Journal of Clinical Microbiology最新文献

Diagnosing malaria using standard techniques is time-consuming. With limited staffing in many laboratories, this may lead to delays in reporting. Innovative technologies are changing the diagnostic landscape and may help alleviate staffing shortages. The miLab MAL, an automated artificial intelligence-driven instrument was compared with standard microscopy at LabCorp reference laboratories. Four hundred eight samples submitted for parasitic examination were prepared with thick and thin smears and Noul's malaria platform miLab MAL, and evaluated for positivity, negativity, percent positivity, and species identification. Of 408 samples, 399 samples were manually negative, while 397 were negative by miLab MAL. Two samples initially classified as negative manually were found positive by miLab MAL. In all nine cases, Plasmodium falciparum was identified by both methods. Percentage parasitemia was higher in the manually calculated method, especially when >1%. miLab MAL was accurate in identifying the absence of Plasmodium falciparum and exhibited higher sensitivity than the manual method. All positive samples detected by microscopy were also identified with miLab MAL. All positive Plasmodium cases were correctly identified by miLab MAL. However, the number of positive samples was limited to only Plasmodium falciparum. Although parasitemia by the manual method was on average six times higher than with miLab MAL, this may be due to sampling variability. The findings show that miLab MAL can be used to screen out negative Plasmodium falciparum samples. Further studies assessing parasitemia between methods and identification of non-falciparum samples are necessary to assess the reliability of this new technology.

This study aimed to evaluate a new protocol of the bile solubility test performed directly on the blood from positive blood culture bottles to identify Streptococcus pneumoniae rapidly. Seventy-five positive blood cultures (PBC) showing Gram-positive cocci in pairs or chains on Gram stain, including 32 S. pneumoniae isolates and three reference American Type Culture Collection (ATCC) isolates were included to evaluate the performance of a modified bile solubility test (MBST). One milliliter of blood from the PBC bottle was mixed with 0.5 mL of 10% desoxycholate or a saline solution. Both suspensions were analyzed after 10 min of incubation through a Gram stain to detect solubilization. This technique was compared with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry identification, performed on PBC following extraction or on colonies after short or standard incubation, and the optochin susceptibility test on colonies. The capsular serotypes were determined for all S. pneumoniae, and the Belgian National Reference Center confirmed the identification. All 32 clinical isolates and the ATCC isolate of S. pneumoniae were solubilized on the desoxycholate-treated slides, while the other species tested remained visually unchanged on both, the test and control slides. The MBST test demonstrated a 100% sensitivity and specificity with a mean turnaround time (TAT) of just 39 min, making it 14 h and 56 min faster than the optochin susceptibility test. This rapid variant of the bile solubility test appears to be a reliable method to identify S. pneumoniae directly from positive blood culture bottles, with a TAT of 39 min. It is a cost-effective, easy-to-perform, and time-efficient technique. Negative results should be interpreted cautiously, as they may result from mixed infections with S. pneumoniae and other Gram-positive cocci.

Cyclosporiasis has historically been underdiagnosed due to a lack of clinical recognition and challenges in laboratory detection. Microscopic detection of Cyclospora requires either the modified Kinyoun's acid-fast or safranin stains, which are not part of the standard ova-and-parasite examination. Alternatively, the use of fluorescence microscopy for the detection of Cyclospora due to the organism's autofluorescent properties has been an available tool in the field of clinical parasitology. However, it is unknown how ultraviolet (UV) fluorescence performs compared to permanent stains in routine diagnostic laboratory settings. In this study, we present a direct head-to-head comparison of UV fluorescence against modified acid-fast (MAF) stains using 50 (35 Cyclospora positive, 5 Cystoisospora positive, and 10 Cystoisospora negative) blinded, concentrated stool specimens. Five technically competent medical laboratory technologists of varying years of experience in clinical parasitology at a large reference laboratory independently read each of the MAF slides and UV wet mounts. Overall, the performance of UV fluorescence was similar to MAF slides with a higher agreement of 88.8% (95% CI 86.3%-91.3%) compared to 85.2% (95% CI 84.2%-86.2%), respectively. A higher false-negative rate was associated with MAF slides (MAF: 11.6%, UV: 6.8%). However, UV fluorescence was found to have a higher proportion of false positives (UF: 3.2%, MAF: 2%) and misidentified coccidia. Discrepant results regardless of methods were more frequently observed with technologists who have less experience in parasite morphology. This study is the first to document evidence to support the increased sensitivity and utility of UV fluorescence to improve laboratory diagnosis of cyclosporiasis.

Importance: This study is important as there is a dearth of studies in our field of clinical parasitology that investigate and establish performance characteristics of classic and newer, non-molecular methods. While these studies may not seem as heavy hitting as some new technologies described in our related disciplines, for our field, such studies are long overdue and critically lacking.

