Mathias Schmelcher

Last Name

Schmelcher

First Name

Mathias

ORCID

0000-0003-3535-3861

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01630 - Lehre HEST

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Publications 1 - 10 of 29

Linker-Improved Chimeric Endolysin Selectively Kills Staphylococcus aureus In Vitro, on Reconstituted Human Epidermis, and in a Murine Model of Skin Infection
Eichenseher, Fritz; Herpers, Bjorn L.; Badoux, Paul; et al. (2022)
Antimicrobial Agents and Chemotherapy
Staphylococcus aureus causes a broad spectrum of diseases in humans and animals. It is frequently associated with inflammatory skin disorders such as atopic dermatitis, where it aggravates symptoms. Treatment of S. aureus-associated skin infections with antibiotics is discouraged due to their broad-range deleterious effect on healthy skin microbiota and their ability to promote the development of resistance. Thus, novel S. aureus-specific antibacterial agents are desirable. We constructed two chimeric cell wall-lytic enzymes, Staphefekt SA.100 and XZ.700, which are composed of functional domains from the bacteriophage endolysin Ply2638 and the bacteriocin lysostaphin. Both enzymes specifically killed S. aureus and were inactive against commensal skin bacteria such as Staphylococcus epidermidis, with XZ.700 proving more active than SA.100 in multiple in vitro activity assays. When surface-attached mixed staphylococcal cultures were exposed to XZ.700 in a simplified microbiome model, the enzyme selectively removed S. aureus and retained S. epidermidis. Furthermore, XZ.700 did not induce resistance in S. aureus during repeated rounds of exposure to sublethal concentrations. Finally, we demonstrated that XZ.700 formulated as a cream is effective at killing S. aureus on reconstituted human epidermis and that an XZ.700-containing gel significantly reduces bacterial numbers compared to an untreated control in a mouse model of S. aureus-induced skin infection.
In Situ Nucleic Acid Amplification and Ultrasensitive Colorimetric Readout in a Paper-Based Analytical Device Using Silver Nanoplates
Suea-Ngam, Akkapol; Choopara, Ilada; Li, Shangkun; et al. (2021)
Advanced Healthcare Materials
A rapid, highly sensitive, and quantitative colorimetric paper‐based analytical device (PAD) based on silver nanoplates (AgNPls) and loop‐mediated isothermal amplification (LAMP) is presented. It is shown that cauliflower‐like concatemer LAMP products can mediate crystal etching of AgNPls, with a threefold signal enhancement versus linear dsDNA. Methicillin‐resistant Staphylococcus aureus (MRSA), an antimicrobial resistant bacterium that poses a formidable risk with persistently high mortality, is used as a model pathogen. Due to the excellent color contrast provided by AgNPls, the PAD allows qualitative analysis by the naked eye and quantitative analysis using a smartphone camera, with detection limits down to a single copy in just 30 min, and a linear response from 1 to 104 copies (R2 = 0.994). The entire assay runs in situ on the paper surface, which drastically simplifies operation of the device. This is the first demonstration of single copy detection using a colorimetric readout, and the developed PAD shows great promise for translation into an ultrasensitive gene‐based point‐of‐care test for any infectious disease target, via modification of the LAMP primer set. © 2020 Wiley‐VCH GmbH.
Chimeric Peptidoglycan Hydrolases Kill Staphylococcal Mastitis Isolates in Raw Milk and within Bovine Mammary Gland Epithelial Cells
Keller, Anja P.; Ly, Shera; Daetwyler, Steven; et al. (2022)
Viruses
Staphylococcus aureus is a major causative agent of bovine mastitis, a disease considered one of the most economically devastating in the dairy sector. Considering the increasing prevalence of antibiotic-resistant strains, novel therapeutic approaches efficiently targeting extra- and intracellular bacteria and featuring high activity in the presence of raw milk components are needed. Here, we have screened a library of eighty peptidoglycan hydrolases (PGHs) for high activity against S. aureus in raw bovine milk, twelve of which were selected for further characterization and comparison in time-kill assays. The bacteriocins lysostaphin and ALE-1, and the chimeric PGH M23LST(L)_SH3b2638 reduced bacterial numbers in raw milk to the detection limit within 10 min. Three CHAP-based PGHs (CHAPGH15_SH3bAle1, CHAPK_SH3bLST_H, CHAPH5_LST_H) showed gradually improving activity with increasing dilution of the raw milk. Furthermore, we demonstrated synergistic activity of CHAPGH15_SH3bAle1 and LST when used in combination. Finally, modification of four PGHs (LST, M23LST(L)_SH3b2638, CHAPK_SH3bLST, CHAPGH15_SH3bAle1) with the cell-penetrating peptide TAT significantly enhanced the eradication of intracellular S. aureus in bovine mammary alveolar cells compared to the unmodified parentals in a concentration-dependent manner.
