Journal of Medical Microbiology

BackgroundPseudomonas aeruginosa (PA) is the most commonly detected pathogen in bronchiectasis. The clinical standard of care for pathogen detection is culture, which has low sensitivity. Early detection and improved monitoring could be used to enhance eradication and long-term suppressive treatments. MethodsA novel bacterial quantification assay (BQA) targeting ribosomal RNA for the detection of viable PA was developed. The BQA sensitivity and specificity in patient samples was assessed utilising culture (n=57); as well as BioFire pneumonia panel (n=111) and 16S rRNA gene sequencing (n=62). The potential clinical impact of the BQA in clinical settings was explored using two international patient cohorts. ResultsThe BQA detected PA in 100% of samples where PA was identified by other methods. The BQA quantification was positively correlated with quantitative culture (r=0.42; p<0.001) and BioFire (r=0.68; p<0.001). The BQA identified PA in an additional 23-44% of cases. To evaluate if this increased detection had clinical implications, we tested sputum samples considered to be PA negative by culture in patients who subsequently tested positive for PA. The BQA detected PA in 8/20 patient samples 8-484 days prior to primary detection by culture. In patients considered to have successful PA eradication by culture, the BQA detected PA in 13 samples from 27 patients who went on to relapse. ConclusionsThe BQA is a highly sensitive detection and quantification method for PA. The BQA demonstrated improved detection and treatment monitoring compared to culture, identifying patients who went on to either isolate a first PA or relapse.

ObjectivesThe aim of this study was to investigate the potential association between the formation of oral bacterial biofilms and the occurrence of antibiotic resistance among 294 oral bacterial isolates. Study designA total of 100; 25-65year old patients were chosen. Buccal mucosa swabs were collected and cultured in appropriate media, then bacteria were isolated and identified. Then 294 bacterial isolates were assessed for biofilm production by the phenotypic method, i.e., the tissue culture plate method (TCPM). Finally, antibiotic susceptibility patterns of the 294 isolates were done by the Kirby-Bauer disc diffusion method for 7 {beta}-lactam antibiotics (ampicillin, penicillin, amoxicillin, cephalothin, oxacillin, cloxacillin, and cefoxitin) and 7 non-{beta}-lactam antibiotics (tetracycline, co-trimoxazole, ciprofloxacin, clindamycin, erythromycin, lincomycin, and vancomycin). ResultsWhen isolates were exposed to biofilm detection by the TCP method, 9 (3.1%) showed high biofilm formation capacity, 213 (72.4%) showed moderate biofilm formation capacity, while 72 (24.5%) showed weak/no biofilm formation. The isolated bacterial biofilms positively showed that the bacterial isolates that showed high and moderate biofilm formation capacity have a higher rate of resistance to most antibiotics with significant difference (p < 0.0001) than weak/no biofilm formation. ConclusionThe present study demonstrates that aerobic bacteria are still the major bacteria isolates from the oral cavity. Antibiotic resistance in the oral bacterial isolates was found to be associated with bacterial biofilm formation.

Background and objectiveDiagnosis of P. jirovecii pneumonia (PJP) is by PCR on lower respiratory tract specimens, the collection of which is not always well-tolerated and requires trained staff and costly equipment not usually available in low-resource settings. We aimed to evaluate P. jirovecii PCR performed on nasopharyngeal swabs (NPS) as a diagnostic test for PJP, as well as the impact of specimen quality on test performance. MethodsPatients with clinically-suspected PJP in public hospitals in Queensland, Australia, who had quantitative P. jirovecii PCR performed on lower respiratory tract specimens from 1 January 2015 to 31 December 2016, and also had NPS collected by healthcare staff within seven days of lower respiratory tract specimen collection were included in this retrospective cohort study. Quantitative P. jirovecii PCR was performed, and sensitivity, specificity, and positive and negative predictive values were calculated. Specimen quality was assessed by quantifying endogenous retrovirus 3 (ERV3) loads, with higher values indicating better specimen quality. ResultsOne hundred and eleven patients were included. The sensitivity of NPS P. jirovecii PCR was 0.66 and specificity was 1.0. The positive predictive value was 1.0 and the negative predictive value was 0.63. Median ERV3 loads in lower respiratory tract specimens and NPS were not significantly different in true positive vs. true negative patients, but was significantly higher in true positives vs. false negatives (7.55x102 vs. 3.67x102; P=0.05). ConclusionP. jirovecii PCR on NPS was highly specific but poorly sensitive. Proper specimen collection is essential to ensure adequate quality and prevent misclassification. Summary at a GlanceUsing nasopharyngeal swabs instead of lower respiratory tract specimens for PCR to diagnose P. jirovecii pneumonia (PJP) may be better tolerated and improve diagnostic accessibility. In this two-year retrospective cohort study of patients with clinically-suspected PJP from Queensland, Australia, P. jirovecii PCR on NPS had high specificity but low sensitivity.

