Hasil untuk "Microbiology"

I. R. Kaplan, S. Rittenberg

W. W. Davis, T. R. Stout

Several factors are investigated that normally cause variation in zone diameters in conventional disc plate diffusion assay procedures. Of these factors the most serious is the unequal exposure of the individual plates at top or bottom of stacks to temperatures above and below room temperature. This unequal temperature exposure is avoided by novel handling and incubation procedures. A major variable, but one which can be controlled, is the varying time interval between pouring seeded agar and the time of applying the pads with antibiotic to the plates. This influence of time of setting and the effects of several other sequential operations are combined into a composite variable. This variable is then accounted for and normalized by interposing “external” reference plates set with a reference solution in the sequence of approximately 100 plates. No “internal” reference zones are employed. Such factors as volume of agar poured, wedge shape of agar in a dish, volumetric errors in dilutions, and timing considerations are studied and discussed. The results of this study form the basis for a test protocol which is presented in a following paper.

Maria Martínez-Ruiz, Miriam Hernández-Porto, Cintia Hernández-Sánchez et al.

Recycled wastewater is vital for the circular economy, especially on water-scarce islands. This study explored the presence of Carbapenem-Resistant Enterobacterales and other emerging pathogens in irrigation water on four Canarian Islands, applying a One Health perspective. Using membrane filtration and MALDI-TOF mass spectrometry, 69 bacterial isolates were identified. The findings revealed that 78% were Gram-negative bacilli like <i>Pseudomonas aeruginosa</i>, <i>Acinetobacter</i> spp., Enterobacteriaceae, etc., while 22% were Gram-positive bacteria, including <i>Enterococcus</i> spp. The main mechanisms of carbapenem resistance in <i>Pseudomonas</i> spp. and <i>Acinetobacter</i> spp. were oxacillinases, followed by metallo-β-lactamases (MBL). In Enterobacteriaceae, characterization of carbapenemase types was less frequent, with oxacillinase 48 (OXA-48) being the most prevalent. The detection of multidrug-resistant organisms in recycled wastewater highlights an urgent need for routine microbiological monitoring in water management to protect both public health and agricultural sustainability.

Qinghua Gao, Jiahao Xu, Yue Ma et al.

IntroductionThis study aims to evaluate the utility of molecular diagnostic techniques in identifying pathogens in bronchiectasis and to investigate the differences in clinical characteristics and pathogen distribution among patients with different microbial infections.MethodsThis retrospective study collected and analyzed clinical data and lower respiratory tract pathogen detection results from 410 patients with bronchiectasis admitted to the Anning First People’s Hospital Affiliated to Kunming University of Science and Technology, between August 2020 and August 2024. By comparing molecular diagnostic methods with conventional culture, we assessed differences in pathogen detection rates and spectrum, evaluated the diagnostic performance of molecular techniques relative to traditional methodologies, and analyzed the clinical characteristics of bronchiectasis patients with different microbiological etiologies.ResultsCompared with conventional microbiological testing (CMT), molecular diagnostics demonstrated significantly higher sensitivity, positive predictive value, and negative predictive value. The most frequently detected bacteria were Haemophilus influenzae (26.83%), Pseudomonas aeruginosa (14.88%), Streptococcus pneumoniae (13.17%), Klebsiella pneumoniae (9.02%), and Staphylococcus aureus (4.39%). Patients infected with P. aeruginosa had significantly lower body mass index (BMI) compared to those infected with H. influenzae, as well as more severe lung function impairment. Inflammatory markers, including white blood cell (WBC) count and C-reactive protein (CRP), were significantly higher in the P. aeruginosa group than in the H. influenzae group. In terms of pathogen detection, the conventional culture positivity rate was significantly higher in the P. aeruginosa group compared to the H. influenzae group, whereas the false-negative rate of culture was markedly higher in the H. influenzae group. Molecular diagnostics showed high true-positive rates in both groups, though slightly lower in the P. aeruginosa group than in the H. influenzae group. Furthermore, patients infected with P. aeruginosa had significantly higher rates of respiratory failure, cystic bronchiectasis, and oxygen therapy requirement compared to those infected with H. influenzae.ConclusionThe application of molecular diagnostic technology has significantly improved the detection rate of pathogens in patients with bronchiectasis, especially for fastidious bacteria and rare pathogens. This method can provide a more comprehensive understanding of the distribution of microorganisms and disease characteristics, shorten the diagnosis cycle, accurately guide anti-infection treatment decisions and assist in prognosis assessment.

Saddaf Razzaq, Aayesha Riaz, Naila Siddique et al.

