Spectrum Of Virulence Research Articles

Commensal-pathogen dynamics structure disease outcomes during Clostridioides difficile colonization.

Gastrointestinal colonization by Clostridioides difficile is common in healthcare settings and ranges in clinical presentation from asymptomatic carriage to lethal C. difficile infection (CDI). We used a systems biology approach to investigate why patients colonized with C. difficile have a range of outcomes. Microbiota-humanization of germ-free mice with fecal samples from toxigenic C. difficile carriers revealed a spectrum of virulence among clade 1 lineages and identified commensal Blautia associated with markers of non-pathogenic colonization. Using gnotobiotic mice engrafted with defined human microbiota, we observed strain-specific CDI severity across clade 1 strains. Yet, mice engrafted with a higher diversity community were protected from severe disease across all strains without suppression of C. difficile colonization. These results indicate that when colonization resistance has been breached without overt infection, commensals can attenuate a diversity of virulent strains without inhibiting pathogen colonization, providing insight into determinants of stable C. difficile carriage.

Phylogenomic and genomic analysis reveals unique and shared genetic signatures of Mycobacterium kansasii complex species.

Species belonging to the Mycobacterium kansasii complex (MKC) are frequently isolated from humans and the environment and can cause serious diseases. The most common MKC infections are caused by the species M. kansasii (sensu stricto), leading to tuberculosis-like disease. However, a broad spectrum of virulence, antimicrobial resistance and pathogenicity of these non-tuberculous mycobacteria (NTM) are observed across the MKC. Many genomic aspects of the MKC that relate to these broad phenotypes are not well elucidated. Here, we performed genomic analyses from a collection of 665 MKC strains, isolated from environmental, animal and human sources. We inferred the MKC pangenome, mobilome, resistome, virulome and defence systems and show that the MKC species harbours unique and shared genomic signatures. High frequency of presence of prophages and different types of defence systems were observed. We found that the M. kansasii species splits into four lineages, of which three are lowly represented and mainly in Brazil, while one lineage is dominant and globally spread. Moreover, we show that four sub-lineages of this most distributed M. kansasii lineage emerged during the twentieth century. Further analysis of the M. kansasii genomes revealed almost 300 regions of difference contributing to genomic diversity, as well as fixed mutations that may explain the M. kansasii's increased virulence and drug resistance.

Darwin review: the evolution of virulence in human pathogens.

By definition, all pathogens cause some level of harm to their hosts. If this harm occurs while the pathogen is transmitting, it can negatively affect the pathogen's fitness by shortening the duration over which transmission can occur. However, many of the factors that increase virulence (i.e. harm to host) also promote transmission, driving the pathogen population towards an optimal state of intermediate virulence. A wider spectrum of virulence may be maintained among pathogen populations which are structured into multiple discrete strains though direct resource and immune-mediated competition. These various evolutionary outcomes, and the effects of medical and public health interventions, are best understood within a framework that recognizes the complex relationship between transmission and virulence in the context of the antigenic diversity of the pathogen population.

Comparative Genomic Analysis of Shrimp-Pathogenic Vibrio parahaemolyticus LC and Intraspecific Strains with Emphasis on Virulent Factors of Mobile Genetic Elements.

Vibrio parahaemolyticus exhibits severe pathogenicity in humans and animals worldwide. In this study, genome sequencing and comparative analyses were conducted for in-depth characterization of the virulence factor (VF) repertoire of V. parahaemolyticus strain LC, which presented significant virulence to shrimp Litopenaeus vannamei. Strain LC, harboring two circular chromosomes and three linear plasmids, demonstrated ≥98.14% average nucleotide identities with 31 publicly available V. parahaemolyticus genomes, including 13, 11, and 7 shrimp-, human-, and non-pathogenic strains, respectively. Phylogeny analysis based on dispensable genes of pan-genome clustered 11 out of 14 shrimp-pathogenic strains and 7 out of 11 clinical strains into two distinct clades, indicating the close association between host-specific pathogenicity and accessory genes. The VFDB database revealed that 150 VFs of LC were mainly associated with the secretion system, adherence, antiphagocytosis, chemotaxis, motility, and iron uptake, whereas no homologs of the typical pathogenic genes pirA, pirB, tdh, and trh were detected. Four genes, mshB, wbfT, wbfU, and wbtI, were identified in both types of pathogenic strains but were absent in non-pathogens. Notably, a unique cluster similar to Yen-Tc, which encodes an insecticidal toxin complex, and diverse toxin-antitoxin (TA) systems, were identified on the mobile genetic elements (MGEs) of LC. Conclusively, in addition to the common VFs, various unique MGE-borne VFs, including the Yen-Tc cluster, TA components, and multiple chromosome-encoded chitinase genes, may contribute to the full spectrum of LC virulence. Moreover, V. parahaemolyticus demonstrates host-specific virulence, which potentially drives the origin and spread of pathogenic factors.

