Bacterial Lipoproteins Shift Cellular Metabolism to Glycolysis in Macrophages Causing Bone Erosion.

Belonging to a group of membrane proteins, bacterial lipoproteins (LPPs) are defined by a unique lipid structure at their N-terminus providing the anchor in the bacterial cell membrane. In Gram-positive bacteria, LPPs play a key role in host immune activation triggered through a Toll-like receptor 2 (TLR2)-mediated action resulting in macrophage stimulation and subsequent tissue damage demonstrated in in vivo experimental models. Yet the physiologic links between LPP activation, cytokine release, and any underlying switches in cellular metabolism remain unclear. In this study, we demonstrate that Staphylococcus aureus Lpl1 not only triggers cytokine production but also confers a shift toward fermentative metabolism in bone marrow-derived macrophages (BMDMs). Lpl1 consists of di- and tri-acylated LPP variants; hence, the synthetic P2C and P3C, mimicking di-and tri-acylated LPPs, were employed to reveal their effect on BMDMs. Compared to P3C, P2C was found to shift the metabolism of BMDMs and the human mature monocytic MonoMac 6 (MM6) cells more profoundly toward the fermentative pathway, as indicated by lactate accumulation, glucose consumption, pH reduction, and oxygen consumption. In vivo, P2C caused more severe joint inflammation, bone erosion, and lactate and malate accumulation than P3C. These observed P2C effects were completely abrogated in monocyte/macrophage-depleted mice. Taken together, these findings now solidly confirm the hypothesized link between LPP exposure, a macrophage metabolic shift toward fermentation, and ensuing bone destruction. IMPORTANCE Osteomyelitis caused by S. aureus is a severe infection of the bone, typically associated with severe bone function impairment, therapeutic failure, high morbidity, invalidity, and occasionally even death. The hallmark of staphylococcal osteomyelitis is the destruction of the cortical bone structures, yet the mechanisms contributing to this pathology are hitherto poorly understood. One bacterial membrane constituent found in all bacteria is bacterial lipoproteins (LPPs). Previously, we have shown that injection of purified S. aureus LPPs into wild-type mouse knee joints caused a TLR2-dependent chronic destructive arthritis but failed to elicit such effect in monocyte/macrophage-depleted mice. This observation stirred our interest in investigating the interaction of LPPs and macrophages and analyzing the underlying physiological mechanisms. This ascertainment of LPP-induced changes in the physiology of macrophages provides an important clue in the understanding of the mechanisms of bone disintegration, opening novel avenues to manage the course of S. aureus disease.

Protein Disulfide Isomerase and Extracellular Adherence Protein Cooperatively Potentiate Staphylococcal Invasion into Endothelial Cells.

Invasion of host cells is an important feature of Staphylococcus aureus. The main internalization pathway involves binding of the bacteria to host cells, e.g., endothelial cells, via a fibronectin (Fn) bridge between S. aureus Fn binding proteins and α5ß1-integrin, followed by phagocytosis. The secreted extracellular adherence protein (Eap) has been shown to promote this cellular uptake pathway of not only S. aureus, but also of bacteria otherwise poorly taken up by host cells, such as Staphylococcus carnosus. The exact mechanisms are still unknown. Previously, we demonstrated that Eap induces platelet activation by stimulation of the protein disulfide isomerase (PDI), a catalyst of thiol-disulfide exchange reactions. Here, we show that Eap promotes PDI activity on the surface of endothelial cells, and that this contributes critically to Eap-driven staphylococcal invasion. PDI-stimulated ß1-integrin activation followed by increased Fn binding to host cells likely accounts for the Eap-enhanced uptake of S. aureus into non-professional phagocytes. Additionally, Eap supports the binding of S. carnosus to Fn-α5ß1 integrin, thereby allowing its uptake into endothelial cells. To our knowledge, this is the first demonstration that PDI is crucial for the uptake of bacteria into host cells. We describe a hitherto unknown function of Eap-the promotion of an enzymatic activity with subsequent enhancement of bacterial uptake-and thus broaden mechanistic insights into its importance as a driver of bacterial pathogenicity. IMPORTANCE Staphylococcus aureus can invade and persist in non-professional phagocytes, thereby escaping host defense mechanisms and antibiotic treatment. The intracellular lifestyle of S. aureus contributes to the development of infection, e.g., in infective endocarditis or chronic osteomyelitis. The extracellular adherence protein secreted by S. aureus promotes its own internalization as well as that of bacteria that are otherwise poorly taken up by host cells, such as Staphylococcus carnosus. In our study, we demonstrate that staphylococcal uptake by endothelial cells requires catalytic disulfide exchange activity by the cell-surface protein disulfide isomerase, and that this critical enzymatic function is enhanced by Eap. The therapeutic application of PDI inhibitors has previously been investigated in the context of thrombosis and hypercoagulability. Our results add another intriguing possibility: therapeutically targeting PDI, i.e., as a candidate approach to modulate the initiation and/or course of S. aureus infectious diseases.

