Hypervirulent Klebsiella pneumoniae employs genomic island encoded toxins against bacterial competitors in the gut.

The hypervirulent lineages of Klebsiella pneumoniae (HvKp) cause invasive infections such as Klebsiella-liver abscess. Invasive infection often occurs after initial colonization of the host gastrointestinal tract by HvKp. Over 80% of HvKp isolates belong to the clonal group 23 sublineage I that has acquired genomic islands (GIs) GIE492 and ICEKp10. Our analysis of 12 361 K. pneumoniae genomes revealed that GIs GIE492 and ICEKp10 are co-associated with the CG23-I and CG10118 HvKp lineages. GIE492 and ICEKp10 enable HvKp to make a functional bacteriocin microcin E492 (mccE492) and the genotoxin colibactin, respectively. We discovered that GIE492 and ICEKp10 play cooperative roles and enhance gastrointestinal colonization by HvKp. Colibactin is the primary driver of this effect, modifying gut microbiome diversity. Our in vitro assays demonstrate that colibactin and mccE492 kill or inhibit a range of Gram-negative Klebsiella species and Escherichia coli strains, including Gram-positive bacteria, sometimes cooperatively. Moreover, mccE492 and colibactin kill human anaerobic gut commensals that are similar to the taxa found altered by colibactin in the mouse intestines. Our findings suggest that GIs GIE492 and ICEKp10 enable HvKp to kill several commensal bacterial taxa during interspecies interactions in the gut. Thus, acquisition of GIE492 and ICEKp10 could enable better carriage in host populations and explain the dominance of the CG23-I HvKp lineage.

Disease progression, treatments, hospitalization, and clinical outcomes in acute GVHD: a multicenter chart review.

Acute graft-versus-host disease (GVHD) remains a barrier to successful allogeneic hematopoietic cell transplantation (HCT) outcomes. This multicenter, retrospective chart review describes disease progression, treatment patterns, hospitalizations, and clinical outcomes among 475 patients (≥12 years old) who developed grades II-IV acute GVHD after their first HCT (January 2014-June 2016). Median (interquartile range) age at HCT was 55 (44-63) years. From the date of acute GVHD diagnosis, 190 patients (40.0%) experienced progression to more severe disease and/or developed new organ involvement. Among 431 patients with grades II-IV acute GVHD at diagnosis, 73.1% received first-line systemic corticosteroids. During follow-up (median 524 days since acute GVHD diagnosis), 23.4% of patients had an increase in steroid dose and 44.4% were unable to taper below 10 mg/day. Over half of patients (54.9%) required ≥1 hospital readmission within 100 days post-HCT (≥2 readmissions in 22.3%); mean inpatient length of stay upon readmission was 27.5 days. Approximately half of patients (52.8%) died during follow-up; 1-year overall mortality from date of acute GVHD diagnosis and nonrelapse mortality rates were 35.2% and 25.5%, respectively. Overall, patients who developed acute GVHD following HCT had poor clinical outcomes, highlighting the unmet need for early and effective treatment strategies.

Disease progression, hospital readmissions, and clinical outcomes for patients with steroid-refractory acute graft-versus-host disease: A multicenter, retrospective study.

Acute graft-versus-host disease (GVHD) is a significant cause of morbidity and mortality following allogeneic hematopoietic cell transplantation (HCT). This analysis of 168 patients (mean age, 54.8 years) from a multicenter, retrospective chart review describes the clinical course, treatment patterns, hospitalizations, and clinical outcomes of patients aged ≥12 years who developed grades II-IV acute GVHD after their first allogeneic HCT (January 1, 2014, to June 30, 2016) and were refractory to or dependent on corticosteroids. Between diagnosis and maximum grade (median, 6.0 days), 53.6% of patients had new organ involvement, particularly lower gastrointestinal tract acute GVHD, or an increase in acute GVHD grade. Eighty-nine patients (53.0%) received additional systemic GVHD therapy (after systemic corticosteroids) within a median of 21.0 days. Hospital readmission(s) was required for 56.5% of patients within 100 days post-HCT (mean inpatient length of readmission stay, 49.5 days); 24.4% had ≥2 readmissions within 100 days post-HCT. From the date of acute GVHD diagnosis, 70.2% of patients died at a median (interquartile range) of 117.5 (49-258) days. In summary, steroid-refractory and steroid-dependent acute GVHD is associated with a rapidly worsening clinical course that leads to high readmission and mortality rates, emphasizing the need for effective and tolerable therapies.