Genetic Control of Pitch Canker Response in Southern Pine and Southern Pine Hybrids

Fusarium circinatum causes pine pitch canker (PPC) disease and associated symptoms such as resinous lesions, mechanical weakness, and crown dieback that may lead to mortality in Pinus and Pseudotsuga spp. There are no ameliorative techniques available for the disease, and the genetic resistance among populations to support commercial plantation deployment has not been well characterized. In this study, we characterize the genetic control of PPC disease tolerance (and/or resistance) and predict the tolerance of families in existing breeding populations: open-pollinated (OP) half-sib and control-pollinated full-sib (FS) slash pine (Pinus elliottii var. elliottii Engelm.), OP loblolly pine (Pinus taeda L.), and advanced-generation OP hybrid slash × P. caribaea (Pinus elliottii var. elliottii Engelm. × Pinus caribaea var. bahamensis, caribaea, and hondurensis) using F. circinatum isolates obtained from three locations in Georgia and FL, USA. We describe a new experimental design that improves the accuracy of breeding value predictions, provides more precise genetic parameter estimates, and facilitates comparisons within and among taxa as well as comparisons among isolates. We found strong evidence for genetic control of the ratio of stem damage by F. circinatum, especially in slash pine and slash × P. caribaea hybrids. Loblolly and slash × P. caribaea hybrids exhibited less damage than slash pine. We observed a spectrum of virulence among F. circinatum isolate sources, which were not equally virulent in different pine taxa.

NSs of the mildly virulent sandfly fever Sicilian virus is unable to inhibit interferon signaling and upregulation of interferon-stimulated genes.

Phleboviruses (order Bunyavirales, family Phenuiviridae) are globally emerging arboviruses with a wide spectrum of virulence. Sandfly fever Sicilian virus (SFSV) is one of the most ubiquitous members of the genus Phlebovirus and associated with a self-limited, incapacitating febrile disease in travellers and military troops. The phleboviral NSs protein is an established virulence factor, acting as antagonist of the antiviral interferon (IFN) system. Consistently, we previously reported that SFSV NSs targets the induction of IFN mRNA synthesis by specifically binding to the DNA-binding domain of the IFN transcription factor IRF3. Here, we further characterized the effect of SFSV and its NSs towards IFN induction, and evaluated its potential to affect the downstream IFN-stimulated signalling and the subsequent transactivation of antiviral interferon-stimulated genes (ISGs). We found that SFSV dampened, but did not entirely abolish type I and type III IFN induction. Furthermore, SFSV NSs did not affect IFN signalling, resulting in substantial ISG expression in infected cells. Hence, although SFSV targets IRF3 to reduce IFN induction, it is not capable of entirely disarming the IFN system in the presence of high basal IRF3 and/or IRF7 levels, and we speculate that this significantly contributes to its low level of virulence.

Complete Genome Sequencing of Leptospira interrogans Isolates from Malaysia Reveals Massive Genome Rearrangement but High Conservation of Virulence-Associated Genes.

