoculum adjustment before inoculation of plates are time-consuming. We, here, showed that unfiltered and unadjusted conidial suspensions do not negatively influence the performance of the E.Def 10.2 method when screening for azole resistance in A. fumigatus sensu stricto.
{"title":"Azole resistance screening in <i>Aspergillus fumigatus sensu stricto</i> using the azole-containing agar method (EUCAST E.Def 10.2): conidial suspension filtration and inoculum adjustment before inoculum preparation may not be needed.","authors":"Julia Serrano-Lobo, Elena Reigadas, Patricia Muñoz, Pilar Escribano, Jesús Guinea","doi":"10.1128/jcm.00369-24","DOIUrl":"10.1128/jcm.00369-24","url":null,"abstract":"<p><p>Azole resistance screening in <i>Aspergillus fumigatus sensu stricto</i> can be routinely carried out by using azole-containing agar plates (E.Def 10.2 procedure); however, conidial suspension filtering and inoculum adjustment before inoculum preparation are time-consuming. We evaluated whether skipping the filtration and inoculum adjustment steps negatively influenced the performance of the E.Def 10.2 procedure. <i>A. fumigatus sensu stricto</i> isolates (<i>n</i> = 98), previously classified as azole susceptible or azole resistant (E.Def 9.4 method), were studied. Azole-resistant isolates had either the wild-type <i>cyp51A</i> gene sequence (<i>n</i> = 1) or the following <i>cyp51A</i> gene substitutions: TR<sub>34</sub>-L98H (<i>n</i> = 41), G54R (<i>n</i> = 5), TR<sub>46</sub>-Y121F-T289A (<i>n</i> = 1), or G448S (<i>n</i> = 1). In-house azole-containing agar plates were prepared according to the EUCAST E.Def 10.2 procedure. Conidial suspensions obtained by adding distilled water (Tween 20 0.1%) were either filtered and the inocula adjusted to 0.5 McFarland or left unfiltered and unadjusted. Agreements between the agar screening methods using inocula prepared by each procedure were high for itraconazole (99%), voriconazole (100%), and posaconazole (94.9%). Sensitivity and specificity (considering the susceptibility category as per the microdilution E.Def 9.4 method as the gold standard) of E.Def 10.2 were 100% to rule in or rule out resistance when unfiltered and unadjusted suspensions were used; the resistance phenotype of isolates harboring the TR<sub>34</sub>-L98H, G54R, or TR<sub>46</sub>-Y121F-T289A substitutions was correctly detected. Unfiltered and unadjusted conidial suspensions do not negatively influence the performance of the E.Def 10.2 method when screening for azole resistance in <i>A. fumigatus sensu stricto</i>.</p><p><strong>Importance: </strong>Azole resistance screening in <i>Aspergillus fumigatus sensu stricto</i> can be routinely carried out by using azole-containing plates (E.Def 10.2 procedure); however, conidial suspension filtering and inoculum adjustment before inoculation of plates are time-consuming. We, here, showed that unfiltered and unadjusted conidial suspensions do not negatively influence the performance of the E.Def 10.2 method when screening for azole resistance in <i>A. fumigatus sensu stricto</i>.</p>","PeriodicalId":15511,"journal":{"name":"Journal of Clinical Microbiology","volume":" ","pages":"e0036924"},"PeriodicalIF":6.1,"publicationDate":"2024-07-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11250424/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141179788","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}