BackgroundLower respiratory tract infections (LRTIs) remain a major cause of morbidity and mortality among hospitalized patients.1 However, isolating organisms from respiratory samples often leads to diagnostic uncertainty due to the coexistence of colonizers, commensals, and contaminants.2 To address this challenge, this study employed a structured, stepwise exploratory model to differentiate true pathogens from non-pathogens in aerobic respiratory cultures and Multiplex PCR (Biofire(R) FilmArray) results. MethodsThis prospective, longitudinal time-bound study was conducted over three months (August- October 2024) at a tertiary care center in northern India. Adult patients ([&ge;] 18 years) with positive lower respiratory tract samples (aerobic culture or Multiplex PCR (Biofire(R) FilmArray) were enrolled. Each isolate was independently classified by the treating clinician, microbiologist, and study investigator using a six-step clinical-pathological algorithm that incorporated clinical signs, Sequential Organ Failure Assessment (SOFA) score trends, alternative infection sources, host factors, and outcome data. The final classification was determined by the investigator. Outcomes, including treatment response and mortality at 28 days, were compared across pathogen and non-pathogen groups. FindingsOf the 145 included cases, 131 (90{middle dot}3%) were classified as pathogens and 14 (9{middle dot}7%) as non-pathogens. Cohens Kappa between investigator and microbiologist classifications was 0{middle dot}28, indicating fair agreement. Among pathogen cases, 68 (51{middle dot}9%) responded to treatment, while 63 (48{middle dot}1%) did not respond to treatment in the pathogenic group. In contrast, 12 of 14 non-pathogen cases (85{middle dot}7%) were not treated, with favourable outcomes in most, and only one unrelated death (7{middle dot}1%). InterpretationThe structured clinico -microbiological model strongly correlates with treatment outcomes, making it useful for differentiating infection from colonization. Crucially, microbiological detection alone doesnt determine pathogenicity. Integrating clinical, laboratory, and outcome data is essential for rational antibiotic use and effective antimicrobial stewardship. FundingNone

Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, Escherichia coli, Streptococcus pneumoniae and Staphylococcus aureus are among the major bacterial causative agents of lower respiratory tract infections (LRTIs), causing substantial morbidity and mortality globally. The rapid increase of antimicrobial resistance (AMR) in these pathogens poses significant challenges for effective antibiotic therapy of LRTIs. In low-resourced settings, the diagnostics of LRTIs relies heavily on microbiological culture and patients are often treated with empirical antibiotics while awaiting several days for culture results. Rapid detection of LRTIs pathogens and AMR genes could prompt early antibiotic switching and inform antibiotic treatment duration. In this study, we developed multiplex quantitative real-time PCRs using EvaGreen dye and melting curve analysis (MCA) to rapidly identify the six major LRTIs pathogens and their AMR genes directly from the tracheal aspirate and sputum samples. The accuracy of RT-PCRs was assessed by comparing its performance against the gold standard, conventional culture method on 50 tracheal aspirate and sputum specimens. Our RT-PCR assays had 100% sensitivity for K. pneumoniae, A. baumannii, P. aeruginosa, E. coli and 63.6% for S. aureus and the specificity ranked from 87.5% to 97.6%. The kappa correlation values of all pathogens between the two methods varied from 0.63 to 0.95. The limit of detection (LOD) of target bacteria in multiplex RT-PCRs was 1600 CFU/mL. Compared to the culture results, PCR assays exhibited higher sensitivity in detecting mixed infections and S. pneumoniae. Our findings also demonstrated a high level of concordance between the detection of AMR gene and AMR phenotype in single infections. We conclude that our multiplex quantitative RT-PCRs with fluorescence MCA is simple but sensitive and specific in detecting six major drug resistant bacterial pathogens of LRTIs and should be further evaluated for clinical utility.