Abstract Newcastle disease virus (NDV) remains a major threat to the poultry industry worldwide. Recombinant DNA vaccine against NDV offers a promising solution to current Newcastle disease (ND) challenges. Present study describes the development of a DNA vaccine (rDNA-NDV-F) using the fusion (F) gene from NDV genotype VII strain isolated from Rawalpindi, Pakistan. While conventional NDV vaccines reduce mortality in commercial poultry, they do not provide complete protection or prevent viral shedding. To address this issue, genotype-matched vaccines have been proposed. Here, we developed and evaluated the efficacy of the rDNA-NDV-F vaccine against genotype VII challenge. NDV was isolated from a field strain and propagated in embryonated chicken eggs (ECE). Virus activity was confirmed using Hemagglutination assay (HA), HA inhibition (HAI), and Mean Death Time (MDT) assay. Polymerase Chain Reaction (PCR) and sequencing confirmed the genotype VII.2 strain. The DNA vaccine was constructed using the fusion (F) protein gene cloned into the expression plasmid pcDNA3.1. Gene insertion was verified by PCR and restriction digestion, while protein expression was confirmed via immunofluorescence assay. To assess vaccine efficacy, 120 chickens (14 days old) were divided into four groups: G1 (rDNA-NDV-F), G2 (empty vector), G3 (PBS control), and G4 (non-vaccinated, non-challenged control). Serological responses were measured using ELISA on days 0, 7, 14, 21, and 28. Birds were challenged with NDV genotype VII (105 EID50). Virus shedding from tracheal and cloacal swabs was analyzed on days 3, 7, and 10 post-challenge. Clinical signs and mortality rates were also recorded. The rDNA-NDV-F vaccine induced strong immune responses, with peak ELISA (6180) titers at 28 days. Virus shedding was detected in three birds on day 3 but was absent by day 10. No virus shedding was observed in cloacal swabs, indicating restriction in the digestive system. Vaccinated birds showed mild clinical signs in only two cases, with no neurological symptoms or mortality. In contrast, negative and vector control groups exhibited severe clinical signs and 90–100% mortality. Statistical analysis confirmed significant differences (P < 0.05). This study highlights the effectiveness of genotype-matched recombinant NDV vaccines in providing effective protection for poultry.

Larissa Leão F. de Sousa, Mariana Dias Guilardi, Junior Olimpio Martins et al.

Abstract Background Rabies, a lethal viral zoonotic disease, remains a significant global public health concern. In northeastern Brazil, in particular, its epidemiology is complex and dynamic, characterized by the presence of several reservoirs associated with human rabies infection. Methods This study, conducted from June 2022 to July 2023, was part of a passive epidemiological surveillance initiative under Brazil’s National Rabies Surveillance Program. It investigated the presence of Rhabdovirus (RhabV) in 356 postmortem chiropteran brain samples using three diagnostic techniques for rabies and conducted an evolutionary study on both pan-RhabV- and pan-LYSSAV-positive PCR samples. The samples were collected from 20 bat species and different locations in the State of Ceará, an endemic region for the rabies virus (RABV). Rabies-positive samples were further explored through Bayesian, genetic distance mapping and recombination analyses. Results From a total of 356 samples collected, 43 (12.07%) were positive for direct immunofluorescence (DIF) and 40 (11.23%) for mouse intracerebral inoculation (MIT) tests. Among the positive results, 40 samples were confirmed by both DIF and MIT, while 13 (3.65%) had inconclusive results for one or both techniques. Molecular assays identified 38 rabies-positive samples (10.67%). Members of the Molossidae and Phyllostomidae families had the highest prevalence, highlighting the role of insectivorous and frugivorous bats in the cycle and dynamics of rabies transmission. Phylogenetic reconstructions revealed three distinct and well-supported clusters and clades, indicating the cocirculation of different RABV lineages in the region and shedding light on both intra- and interhost diversity. We also demonstrated genetic distance among the RABV clusters and inferred that their common ancestor originated in Europe, later diversifying across continents. No recombination breakpoints were identified. Conclusions This study highlights the dynamic nature of RABV evolution within individual bat hosts, contributing to the understanding of the genetic diversity of RABV variants found in several bat species in northeastern Brazil. This study provides crucial insights into viral transmission dynamics within and between different host species and is essential for designing effective rabies control and prevention strategies tailored to endemic regions.

R. Novick, R. Clowes, S. Cohen et al.

Hiroki Futatsusako, Rina Hashimoto, Masaki Yamamoto et al.