The ability of Leptospirae to persist in environments and animal hosts but to cause clinically highly variable disease in humans has made leptospirosis the most common zoonotic disease. Considering the paucity of data on variation in complete genomes of human pathogenic Leptospirae, we have used a combination of Single Molecule Real-Time (SMRT) and Illumina sequencing to obtain complete genome sequences of six human clinical L. interrogans isolates from Malaysia. All six contained the larger (4.28–4.56 Mb) and smaller (0.34–0.395 Mb) chromosome typical of human pathogenic Leptospirae and 0–7 plasmids. Only 24% of the plasmid sequences could be matched to databases. We identified a chromosomal core genome of 3318 coding sequences and strain-specific accessory genomes of 49–179 coding sequences. These sequences enabled detailed genomic strain typing (Genome BLAST Distance Phylogeny, DNA–DNA hybridization, and multi locus sequence typing) and phylogenetic classification (whole-genome SNP genotyping). Even though there was some shared synteny and collinearity across the six genomes, there was evidence of major genome rearrangement, likely driven by horizontal gene transfer and homologous recombination. Mobile genetic elements were identified in all strains in highly varying numbers, including in the rfb locus, which defines serogroups and contributes to immune escape and pathogenesis. On the other hand, there was high conservation of virulence-associated genes including those relating to sialic acid, alginate, and lipid A biosynthesis. These findings suggest (i) that the antigenic variation, adaption to various host environments, and broad spectrum of virulence of L. interrogans are in part due to a high degree of genomic plasticity and (ii) that human pathogenic strains maintain a core set of genes required for virulence.

A CASE OF NATIVE AORTIC VALVE STAPHYLOCOCCUS LUGDUNENSIS ENDOCARDITIS

Staphylococcus lugdunensis is a coagulase negative staphylococcus (CoNS). The spectrum of virulence ranges from mild to life threatening systemic infections. A 54 year old female presented with complaints of dyspnea, orthopnea and substernal chest pain. Her past medical history included bicuspid

Integration of radiotherapy and immunotherapy for treatment of oligometastases.

Metastasis is the leading cause of cancer-related mortality and remains one of the prevailing challenges in cancer treatment. Most patients with metastatic disease are treated with systemic agents, which prolong survival and improve symptoms but are typically not curative. The oligometastatic hypothesis challenges the perspective that metastasis is an invariably disseminated process, and proposes a biological spectrum of metastatic virulence. Mounting evidence supports the idea that patients with numerically and spatially restricted sites of metastases, termed oligometastases, can achieve prolonged survival following metastasis-directed therapies, such as surgery or radiotherapy. Improvements in clinical and molecular staging of metastatic disease, as well as integration of effective systemic therapies with localised interventions, might achieve better outcomes for patients with diverse metastatic states. In this Series paper, we propose a rationale for the integration of immune checkpoint inhibitors with radiotherapy to advance the potential for effective treatment along the spectrum of disease, with emphasis on how immunotherapy can potentiate radiotherapy treatment in the oligometastatic setting.

Integrated molecular and clinical staging defines the spectrum of metastatic cancer.

The elimination of metastases remains one of the major challenges in the curative treatment of patients with cancer. Therefore, most patients with metastatic disease typically receive systemic agents, which prolong survival and alleviate symptoms but are rarely curative. The oligometastatic paradigm challenges the prevailing view of metastasis as a disseminated process and proposes the existence of a spectrum of biological virulence within metastatic lesions. In this Perspectives article, we present evidence supporting our hypothesis that integrated clinical and molecular classification of metastases along the spectrum of disease influences the staging and treatment of patients with cancer.

A Controlled Human Infection Model of Group A Streptococcus Pharyngitis: Which Strain and Why?

Group A Streptococcus (GAS) is a major cause of global infection-related morbidity and mortality. A modern controlled human infection model (CHIM) of GAS pharyngitis can accelerate vaccine development and pathogenesis research. A robust rationale for strain selection is central to meeting ethical, scientific, and regulatory requirements. Multifaceted characterization studies were done to compare a preferred candidate emm75 (M75) GAS strain to three other strains: an alternative candidate emm12 (M12) strain, an M1 strain used in 1970s pharyngitis CHIM studies (SS-496), and a representative (5448) of the globally disseminated M1T1 clone. A range of approaches were used to explore strain growth, adherence, invasion, delivery characteristics, short- and long-term viability, phylogeny, virulence factors, vaccine antigens, resistance to killing by human neutrophils, and lethality in a murine invasive model. The strains grew reliably in a medium without animal-derived components, were consistently transferred using a swab method simulating the CHIM protocol, remained viable at -80°C, and carried genes for most candidate vaccine antigens. Considering GAS molecular epidemiology, virulence factors, in vitro assays, and results from the murine model, the contemporary strains show a spectrum of virulence, with M75 appearing the least virulent and 5448 the most. The virulence profile of SS-496, used safely in 1970s CHIM studies, was similar to that of 5448 in the animal model and virulence gene carriage. The results of this multifaceted characterization confirm the M75 strain as an appropriate choice for initial deployment in the CHIM, with the aim of safely and successfully causing pharyngitis in healthy adult volunteers.IMPORTANCE GAS (Streptococcus pyogenes) is a leading global cause of infection-related morbidity and mortality. A modern CHIM of GAS pharyngitis could help to accelerate vaccine development and drive pathogenesis research. Challenge strain selection is critical to the safety and success of any CHIM and especially so for an organism such as GAS, with its wide strain diversity and potential to cause severe life-threatening acute infections (e.g., toxic shock syndrome and necrotizing fasciitis) and postinfectious complications (e.g., acute rheumatic fever, rheumatic heart disease, and acute poststreptococcal glomerulonephritis). In this paper, we outline the rationale for selecting an emm75 strain for initial use in a GAS pharyngitis CHIM in healthy adult volunteers, drawing on the findings of a broad characterization effort spanning molecular epidemiology, in vitro assays, whole-genome sequencing, and animal model studies.