BackgroundHypervirulent Klebsiella pneumoniae (hvKp) infections have mainly been described in Asia. Two patients in the age group of 30 to 50 years presented within a two month period to a tertiary referral hospital in Texas with septic shock, hepatic abscess and septic thrombophlebitis. Blood cultures were positive for Klebsiella pneumoniae (isolates 2020CK-00441 and 2021CK-00720 respectively). MethodsWhole genome sequencing was performed using paired-end Illumina MiSeq reads for both isolates. Nanopore sequencing to obtain a closed genome was performed for 2020CK-00441. Results2020CK-00441 belonged to ST23 type while 2021CK-00720 was a ST65 type isolate. Kleborate analyses predicted with high confidence that both the isolates were hvKp. Phylogenetic analyses showed that the two strains are not closely related to each other or to any other known hvKp isolates. Both the isolates had yersiniabactin, colibactin, aerobactin and salmochelin producing loci which likely confer these isolates hvKp phenotype. 2020CK-00441 had a unique pK2044 like plasmid. ConclusionsHvKp strains capable of causing devastating metastatic septic infections have emerged in Texas. These isolates are unique when compared to other hvKp strains of the world. Country wide surveillance and whole genome sequencing of these strains is essential to prevent a major public health emergency in USA.

BackgroundThe microbial aetiology of early childhood caries (ECC) in sub-Saharan African populations remains poorly characterised, with most studies focusing on conventional cariogenic pathogens like Streptococcus mutans. This study aimed to characterise the salivary microbial profile of children with ECC in urban Kano, northern Nigeria. MethodsIn this cross-sectional study of 162 children aged 3-5 years in urban Kano, unstimulated saliva samples were collected and analysed using standard bacteriological culture methods. Caries status was assessed using decayed, missing, and filled teeth (dmft) index and International Caries Detection and Assessment System (ICDAS). Microbial isolates were identified through Gram staining, colony morphology, and biochemical tests (catalase, coagulase, oxidase). ResultsOf 32 microbial isolates obtained, Staphylococcus aureus was the most prevalent (43.8%, n=14), followed by Streptococcus species (28.1%, n=9), Klebsiella species (12.5%, n=4), non-aureus staphylococci (6.3%, n=2), yeast (6.3%, n=2), and Pseudomonas species (3.1%, n=1). Only one isolate demonstrated direct association with dmft-detectable caries. Polymicrobial colonisation occurred in four cases (12.5%), predominantly featuring S. aureus-yeast combinations (n=2). White spot lesions (ICDAS 1-2) were associated with S. aureus and Klebsiella species in two separate cases. ConclusionThis study reveals an unexpected predominance of S. aureus in the salivary microbiome of children in northern Nigeria, challenging conventional paradigms of ECC microbiology. The low correlation between microbial isolates and clinical caries suggests complex, multifactorial aetiology. These findings highlight the need for molecular characterisation of oral microbiomes in African populations and reconsideration of caries pathogenesis models in this unique epidemiological context.