Antibiotic resistance, particularly carbapenem resistance, poses a significant global health threat due to the limited availability of effective antibiotics. Carbapenem-resistant Aeromonas are increasingly recognized for their role in various infections, necessitating rapid and accurate detection methods. This study aimed to evaluate several phenotypic tests, including the Carba NP test (CNPt), Carba NP-direct test (CNPd), and Blue-Carba test (BCT), for their effectiveness in rapidly detecting carbapenemase production in Aeromonas. These tests target both the chromosomally encoded CphA metallo-β-lactamase (MBL) and acquired carbapenemases. Additionally, a modified phenotypic test called the Colony-Carba NP test (c-CNPt) was introduced to enhance sensitivity and specificity. A retrospective analysis was conducted on 131 clinically conserved Aeromonas strains harboring identified carbapenem resistance genes, using CNPt, CNPd, BCT, and the newly developed c-CNPt and EDTA-Colony-Carba NP test (ec-CNPt). The stability of c-CNPt reagents stored at -80°C was also assessed. Additionally, a prospective study conducted from July 2021 to November 2023 evaluated 152 Aeromonas isolates to determine the clinical applicability of these tests. Our results demonstrated that CNPd and BCT achieved 100% sensitivity and specificity, surpassing the traditional CNPt, which showed only 63.6% sensitivity for Aeromonas strains. The c-CNPt also showed 100% sensitivity and specificity, with the ec-CNPt effectively differentiating between MBL and serine carbapenemase types. Stability tests confirmed that c-CNPt reagents could be stored at -80℃ for up to 1 year without performance degradation. These findings highlight the practicality and reliability of these phenotypic tests for routine laboratory use, providing a rapid and cost-effective method for detecting carbapenemase production.The rapid detection of carbapenemase production in Aeromonas is of paramount importance due to the significant clinical and public health implications associated with antibiotic resistance. The development and validation of rapid phenotypic tests such as the Colony-Carba NP test (c-CNPt) and the EDTA-Colony-Carba NP test (ec-CNPt) are crucial advancements in the field. These tests offer a highly sensitive and specific method for detecting carbapenemase production in Aeromonas, including the differentiation between metallo-β-lactamase and serine carbapenemases. The c-CNPt and ec-CNPt are cost-effective, easy to perform, and provide rapid results, making them suitable for routine clinical use. Additionally, the stability of the reagents ensures their practicality for long-term application in various healthcare settings. Implementing these phenotypic tests in clinical laboratories can significantly enhance the early detection and appropriate treatment of carbapenem-resistant Aeromonas infections.

Invasive aspergillosis (IA) is a life-threatening infection. Early and specific diagnosis is pivotal to ensure adequate therapy. Antigen testing from blood is a widespread and convenient diagnostic approach. Various tests for the detection of Aspergillus antigen as well as for the panfungal antigen β-1,3-D-glucan (BDG) are available, for which comprehensive comparisons are still lacking. Blood samples of 82 proven/probable (11/71) IA patients and 52 controls were tested using two enzyme-linked immunosorbent assays (ELISAs) (Bio-Rad and Euroimmun), one chemiluminescent immunoassay (CLIA) (Vircell), one BDG assay (Fujifilm Wako), and two point of care (PoC) assays (Immy sōna and OLM). PoC assays were evaluated visually and used automated read out systems. Of the 82 IA patients, 37 had received solid organ transplantation (SOT) and 25 hematopoietic stem cell transplant (HSCT). Sensitivities and specificities for the eight test systems ranged from 27% to 71% and from 64% to 100%. Estimating a 10% prevalence of IA, test performance would have resulted in positive and negative predictive values of 14%-100% and 91%-95%. Areas under the curve (AUCs) for all tests except GM were below 0.7. When the cut-off values for quantitative tests were normalized to a specificity close to 95%, sensitivities ranged from 14% to 40%. The use of automated read out systems for the PoC assays had a significant impact. Combining different tests did not result in better test strategies. Sensitivity of Aspergillus antigen testing from single serum samples is low. Due to specificity issues, the majority of tests is not suited for screening purposes. The different assays can meet different needs in different diagnostic settings.