Shared signatures and divergence in skin microbiomes of children with atopic dermatitis and their caregivers.

BACKGROUND: Atopic dermatitis (AD) is a common chronic skin condition in children (15-20%) that can significantly impair their quality of life. As a result of its relapsing nature and enrichment of Staphylococcus aureus during flares, clinical management can include eradicating S aureus from the skin of children; however, this does not extend to their healthy caregivers, who are potential reservoirs. OBJECTIVE: Our aim was to understand skin microbiome sharing and microbial features in children with AD and their healthy adult caregivers. METHODS: We utilized whole-metagenome profiling at 4 body sites (volar forearm, antecubital fossae, cheeks, and lesions) in combination with sequencing of S aureus isolates to characterize a cohort of children with AD and their healthy caregivers (n = 30 families) compared to matched pairs from control households (n = 30 families). RESULTS: Metagenomic analysis revealed distinct microbiome configurations in the nonlesional skin of AD children and their healthy caregivers versus controls, which were sufficient to accurately predict case-control status (area under the receiver operating characteristic curve > 0.8). These differences were accompanied by significant microbiome similarity between children and their caregivers, indicating that microbiome sharing may play a role in recurrent disease flares. Whole-genome comparisons with high-quality S aureus isolate genomes (n = 55) confirmed significant strain sharing between AD children and their caregivers and AD-specific enrichment of strains expressing enterotoxins Q and K/K2. CONCLUSION: Our results highlight the distinctive skin microbiome features of healthy caregivers for children with AD and support their inclusion in strategies for the treatment of recurrent pediatric AD.

Biosorption of Cr(VI) Using Cellulose Nanocrystals Isolated from the Waterless Pulping of Waste Cotton Cloths with Supercritical CO2: Isothermal, Kinetics, and Thermodynamics Studies.

In the present study, supercritical carbon dioxide (scCO2) was utilized as a waterless pulping for the isolation of cellulose nanocrystals (CNCs) from waste cotton cloths (WCCs). The isolation of CNCs from the scCO2-treated WCCs' fiber was carried out using sulphuric acid hydrolysis. The morphological and physicochemical properties analyses showed that the CNCs isolated from the WCCs had a rod-like structure, porous surface, were crystalline, and had a length of 100.03 ± 1.15 nm and a width of 7.92 ± 0.53 nm. Moreover, CNCs isolated from WCCs had a large specific surface area and a negative surface area with uniform nano-size particles. The CNCs isolated from WCCs were utilized as an adsorbent for the hexavalent chromium [Cr(VI)] removal from aqueous solution with varying parameters, such as treatment time, adsorbent doses, pH, and temperature. It was found that the CNCs isolated from the WCCs were a bio-sorbent for the Cr(VI) removal. The maximum Cr(VI) removal was determined to be 96.97% at pH 2, 1.5 g/L of adsorbent doses, the temperature of 60 °C, and the treatment time of 30 min. The adsorption behavior of CNCs for Cr(VI) removal was determined using isothermal, kinetics, and thermodynamics properties analyses. The findings of the present study revealed that CNCs isolated from the WCCs could be utilized as a bio-sorbent for Cr(VI) removal.

Influence of Fresh Palm Fruit Sterilization in the Production of Carotenoid-Rich Virgin Palm Oil.