Summary: A primary reason for the ongoing spread of coronavirus disease 2019 (COVID-19) is the continuous acquisition of mutations by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). However, the mechanism of acquiring mutations is not fully understood. In this study, we isolated SARS-CoV-2 from an immunocompromized patient persistently infected with Omicron strain BF.5 for approximately 4 months to analyze its genome and evaluate drug resistance. Although the patient was administered the antiviral drug remdesivir (RDV), there were no acquired mutations in RDV binding site, and all isolates exhibited susceptibility to RDV. Notably, upon analyzing the S protein sequence of the day 119 isolate, we identified mutations acquired by mutant strains emerging from the BF.5 variant, suggesting that viral genome analysis in persistent COVID-19 patients may be useful in predicting viral evolution. These results suggest mutations in SARS-CoV-2 are acquired during long-term viral replication rather than in response to antiviral drugs.

Cornelia-Ioana Ilie, Angela Spoiala, Cristina Chircov et al.

The gut microbiota dysbiosis that often occurs in cancer therapy requires more efficient treatment options to be developed. In this concern, the present research approach is to develop drug delivery systems based on magnetite nanoparticles (MNPs) as nanocarriers for bioactive compounds. First, MNPs were synthesized through the spraying-assisted coprecipitation method, followed by loading bee pollen or bee bread extracts and an antitumoral drug (5-fluorouracil/5-FU). The loaded-MNPs were morphologically and structurally characterized through transmission electron microscopy (TEM), selected area electron diffraction (SAED), scanning electron microscopy (SEM), X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FT-IR), Dynamic Light Scattering (DLS), and thermogravimetric analysis. UV-Vis spectroscopy was applied to establish the release profiles and antioxidant activity. Furthermore, the antibacterial and antitumoral activity of loaded-MNPs was assessed. The results demonstrate that MNPs with antioxidant, antibacterial, antiproliferative, and prebiotic properties are obtained. Moreover, the data highlight the improvement of 5-FU antibacterial activity by loading on the MNPs’ surface and the synergistic effects between the anticancer drug and phenolic compounds (PCs). In addition, the prolonged release behavior of PCs for many hours (70–75 h) after the release of 5-FU from the developed nanocarriers is an advantage, at least from the point of view of the antioxidant activity of PCs. Considering the enhancement of <i>L. rhamnosus</i> MF9 growth and antitumoral activity, this study developed promising drug delivery alternatives for colorectal cancer therapy.

Tian Gao, Hua Qu, Zai-Wei Ge

Species of Micropsalliota generally grow in the tropics and are characterised by small, slender basidiomes, brown basidiospores, and cheilocystidia that vary in shape with capitate or subcapitate apex, and pigmented pileipellis. Based on morphological characters and molecular evidence, here we describe seven new species from southern China, viz. Micropsalliota ferruginea, M. fimbriata, M. gigaspora, M. longicystis, M. nana, M. squarrosa, and M. umbonata. Micropsalliota appendiculata, a species recently described from Vietnam, was first recorded in China. The Maximum likelihood and Bayesian analyses based on multi-locus sequence datasets (the nuc rDNA internal transcribed spacer region ITS1-5.8S-ITS2, nrITS; the D1–D2 domains of nuc 28S rDNA, LSU; partial sequences of the most variable region of the second-largest subunit of RNA polymerase II, rpb2, and a portion of the translation-elongation factor 1-α, tef1) shows that the genus is separated into 11 major clades and subclades. To aid in diagnosis, a key to 32 species of Micropsalliota in China is provided.

Cheryl S. Rosenfeld

The placenta operates during gestation as the primary communication organ between the mother and fetus. It is essential for gas, nutrient exchange, and fetal waste transfer. The placenta also produces a wide range of hormones and other factors that influence maternal physiology, including survival and activity of the corpus luteum of the ovary, but the means whereby the placenta shapes fetal development remain less clear, although the fetal brain is thought to be dependent upon the placenta for factors that play roles in its early differentiation and growth, giving rise to the term “placenta–brain axis”. Placental hormones transit via the maternal and fetal vasculature, but smaller placental molecules require protection from fetal and maternal metabolism. Such biomolecules include small RNA, mRNA, peptides, lipids, and catecholamines that include serotonin and dopamine. These compounds presumably shuttle to maternal and fetal systems via protective extracellular vesicles (EVs). Placental EVs (pEVs) and their components, in particular miRNA (miRs), are known to play important roles in regulating maternal systems, such as immune, cardiovascular, and reproductive functions. A scant amount is known about how pEVs affect fetal cells and tissues. The composition of pEVs can be influenced by gestational diseases. This review will provide critical insight into the roles of pEVs as the intermediary link between maternal and fetal systems, the impact of maternal pathologies on pEV cargo contents, and how an understanding of biomolecular changes within pEVs in health and disease might be utilized to design early diagnostic and mitigation strategies to prevent gestational diseases and later offspring disorders.