Investigation on the Curvature and Stress Distribution of Laminates Based on an Analytic Solution and FE Simulation.

The potential combinations of favorable properties give metal-ceramic laminates (MCLs) a high degree of application flexibility. However, the different thermal expansion coefficients (CTEs) and shrinkage rates of the metals and ceramics during the co-sintering process often lead to large internal stresses that cause undesired deformation or even production failures. In practice, the identification of manufacturable MCLs relies on the "trial and error" principle, which usually requires a long development period. Therefore, there is a great demand for analytic and numerical methods that allow the prediction of the deformation and manufacturability of MCLs during the co-sintering process. The main objective of this study is to investigate the curvature and stress distribution in the MCLs (steel 17-4PH/ ceramic 3Y-TZP) based on the analytic solution and finite element (FE) simulation. To achieve this, the Young's moduli (E) and shear moduli (G) at high temperatures and the CTEs of both materials were measured. In addition, the curvatures and stress distributions of the two-layer and three-layer laminates were obtained based on the analytic method and FE simulation, which were in very good agreement. Furthermore, the influence of the CTE, Young's modulus, height ratio, and interface on the curvature were studied. The results showed that the CTE and height ratio have a higher influence on the curvature in comparison to the Young's modulus. The interface prevents the curvature significantly by assuming it to be a cohesive surface in the FE simulation. This provides hints to avoid delamination during the manufacturing process.

Li-Ion Conductive Li1.3Al0.3Ti1.7(PO4)3 (LATP) Solid Electrolyte Prepared by Cold Sintering Process with Various Sintering Additives.

The density, microstructure, and ionic conductivity of solid electrolyte Li1.3Al0.3Ti1.7(PO4)3 (LATP) ceramics prepared by cold sintering using liquid and solid sintering additives are studied. The effects of both liquid (water and water solutions of acetic acid and lithium hydroxide) and solid (lithium acetate) additives on densification are investigated. The properties of cold-sintered LATP are compared to those of conventionally sintered LATP. The materials cold-sintered at temperatures 140-280 °C and pressures 510-600 MPa show relative density in the range of 90-98% of LATP's theoretical value, comparable or higher than the density of conventionally sintered ceramics. With the relative density of 94%, a total ionic conductivity of 1.26 × 10-5 S/cm (room temperature) is achieved by cold sintering at the temperature of 200 °C and uniaxial pressure of 510 MPa using water as additive. The lower ionic conductivities of the cold-sintered ceramics compared to those prepared by conventional sintering are attributed to the formation of amorphous secondary phases in the intergranular regions depending on the type of additives used and on the processing conditions selected.

Staphylococcus schweitzeri-An Emerging One Health Pathogen?

The Staphylococcus aureus-related complex is formed by the Staphylococcus aureus, Staphylococcus schweitzeri, Staphylococcus argenteus, Staphylococcus roterodami and Staphylococcus singaporensis. Within this complex, S. schweitzeri is the only species mainly found in African wildlife, but it is rarely detected as a colonizer in humans or as a contaminant of fomites. The few detections in humans are most likely spillover events after contact with wildlife. However, since S. schweitzeri can be misidentified as S. aureus using culture-based routine techniques, it is likely that S. schweitzeri is under-reported in humans. The low number of isolates in humans, though, is consistent with the fact that the pathogen has typical animal adaptation characteristics (e.g., growth kinetics, lack of immune evasion cluster and antimicrobial resistance); however, evidence from selected in vitro assays (e.g., host cell invasion, cell activation, cytotoxicity) indicate that S. schweitzeri might be as virulent as S. aureus. In this case, contact with animals colonized with S. schweitzeri could constitute a risk for zoonotic infections. With respect to antimicrobial resistance, all described isolates were found to be susceptible to all antibiotics tested, and so far no data on the development of spontaneous resistance or the acquisition of resistance genes such the mecA/mecC cassette are available. In summary, general knowledge about this pathogen, specifically on the potential threat it may incur to human and animal health, is still very poor. In this review article, we compile the present state of scientific research, and identify the knowledge gaps that need to be filled in order to reliably assess S. schweitzeri as an organism with global One Health implications.