Integrated molecular subtyping defines a curable oligometastatic state in colorectal liver metastasis

The oligometastasis hypothesis suggests a spectrum of metastatic virulence where some metastases are limited in extent and curable with focal therapies. A subset of patients with metastatic colorectal cancer achieves prolonged survival after resection of liver metastases consistent with oligometastasis. Here we define three robust subtypes of de novo colorectal liver metastasis through integrative molecular analysis. Patients with metastases exhibiting MSI-independent immune activation experience the most favorable survival. Subtypes with adverse outcomes demonstrate VEGFA amplification in concert with (i) stromal, mesenchymal, and angiogenic signatures, or (ii) exclusive NOTCH1 and PIK3C2B mutations with E2F/MYC activation. Molecular subtypes complement clinical risk stratification to distinguish low-risk, intermediate-risk, and high-risk patients with 10-year overall survivals of 94%, 45%, and 19%, respectively. Our findings provide a framework for integrated classification and treatment of metastasis and support the biological basis of curable oligometastatic colorectal cancer. These concepts may be applicable to many patients with metastatic cancer.

Aiding and abettingStaphylococcus aureus

Infection Human skin hosts an ecosystem of microorganisms. Infections can therefore involve more than one organism, but antibiotic therapy options rarely consider the possibility of interference by a nontarget species. Staphylococcus aureus is a member of the normal skin microbiota that can show a spectrum of virulence and cause persistent and intractable infections. Radlinski et al. show that co-infections of S. aureus and Pseudomonas aeruginosa can synergize in multiple ways to antagonize or potentiate antibiotic susceptibility in S. aureus. P. aeruginosa secretes a series of secondary products in a strain-specific manner. Some, such as rhamnolipids or LasA endopeptidase, increase S. aureus susceptibility to antibiotics. Others, such as HQNO, induce multidrug tolerance and potentiate antibiotic resistance in S. aureus . Some of these interactions could theoretically be exploited to assist treatment, but the reciprocal effects of S. aureus on P. aeruginosa , coupled with strain variation, mean each case is different. PLOS Biol. 10.1371/journal.pbio.2003981 (2017).

Real time PCR for the rapid identification and drug susceptibility of Mycobacteria present in Bronchial washings.

BackgroundMycobacteria have a spectrum of virulence and different susceptibilities to antibiotics. Distinguishing mycobacterial species is vital as patients with non-tuberculous mycobacterial (NTM) infections present clinical features that are similar to those of patients with tuberculosis. Thus, rapid differentiation of Mycobacterium tuberculosis complex from NTM is critical to administer appropriate treatment. Hence the aim of the study was to rapid identification of mycobacterial species present in bronchial washings using multiplex real time Polymerase Chain Reaction (PCR) and to determine the drug susceptibility in identified mycobacterial species.MethodsSputum smear negative bronchoscopy specimens (n = 150) were collected for a period of one year, from patients attending the General Hospital Kandy, Sri Lanka. The specimens were processed with modified Petroff’s method and were cultured on Löwenstein– Jensen medium. DNA, extracted from the mycobacterial isolates were subjected to a SYBR green mediated real time multiplex, PCR assay with primers specific for the M. tuberculosis complex, M. avium complex, M. chelonae-M.abscessus group and M. fortuitum group. DNA sequencing was performed for the species confirmation, by targeting the 16S rRNA gene and the drug susceptibility testing was performed for the molecularly identified isolates of M. tuberculosis and NTM.ResultsThe optimized SYBR Green mediated multiplex real-time PCR assay was able to identify the presence of genus Mycobacterium in 25 out of 26 AFB positive isolates, two M. tuberculosis complex, three M. avium complex and two isolates belonging to M. chelonae-M. abscessus group. DNA sequencing confirmed the presence of M. tuberculosis, M. chelonae-M. abscessus, M. intracellulare, M. avium, Rhodococcus sp. and M. celatum. Remaining isolates were identified as Mycobacterium sp. All the NTM isolates were sensitive to amikacin and seven were resistant to ciproflaxacin. Twenty two of the NTM isolates and the isolate Rhodococcus was resistant to clarithromycin. The two isolates of M. tuberculosis were sensitive to all first line anti tuberculosis drugs.ConclusionThe optimized SYBR Green mediated multiplex real time PCR assay could be an effective tool for the rapid differentiation of pathogenic M. tuberculosis complex from the opportunistic nontuberculous mycobacteria and also it confirmed the presence of NTM in 15.3 % of the study population.

