Conventional Culture Research Articles

Diagnostic Benefit of Molecular Imaging in Patients Undergoing Heart Valve Surgery for Infective Endocarditis.

(1) Background: The successful treatment of infective endocarditis (IE) relies on detecting causative pathogens to administer targeted antibiotic therapy. In addition to standard microbiological cultivation of pathogens from tissue obtained during heart valve surgery, the potential of molecular biological methods was evaluated. (2) Methods: A retrospective study was performed on heart valve tissue from 207 patients who underwent heart valve surgery for IE. FISHseq (fluorescence in situ hybridization combined with 16S rRNA gene PCR and sequencing) was performed in addition to conventional culture-based microbiological diagnostics. The diagnostic performance of FISHseq was compared with the conventional methods and evaluated in the clinical context. (3) Results: Overall, FISHseq provided a significantly higher rate of specific pathogen detection than conventional valve culture (68.1% vs. 33.3%, p < 0.001). By complementing the findings from blood culture and valve culture, FISHseq was able to provide a new microbiological diagnosis in 10% of cases, confirm the cultural findings in 24.2% of cases and provide greater diagnostic accuracy in 27.5% of cases. FISHseq could identify a pathogen in blood-culture-negative IE in 46.2% of cases, while valve culture provided only 13.5% positive results (p < 0.001). (4) Conclusions: This study demonstrates that using FISHseq as an additional molecular biological technique for diagnostics in IE adds substantial diagnostic value, with potential implications for the treatment of IE. It provides pathogen detection, especially in cases where conventional microbiological cultivation is negative or inconclusive.

Improved detection of mycobacteria in CF and tissue samples grown in mycobacteria growth indicator tube incubated at 30°C compared to conventional growth conditions of liquid and solid media.

Our study shows that the addition of mycobacteria growth indicator tube (MGIT) liquid culture incubated at 30°C improves the detection of mycobacteria from CF and tissue samples. MGIT incubated at 30°C recovered significantly more mycobacterial isolates than MGIT incubated at 37°C and significantly more isolates than either Lowenstein Jensen or Stonebrink solid media incubated at either 36°C or 30°C. Of 202 mycobacterial isolates recovered from 1,549 specimens, 170 were recovered from MGIT incubated at 30°C, followed by MGIT incubated at 37°C with 124 isolates and solid media culture conditions that recovered between 95 and 107 mycobacterial isolates. All conventional culture conditions combined without MGIT incubated at 30°C recovered 166 isolates. MGIT incubated at 30°C recovered the highest number of isolates detected exclusively by a single culture condition and recovered mycobacterial isolates of highly relevant mycobacterial species, including Mycobacterium abscessus and Mycobacterium tuberculosis.

Comparison of targeted next-generation sequencing and traditional microbial culture in the diagnosis of pulmonary infections

This study investigated the diagnostic potential of targeted next-generation sequencing (tNGS) for pulmonary infections. The positivity rate of tNGS was significantly higher than that of traditional microbial culture (92.6 % vs 25.2 %, χ2 = 378.272, P < 0.001). The proportion of two or more species of pathogens detected using tNGS exceeded that detected using microbial culture (χ2 = 337.283, P < 0.001). There were inconsistencies between the results of the tNGS antibiotic resistance gene and the drug susceptibility test resistance phenotype. The tNGS technique demonstrates rapid and effective capabilities in identifying bacteria, fungi, viruses, and specific pathogens, with a detection sensitivity that surpasses that of conventional culture methodologies. Microbial drug resistance genotypes detected by tNGS cannot accurately predict drug resistance phenotypes and require further improvement or integration with traditional microbial culture to establish a foundation for effective clinical treatment.