Quiescence of Human Monocytes after Affinity Purification: A Novel Method Apt for Monocyte Stimulation Assays.

Several methods to isolate monocytes from whole blood have been previously published, with different advantages and disadvantages. For the purpose of cytokine release assessment upon external stimulation, the use of monocyte preparations consisting of non-activated cells is prerequisite. Affinity-isolated monocyte preparations from peripheral blood mononuclear cells (PBMCs), obtained via positive or negative selection using magnetic beads, released pro-inflammatory cytokines such as TNF-α and IL-6 even without adding external stimuli, hindering any assessment of an effect of bacterial lipoproteins on cell stimulation. Hence, the cell preparation protocol was modified by adding a quiescence step on repellent surface culture plates, dampening any monocyte pre-activation. This protocol now provides a robust method to prepare silent yet fully activatable, pure monocyte populations for further use in stimulus-elicited activation experiments.

Influence of R=Y, Gd, Sm on Crystallization and Sodium Ion Conductivity of Na5RSi4O12 Phase.

New sodium-based battery concepts require solid electrolytes as ion conducting separators. Besides NaSICON and ß-Al2O3 in the Na2O-R2O3-SiO2 system (R = rare earth), a rarely noticed glass-ceramic solid electrolyte with the composition Na5RSi4O12 (N5-type) exists. The present study addresses the investigation of the ionic conductivity of Na5RSi4O12 solid electrolytes sintered from pre-crystallized glass-ceramic powders. The sintering behavior (optical dilatometry), the microstructure (SEM/EDX), and phase composition (XRD), as well as electrochemical properties (impedance spectroscopy), were investigated. To evaluate the effect of the ionic radii, Y, Sm and Gd rare elements were chosen. All compositions were successfully synthesized to fully densified compacts having the corresponding conducting N5-type phase as the main component. The densification behavior was in agreement with the melting point, which decreased with increasing ionic radii and specific cell volume. Alternatively, the ionic conductivities of N5-phases decreased from Y to Gd and Sm containing samples. The highest ionic conductivity of 1.82 × 10-3 S cm-1 at 20 °C was obtained for Na5YSi4O12 composition. The impact of grain boundaries and bulk conductivity on measured values is discussed. A powder-based synthesis method of this glass-ceramic solid electrolyte using different rare earth elements opens possibilities for optimizing ionic conductivity and scalable technological processing by tape casting.

More Is Not Always Better-the Double-Headed Role of Fibronectin in Staphylococcus aureus Host Cell Invasion.

While Staphylococcus aureus has classically been considered an extracellular pathogen, these bacteria are also capable of being taken up by host cells, including nonprofessional phagocytes such as endothelial cells, epithelial cells, or osteoblasts. The intracellular S. aureus lifestyle contributes to infection development. The predominant recognition and internalization pathway appears to be the binding of the bacteria via a fibronectin bridge to the α5ß1-integrin on the host cell membrane, followed by phagocytosis. Although osteoblasts showed high expression of α5ß1-integrin and fibronectin, and bacteria adhered to osteoblasts to a high proportion, here we demonstrate by internalization assays and immunofluorescence microscopy that S. aureus was less engulfed in osteoblasts than in epithelial cells. The addition of exogenous fibronectin during the infection of cells with S. aureus resulted in an increased uptake by epithelial cells but not by osteoblasts. This contrasts with the previous conception of the uptake mechanism, where high expression of integrin and fibronectin would promote the bacterial uptake into host cells. Extracellular fibronectin surrounding osteoblasts, but not epithelial cells, is organized in a fibrillary network. The inhibition of fibril formation, the short interfering RNA-mediated reduction of fibronectin expression, and the disruption of the fibronectin-fibril meshwork all resulted in a significant increase in S. aureus uptake by osteoblasts. Thus, the network of fibronectin fibrils appears to strongly reduce the uptake of S. aureus into a given host cell, indicating that the supramolecular structure of fibronectin determines the capacity of particular host cells to internalize the pathogen. IMPORTANCE Traditionally, Staphylococcus aureus has been considered an extracellular pathogen. However, among other factors, the frequent failure of antimicrobial therapy and the ability of the pathogen to cause recurrent disease have established the concept of eukaryotic invasion of the pathogen, thereby evading the host's immune system. In the current model of host cell invasion, bacteria initially bind to α5ß1 integrin on the host cell side via a fibronectin bridge, which eventually leads to phagocytosis of S. aureus by host cells. However, in this study, we demonstrate that not the crude amount but the supramolecular structure of fibronectin molecules deposited on the eukaryotic cell surface plays an essential role in bacterial uptake by host cells. Our findings explain the large differences of S. aureus uptake efficacy in different host cell types as well as in vivo differences between courses of bacterial infections and the localization of bacteria in different clinical settings.