Rapid antimicrobial drug administration is crucial for the efficient treatment of sepsis or septic shock, but empirical therapy is limited by the increasing prevalence of multidrug-resistant bacteria. Thus, rapid and reliable antimicrobial susceptibility testing (AST) is needed to start appropriate antimicrobial drug administration as quickly as possible. In the present study, we evaluated the performance of the Reveal rapid AST system. From February to April 2021, 102 positive blood culture bottles (BCBs) from hospitalized patients with bacteremia caused by Gram-negative bacteria were included in the study. All isolates were tested by the Reveal system directly from the positive BCBs in comparison to the DxM MicroScan WalkAway. Essential agreement (EA) and category agreement (CA) were high with 98.5% and 97.1%, respectively. We also determined the susceptibility of 10 highly resistant CDC & FDA AR strains in duplicate. Here, EA was 99.6% and CA 97.9%. The average time to result by Reveal was 5.4 h ± 1.2 h compared to an average of 16 h by DxM MicroScan WalkAway for clinical strains and 3.8 h ± 1.2 h for more resistant CDC & FDA AR strains. Susceptibility determination with the Reveal rapid AST system directly from positive BCBs is for the frequently represented bug-drug combinations a reliable and accurate approach, meeting the European ISO guideline for the performance of AST systems. Moreover, AST directly from blood cultures performed with the Reveal system saves time when compared to the conventional AST, as no subculturing is needed and time to result is very short.

The promising diagnostic performance of molecular testing for syphilis using saliva and urine samples has been reported; however, further evaluation of its possible application for diagnosis and molecular surveillance is required. In addition, the development of a rapid and easy-to-perform molecular test for syphilis is important for its use in the clinical setting. We comprehensively evaluated the diagnostic and surveillance performance of two novel loop-mediated isothermal amplification (LAMP) assays using saliva and urine samples. Saliva, urine, and whole blood were collected from patients who underwent serological testing for syphilis at outpatient clinics. Treponema pallidum DNA in specimens was detected using quantitative PCR (qPCR), nested PCR, and novel LAMP assays. T. pallidum genotyping was conducted by multi-locus sequence typing (MLST). Of the 163 patients recruited, 98 were diagnosed with syphilis (primary: n = 35; secondary: n = 40; latent: n = 23). qPCR showed the highest sensitivity among the molecular tests performed with a sensitivity of 54.1% and 30.3% for all syphilis patients using saliva and urine samples, respectively. A novel method of LAMP combined with dry reagents and crude DNA extraction (Dry-LAMP) showed a probit detection limit of 37.4 copies/reaction within 45 min. The agreement rate between Dry-LAMP and qPCR for saliva was 95.7% (κ coefficient 0.90). The T. pallidum genotype was identified in 48 patients by MLST using saliva samples. Molecular analysis of saliva could be used as a supplementary diagnostic test for syphilis and molecular surveillance of the T. pallidum genotype. Dry-LAMP is expected to be helpful in the clinical diagnosis of syphilis.

The Gram-positive bacteria Streptococcus pneumoniae is part of the Streptococcus mitis group (SMG) and causes life-threatening infections, such as pneumonia, sepsis, and meningitis. The closely related Streptococcus pseudopneumoniae has recently been shown to cause respiratory tract infections, as well as invasive infections, especially in patients with comorbidities. Due to the genetic and phenotypic similarities of species belonging to the SMG, the identification of S. pneumoniae and S. pseudopneumoniae is difficult and unreliable using phenotypic tests, as well as matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) and 16S rRNA sequencing. In this study, a loop-mediated isothermal amplification (LAMP)-based assay was developed using molecular markers specific for S. pneumoniae (SPN0001) and S. pseudopneumoniae (SPPN_RS10375). The LAMP assay was evaluated using a collection of SMG clinical isolates, concluding that the method provides a correct, reliable, and fast identification of both S. pneumoniae and S. pseudopneumoniae clinical isolates.

The definition of the microbiology of pleural space infection has been challenging due to the poor yield of conventional culture. Here, the results of a 16S ribosomal RNA gene PCR/sequencing assay performed on pleural fluid in routine clinical practice between August 2020 and January 2023 were evaluated. Amplified 16S rRNA gene DNA was submitted to Sanger sequencing and/or next-generation sequencing or results were reported as negative, depending on PCR crossing threshold value. In all, 496 pleural fluids were tested at Mayo Clinic Laboratories, with 227 positive results, including 57 from Mayo Clinic patients. Among the 57 Mayo Clinic patients, pleural space infection was community acquired in 48 (84%); Fusobacterium nucleatum group and/or Streptococcus intermedius were detected in 31/57 (54%) cases [including 28/48 (58%) community-acquired cases], with additional facultative and/or anaerobic species also found in various combinations in 17/31 (55%). Results of this study suggest that the most frequent microorganism profile involved in community-acquired pleural space infection may be a combination of F. nucleatum group and/or S. intermedius, with or without other normal microbiota.

Importance: We describe here the most frequent microorganisms detected in community-acquired pleural space infection using a clinically performed sequencing-based assay. We found that the most common detection was the Fusobacterium nucleatum group and/or Streptococcus intermedius, with or without other normal microbiota. We propose the term e-FuSion (effusion with Fusobacterium nucleatum group, Streptococcus intermedius, and other oral normal microbiota) for this entity.