Etiology and antimicrobial susceptibility patterns of bacteria causing pneumonia among adult patients with signs and symptoms of lower respiratory tract infections during the COVID-19 pandemic in Mwanza, Tanzania: a cross-sectional study

BackgroundBacterial pneumonia is among the leading causes of morbidity and mortality worldwide. The extensive misuse and overuse of antibiotics observed during the Corona Virus Disease 2019 (COVID-19) pandemic may have changed the patterns of pathogens causing bacterial pneumonia and their antibiotic susceptibility profiles. This study was designed to establish the prevalence of culture-confirmed bacterial pneumonia and describe their antimicrobial susceptibility profile in adult patients who presented with signs and symptoms of lower respiratory tract infections (LRTIs) during the COVID-19 pandemic.MethodologyThis hospital-based cross-sectional study was conducted from July 2021 to July 2022 at a zonal referral hospital and two district hospitals in Mwanza, Tanzania. Demographic and clinical data were collected using a standardized questionnaire. Sputum samples were processed by conventional culture followed by the identification of isolates and antibiotic susceptibility testing. Descriptive data analysis was performed using STATA version 15.0.ResultsA total of 286 patients with a median age of 40 (IQR 29–60) years were enrolled in the study. More than half of the patients enrolled were females (52.4%, n = 150). The overall prevalence of bacterial pneumonia was 34.3% (n = 98). The majority of the bacterial pathogens isolated were Gram-negative bacteria (GNB) (61.2%, 60/98), with a predominance of Klebsiella spp., 38.8% (38/98), followed by Streptococcus pyogenes (21.4%, 21/98). Multi drug resistant (MDR) bacteria were detected in 72/98 (73.5%) of the isolates. The proportions of GNB-resistant strains were 60.0% (36/60) for ciprofloxacin, 60% (36/60) for amoxicillin, 60% (36/60) for amoxicillin, 68.3% (41/60) for trimethoprim-sulfamethoxazole and 58.3% (35/60) for ceftriaxone.ConclusionOne-third of the patients with signs and symptoms of LRTIs had laboratory-confirmed bacterial pneumonia with a predominance of Gram negative MDR bacteria. This calls for continuous antimicrobial resistance (AMR) surveillance and antimicrobial stewardship programs in the study setting and other settings in developing countries as important strategies for tackling AMR.

Urinary tract infection among people living with human immunodeficiency virus attending selected hospitals in Addis Ababa and Adama, central Ethiopia.

Urinary tract infections (UTIs) and antibacterial resistance (ABR) are important public health problems, but they are not well-studied among people living with human immunodeficiency virus (PLHIV) globally, especially in low-income countries. Therefore, it is important to regularly measure the extent of UTIs and ABR in the most susceptible populations. This study aimed to investigate the prevalence of UTIs, associated factors, bacterial causal agents, and their antibiotic susceptibility profile among PLHIV in central Ethiopia. A hospital-based cross-sectional study was conducted to recruit 688 PLHIV by a simple random sampling method. Background information was gathered through interviews, while clinical information was gathered from recent information sheets of patient charts using organized, pretested, and validated study tools. Midstream urine was collected aseptically and transported to the Microbiology Laboratory of Aklilu Lemma Institute of Pathobiology within 4 h of collection, maintaining its cold chain. Standard conventional microbial culture methods and matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry were used to identify the bacterial isolates at the species level. Kirby Bauer's disk diffusion method was used to determine the antibiotic susceptibility profile of the bacterial isolates based on the interpretation guidelines of the Clinical Laboratory Standard Institute. Logistic regression models were used to examine factors associated with the occurrence of UTIs among PLHIV attending selected hospitals in Addis Ababa, and Adama. Out of 688 PLHIVs involved in the current study, 144 (20.9%) were positive for UTIs, whereas the majority were asymptomatic for UTIs. In the multivariable logistic regression analysis, only HIV RNA ≥ 200 copies/ml [AOR = 12.24 (95% CI, 3.24, 46.20), p < 0.01] and being symptomatic for UTIs during the study period [AOR = 11.57 (95% CI, 5.83, 22.97), p < 0.01] were associated with the occurrence of UTIs. The dominant bacterial species isolated were Escherichia coli (E. coli; n = 65; 43%), followed by Enterococcus faecalis (E. faecalis; n = 16; 10.6%) and Klebsiella pneumoniae (K. pneumoniae; n = 11; 7.3%). Over half of the E. coli isolates were resistant to antibiotics such as gentamicin (GM; n = 44; 67.7%), amikacin (AN; n = 46; 70.8%), nalidixic acid (NA; n = 42; 64.6%), ciprofloxacin (CIP; n = 40; 61.5%), and azithromycin (AZM; n = 45; 69.2%). All of the K. pneumoniae isolates (n = 11; 100%), (n = 6; 54.5%), and (n = 7; 63.6%) were resistant to [amoxicillin as well as amoxicillin + clavulanic acid], ceftriaxone, and sulfamethoxazole + trimethoprim, respectively. All the Staphylococcus aureus (S. aureus) isolates were resistant to cefoxitin, which implies methicillin-resistant S. aureus (MRSA). The high prevalence of UTIs and antibiotic resistance revealed in the current study needs public health interventions such as educating the population about preventive measures and the importance of early treatment of UTIs. Our findings also highlight the need to provide UTI screening services for PLHIV, and healthcare providers should adopt antibiotic stewardship programs to promote and ensure their appropriate and judicious use.