BackgroundDespite advances in molecular diagnostics, only 48% of newly diagnosed tuberculosis (TB) cases were confirmed using nucleic acid amplification tests (NAATs) in 2023. The BD MAXTM MDR-TB (MAX MDR-TB) assay, a moderate complexity NAAT, detects Mycobacterium tuberculosis complex (MTBC) and resistance to rifampicin (RIF) and isoniazid (INH), but data on clinical performance is limited. This study assessed assay performance on raw sputum, NALC/NaOH decontaminated sputum, and tongue swab (TS) specimens. MethodsThis evaluation assessed the MAX MDR-TB assay for MTBC detection and RIF and INH resistance profiling on sputum, using liquid culture as the reference standard. Additionally, diagnostic accuracy for MTBC detection in TS specimens was evaluated under different transport and processing conditions. ResultsAssay sensitivity was similar for sputum pellet (87%) and raw sputum (89%), with one additional case detected using raw sputum. Two false RIF-resistant results were observed. INH resistance was missed in two cases. Although specimen numbers were small, TS demonstrated better diagnostic accuracy when using a diluted (66%) STR buffer. A total of 15/55 (27%) were classified as "MTB Low POS." ConclusionThese findings suggest that MAX MDR-TB assay performance is comparable between sputum pellet and raw sputum. While TS showed promise, further validation in larger studies is warranted. The high rate of "MTB Low POS" results across specimen types underscores the importance of assay optimisation to reduce the burden of repeat testing and improve diagnostic reliability. Future research should enhance sensitivity and integration into diagnostic algorithms to improve patient outcomes.

BackgroundCandida auris is a worldwide emerging pathogen known for causing infections and outbreaks in health care settings. In Orange County, CA, C. auris has been circulating and causing outbreaks in long term care facilities since 2019, with a total of 1,017 cases detected from February 2019 to December 2021. ObjectiveTo evaluate an rt-PCR assay for rapid identification and assessment of colonization of C. auris from patient surveillance samples at the OC Public Health Laboratory. MethodAn extraction protocol using the MasterPure kit and automated EZ1 Advanced XL followed by rt-PCR using PerfeCTa Multiplex qPCR TaqMan, and the 7500 Fast Dx was conducted. The assay was evaluated using 131 previously confirmed patient samples and 123 prospective fresh samples from different body sites. ResultsThe assay was highly reproducible at an LOD of 230 CFU/ml. The sensitivity and specificity of the 131 samples was 90 % and 86 % respectively and for the 123 samples it was 93% and 90% respectively. ROC analysis was calculated using all samples (n = 254) to determine the most ideal diagnostic value. The AUC was 0.922 and optimal cutoff was a cycle threshold 36.78. A CT [&le;]37.00 is the ideal diagnostic value for patient surveillance samples. ConclusionThe successful implementation of the rt-PCR assay for C. auris in a local public health laboratory allows for accurate and rapid screening and identification of C. auris which can contribute to enhanced surveillance and control of C. auris spread in local health care facilities.

The study explores the polymicrobial nature of primary endodontic infections using Illumina Next Generation Sequencing. Samples involved in research have been collected from root canals of the patients suffering from pulp and periapical inflammations with no history of endodontic interventions on affected teeth. The study revealed prevalence of different bacterial phyla, classes, orders, and species. Further work will show potential correlations between individual microbiotas and clinical diagnosis.

Tooth brushing and flossing are usually both hallmarks of a good oral hygiene routine to prevent decay, gingivitis, and periodontitis. While brushing removes much of the oral bacteria from the front and backs of the teeth, flossing is believed to be necessary to remove bacteria between the teeth. However, the effectiveness of self-flossing has not been established very well. Flossing effectiveness was evaluated two ways in this study: adults and 12 year old children were instructed on how to floss and bacterial colonies were determined before or after 7 days or pediatric and family dentists were blinded to patient surveys that asked about flossing frequency and evaluated the patient for gingivitis. We found a significant number of children did not floss at all despite brushing daily. However, flossing had no effect on the number of bacteria in their mouths nor did flossing have any correlation with reducing gingivitis development. Taken together, self-flossing did not appear to be an effective strategy for reduction of gingivitis in children or adults but could stem from improper technique or simply a lack of doing it.