Fluorometric Paper-Based, Loop-Mediated Isothermal Amplification Devices for Quantitative Point-of-Care Detection of Methicillin-Resistant Staphylococcus aureus (MRSA)
Choopara, Ilada; Suea-Ngam, Akkapol; Teethaisong, Yothin; et al. (2021)
ACS Sensors
Loop-mediated isothermal amplification (LAMP) has been widely used to detect many infectious diseases. However, minor inconveniences during the steps of adding reaction ingredients and lack of simple color results hinder point-of-care detection. We therefore invented a fluorometric paper-based LAMP by incorporating LAMP reagents, including a biotinylated primer, onto a cellulose membrane paper, with a simple DNA fluorescent dye incubation that demonstrated rapid and accurate results parallel to quantitative polymerase chain reaction (qPCR) methods. This technology allows for instant paper strip detection of methicillin-resistant Staphylococcus aureus (MRSA) in the laboratory and clinical samples. MRSA represents a major public health problem as it can cause infections in different parts of the human body and yet is resistant to commonly used antibiotics. In this study, we optimized LAMP reaction ingredients and incubation conditions following a central composite design (CCD) that yielded the shortest reaction time with high sensitivity. These CCD components and conditions were used to construct the paper-based LAMP reaction by immobilizing the biotinylated primer and the rest of the LAMP reagents to produce the ready-to-use MRSA diagnostic device. Our paper-based LAMP device could detect as low as 10 ag (equivalent to 1 copy) of the MRSA gene mecA within 36–43 min, was evaluated using both laboratory (individual cultures of MRSA and non-MRSA bacteria) and clinical blood samples to be 100% specific and sensitive compared to qPCR results, and had 35 day stability under 25 °C storage. Furthermore, the color readout allows for quantitation of MRSA copies. Hence, this device is applicable for point-of-care MRSA detection.
Bacteriophage endolysins — extending their application to tissues and the bloodstream
Schmelcher, Mathias; Loessner, Martin J. (2021)
Current Opinion in Biotechnology
The rapid emergence of antibiotic-resistant bacteria and the lack of novel antibacterial agents pose a serious threat for patients and healthcare systems. Bacteriophage-encoded peptidoglycan hydrolases (endolysins) represent a promising new class of antimicrobials. Over the past two decades, research on these enzymes has evolved from basic in vitro characterization to sophisticated protein engineering approaches, including advanced preclinical and clinical testing. In recent years, increasingly specific animal models have shown efficacy of endolysins against bacterial infections of various different organs and tissues of the body. Despite these advances, some challenges with regard to systemic application of endolysins remain to be addressed. These include immunogenicity, circulation half-life, and cell and tissue-specific targeting and penetration properties.
Peptidoglycan Hydrolases: Powerful Tools for Detection and Control of Bacterial Pathogens
Schmelcher, Mathias (2020)
CkP1 bacteriophage, a S16-like myovirus that recognizes Citrobacter koseri lipopolysaccharide through its long tail fibers
Oliveira, Hugo; Santos, Sílvio; Pires, Diana P.; et al. (2023)
Applied Microbiology and Biotechnology