Molecular diagnostics in a tuberculous patient with clinical non response to standard treatment

Rapid distinguishing of mycobacterial species and knowledge about their susceptibility is vital for the management of patients with pulmonary disease who are not responding to initial treatment as the mycobacteria have a spectrum of virulence and different susceptibilities to antibiotics. Most biochemical techniques used for species differentiation are laborious and time consuming. Hence molecular techniques are becoming an alternative to existing conventional testing. This study demonstrates the utility of molecular diagnostics for patients with mycobacterial infections who are not responding to initial anti TB therapy.

Utilizing "Omic" Technologies to Identify and Prioritize Novel Sources of Resistance to the Oomycete Pathogen Phytophthora infestans in Potato Germplasm Collections.

The greatest threat to potato production world-wide is late blight, caused by the oomycete pathogen Phytophthora infestans. A screen of 126 wild diploid Solanum accessions from the Commonwealth Potato Collection (CPC) with P. infestans isolates belonging to the genotype 13-A2 identified resistances in the species S. bulbocastanum, S. capsicibaccatum, S. microdontum, S. mochiquense, S. okadae, S. pinnatisectum, S. polyadenium, S. tarijense, and S. verrucosum. Effector-omics, allele mining, and diagnostic RenSeq (dRenSeq) were utilized to investigate the nature of resistances in S. okadae accessions. dRenSeq in resistant S. okadae accessions 7129, 7625, 3762, and a bulk of 20 resistant progeny confirmed the presence of full-length Rpi-vnt1.1 under stringent mapping conditions and corroborated allele mining results in the accessions 7129 and 7625 as well as Avr-vnt1 recognition in transient expression assays. In contrast, susceptible S. okadae accession 3761 and a bulk of 20 susceptible progeny lacked sequence homology in the 5′ end compared to the functional Rpi-vnt1.1 gene. Further evaluation of S. okadae accessions with P. infestans isolates that have a broad spectrum of virulence demonstrated that, although S. okadae accessions 7129, 7625, and 7629 contain functional Rpi-vnt1.1, they also carry a novel resistance gene. We provide evidence that existing germplasm collections are important sources of novel resistances and that “omic” technologies such as dRenSeq-based genomics and effector-omics are efficacious tools to rapidly explore the diversity within these collections.

Current Perspectives in the Diagnosis and Treatment of Nontuberculous Mycobacterial Pulmonary Disease

Nontuberculous mycobacteria (NTM), an extremely diverse group with a wide spectrum of virulence, appear to be rising in prevalence. This may be due to both increased awareness and improved diagnosis, among other factors. Most NTM-pulmonary disease (NTM-PD) infections are caused by the slow growingMycobacterium avium complex(MAC) however, other species can also cause pulmonary disease includingM. abscessus, M. xenopi, M. malmoenseandM. kansasii.Symptoms of NTM-PD vary considerably and may include chronic cough, with or without sputum production or haemoptysis. Treatment options are limited at present and associated with significant intolerance, treatment failures and drug–drug interactions. Further prospective studies are needed to define suitable drug regimens for different NTM species and to improve knowledge of the natural history and epidemiology of NTM-PD.