Verification and application of qPCR and viability-qPCR for Legionella monitoring in evaporative cooling systems complementing the conventional culture method

To date, in many countries the only legally valid method for evaporative cooling system (ECS) monitoring is the culture method. However, a duration of up to 14 days and a risk of underestimation of Legionella concentrations are seen as limitations of cultivation methods. Rapid cultivation-independent methods are an important step towards a more practicable monitoring of ECS to quickly control interventions if elevated concentrations of Legionella are found.Two commercial kits for quantitative polymerase chain reaction (qPCR) and viability-qPCR (v-qPCR) were studied, comprising sample filtration and DNA extraction. Cryopreserved Legionella pneumophila were established as calibration standard with intact (ILC) and total Legionella count (TLC) determined by flow cytometry before conducting spiking experiments in commercial mineral water and artificial process water. Final assessment was carried out using real ECS samples.Recovery and robustness ranged from 86 to 108 % for qPCR with a drop to 40–60 % for v-qPCR when compared to direct extraction, possibly attributable to cell damage during sample concentration. All methods including culture did perform well regarding linearity with R2 ≥ 0.95 for most trials. Detected concentrations in comparison to spiked Legionella counts differed with culture averaging 25 ± 7 % of spiked ILC and v-qPCR being closest to spiked concentrations with 65–144 %. In comparison, qPCR was several fold above spiked TLC concentrations. For real ECS samples Legionella spp. were detected in concentrations above 103 GU/100 mL by v-qPCR in 70–92 % of samples, depending on the kit used. Most of these samples were either culture-negative or not evaluable on agar plates.This study showed that a cryopreserved bacterial standard based examination is applicable and can be used for future v-qPCR verification. For assessment of differences in results between culture and v-qPCR/qPCR in ECS samples expert knowledge about the operating mode and used analytical methods is required. Guidelines addressing this issue could be a solution.

Agricultural Mitigation Strategies to Reduce the Impact of Romaine Lettuce Contamination.

Leafy greens, particularly romaine lettuce, are often associated with outbreaks due to their susceptibility to contamination from various environmental sources. This study aimed to evaluate the presence of E. coli, Salmonella, copper, nickel, zinc, and manganese in irrigation water, lettuce leaves, and agricultural soil in the Litani River Basin (LRB), Lebanon. Samples were collected from five demonstration plots employing different agricultural practices. Heavy metal concentrations were determined using atomic absorption spectrometry, while E. coli and Salmonella testing were conducted through conventional culturing techniques. The impact of E. coli contamination on seed germination and the interaction effects between E. coli and heavy metals were also examined. The study also compared the effectiveness of various irrigation systems in reducing bacterial contamination. The results demonstrated that contamination levels varied significantly across the plots and irrigation types. This variation underscores the necessity of site-specific mitigation strategies to enhance food safety. Our findings highlight the importance of selecting appropriate irrigation methods and implementing tailored agricultural practices to minimize the risk of contamination. This research provides valuable insights for optimizing agricultural practices in the LRB to ensure food safety and environmental sustainability.