Thermal Stability of TiN Coated Cubic Boron Nitride Powder.

Wear-resistant, super hard ceramic composites based on cubic boron nitride (cBN) are of great interest to industry. However, cBN is metastable under sintering conditions at normal pressure and converts into the soft hexagonal BN (hBN). Therefore, efforts are being made to avoid this process. Besides short sintering times, the use of coated cBN-particles is a way to minimize this process. Therefore, the thermal stability of TiN coated cBN powders in high purity argon and nitrogen atmospheres up to temperatures of 1600 °C was investigated by thermogravimetry, X-ray phase analysis, scanning electron microscopy and Raman spectroscopy. The TiN coating was prepared by the atomic layer deposition (ALD)-method. The investigations showed that the TiN layer reacts in Ar at T ≥ 1200 °C with the cBN and forms a porous TiB2 layer. No reaction takes place in nitrogen up to temperatures of 1600 °C. Nevertheless, the 20 and 50 nm thin coatings also undergo a recrystallization process during heat treatment up to temperatures of 1600 °C.

Microstructure and Fracture Mechanism Investigation of Porous Silicon Nitride-Zirconia-Graphene Composite Using Multi-Scale and In-Situ Microscopy.

Silicon nitride-zirconia-graphene composites with high graphene content (5 wt.% and 30 wt.%) were sintered by gas pressure sintering (GPS). The effect of the multilayer graphene (MLG) content on microstructure and fracture mechanism is investigated by multi-scale and in-situ microscopy. Multi-scale microscopy confirms that the phases disperse evenly in the microstructure without obvious agglomeration. The MLG flakes well dispersed between ceramic matrix grains slow down the phase transformation from α to ß-Si3N4, subsequent needle-like growth of ß-Si3N4 rods and the densification due to the reduction in sintering additives particularly in the case with 30 wt.% MLG. The size distribution of Si3N4 phase shifts towards a larger size range with the increase in graphene content from 5 to 30 wt.%, while a higher graphene content (30 wt.%) hinders the growth of the ZrO2 phase. The composite with 30 wt.% MLG has a porosity of 47%, the one with 5 wt.% exhibits a porosity of approximately 30%. Both Si3N4/MLG composites show potential resistance to contact or indentation damage. Crack initiation and propagation, densification of the porous microstructure, and shift of ceramic phases are observed using in-situ transmission electron microscopy. The crack propagates through the ceramic/MLG interface and through both the ceramic and the non-ceramic components in the composite with low graphene content. However, the crack prefers to bypass ceramic phases in the composite with 30 wt.% MLG.

Influence of Cemented Carbide Composition on Cutting Temperatures and Corresponding Hot Hardnesses.

During metal cutting, high temperatures of several hundred-degree Celsius occur locally at the cutting edge, which greatly impacts tool wear and life. Not only the cutting parameters, but also the tool material's properties influence the arising cutting temperature which in turn alters the mechanical properties of the tool. In this study, the hardness and thermal conductivity of cemented tungsten carbides were investigated in the range between room temperature and 1000 °C. The occurring temperatures close to the cutting edge were measured with two color pyrometry. The interactions between cemented carbide tool properties and cutting process parameters, including cutting edge rounding, are discussed. The results show that cemented carbides with higher thermal conductivities lead to lower temperatures during cutting. As a result, the effective hardness at the cutting edge can be strongly influenced by the thermal conductivity. The differences in hardness measured at room temperature can be equalized or evened out depending on the combination of hardness and thermal conductivity. This in turn has a direct influence on tool wear. Wear is also influenced by the softening of the workpiece, so that higher cutting temperatures can lead to less wear despite the same effective hardness.

Lipoproteins in Gram-Positive Bacteria: Abundance, Function, Fitness.