Palm oil is known to be rich in carotenoids and other phytonutrients. However, the carotenoids and phytonutrients degrade due to high heat sterilization of oil palm fruits. The present study was conducted to produce carotenoid-rich virgin palm oil (VPO) using cold-press extraction. Herein, the influence of sterilization of oil palm fresh fruits in the production of cold-pressed VPO was determined with varying sterilization temperatures, times, and amounts of palm fruits in sterilization. The experimental sterilization conditions were optimized using response surface methodology (RSM) based on the maximum VPO yield and minimum FFAs in cold-pressed VPO. The optimal sterilization experimental conditions of oil palm fruits were determined to be a sterilization temperature of 62 °C, a time of 90 min, and an amount of oil palm fruits of 8 kg. Under these experimental conditions, the maximum cold-pressed VPO yield and the minimal content of free fatty acids (FFAs) obtained were 27.94 wt.% and 1.32 wt.%, respectively. Several analytic methods were employed to determine cold-pressed VPO quality and fatty acids compositions and compared with the crude palm oil. It was found that cold-pressed VPO contains higher carotenoids (708 mg/g) and unsaturated fatty acids compared with the carotenoid (343 mg/g) and fatty acid compositions in CPO. The findings of the present study reveal that the sterilization temperature potentially influences the carotenoid and nutrient contents in VPO; therefore, the optimization of the sterilization conditions is crucial to producing carotenoid- and phytonutrient-rich VPO.

Developing a JAK Inhibitor for Targeted Local Delivery: Ruxolitinib Cream.

Named after the two-faced Roman god of doorways, Janus kinases (JAKs) represent a class of tyrosine kinases. The JAK signaling pathway is pivotal for the downstream signaling of inflammatory cytokines, including interleukins, interferons, and multiple growth factors. This article provides an overview of the JAK pathway and signaling, its significance in immune-mediated dermatologic diseases and the development of a targeted, localized option of a selective JAK inhibitor, ruxolitinib cream. In the early 1990s, various discovery and clinical development programs were initiated to explore pharmaceutical inhibition of the JAK-STAT pathway. Incyte Corporation launched a strategy to identify molecules suitable for both topical as well as oral delivery. Ruxolitinib was designed as a molecule with low nanomolar potency selective for JAK1 and 2 enzymes, but without significant inhibition of non-JAK kinases, as well as physicochemical properties for both topical and oral administration. An oil-in-water emulsified ruxolitinib cream formulation was developed for topical application and was studied in multiple immune-mediated dermatologic diseases including psoriasis, alopecia areata, atopic dermatitis and vitiligo. Ruxolitinib cream represents a novel, JAK1/2 selective therapy that can be delivered directly to the skin to treat a number of cytokine-driven, inflammatory dermatoses.

Recycling Waste Cotton Cloths for the Isolation of Cellulose Nanocrystals: A Sustainable Approach.

There is an interest in the sustainable utilization of waste cotton cloths because of their enormous volume of generation and high cellulose content. Waste cotton cloths generated are disposed of in a landfill, which causes environmental pollution and leads to the waste of useful resources. In the present study, cellulose nanocrystals (CNCs) were isolated from waste cotton cloths collected from a landfill. The waste cotton cloths collected from the landfill were sterilized and cleaned using supercritical CO2 (scCO2) technology. The cellulose was extracted from scCO2-treated waste cotton cloths using alkaline pulping and bleaching processes. Subsequently, the CNCs were isolated using the H2SO4 hydrolysis of cellulose. The isolated CNCs were analyzed to determine the morphological, chemical, thermal, and physical properties with various analytical methods, including attenuated total reflection-Fourier transform-infrared spectroscopy (ATR-FTIR), field-emission scanning electron microscopy (FE-SEM), energy-filtered transmission electron microscopy (EF-TEM), X-ray diffraction (XRD), thermogravimetric analysis (TGA), and differential scanning calorimetry (DSC). The results showed that the isolated CNCs had a needle-like structure with a length and diameter of 10-30 and 2-6 nm, respectively, and an aspect ratio of 5-15, respectively. Additionally, the isolated CNCs had a high crystallinity index with a good thermal stability. The findings of the present study revealed the potential of recycling waste cotton cloths to produce a value-added product.