BackgroundTo describe whether SARS-CoV-2 viral loads (VLs) and cycle thresholds (CTs) vary by sample type, disease severity and symptoms duration. MethodsSystematic searches were conducted in MEDLINE, EMBASE, BioRxiv and MedRxiv. Studies reporting individual SARS-CoV-2 VLs and/or CT values from biological samples. Paired reviewers independently screened potentially eligible articles. CT values and VLs distributions were described by sample type, disease severity and time from symptom onset. Differences between groups were examined using Kruskal-Wallis and Dunn s tests (post-hoc test). The risk of bias was assessed using the Joanna Briggs Critical Appraisal Tools. Results14 studies reported CT values, 8 VLs and 2 CTs and VLs, resulting in 432 VL and 873 CT data points. VLs were higher in saliva and sputum (medians 4.7x108 and 6.5x104 genomes per ml, respectively) than in nasopharyngeal and oropharyngeal swabs (medians 1.7x102 and 4.8x103). Combined naso/oropharyngeal swabs had lower CT values (i.e. higher VLs) than single site samples (p=<0.0001). CT values were also lower in asymptomatic individuals and patients with severe COVID-19 (median CT 30 for both) than among patients with moderate and mild symptoms (31.4 and 31.3, respectively). Stool samples were reported positive for a longer period than other specimens. ConclusionVLs are higher in saliva and sputum and in individuals who are asymptomatic of with severe COVID-19. Diagnostic testing strategies should consider that VLs vary by sample type, disease severity and time since symptoms onset. SummaryThis systematic review found a higher viral load in saliva and sputum than in nasopharyngeal swabs, in asymptomatic individuals and patients with severe COVID-19. Diagnostic testing strategies should consider the type of sample, disease severity and the time since symptoms onset.

BackgroundReliable detection of latent Mycobacterium tuberculosis (Mtb) infection (LTBI) remains challenging, particularly in TB contacts and immunocompromised individuals, where interferon-{gamma} release assays (IGRAs) demonstrate variable sensitivity. IP-10, a chemokine produced at substantially higher concentrations than IFN-{gamma}, represents a promising immune marker. This study aimed to evaluate the diagnostic performance of two IP-10 based assays RIDA(R)QUICK TB (lateral flow) and RIDASCREEN(R) TB (ELISA), by comparison with QuantiFERON-TB Gold Plus (QFT-Plus) assay or a composite reference standard. MethodsA cross-sectional diagnostic accuracy study enrolled 99 adults: 49 with culture-confirmed active pulmonary TB, 30 close TB contacts and 20 individuals with autoimmune disease, in Bucharest, Romania. All participants underwent RIDA Quick, RIDA Screen and QFT-Plus testing. Indeterminate results for all assays were reclassified using a composite reference standard. ResultsAgainst culture in active TB cases, RIDA(R)QUICK TB demonstrated a sensitivity of 85.7% (95% CI: 72.8-94.1) and PPV of 97.7%, while RIDA(R)SCREEN TB achieved 91.8% sensitivity (95% CI: 80.4-97.7) and 97.8% PPV. Specificity and NPV could not be reliably estimated due to the near-absence of true-negative individuals. Agreement with QFT-Plus was moderate to good ({kappa}=0.47-0.93).ROC analysis performed against QFT-Plus as a comparator demonstrated good immunological discrimination for RIDA(R)QUICK TB (AUC = 0.828) and RIDA(R)SCREEN TB (AUC = 0.767), reflecting concordance with QFT-Plus rather than diagnostic accuracy against confirmed infection. ConclusionIP-10 based assays demonstrated higher sensitivity than QFT-Plus and excellent PPV across bacteriological standard, supporting their use as complementary tools for LTBI detection. Larger, more heterogeneous cohorts are needed to accurately define specificity and operational integration.