When one thinks of the Gram+ cell wall, the peptidoglycan (PG) scaffold in particular comes to mind. However, the cell wall also consists of many other components, for example those that are covalently linked to the PG: the wall teichoic acid and the cell wall proteins tethered by the sortase. In addition, there are completely different molecules that are anchored in the cytoplasmic membrane and span the cell wall. These are lipoteichoic acids and bacterial lipoproteins (Lpp). The latter are in the focus of this review. Lpp are present in almost all bacteria. They fulfill a wealth of different tasks. They represent the window to the outside world by recognizing nutrients and incorporating them into the bacterial cell via special transport systems. Furthermore, they perform very diverse and special tasks such as acting as chaperonin, as cyclomodulin, contributing to invasion of host cells or uptake of plasmids via conjugation. All these functions are taken over by the protein part. Nevertheless, the lipid part of the Lpp plays an as important role as the protein part. It is the released lipoproteins and derived lipopeptides that massively modulate our immune system and ultimately play an important role in immune tolerance or non-tolerance. All these varied activities of the Lpp are considered in this review article.

Analysis of the dynamics of Staphylococcus aureus binding to white blood cells using whole blood assay and geno-to-pheno mapping.

Given that binding and internalization of bacteria to host cells promotes infections and invasion, we aimed at characterizing how various S. aureus isolates adhere to and are internalized by different white blood cells. In particular, the role of genetic determinants on the association kinetics should be unveiled. A flow cytometric (FACS) whole blood assay with fluorescently labelled isolates was applied to 56 clinical S. aureus isolates. This phenotypic data was then linked to previously obtained genotyping data (334 genes) with the help of a redescription mining algorithm. Professional phagocytes showed a time-dependent increase of bacterial adhesion and internalization. Isolates showing higher affinity to granulocytes were associated with lower binding to monocytes. In contrast binding activity between S. aureus and lymphocytes could be subdivided into two phases. Preliminary binding (phase 1) was highest directly after co-incubation and was followed by S. aureus detachment or by sustained binding of a small lymphocyte subset (phase 2). Strain-dependent low granulocyte binding was observed for clonal complex 5 (CC5) isolates (MRSA), as compared to CC30 and CC45 (MSSA). S. aureus isolates associated with low granulocyte phagocytosis were characterized by the presence (cap8, can) and the absence (sasG, lukD, isdA, splA, setC) of specific hybridization signals.

ClpC affects the intracellular survival capacity of Staphylococcus aureus in non-professional phagocytic cells.

Invasion and persistence of bacteria within host cells requires that they adapt to life in an intracellular environment. This adaptation induces bacterial stress through events such as phagocytosis and enhanced nutrient-restriction. During stress, bacteria synthesize a family of proteins known as heat shock proteins (HSPs) to facilitate adaptation and survival. Previously, we determined the Staphylococcus aureus HSP ClpC temporally alters bacterial metabolism and persistence. This led us to hypothesize that ClpC might alter intracellular survival. Inactivation of clpC in S. aureus strain DSM20231 significantly enhanced long-term intracellular survival in human epithelial (HaCaT) and endothelial (EA.hy926) cell lines, without markedly affecting adhesion or invasion. This phenotype was similar across a genetically diverse collection of S. aureus isolates, and was influenced by the toxin/antitoxin encoding locus mazEF. Importantly, MazEF alters mRNA synthesis and/or stability of S. aureus virulence determinants, indicating ClpC may act through the mRNA modulatory activity of MazEF. Transcriptional analyses of total RNAs isolated from intracellular DSM20231 and isogenic clpC mutant cells identified alterations in transcription of α-toxin (hla), protein A (spa), and RNAIII, consistent with the hypothesis that ClpC negatively affects the intracellular survival of S. aureus in non-professional phagocytic cells, via modulation of MazEF and Agr.

Integrating Culture-based Antibiotic Resistance Profiles with Whole-genome Sequencing Data for 11,087 Clinical Isolates.

Emerging antibiotic resistance is a major global health threat. The analysis of nucleic acid sequences linked to susceptibility phenotypes facilitates the study of genetic antibiotic resistance determinants to inform molecular diagnostics and drug development. We collected genetic data (11,087 newly-sequenced whole genomes) and culture-based resistance profiles (10,991 out of the 11,087 isolates comprehensively tested against 22 antibiotics in total) of clinical isolates including 18 main species spanning a time period of 30 years. Species and drug specific resistance patterns were observed including increased resistance rates for Acinetobacter baumannii to carbapenems and for Escherichia coli to fluoroquinolones. Species-level pan-genomes were constructed to reflect the genetic repertoire of the respective species, including conserved essential genes and known resistance factors. Integrating phenotypes and genotypes through species-level pan-genomes allowed to infer gene-drug resistance associations using statistical testing. The isolate collection and the analysis results have been integrated into GEAR-base, a resource available for academic research use free of charge at https://gear-base.com.