Design and Implementation of Fast Spoken Foul Language Recognition with Different End-to-End Deep Neural Network Architectures.

Given the excessive foul language identified in audio and video files and the detrimental consequences to an individual's character and behaviour, content censorship is crucial to filter profanities from young viewers with higher exposure to uncensored content. Although manual detection and censorship were implemented, the methods proved tedious. Inevitably, misidentifications involving foul language owing to human weariness and the low performance in human visual systems concerning long screening time occurred. As such, this paper proposed an intelligent system for foul language censorship through a mechanized and strong detection method using advanced deep Convolutional Neural Networks (CNNs) and Recurrent Neural Networks (RNNs) through Long Short-Term Memory (LSTM) cells. Data on foul language were collected, annotated, augmented, and analysed for the development and evaluation of both CNN and RNN configurations. Hence, the results indicated the feasibility of the suggested systems by reporting a high volume of curse word identifications with only 2.53% to 5.92% of False Negative Rate (FNR). The proposed system outperformed state-of-the-art pre-trained neural networks on the novel foul language dataset and proved to reduce the computational cost with minimal trainable parameters.

Treatment of Palm Oil Refinery Effluent Using Tannin as a Polymeric Coagulant: Isotherm, Kinetics, and Thermodynamics Analyses.

The refining of the crude palm oil (CPO) generates the palm oil refinery effluent (PORE). The presence of high contents of biochemical oxygen demand (BOD), chemical oxygen demand (COD), turbidity, and suspended solids (SS) in PORE encourages the determination of an effective treatment process to minimize the environmental pollution and preserve aquatic life. In the present study, a biodegradable natural polymer, namely tannin, was utilized as a coagulant to treat PORE. The coagulation experiment was conducted using a jar test apparatus. The tannin coagulation efficiency was evaluated based on the BOD, COD, turbidity, and SS removal from PORE by varying the tannin dose (50-300 mg/L), pH (pH 4-10), treatment time (15-90 min), and sedimentation time (15-90 min). It was found that the maximum removal of BOD, COD, turbidity, and SS was 97.62%, 88.89%, 93.01%, and 90.21%, respectively, at pH 6, a tannin dose of 200 mg/L, 60 min of coagulation time, and 60 min of sedimentation time. Analyses of isotherm models revealed that the Freundlich isotherm model was well fitted with the coagulation study. Kinetics studies show that the pseudo-second-order kinetics model was the well-fitted kinetics model for the BOD, COD, turbidity, and SS removal from PORE using tannin as a polymeric coagulant. The determination of thermodynamics parameters analyses revealed that BOD, COD, turbidity, and SS removal from PORE was spontaneous, exothermic, and chemical in nature. The finding of the present study shows that tannin as a natural polymeric coagulant would be utilized in PORE treatment to avoid toxic sludge generation.

Itacitinib (INCB039110), a JAK1 Inhibitor, Reduces Cytokines Associated with Cytokine Release Syndrome Induced by CAR T-cell Therapy.