High-yield extracellular vesicle production from HEK293T cells encapsulated in 3D auxetic scaffolds with cyclic mechanical stimulation for effective drug carrier systems

Extracellular vesicles (EVs) show promise in drug loading and delivery for medical applications. However, the lack of scalable manufacturing processes hinders the generation of clinically suitable quantities, thereby impeding the translation of EV-based therapies. Current EV production relies heavily on non-physiological two-dimensional (2D) cell culture or bioreactors, requiring significant resources. Additionally, EV-derived ribonucleic acid cargo in three-dimensional (3D) and 2D culture environments remains largely unknown. In this study, we optimized the biofabrication of 3D auxetic scaffolds encapsulated with human embryonic kidney 293 T (HEK293 T) cells, focusing on enhancing the mechanical properties of the scaffolds to significantly boost EV production through tensile stimulation in bioreactors. The proposed platform increased EV yields approximately 115-fold compared to conventional 2D culture, possessing properties that inhibit tumor progression. Further mechanistic examinations revealed that this effect was mediated by the mechanosensitivity of YAP/TAZ. EVs derived from tensile-stimulated HEK293 T cells on 3D auxetic scaffolds demonstrated superior capability for loading doxorubicin compared to their 2D counterparts for cancer therapy. Our results underscore the potential of this strategy for scaling up EV production and optimizing functional performance for clinical translation.

Human Umbilical Cord Mesenchymal Stem Cells Derived Exosomes Improved The Aged Mouse IVM Oocytes Quality.

During assisted reproductive technology (ART) treatment, the aged women, especially those over 35years old, have fewer mature oocytes and poorer quality of the oocytes comparing with the young women. In vitro maturation (IVM) technology facilitates the usage of immature oocytes, which is clinically important for the aged women. However, the maturation rate is low for the oocytes from the aged women. Human umbilical cord mesenchymal stem cells derived exosomes (HUCMSCs-exosomes), as important mediators of intercellular communication, have been widely used to restore ovarian function and improve female fertility. In this study, we isolated HUCMSCs-exosomes and collected the immature germinal vesicle oocytes from the naturally aged mouse model. And we added these HUCMSCs-exosomes to the conventional IVM culture system. The effects of HUCMSCs-exosomes on IVM oocytes were observed and analyzed from multiple aspects including maturation rate, spindle morphology, mitochondria function, and development potential. We found the quality of oocytes was improved by HUCMSCs-exosomes. Based on the results, we propose that HUCMSCs-exosomes may provide a novel and cell free strategy in the improvement of the IVM in elderly infertile women in the future.

Assessing Diagnostic Performance of Molecular Culture for Neonatal Sepsis: Protocol of the CHAMPIONS Study.

Managing neonatal sepsis is challenging due to nonspecific clinical signs, hematological markers with poor accuracy, and a lengthy turnaround time for the identification of microorganisms. Delaying the initiation of antibiotics in truly infected infants can lead to severe morbidity and mortality. Therefore, decisions regarding empiric antibiotic treatment are risk stratified, which exposes many uninfected infants to antibiotics. This causes gut microbiota perturbation, unnecessary hospital admissions, and the generation of multi-resistant organisms. High-speed diagnostic assays could expedite discontinuation or avert the initiation of antibiotics in uninfected infants. This study will evaluate the diagnostic performance of molecular culture (MC), a rapid broad-range PCR-based bacterial profiling technique, for diagnosing neonatal sepsis in infants below 90 days old. A multi-center prospective observational cohort study will include infants evaluated for early and late-onset sepsis. Routine evaluation for suspected sepsis includes microbiological cultures of blood. Additionally, blood for MC will be collected. For early-onset sepsis, umbilical cord blood may be used alternatively. Primary outcome is the agreement between MC and conventional blood culture results. Secondary outcome is the agreement of both assays with clinical sepsis using four different, commonly used definitions. Faster diagnostic pathways for sepsis may reduce antibiotic exposure time. Broad-range molecular assays may identify pathogens undetectable by conventional methods. Employment of umbilical cord blood samples for early-onset sepsis diagnosis can resolve challenges in collecting adequate blood volume and could further expedite treatment decisions.

Liver-on-a-Chip Integrated with Label-Free Optical Biosensors for Rapid and Continuous Monitoring of Drug-Induced Toxicity.