The Staphylococcus aureus Extracellular Adherence Protein Eap Is a DNA Binding Protein Capable of Blocking Neutrophil Extracellular Trap Formation.

The extracellular adherence protein (Eap) of Staphylococcus aureus is a secreted protein known to exert a number of adhesive and immunomodulatory properties. Here we describe the intrinsic DNA binding activity of this multifunctional secretory factor. By using atomic force microscopy, we provide evidence that Eap can bind and aggregate DNA. While the origin of the DNA substrate (e.g., eukaryotic, bacterial, phage, and artificial DNA) seems to not be of major importance, the DNA structure (e.g., linear or circular) plays a critical role with respect to the ability of Eap to bind and condense DNA. Further functional assays corroborated the nature of Eap as a DNA binding protein, since Eap suppressed the formation of "neutrophil extracellular traps" (NETs), composed of DNA-histone scaffolds, which are thought to function as a neutrophil-mediated extracellular trapping mechanism. The DNA binding and aggregation activity of Eap may thereby protect S. aureus against a specific anti-microbial defense reaction from the host.

Management of superficial and deep-seated Staphylococcus aureus skin and soft tissue infections in sub-Saharan Africa: a post hoc analysis of the StaphNet cohort.

PURPOSE: The incidence of Staphylococcus aureus skin and soft tissue infection (SSTI) is high in sub-Saharan Africa. This is fueled by a high prevalence of Panton-Valentine leukocidin (PVL), which can be associated with necrotizing disease. The aim was to describe the clinical presentation and the treatment of SSTI in the African setting and to identify challenges in the management. METHODS: Patients (n = 319) were recruited in DR Congo (n = 56, 17.6%), Gabon (n = 89, 27.9%), Mozambique (n = 79, 24.8%) and Tanzania (n = 95, 29.8%) during the prospective observational StaphNet cohort study (2010-2015). A physician recorded the clinical management in standardized questionnaires and stratified the entity of SSTI into superficial (sSSTI) or deep-seated (dSSTI). Selected virulence factors (PVL, ß hemolysin) and multilocus sequence types (MLST) were extracted from whole genome sequencing data. RESULTS: There were 220/319 (69%) sSSTI and 99/319 (31%) dSSTI. Compared to sSSTI, patients with dSSTI were more often hospitalized (13.2 vs. 23.5%, p = 0.03), HIV-positive (7.6 vs. 15.9%, p = 0.11), and required more often incision and drainage (I&D, 45.5 vs. 76.5%, p = 0.04). The proportion of an adequate antimicrobial therapy increased marginally from day 1 (empirical therapy) to day 3 (definite therapy), for sSSTI (70.7 to 72.4%) and dSSTI (55.4 to 58.9%). PVL was a risk factor for I&D (OR = 1.7, p = 0.02) and associated with MLST clonal complex CC121 (OR = 2.7, p < 0.001). CONCLUSION: Appropriate antimicrobial agents and surgical services to perform I&D were available for the majority of patients. Results from susceptibility testing should be considered more efficiently in the selection of antimicrobial therapy.

On the Stability of c-BN-Reinforcing Particles in Ceramic Matrix Materials.

Cubic boron nitride (c-BN) composites produced at high pressures and temperatures are widely used as cutting tool materials. The advent of new, effective pressure-assisted densification methods, such as spark plasma sintering (SPS), has stimulated attempts to produce these composites at low pressures. Under low-pressure conditions, however, transformation of c-BN to the soft hexagonal BN (h-BN) phase can occur, with a strong deterioration in hardness and wear. In the present work, the influence of secondary phases (B2O3, Si3N4, and oxide glasses) on the transformation of c-BN was studied in the temperature range between 1100 °C and 1575 °C. The different heat treated c-BN particles and c-BN composites were analyzed by SEM, X-ray diffraction, and Raman spectroscopy. The transformation mechanism was found to be kinetically controlled solution-diffusion-precipitation. Given a sufficiently low liquid phase viscosity, the transformation could be observed at temperatures as low as 1200 °C for the c-BN-glass composites. In contrast, no transformation was found at temperatures up to 1575 °C when no liquid oxide phase is present in the composite. The results were compared with previous studies concerning the c-BN stability and the c-BN phase diagram.