PURPOSE: T cells engineered to express a chimeric antigen receptor (CAR) are a promising cancer immunotherapy. Such targeted therapies have shown long-term relapse-free survival in patients with B-cell leukemia and lymphoma. However, cytokine release syndrome (CRS) represents a serious, potentially life-threatening side effect often associated with CAR T-cell therapy. CRS manifests as a rapid (hyper)immune reaction driven by excessive inflammatory cytokine release, including IFNγ and IL6. EXPERIMENTAL DESIGN: Many cytokines implicated in CRS are known to signal through the JAK-STAT pathway. Here we study the effect of blocking JAK pathway signaling on CAR T-cell proliferation, antitumor activity, and cytokine levels in in vitro and in vivo models. RESULTS: We report that itacitinib, a potent, selective JAK1 inhibitor, was able to significantly and dose-dependently reduce levels of multiple cytokines implicated in CRS in several in vitro and in vivo models. Importantly, we also report that at clinically relevant doses that mimic human JAK1 pharmacologic inhibition, itacitinib did not significantly inhibit proliferation or antitumor killing capacity of three different human CAR T-cell constructs (GD2, EGFR, and CD19). Finally, in an in vivo model, antitumor activity of CD19-CAR T cells adoptively transferred into CD19+ tumor-bearing immunodeficient animals was unabated by oral itacitinib treatment. CONCLUSIONS: Together, these data suggest that itacitinib has potential as a prophylactic agent for the prevention of CAR T cell-induced CRS, and a phase II clinical trial of itacitinib for prevention of CRS induced by CAR T-cell therapy has been initiated (NCT04071366).

Characterization of Polyethylene Branching by Thermal Analysis-Photoionization Mass Spectrometry.

The rising demand for more and more specialized polyethylene represents a challenge for synthesis and analysis. The desired properties are dependent on the structure, but its elucidation is still intricate. For this purpose, we applied thermal analysis hyphenated to single photon ionization mass spectrometry (STA-SPI-MS). The melting and pyrolysis behavior of different types of polyethylene were tracked by DSC and mass loss. Crystallinity and melting point give hints about the branching but are also influenced by the molecular weight distribution. The evolving gas analysis patterns obtained by SPI-MS however, contain specific molecular information about the samples. Shifts in the summed spectra, which can be clearly observed with our technique, result from differently favored degradation reactions due to the respective structure. Pyrolysis gas chromatography mass spectrometry (Py-GC-EI-MS) was used to support the assignment of pyrolysis products. Principal component analysis was successfully applied to reduce the complexity of data and find suitable markers. The obtained grouping is based on the molecular fingerprint of the samples and is strongly influenced by short-chain branching. Short and medium alkenes and dienes have the strongest impact on the first four principal components. Thus, two marker ratios could be defined, which also give a comprehensible and robust grouping. Butene and pentene were the most abundant signals in our set of samples. With STA-PI-MS, a broad range of pyrolysis products can be measured at the same time, possibly extending the range for quantifiable short-chain branches to more than six carbon atoms for PE. Unfortunately, no clear trend between long-chain branching and any grouping was observed. The quite universal and soft single photon ionization enables access to many different compound classes and hence other polymers can be studied.

A blood RNA transcript signature for TB exposure in household contacts.

BACKGROUND: Current tools for diagnosing latent TB infection (LTBI) detect immunological memory of past exposure but are unable to determine whether exposure is recent. We sought to identify a whole-blood transcriptome signature of recent TB exposure. METHODS: We studied household contacts of TB patients; healthy volunteers without recent history of TB exposure; and patients with active TB. We performed whole-blood RNA sequencing (in all), an interferon gamma release assay (IGRA; in contacts and healthy controls) and PET/MRI lung scans (in contacts only). We evaluated differentially-expressed genes in household contacts (log2 fold change ≥1 versus healthy controls; false-discovery rate < 0.05); compared these to differentially-expressed genes seen in the active TB group; and assessed the association of a composite gene expression score to independent exposure/treatment/immunological variables. RESULTS: There were 186 differentially-expressed genes in household contacts (n = 26, age 22-66, 46% male) compared with healthy controls (n = 5, age 29-38, 100% male). Of these genes, 141 (76%) were also differentially expressed in active TB (n = 14, age 27-69, 71% male). The exposure signature included genes from inflammatory response, type I interferon signalling and neutrophil-mediated immunity pathways; and genes such as BATF2 and SCARF1 known to be associated with incipient TB. The composite gene-expression score was higher in IGRA-positive contacts (P = 0.04) but not related to time from exposure, isoniazid prophylaxis, or abnormalities on PET/MRI (all P > 0.19). CONCLUSIONS: Transcriptomics can detect TB exposure and, with further development, may be an approach of value for epidemiological research and targeting public health interventions.