Drug toxicity assays using conventional 2D static cultures and animal studies have limitations preventing the translation of potential drugs to the clinic. The recent development of organs-on-a-chip platforms provides promising alternatives for drug toxicity/screening assays. However, most studies conducted with these platforms only utilize single endpoint results, which do not provide real-time/ near real-time information. Here, a versatile technology is presented that integrates a 3Dliver-on-a-chip with a label-free photonic crystal-total internal reflection (PC-TIR) biosensor for rapid and continuous monitoring of the status of cells. This technology can detect drug-induced liver toxicity by continuously monitoring the secretion rates and levels of albumin and glutathione S-transferase α (GST-α) of a 3D liver on-a-chip model treated with Doxorubicin. The PC-TIR biosensor is based on a one-step antibody functionalization with high specificity and a detection range of 21.7ngmL-1 to 7.83 x 103ngmL-1 for albumin and 2.20ngmL-1 to 7.94 x 102ngmL-1 for GST-α. This approach provides critical advantages for the early detection of drug toxicity and improved temporal resolution to capture transient drug effects. The proposed proof-of-concept study introduces a scalable and efficient plug-in solution for organ-on-a-chip technologies, advancing drug development and in vitro testing methods by enabling timely and accurate toxicity assessments.

Establishing Three-Dimensional Explant Culture of Human Dental Pulp Tissue.

Mesenchymal stem cells in the dental tissue indicate a disposition for differentiation into diverse dental lineages and contain enormous potential as the important means for regenerative medicine in dentistry. Among various dental tissues, the dental pulp contains stem cells, progenitor cells and odontoblasts for maintaining dentin homeostasis. The conventional culture of stem cells holds a limit as the living tissue constitutes the three-dimensional (3D) structure. Recent development in the organoid cultures have successfully recapitulated 3D structure and advanced to the assembling of different types. In the current study, the protocol for 3D explant culture of the human dental pulp tissue has been established by adopting the organoid culture. After isolating dental pulp from human tooth, the intact tissue was placed between two layers for Matrigel with addition of the culture medium. The reticular outgrowth of pre-odontoblast layer continued for a month and the random accumulation of dentin was observed near the end. Electron microscopy showed the cellular organization and in situ development of dentin, and immunohistochemistry exhibited the expression of odontoblast and stem cell markers in the outgrowth area. Three-dimensional explant culture of human dental pulp will provide a novel platform for understanding stem cell biology inside the tooth and developing the regenerative medicine.

Clinical utility of metagenomic next-generation sequencing on bronchoalveolar lavage fluid in diagnosis of lower respiratory tract infections

BackgroundIn this study, we aimed to evaluate the clinical utility of Metagenomic Next-Generation sequencing (mNGS) on bronchoalveolar lavage fluid (BALF) in diagnosis of Lower Respiratory Tract Infections (LRTIs).MethodsIn this study, we retrospectively analyzed 186 hospitalized patients who were suspected with LRTIs and performed mNGS (DNA) test of BALF simultaneously at The Fifth Affiliated Hospital of Sun Yat-Sen University from March 2023 to August 2023. Suspected LRTI was based on LRTI related clinical manifestations or imaging examination. Among them, 155 patients had undergone conventional culture and mNGS (DNA) testing simultaneously. Finally, 138 cases (89.03%,138/155) were diagnosed as LRTI and 17 cases (10.97%,17/155) were diagnosed as non-LRTI. Both detecting rate and diagnostic efficacy of mNGS and conventional culture were compared.ResultsThe positive detection rates of pathogens between mNGS and conventional culture were significant different (81.29% VS 39.35%, P < 0.05). Compared with paired conventional culture result, the sensitivity of mNGS in diagnosis of LRTIs was more superior (88.41% VS 43.48%; P < 0.05), the specificity was opposite (76.47% VS 94.12%; P > 0.05). Furthermore, 77.54% and 35.51% of LRTI cases were being etiologically diagnosed by mNGS and culture respectively. Importantly, mNGS directly led to a change of treatment regimen in 58 (37.42%) cases, including antibiotic adjustment (29.68%) and ruling out active infection (7.74%). Moreover, treatment regimen remained unchanged in 97 (62.58%) cases, considering the current antibiotic therapy already covered the detected pathogens (36.13%) or empirical treatment was effective (11.61%).ConclusionsmNGS can identify a wide range of pathogens in LRTIs, with improved sensitivity and being more superior at diagnosing LRTIs etiologically. mNGS has the potential to enhance clinical outcomes by optimizing the treatment regimens.