BackgroundAntibody testing can help define how protective immunity to SARS-CoV-2 is and how long this immunity lasts. Many antibody tests have been evaluated in hospitalised rather than community based COVID-19 cases. Virtus Respiratory Research Ltd (Virtus) has developed its own quantitative IgM and IgG SARS CoV-2 antibody assay. We report its validation and performance characteristics and compare its performance with the Abbott Architect and Roche Elecsys assays in community COVID cases. MethodsWe developed a quantitative antibody test to detect IgM and IgG to the SARS-CoV-2 S1 spike protein (the Virtus test) and validated this test in 107 "true positive" sera from 106 community-managed and 1 hospitalised COVID-19 cases and 208 "true negative" serum samples. We validated the Virtus test against a neutralising antibody test. We determined sensitivities of the Abbott test in the 107 true positive samples and the Roche test in a subset of 75 true positive samples. ResultsThe Virtus quantitative test was positive in 93 of 107 (87%) community cases of COVID-19 and both IgM and IgG levels correlated strongly with neutralising antibody titres (r=0.75 for IgM, r=0.71 for IgG, P<0.0001 for both antibodies). The specificity of the Virtus test was 98.6% for low level antibody positives, 99.5% for moderate positives and 100% for high or very high positives. The Abbott test had a sensitivity of 68%. In the 75 sample subset, the Virtus test was positive in 91%, the Roche test in 69%. ConclusionsThe Abbott and Roche tests had sensitives of 68% and 69% respectively in this community set of COVID-19 sera, while the Virtus test had sensitivities of 87% and 91% in the same sample sets. The strong positive correlation with virus neutralization suggests a positive Virtus quantitative antibody test is likely predictive of protective against recurrent COVID-19. FundingThe development of the Virtus test and sample testing with all antibody tests was funded by Virtus Respiratory Research Ltd. The research studies providing 111 of the 208 of the "true negative" samples was supported by MRC Grant numbers MR/M025330/1 and G1100238 and by the National Institute of Health Research (NIHR) Imperial Biomedical Research Centre (BRC), SLJ is a NIHR Emeritus Senior Investigator and is funded in part by European Research Council Advanced Grant 788575 and the Asthma UK Clinical Chair (grant CH11SJ). The views expressed are those of the author(s) and not necessarily those of the NIHR or the Department of Health and Social Care.

ObjectiveTo evaluate the diagnostic efficiency of different methods in detecting COVID-19 to provide preliminary evidence on choosing favourable method for COVID-19 detection. MethodsPubMed, Web of Science and Embase databases were searched for identifing eligible articles. All data were calculated utilizing Meta Disc 1.4, Revman 5.3.2 and Stata 12. The diagnostic efficiency was assessed via these indicators including summary sensitivity and specificity, positive likelihood ratio (PLR), negative LR (NLR), diagnostic odds ratio (DOR), summary receiver operating characteristic curve (sROC) and calculate the AUC. Results18 articles (3648 cases) were included. The results showed no significant threshold exist. EPlex: pooled sensitivity was 0.94; specificity was 1.0; PLR was 90.91; NLR was 0.07; DOR was 1409.49; AUC=0.9979, Q*=0.9840. Panther Fusion: pooled sensitivity was 0.99; specificity was 0.98; PLR was 42.46; NLR was 0.02; DOR was 2300.38; AUC=0.9970, Q*=0.9799. Simplexa: pooled sensitivity was 1.0; specificity was 0.97; PLR was 26.67; NLR was 0.01; DOR was 3100.93; AUC=0.9970, Q*=0.9800. Cobas(R): pooled sensitivity was 0.99; specificity was 0.96; PLR was 37.82; NLR was 0.02; DOR was 3754.05; AUC=0.9973, Q*=0.9810. RT-LAMP: pooled sensitivity was 0.98; specificity was 0.99; PLR was 36.22; NLR was 0.04; DOR was 751.24; AUC=0.9905, Q*=0.9596. Xpert Xpress: pooled sensitivity was 0.99; specificity was 0.97; PLR was 27.44; NLR was 0.01; DOR was 3488.15; AUC=0.9977, Q*=0.9829. ConclusionsThese methods (ePlex, Panther Fusion, Simplexa, Cobas(R), RT-LAMP and Xpert Xpress) bear higher sensitivity and specificity, and might be efficient methods complement to the gold standard.