Aurélien Dinh, Alexandre Bleibtreu, Clara Duran et al.

Background: Meropenem–vaborbactam (MEM-VAB) is a novel carbapenem-beta-lactamase-inhibitor combination that demonstrates activity against carbapenem-resistant (CR) Gram-negative bacteria, and more specifically KPC-producers, since vaborbactam is an effective inhibitor of KPC enzymes in vitro. This study aimed to describe the initial uses and efficacy of MEM-VAB for compassionate treatment during the first 21 months following its early access in France. Method: A national multicenter retrospective study was conducted, including all patients who received at least one dose of MEM-VAB between 20 July 2020, and 5 April 2022. Clinical characteristics and outcomes were collected using a standardized questionnaire. The minimum inhibitory concentration (MIC) of antimicrobials, and complete genome sequencing of bacteria were performed when bacterial isolates were available. Results: Ultimately, 21 patients from 15 French hospitals were included in the study. The main indication for MEM-VAB treatment was respiratory tract infections (<i>n</i> = 9). The targeted bacteria included <i>Pseudomonas aeruginosa</i> (<i>n</i> = 12), <i>Klebsiella pneumoniae</i> (<i>n</i> = 3), <i>Enterobacter spp</i> (<i>n</i> = 3), <i>Citrobacter freundii</i> (<i>n</i> = 1), <i>Escherichia coli</i> (<i>n</i> = 1), and <i>Burkholderia multivorans</i> (<i>n</i> = 1). Overall, no significant advantage of vaborbactam over meropenem alone was observed across all strains of <i>P. aeruginosa</i> in terms of in vitro susceptibility. However, MEM-VAB demonstrated a notable impact, compared to carbapenem alone, on the MIC for the two KPC-3-producing <i>K. pneumoniae</i> and <i>B. multivorans</i>. Conclusions: MEM-VAB seems effective as a salvage treatment in compassionate use, but vaborbactam was shown to lack benefits compared to meropenem in treating <i>P. aeruginosa</i>-related infections. Therefore, it is crucial to compare meropenem to MEM-VAB MICs, particularly for <i>P. aeruginosa</i>, before prescribing MEM-VAB.

A. Gristina, J. Costerton

Devaki Pilapitiya, Adam K. Wheatley, Hyon-Xhi Tan

Summary: Currently approved COVID-19 vaccines administered parenterally induce robust systemic humoral and cellular responses. While highly effective against severe disease, there is reduced effectiveness of these vaccines in preventing breakthrough infection and/or onward transmission, likely due to poor immunity elicited at the respiratory mucosa. As such, there has been considerable interest in developing novel mucosal vaccines that engenders more localised immune responses to provide better protection and recall responses at the site of virus entry, in contrast to traditional vaccine approaches that focus on systemic immunity. In this review, we explore the adaptive components of mucosal immunity, evaluate epidemiological studies to dissect if mucosal immunity conferred by parenteral vaccination or respiratory infection drives differential efficacy against virus acquisition or transmission, discuss mucosal vaccines undergoing clinical trials and assess key challenges and prospects for mucosal vaccine development.

C. Triffault-Fillit, T. Ferry, F. Laurent et al.

OBJECTIVES The high microbiologic diversity encountered in prosthetic joint infection (PJI) makes the choice of empirical antimicrobial therapies challenging, especially in cases of implant retention or one-stage exchange. Despite the risk of dysbiosis and toxicity, the combination of vancomycin with a broad-spectrum β-lactam is currently recommended in all cases, even if Gram-negative bacilli (GNB) might be less represented in late PJI. In this context, this study aimed to describe the microbiologic epidemiology of PJI according to the chronology of infection. METHODS This prospective cohort study (2011-2016) evaluated the microbiologic aetiology of 567 PJI according to time of occurrence from prosthesis implantation-early (12 months)-as well as mechanism of acquisition. RESULTS Initial microbiologic documentation (n = 511; 90.1%) disclosed 164 (28.9%) Staphylococcus aureus (including 26 (16.1%) methicillin-resistant S. aureus), 162 (28.6%) coagulase-negative staphylococci (including 81 (59.1%) methicillin-resistant coagulase-negative staphylococci), 80 (14.1%) Enterobacteriaceae, 74 (13.1%) streptococci and 60 (10.6%) Cutibacterium acnes. Considering nonhaematogenous late PJI (n = 182), Enterobacteriaceae (n = 7; 3.8%) were less represented than in the first year after implantation (n = 56; 17.2%; p <0.001), without difference regarding nonfermenting GNB (4.6% and 2.7%, respectively). The prevalence of anaerobes (n = 40; 21.9%; including 32 (80.0%) C. acnes) was higher in late PJI (p <0.001). Consequently, a broad-spectrum β-lactam might be useful in 12 patients (6.6%) with late PJI only compared to 66 patients (20.3%) with early/delayed PJI (p <0.001). CONCLUSIONS Considering the minority amount of GNB in late postoperative PJI, the empirical use of a broad-spectrum β-lactam should be reconsidered, especially when a two-stage exchange is planned.

