During the early phase of the coronavirus disease 2019 (COVID-19) pandemic, design, development, validation, verification and implementation of diagnostic tests were actively addressed by a large ...number of diagnostic test manufacturers. Hundreds of molecular tests and immunoassays were rapidly developed, albeit many still await clinical validation and formal approval. In this Review, we summarize the crucial role of diagnostic tests during the first global wave of COVID-19. We explore the technical and implementation problems encountered during this early phase in the pandemic, and try to define future directions for the progressive and better use of (syndromic) diagnostics during a possible resurgence of COVID-19 in future global waves or regional outbreaks. Continuous global improvement in diagnostic test preparedness is essential for more rapid detection of patients, possibly at the point of care, and for optimized prevention and treatment, in both industrialized countries and low-resource settings.
ABSTRACT
Antimicrobial resistance (AMR) has become a global medical priority that needs urgent resolution. Pseudomonas aeruginosa is a versatile, adaptable bacterial species with widespread ...environmental occurrence, strong medical relevance, a diverse set of virulence genes and a multitude of intrinsic and possibly acquired antibiotic resistance traits. Pseudomonas aeruginosa causes a wide variety of infections and has an epidemic-clonal population structure. Several of its dominant global clones have collected a wide variety of resistance genes rendering them multi-drug resistant (MDR) and particularly threatening groups of vulnerable individuals including surgical patients, immunocompromised patients, Caucasians suffering from cystic fibrosis (CF) and more. AMR and MDR especially are particularly problematic in P. aeruginosa significantly complicating successful antibiotic treatment. In addition, antimicrobial susceptibility testing (AST) of P. aeruginosa can be cumbersome due to its slow growth or the massive production of exopolysaccharides and other extracellular compounds. For that reason, phenotypic AST is progressively challenged by genotypic methods using whole genome sequences (WGS) and large-scale phenotype databases as a framework of reference. We here summarize the state of affairs and the quality level of WGS-based AST for P. aeruginosa mostly from clinical origin.
The authors review part of the clinical biology of Pseudomonas aeruginosa which is an often drug resistant human pathogen for which antimicrobial susceptibility can be increasingly assessed by using next generation genome sequencing.
Antimicrobial resistance (AMR) is a major threat to human health worldwide, and the rapid detection and quantification of resistance, combined with antimicrobial stewardship, are key interventions to ...combat the spread and emergence of AMR. Antimicrobial susceptibility testing (AST) systems are the collective set of diagnostic processes that facilitate the phenotypic and genotypic assessment of AMR and antibiotic susceptibility. Over the past 30 years, only a few high-throughput AST methods have been developed and widely implemented. By contrast, several studies have established proof of principle for various innovative AST methods, including both molecular-based and genome-based methods, which await clinical trials and regulatory review. In this Review, we discuss the current state of AST systems in the broadest technical, translational and implementation-related scope.
Vancomycin-resistant Staphylococcus aureus (VRSA), Vancomycin-intermediate S. aureus (VISA) and heterogeneous VISA (hVISA) are subject to vancomycin treatment failure. The aim of the present study ...was to determine their precise prevalence and investigate prevalence variability depending on different years and locations. Several international databases including Medline (PubMed), Embase and Web of Sciences were searched (data from 1997 to 2019) to identify studies that addressed the prevalence of VRSA, VISA and hVISA among human clinical isolates around the world. Subgroup analyses and meta-regression were conducted to indicate potential source of variation. Publication bias was assessed using Egger's test. Statistical analyses were conducted using STATA software (version 14.0). Data analysis showed that VRSA, VISA and hVISA isolates were reported in 23, 50 and 82 studies, with an overall prevalence of 1.5% among 5855 S. aureus isolates, 1.7% among 22,277 strains and 4.6% among 47,721 strains, respectively. The overall prevalence of VRSA, VISA, and hVISA before 2010 was 1.2%, 1.2%, and 4%, respectively, while their prevalence after this year has reached 2.4%, 4.3%, and 5.3%. The results of this study showed that the frequency of VRSA, VISA and hVISA after 2010 represent a 2.0, 3.6 and 1.3-fold increase over prior years. In a subgroup analysis of different strain origins, the highest frequency of VRSA (3.6%) and hVISA (5.2%) was encountered in the USA while VISA (2.1%) was more prevalent in Asia. Meta-regression analysis showed significant increasing of VISA prevalence in recent years (p value ≤ 0.05). Based on the results of case reports (which were not included in the calculations mentioned above), the numbers of VRSA, VISA and hVISA isolates were 12, 24 and 14, respectively, among different continents. Since the prevalence of VRSA, VISA and hVISA has been increasing in recent years (especially in the Asian and American continents), rigorous monitoring of vancomycin treatment, it's the therapeutic response and the definition of appropriate control guidelines depending on geographical regions is highly recommended and essential to prevent the further spread of vancomycin-resistant S. aureus.