Aminoglycoside drugs, amikacin, streptomycin, and amikacin liposome inhalation suspension are crucial for treating refractory Mycobacterium avium-intracellulare complex pulmonary disease. In Mycobacterium tuberculosis, cross-resistance occurs between amikacin and kanamycin, but not between amikacin and streptomycin in genetic drug susceptibility testing. However, the occurrence of cross-resistance among aminoglycosides remains unclear in M. avium-intracellulare complex. We aimed to evaluate cross-resistance among aminoglycosides to determine whether streptomycin or kanamycin remains effective after the development of amikacin resistance. This single-center retrospective study included 20 patients with amikacin-resistant M. avium-intracellulare complex harboring rrs mutations. Paired analyses of streptomycin and kanamycin minimum inhibitory concentration values before and after amikacin resistance development were performed. In addition, streptomycin- and kanamycin-resistant strains were generated in vitro and resistance-associated mutations were identified using whole-genome sequencing. No significant increase was observed in streptomycin minimum inhibitory concentration values following amikacin resistance. In contrast, kanamycin values uniformly increased to >256 g/mL after the acquisition of amikacin resistance. Furthermore, amikacin- and kanamycin-resistant isolates shared mutations at position 1408 in the rrs gene, whereas streptomycin-resistant isolates exhibited mutations at position 20 in the rrs gene. These results suggest that amikacin and kanamycin exhibit cross-resistance in M. avium-intracellulare complex, whereas amikacin and streptomycin may not. Two cases in our cohort in which streptomycin treatment was effective after the acquisition of amikacin resistance further support these findings. In conclusion, streptomycin may be a potential therapeutic alternative for amikacin-resistant M. avium-intracellulare complex pulmonary disease. Future studies correlating streptomycin minimum inhibitory concentration values with clinical outcomes are required.

IntroductionC-reactive protein (CRP) is an established acute-phase marker for infection and inflammation, which can help guide clinical decision-making in primary and secondary care. Many European guidelines recommend point-of-care (POC) CRP testing to improve antimicrobial stewardship in primary care. This performance evaluation study assessed the equivalence of the quantitative POC LumiraDx CRP Test compared to a laboratory-based reference method. MethodsMethod comparison, matrix equivalency, and precision were evaluated. Plasma samples from secondary care patients presenting with symptoms of infection or inflammation were analyzed centrally using the LumiraDx CRP Test and the reference test (Siemens CRP Extended Range for Dimension(R) Clinical Chemistry System). The method comparison was conducted used Passing-Bablok regression analysis with prespecified criteria of r[&ge;]95 and a slope of 0.95-1.05. The REACT study (NCT05180110) evaluated the equivalence and precision of the testing modalities (fingerstick, venous blood, and plasma samples from the same secondary care patient) using Passing-Bablok regression analysis of the results of the POC LumiraDx CRP Test. ResultsIn analysis of 320 plasma samples from 110 patients, the POC LumiraDx CRP Test demonstrated close agreement with the reference method, meeting the prespecified performance criteria (r=0.99, slope of 1.05, N=110). Paired replicate precision of the testing modalities was high, with mean %CV of 6.4 (plasma), 6.6 (capillary direct), and 8.1 (venous blood). Passing-Bablok regression showed matrix equivalency for all replicate pairs of the testing modalities, with r values across all sample types of 0.97-0.98. ConclusionThe quantitative POC LumiraDx CRP Test showed very close agreement with the established laboratory-based test when using capillary blood, venous blood, or plasma. The use of capillary blood testing in particular is beneficial in both primary and secondary care, with this portable test system providing rapid quantitative results within 4 minutes, potentiating the ability to help guide clinical decision-making. Data from two study collections, the NOVEL study and the REACT study with a trial registration: ClinicalTrials.gov identifier, NCT05180110, were used in this performance evaluation. Key summary pointsO_LIC-reactive protein (CRP) measurements are clinical markers for infection and inflammation, commonly used in primary and secondary care C_LIO_LIPoint-of-care (POC) CRP testing can assist primary care clinicians in making an immediate decision as to whether to prescribe antibiotics while the patient is still at the clinic C_LIO_LIPOC CRP testing that provides quantitative results near to the patient can be useful in emergency care assessment of patients and in hospital monitoring of antibiotic therapy C_LIO_LIThe POC LumiraDx CRP Test has demonstrated quantitative results comparable to those obtained using a recognized laboratory system using plasma C_LIO_LIThe POC LumiraDx CRP Test has also demonstrated matrix equivalence of capillary blood (both direct application and transfer tube), venous blood, and plasma C_LI