RNA-seq transcriptomic profiling of TGF-β2-exposed human trabecular meshwork explants: Advancing insights beyond conventional cell culture models

Primary open-angle glaucoma (POAG), a leading cause of irreversible vision loss, is closely linked to increased intraocular pressure (IOP), with the trabecular meshwork (TM) playing a critical role in its regulation. The TM, located at the iridocorneal angle, acts as a sieve, filtering the aqueous humor from the eye into the collecting ducts, thus maintaining proper IOP levels. The transforming growth factor-beta 2 (TGF-β2) signaling pathway has been implicated in the pathophysiology of primary open-angle glaucoma POAG particularly, in the dysfunction of the TM. This study utilizes human TM explants to closely mimic in vivo conditions, thereby minimizing transcriptional changes that could arise from cell culture enabling an exploration of the transcriptomic impacts of TGF-β2. Through bulk RNA sequencing and immunohistological analysis, we identified distinct gene expression patterns and morphological changes induced by TGF-β2 exposure (5 ng/ml for 48 h). Bulk RNA sequencing identified significant upregulation in genes linked to extracellular matrix (ECM) regulation and fibrotic signaling. Immunohistological analysis further elucidated the morphological alterations, including cytoskeletal rearrangements and ECM deposition, providing a visual confirmation of the transcriptomic data. Notably, the enrichment analysis unveils TGF-β2’s influence on both bone morphogenic protein (BMP) and Wnt signaling pathways, suggesting a complex interplay of molecular mechanisms contributing to TM dysfunction in glaucoma. This characterization of the transcriptomic modifications on an explant model of TM obtained under the effect of this profibrotic cytokine involved in glaucoma is crucial in order to develop and test new molecules that can block their signaling pathways.

Antibiogram and Salmonella typhi detection in suspected patients using advanced PCR technique, serology, and conventional blood culture

Background &amp; Objectives: Salmonella enterica serovar Typhi is a diverse gram-negative bacillus that causes life-threatening invasive bloodstream infection and remains a public health burden in developing countries. The antimicrobial resistance in Salmonella typhi has led to widespread multidrug resistance strains making complication in the treatment and demanding alternative therapies. Various diagnostics test such as blood culture, serological assay, and molecular assay are available. PCR is a newer gold standard method, but blood culture is still considered a definitive diagnosis of typhoid fever. Materials and Methods: A cross-sectional study was conducted. Venous blood was collected, and a widal test was performed to detect antibodies against O and H antigens in serum. Blood culture, isolation, and identification were performed using the standard techniques. The antibiogram of isolates was performed by the Kirby Bauer method. Amplification of the hilA gene through PCR was used for the molecular diagnosis. Results: Out of 141 suspected typhoid patients, 78 were males, and 63 were females. PCR detected maximum positivity rate (70%) followed by Widal test (43.26%) and blood culture (16.25%). PCR positive cases were highly prevalent in the male in less than 15 years of age group (14.8%), followed by 15-30 (12.7%) and 30-45 (9.1%). The isolated organism was found to be 100% sensitive to most antibiotics studied, except ofloxacin, which showed 71.4% sensitivity. In contrast, all the isolates were 100% resistant to nalidixic acid. None of the isolates showed multi-drug resistance. Conclusion: PCR is a sensitive, rapid, and better alternative than conventional methods that can be used in clinical diagnosis for early detection of the Salmonella typhi.

Metagenomic next-generation sequencing promotes pathogen detection over culture in joint infections with previous antibiotic exposure.