Physiologic specialization of Puccinia triticina Erikss. and effectiveness of Lr-genes in the south of Ukraine during 2013-2014

Leaf rust is the most widespread and frequently occurring fungal disease of wheat (Triticum aestivum L.) in Ukraine and worldwide. The information about the effectiveness of Lr-genes and also the consequent monitoring of virulence dynamics is necessary for the successful wheat breeding for leaf rust resistance. In 2013-2014 pathotype composition and virulence analysis was studied both on the standard differential set and on the North American System of Nomenclature. According to the standard differential set, 12 phenotypes were identified, of which the most common were 77 (75%) and 144 (6%). A total of 40 phenotypes were identified on the North American Nomenclature. Phenotypes TGTT (24%) and TJTT (8%) were the most frequent, TRTT (1.5%) and TSTT (1.5%) were within the broadest spectrum of virulence among the isolates found in the south of Ukraine. For virulence analysis we used wheat lines of 'Thatcher' that are near-isogenic for 24 leaf rust resistance genes and additionally four cultivars/lines. No virulence to Lr19 was found, whereas increasing virulence to Lr9 was detected (13%). Low frequency of virulence was observed to Lr29 (11%) and Lr47 (21%), high level of virulence was detected to other genes. The effectiveness of 53 known Lr-genes was studied at the seedling and the adult plant stages. Most of them were not effective against leaf rust. Genes Lr9, Lr19, Lr29, and Lr47 were highly effective both at the seedling stage and at adult plant stage. Genes Lr24, Lr42, Lr50, Lr51, and Lr56 were effective only at the adult plant stage.

ID: 107: Inhibition of type I IFN induction by Sandfly fever Sicilian virus virulence factor NSs

Phleboviruses are a group of emerging viruses with a wide spectrum of virulence: For example, Rift Valley Fever virus (RVFV) is highly pathogenic, whereas Sandfly fever Sicilian virus (SFSV) causes intermediate pathogenicity. The major virulence factor is the non-structural protein NSs, a suppressor of the type I interferon (IFN) system. While the NSs protein of highly pathogenic RVFV inhibits global host cell transcription via sequestration and degradation of the TFIIH subunits p44 and p62, respectively, the mode of action of the NSs protein of intermediately pathogenic SFSV has remained elusive. We therefore aimed to characterize the IFN-inhibitory function employed by the NSs of SFSV. Using mass spectrometry, our group has identified multiple candidate host interactors, among them interferon regulatory factor 3 (IRF3). Indeed, co-immunoprecipitation confirmed the interaction of SFSV NSs with IRF3, but not with other IRF family members like IRF2, IRF5 and IRF9. Furthermore, SFSV NSs specifically abrogated IRF3 activation and thus IFN- β promoter activity, but not general host cell transcription. Hence, SFSV encodes a NSs protein that efficiently inhibits the induction of type I IFN. Different to the highly pathogenic RVFV, which induces a general host transcription shutoff, however, SFSV specifically targets IRF3-dependent IFN promoter activation. Although RVFV and SFSV are highly related, their NSs proteins display remarkably diverse strategies of counteracting the type I IFN system. Possibly, the quality of NSs-host factor interactions correlates with the virulence levels of phleboviruses.

Evaluation of a D-amino-acid-containing fluorescence resonance energy transfer peptide library for profiling prokaryotic proteases

Bacterial proteases play an important role in a broad spectrum of processes, including colonization, proliferation, and virulence. In this respect, bacterial proteases are potential biomarkers for bacterial diagnosis and targets for novel therapeutic protease inhibitors. To investigate these potential functions, the authors designed and used a protease substrate fluorescence resonance energy transfer (FRET) library comprising 115 short d- and l-amino-acid-containing fluorogenic substrates as a tool to generate proteolytic profiles for a wide range of bacteria. Bacterial specificity of the d-amino acid substrates was confirmed using enzymes isolated from both eukaryotic and prokaryotic organisms. Interestingly, bacterial proteases that are known to be involved in housekeeping and nutrition, but not in virulence, were able to degrade substrates in which a d-amino acid was present. Using our FRET peptide library and culture supernatants from a total of 60 different bacterial species revealed novel, bacteria-specific, proteolytic profiles, although in-species variation was observed for Pseudomonas aeruginosa, Porphyromonas gingivalis, and Staphylococcus aureus. Overall, the specific characteristic of our substrate peptide library makes it a rapid tool to high-throughput screen for novel substrates to detect bacterial proteolytic activity.