Colonization efficiency varies considerably between Streptococcus pneumoniae (pneumococcus) strains. The microbial characteristics that influence those differences are still largely unknown. Here, we report rates and kinetics of colonization of four pneumococcal strains upon experimental human pneumococcal challenge. Healthy adults were intranasally challenged with one of four pneumococcal strains (serotype/clonal name: 6B/BHN418, 15B/SH8286, 23F/P1121 and 23F/P833) over a range of doses. Maximum colonization achieved was 60%, 31%, 16% and 10%, respectively. Density and duration of colonization did not differ significantly between the tested strains. We further evaluated murine colonization, non-opsonic neutrophil mediated killing, epithelial cell adherence and average chain length of these four pneumococcal strains. Of these, only chain length was found to be associated with colonization efficiency in the human challenge model. Our data demonstrate that colonization rates following experimental challenge vary with the strain used and suggest that efficiency in colonization is related to pneumococcal chain length.

Nasopharyngeal (NP), nasal and throat swabs are the most practical specimen sources to test for upper respiratory pathogens. We compared the sensitivity of NP, nasal and throat swabs to detect SARS-CoV-2 in community patients. Using detection at any site as the standard, the sensitivities were 90%, 80% and 87% for NP, nasal and throat respectively (n=30 positive at any site). Throat swabs are likely a suitable alternative to NP swabs for the detection of COVID-19 infections.

Background Real time PCR provides rapid and accurate laboratory confirmation of clinically suspected pertussis and pertussis like illness (PLI). However, it does not inform the viability as it detects bacterial DNA from both viable and degraded bacterial cells. We report development of platinum based viability real time PCR assays for Bordetella pertussis, B.parapertussis and B.holmesii which cause pertussis and PLI. Material and MethodsATCC strains of the three organisms were grown on selective media, and cell counts determined. Untreated (control) and Sodium dodecyl sulphate (SDS) lysed cultures were treated with varying concentrations of platinum chloride (PtCl4) and genomic DNA extracted. Real time PCR was performed for the targets of B.pertussis (IS481), B.parapertussis (pIS1001) and B.holmesii (hIS1001) respectively, and quantitative results obtained. threshold cycle values after PtCl4 exposure and before PtCl4 exposure ({Delta} Cq) were calculated for both untreated and SDS treated cultures. ResultsFor all three species, increase in {Delta}Cq values was noted with corresponding reduction in DNA copy numbers, as PtCl4 concentration was increased, in untreated and SDS treated cultures. Untreated cultures of all three species exposed to PtCl4showed increase in Cq values, suggesting that liquid cultures contain a mixture of live and dead bacteria. ConclusionPerformance of viability q PCR assay incorporating PtCl4 at a concentration range of 6 to 10 mM would inform the viability of the infecting organism in the clinical sample. This assay would improve the diagnostic reliability, help select samples for culture as well as adoption of mitigation strategies among contacts of pertussis cases. ImportanceO_LIPertussis and pertussis like illness (PLI) are highly contagious respiratory tract infections for which Real time PCR is the frontline diagnostic tool. C_LIO_LIThis test however does not inform the viability of the infecting pathogen, as it detects nucleic acid which can be present even after degradation of the cell. C_LIO_LIPerforming a viability qPCR incorporating platinum chloride (PtCl4) at a concentration range of 6 to 10 mM would inform the viability of B. pertussis, B. parapertussis and B. holmesii in the clinical sample. C_LIO_LIIt is critical to know the viability of the infecting organism to determine which cases of pertussis and PLI which are actually infectious, to adopt mitigation strategies including prophylactic antibiotics among close contacts C_LIO_LIA viability assay can be used for quantitative measurement of bacterial number and viability and will help select the correct specimens for culture and isolation of the pathogen. C_LI