To investigate the diagnostic value of metagenomic next-generation sequencing (mNGS) in detecting pathogens from joint infection (JI) synovial fluid (SF) samples with previous antibiotic exposure. From January 2019 to January 2022, 59 cases with suspected JI were enrolled. All cases had antibiotic exposure within 2 weeks before sample collection. mNGS and conventional culture were performed on SF samples. JI was diagnosed based on history and clinical symptoms in conjunction with MSIS criteria. The diagnostic values, including sensitivity, specificity, positive/negative predictive values (PPV/NPV), and accuracy, were in comparison with mNGS and culture. There were 47 of the 59 cases diagnosed with JI, while the remaining 12 were diagnosed with non-infectious diseases. The sensitivity of mNGS was 68.1%, which was significantly higher than that of culture (25.5%, p<0.01). The accuracy of mNGS was significantly higher at 71.2% compared to the culture at 39.0% (p <0.01). Eleven pathogenic strains were detected by mNGS but not by microbiological culture, which included Staphylococcus lugdunensis, Staphylococcus cohnii, Finegoldia magna, Enterococcus faecalis, Staphylococcus saprophytics, Escherichia coli, Salmonella enterica, Pseudomonas aeruginosa, Acinetobacter pittii, Brucella ovis, andCoxiella burnetii. Antibiotic therapy was adjusted based on the mNGS results in 32 (68.1%) patients, including 12 (25.5%) and 20 (42.6%) patients, in whom treatment was upgraded and changed, respectively. All JI patients underwent surgery and received subsequent antibiotic therapy. They were followed up for an average of 23 months (20-27 months), and the success rate of treatment was 89.4%. Out of the 33 patients who had positive results for pathogens, reoperation was performed in 1 case (3.03%), while out of the 14 cases with negative results for both mNGS and cultures, reoperation was performed in 4 cases (28.6%). mNGS has advantages over conventional culture in detecting pathogens in SF samples from JI patients previously treated with antibiotics, potentially improving clinical outcomes.

Standardizing designed and emergent quantitative features in microphysiological systems.

Microphysiological systems (MPSs) are cellular models that replicate aspects of organ and tissue functions in vitro. In contrast with conventional cell cultures, MPSs often provide physiological mechanical cues to cells, include fluid flow and can be interlinked (hence, they are often referred to as microfluidic tissue chips or organs-on-chips). Here, by means of examples of MPSs of the vascular system, intestine, brain and heart, we advocate for the development of standards that allow for comparisons of quantitative physiological features in MPSs and humans. Such standards should ensure that the in vivo relevance and predictive value of MPSs can be properly assessed as fit-for-purpose in specific applications, such as the assessment of drug toxicity, the identification of therapeutics or the understanding of human physiology or disease. Specifically, we distinguish designed features, which can be controlled via the design of the MPS, from emergent features, which describe cellular function, and propose methods for improving MPSs with readouts and sensors for the quantitative monitoring of complex physiology towards enabling wider end-user adoption and regulatory acceptance.

Assessment of the presence of multidrug-resistant Escherichiacoli, Salmonella and Staphylococcus in chicken meat, eggs and faeces in Mymensingh division of Bangladesh

The emergence of bacteria that is resistant to several drugs of clinical importance poses a threat to successful treatment, a phenomenon known as multidrug resistance that affects diverse classes of antibiotics. The purpose of this study was to evaluate the prevalence of multidrug-resistant Escherichiacoli, Salmonella spp. and Staphylococcusaureus in chicken egg, meat and faeces from four districts of Bangladesh. A total of 120 chicken samples were collected from different poultry farms. Conventional culture and molecular detection methods were used for identification of bacterial isolates from the collected samples followed by antibiotic susceptibility test through the disc diffusion method, finally antibiotic resistant genes were detected by PCR. E. coli, Salmonella spp. and Staphylococcusaureus were detected in meat, egg and faecal samples. Antimicrobial susceptibility results revealed isolates from faeces were 100 % resistant to amoxicillin, while all S.aureus and Salmonella sp. from faeces were resistant to doxycycline, tetracycline and erythromycin. Salmonella spp. isolates from eggs indicated 100 % resistance to erythromycin, amoxycillin, while E.coli were 100 % resistant to erythromycin. E.coli and S.aureus from meat were 100 % resistant to amoxicillin and erythromycin. However, Salmonella spp. from eggs were 100 % susceptible to doxycycline, gentamicin, levofloxacin and tetracycline. The mecA and aac(3)-IV genes were only found in S.aureus and E.coli, respectively. The Sul1,tetB, and aadA1 were highest in Salmonella spp. and S.aureus, while the sul1,tetA and blaSHV were higher in E.coli. Isolates from all samples were multidrug resistant. These findings indicate a high risk of transmission of resistance genes from microbial contamination to food of animal origin. The study emphasizes the need for effective biosecurity measures, responsible antibiotic use, and strict regulations in poultry production to prevent the spread of antibiotic resistance.