Multidrug-resistant
infection has recently emerged as a worldwide clinical problem, and colistin is increasingly being used as a last-resort therapy. Despite its favorable bacterial killing, ...resistance and heteroresistance (HR) to colistin have been described. The purpose of the present study was to investigate the role of the PmrAB regulatory pathway in laboratory-selected mutants representative of global epidemic strains. From three unrelated
clinical strains (sequence types 2, 3, and 20), eight colistin-resistant mutants were selected. Half of the mutants showed HR to colistin according to the reference method (population analysis profiling), whereas the other half exhibited stable resistance. M12I mutation within
and M308R, S144KLAGS, and P170L mutations for
were associated with HR to colistin, while T235I, A226T, and P233S mutations within
were associated with stable resistance. The transcript levels of the
operon were upregulated in all the mutants. Compensatory mutations were explored for some mutants. A single mutant (T235I mutant) displayed a compensatory mutation through IS
mobilization within the
gene that was associated with the loss of colistin resistance. The mutant resistance phenotype associated with T235I was partially restored in a
-complementation assay turning to HR. The level of colistin resistance was correlated with the level of expression of
in the
-complemented strains. This report shows the role of different mutations in the PmrAB regulatory pathway and warns of the development of colistin HR that could be present but not easily detected through routine testing.
Many of the human infectious pathogens - especially the zoonotic or vector-borne bacteria - are fastidious organisms which are difficult to cultivate because of their strong adaption to the infected ...host culminating in their near complete physiological dependence on this environment. These bacterial species exhibit reduced multiplication rates once they are removed from their optimal ecological niche. This fact complicates the laboratory diagnosis of the disease and hinders the detection and further characterization of the underlying organisms, e.g. at the level of their resistance to antibiotics due to their slow growth. Here, we describe the current state of microbiological diagnostics for five genera of human pathogens with a fastidious laboratory lifestyle. For Anaplasma spp., Bartonella spp., Coxiella burnetii, Orientia spp., and Rickettsia spp. we will summarize the existing diagnostic protocols, the specific limitations for implementation of novel diagnostic approaches, and the need for further optimization or expansion of the diagnostic armamentarium. We will reflect upon the diagnostic opportunities provided by new technologies including mass spectrometry and next-generation nucleic acid sequencing. Finally, we will review the (im)possibilities of rapidly developing new in vitro diagnostic tools for diseases of which the causative agents are fastidiously growing and therefore hard to detect.
The integration of matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) in clinical microbiology has revolutionized species identification of bacteria, yeasts, ...and molds. However, beyond straightforward identification, the method has also been suggested to have the potential for subspecies-level or even type-level epidemiological analyses. This minireview explores MALDI-TOF MS-based typing, which has already been performed on many clinically relevant species. We discuss the limits of the method's resolution and we suggest interpretative criteria allowing valid comparison of strain-specific data. We conclude that guidelines for MALDI-TOF MS-based typing can be developed along the same lines as those used for the interpretation of data from pulsed-field gel electrophoresis (PFGE).
Abstract
Many of the human infectious pathogens—especially the zoonotic or vector-borne bacteria—are fastidious organisms that are difficult to cultivate because of their strong adaption to the ...infected host culminating in their near-complete physiological dependence on this environment. These bacterial species exhibit reduced multiplication rates once they are removed from their optimal ecological niche. This fact complicates the laboratory diagnosis of the disease and hinders the detection and further characterization of the underlying organisms, e.g. at the level of their resistance to antibiotics due to their slow growth. Here, we describe the current state of microbiological diagnostics for five genera of human pathogens with a fastidious laboratory lifestyle. For Anaplasma spp., Bartonella spp., Coxiella burnetii, Orientia spp. and Rickettsia spp., we will summarize the existing diagnostic protocols, the specific limitations for implementation of novel diagnostic approaches and the need for further optimization or expansion of the diagnostic armamentarium. We will reflect upon the diagnostic opportunities provided by new technologies including mass spectrometry and next-generation nucleic acid sequencing. Finally, we will review the (im)possibilities of rapidly developing new in vitro diagnostic tools for diseases of which the causative agents are fastidiously growing and therefore hard to detect.
Some zoonotic and vector-transmitted bacteria are strongly adapted to the infected host hindering pathogen cultivation and further identification: an overview of the current and future perspectives for diagnosis of medically relevant fastidious Gram-negative bacteria.
Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) has revolutionized the identification of microbial species in clinical microbiology laboratories. ...MALDI-TOF-MS has swiftly become the new gold-standard method owing to its key advantages of simplicity and robustness. However, as with all new methods, adoption of the MALDI-TOF MS approach is still not widespread. Optimal sample preparation has not yet been achieved for several applications, and there are continuing discussions on the need for improved database quality and the inclusion of additional microbial species. New applications such as in the field of antimicrobial susceptibility testing have been proposed but not yet translated to the level of ease and reproducibility that one should expect in routine diagnostic systems. Finally, during routine identification testing, unexpected results are regularly obtained, and the best methods for transmitting these results into clinical care are still evolving. We here discuss the success of MALDI-TOF MS in clinical microbiology and highlight fields of application that are still amenable to improvement.
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