Oliver Razum, Raffaella Ravinetto, Veronika J. Wirtz et al.

Francesca Malagrinò, Valeria Pennacchietti, Daniele Santorelli et al.

The vast majority of our current knowledge about the biochemical and biophysical properties of proteins derives from in vitro studies conducted on isolated globular domains. However, a very large fraction of the proteins expressed in the eukaryotic cell are structurally more complex. In particular, the discovery that up to 40% of the eukaryotic proteins are intrinsically disordered, or possess intrinsically disordered regions, and are highly dynamic entities lacking a well-defined three-dimensional structure, revolutionized the structure–function paradigm and our understanding of proteins. Moreover, proteins are mostly characterized by the presence of multiple domains, influencing each other by intramolecular interactions. Furthermore, proteins exert their function in a crowded intracellular milieu, transiently interacting with a myriad of other macromolecules. In this review we summarize the literature tackling these themes from both the theoretical and experimental perspectives, highlighting the effects on protein folding and function that are played by (i) flanking disordered tails; (ii) contiguous protein domains; (iii) interactions with the cellular environment, defined as quinary structures. We show that, in many cases, both the folding and function of protein domains is remarkably perturbed by the presence of these interactions, pinpointing the importance to increase the level of complexity of the experimental work and to extend the efforts to characterize protein domains in more complex contexts.

Zi-da Huang, Zi-Jie Zhang, Bin Yang et al.

Objectives: To evaluate metagenomic next-generation sequencing (mNGS) as a diagnostic tool in detecting pathogens from osteoarticular infection (OAI) samples. Methods: 130 samples of joint fluid, sonicate fluid, and tissue were prospectively collected from 92 patients with OAI. The performance of mNGS and microbiology culture was compared pairwise. Results: The overall sensitivity of mNGS was 88.5% (115/130), significantly higher than that of microbiological culture, which had a sensitivity of 69.2% (90/130, p 0.05), and Mycoplasma (OR = ∞, 95% confidence interval, 5.12–∞, p < 0.001). Additionally, sensitivity by mNGS was higher in antibiotic-treated samples compared to microbiological culture (89.7 vs. 61.5%, p < 0.01). Conclusions: mNGS is a robust diagnostic tool for pathogenic detection in samples from OAI patients, compared to routine cultures. The mNGS technique is particularly valuable to diagnose pathogens that are difficult to be cultured, or to test samples from patients previously treated with antibiotics.

A. Bull, A. Ward, M. Goodfellow

Jiahui Huang, Feng Zhang, Jumei Zhang et al.

This study investigated the molecular characteristics of rifampin-resistant (RIF-R) <i>Staphylococcus aureus</i> isolates recovered from 4300 retail food samples covering most provincial capitals in China, from 2011 to 2016. Of the 1463 <i>S. aureus</i> enrolled, 149 isolates (142 MSSA and 7 MRSA) were identified as rifampicin-resistant, including 20 high-level (MICs ≥ 8 μg/mL) and 129 low-level (MICs between 2 and 4 μg/mL) rifampicin-resistant strains. Most of the RIF-R <i>S. aureus</i> isolates were resistant to more than three antibiotics. The mutations in the rifampicin resistance-determining region of the <i>rpoB</i> gene were studied in all RIF-R strains. All of the strains presented the mutational change 481 His/Asn and five isolates presented an additional mutation, including 477 Asp/Tyr, 527 Ile/Met, and 466 Leu/Ser, respectively. Thirteen STs and twenty-one <i>spa</i> types were represented, in which five MRSA showed non-type SCC<i>mec</i> and the remaining MRSA belonged to SCC<i>mec</i> type IV—where, ST1-t127 was the predominant type from all of the isolates, while ST398-t034 was the predominant type for the MRSA isolates. In this study, we found that the food-related RIF-R <i>S. aureus</i> may have a unique genetic background selection. However, the scenario regarding the presence of RIF-R <i>S. aureus,</i> especially MRSA, in retail food in China is not favorable and warrants public attention.