Citrobacter koseri is an emerging Gram-negative bacterial pathogen, which causes urinary tract infections. We isolated and characterized a novel S16-like myovirus CKP1 (vB_CkoM_CkP1), infecting C. koseri. CkP1 has a host range covering the whole C. koseri species, i.e., all strains that were tested, but does not infect other species. Its linear 168,463-bp genome contains 291 coding sequences, sharing sequence similarity with the Salmonella phage S16. Based on surface plasmon resonance and recombinant green florescence protein fusions, the tail fiber (gp267) was shown to decorate C. koseri cells, binding with a nanomolar affinity, without the need of accessory proteins. Both phage and the tail fiber specifically bind to bacterial cells by the lipopolysaccharide polymer. We further demonstrate that CkP1 is highly stable towards different environmental conditions of pH and temperatures and is able to control C. koseri cells in urine samples. Altogether, CkP1 features optimal in vitro characteristics to be used both as a control and detection agent towards drug-resistant C. koseri infections.
Point-of-care testing system for digital single cell detection of MRSA directly from nasal swabs
Schulz, Martin; Calabrese, Silvia; Wurm, Holger; et al. (2020)
Lab on a Chip
We present an automated point-of-care testing (POCT) system for rapid detection of species- and resistance markers in methicillin-resistant Staphylococcus aureus (MRSA) at the level of single cells, directly from nasal swab samples. Our novel system allows clear differentiation between MRSA, methicillin-sensitive S. aureus (MSSA) and methicillin-resistant coagulase-negative staphylococci (MR-CoNS), which is not the case for currently used real-time quantitative PCR based systems. On top, the novel approach outcompetes the culture-based methods in terms of its short time-to-result (1 h vs. up to 60 h) and reduces manual labor. The walk-away test is fully automated on the centrifugal microfluidic LabDisk platform. The LabDisk cartridge comprises the unit operations swab-uptake, reagent pre-storage, distribution of the sample into 20 000 droplets, specific enzymatic lysis of Staphylococcus spp. and recombinase polymerase amplification (RPA) of species (vicK) – and resistance (mecA) -markers. LabDisk actuation, incubation and multi-channel fluorescence detection is demonstrated with a clinical isolate and spiked nasal swab samples down to a limit of detection (LOD) of 3 ± 0.3 CFU μl−1 for MRSA. The novel approach of the digital single cell detection is suggested to improve hospital admission screening, timely decision making, and goal-oriented antibiotic therapy. The implementation of a higher degree of multiplexing is required to translate the results into clinical practice. © 2020 The Royal Society of Chemistry.
Domain shuffling and module engineering of Listeria phage endolysins for enhanced lytic activity and binding affinity
Schmelcher, Mathias; Tchang, Vincent S.; Loessner, Martin J. (2011)
Microbial Biotechnology
Targeting Hidden Pathogens: Cell-Penetrating Enzybiotics Eradicate Intracellular Drug-Resistant Staphylococcus aureus
Röhrig, Christian; Huemer, Markus; Lorgé, Dominique; et al. (2020)
mBio
Staphylococcus aureus is a major concern in human health care, mostly due to the increasing prevalence of antibiotic resistance. Intracellular localization of S. aureus plays a key role in recurrent infections by protecting the pathogens from antibiotics and immune responses. Peptidoglycan hydrolases (PGHs) are highly specific bactericidal enzymes active against both drug-sensitive and -resistant bacteria. However, PGHs able to effectively target intracellular S. aureus are not yet available. To overcome this limitation, we first screened 322 recombineered PGHs for staphylolytic activity under conditions found inside eukaryotic intracellular compartments. The most active constructs were modified by fusion to different cell-penetrating peptides (CPPs), resulting in increased uptake and enhanced intracellular killing (reduction by up to 4.5 log units) of various S. aureus strains (including methicillin-resistant S. aureus [MRSA]) in different tissue culture infection models. The combined application of synergistic PGH-CPP constructs further enhanced their intracellular efficacy. Finally, synergistically active PGH-CPP cocktails reduced the total S. aureus by more than 2.2 log units in a murine abscess model after peripheral injection. Significantly more intracellular bacteria were killed by the PGH-CPPs than by the PGHs alone. Collectively, our findings show that CPP-fused PGHs are effective novel protein therapeutics against both intracellular and drug-resistant S. aureus.

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Mathias Schmelcher

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