Integrating omics techniques and culture-independent systems may improve the detection of persistent candidemia: data from an observational study

IntroductionBlood cultures have low sensitivity for candidemia. Sensitivity can be improved by the culture-independent system T2 Magnetic Resonance (T2). SeptiCyte RAPID is a host response assay quantifying the risk of infection-related inflammation through a scoring system (SeptiScore). We investigate the performance of SeptiScore in detecting persistent candidemia as defined by conventional cultures and T2.MethodsThis is a prospective multicentre observational study on patients with candidemia. Blood cultures and blood samples for assessment by T2 and SeptiCyte were collected for 4 consecutive days after the index culture. The performance of SeptiScore was explored to predict persistent candidemia as defined by (1) positive follow-up blood culture (2) either positive follow-up blood culture or T2 sample.Results10 patients were enrolled including 34 blood collections assessed with the 3 methods. Overall, 4/34 (12%) follow-up blood cultures and 6/34 (18%) T2 samples were positive. A mixed model showed significantly higher SeptiScores associated with persistent candidemia when this was defined as either a positive follow-up blood culture or T2 sample (0.82, 95%CI 0.06 to 1.58) but not when this was defined as a positive follow-up blood culture only (-0.57, 95%CI -1.28 to 0.14). ROC curve for detection of persistent candidemia by SeptiScore at day 1 follow-up showed an AUC of 0.85 (95%CI 0.52-1.00) when candidemia was defined by positive follow-up blood culture, and an AUC of 1.00 (95%CI 1.00–1.00) when candidemia was defined according to both methods.ConclusionIntegrating transcriptome profiling with culture-independent systems and conventional cultures may increase our ability to diagnose persistent candidemia.

Performance comparison of three commercial multiplex molecular panels for respiratory viruses at a South African academic hospital.

Respiratory infections are a major contributor to hospital admissions. Identification of respiratory pathogens by means of conventional culture and serology methods remains challenging. Multiplex molecular assays are an appealing alternative that endeavours to be rapid, more accurate and less arduous. The study aimed to compare the clinical performance of three commercial multiplex molecular assays for respiratory viruses. Forty-eight respiratory specimens obtained from patients at Tygerberg Hospital in the Western Cape province of South Africa were studied. These specimens were collected between May 2020 and August 2020. The results of the Seegene Anyplex™ II RV16, FilmArray® Respiratory 2.1 plus Panel (FARP), and QIAstat-Dx® Respiratory SARS-CoV-2 Panel (QRP) were analysed based on the overlapping targets. A composite reference standard was applied to provide a standard reference for comparison. The overall sensitivity of the Seegene Anyplex™ II RV16 was 96.6% (57/59), the FARP 98.2% (56/57) and the QRP 80.7% (46/57). The overall specificities were 99.8% (660/661), 99.0% (704/711) and 99.7% (709/711), respectively. The QRP failed to detect coronaviruses and parainfluenza viruses in 41.7% (5/12) and 28.6% (4/14) of positive specimens, respectively, while the FARP produced the lowest target specificity of 88.4% (38/43) for rhinovirus/enterovirus. The overall specificity of all three platforms was comparable; however, the sensitivity of the QRP was inferior to that of the ARV and FARP. This study adds to the body of performance characteristics described for respiratory multiplex panels, especially in the African context where molecular diagnostics for infectious diseases are gaining momentum.