Gene expression responses to anti-tuberculous drugs in a whole blood model.

BACKGROUND: There is a need for better tools to evaluate new or repurposed TB drugs. The whole blood bactericidal activity (WBA) assay has been advocated for this purpose. We investigated whether transcriptional responses in the WBA assay resemble TB responses in vivo, and whether the approach might additionally reveal mechanisms of action. RESULTS: 1422 of 1798 (79%) of differentially expressed genes in WBA incubated with the standard combination of rifampicin, isoniazid, pyrazinamide and ethambutol were also expressed in sputum (P < 0.0001) obtained from patients taking the same combination of drugs; these comprised well-established treatment-response genes. Gene expression profiles in WBA incubated with the standard drugs individually, or with moxifloxacin or faropenem (with amoxicillin and clavulanic acid) clustered by individual drug exposure. Distinct pathways were detected for individual drugs, although only with isoniazid did these relate to known mechanisms of drug action. CONCLUSIONS: Substantial agreement between whole blood cultures and sputum and the ability to differentiate individual drugs suggest that transcriptomics may add value to the whole blood assay for evaluating new TB drugs.

Isolation and Characterization of Magnetic Oil Palm Empty Fruits Bunch Cellulose Nanofiber Composite as a Bio-Sorbent for Cu(II) and Cr(VI) Removal.

In the present study, magnetic oil palm empty fruits bunch cellulose nanofiber (M-OPEFB-CNF) composite was isolated by sol-gel method using cellulose nanofiber (CNF) obtained from oil palm empty fruits bunch (OPEFB) and Fe3O4 as magnetite. Several analytical methods were utilized to characterize the mechanical, chemical, thermal, and morphological properties of the isolated CNF and M-OPEFB-CNF. Subsequently, the isolated M-OPEFB-CNF composite was utilized for the adsorption of Cr(VI) and Cu(II) from aqueous solution with varying parameters, such as pH, adsorbent doses, treatment time, and temperature. Results showed that the M-OPEFB-CNF as an effective bio-sorbent for the removal of Cu(II) and Cr(VI) from aqueous solution. The adsorption isotherm modeling revealed that the Freundlich equation better describes the adsorption of Cu(II) and Cr(VI) on M-OPEFB-CNF composite. The kinetics studies revealed the pseudo-second-order kinetics model was a better-described kinetics model for the removal of Cu(II) and Cr(VI) using M-OPEFB-CNF composite as bio-sorbent. The findings of the present study showed that the M-OPEFB-CNF composite has the potential to be utilized as a bio-sorbent for heavy metals removal.

Paraffin-Inert Atmospheric Solid Analysis Probe: A Fast and Easy Approach To Characterize Extremely Air-Sensitive Organometallic Complexes by Mass Spectrometry.

Rational characterization of most organometallic compounds is hampered by their high reactivity, in particular, toward oxygen and water. Mass spectrometry experiments require physical introduction of the sample in the ionization source. So, the main challenge is to transfer air-sensitive organometallic compounds from inert atmosphere to the ionization source. In this aim, we have developed an easy technique that allows the analysis of air-sensitive compounds using the atmospheric solid analysis probe (ASAP). This method consists of a glass capillary filled with the sample (solid or liquid) and sealed by a paraffin plug to maintain the inert sample until the ionization process. It is illustrated through the structural characterization of a new highly air-sensitive dinuclear zirconium complex supported by an original switchable stilbene platform.

Erratum: Author Correction: Mimicking immune signatures of flavivirus infection with targeted adjuvants improves dengue subunit vaccine immunogenicity.

[This corrects the article DOI: 10.1038/s41541-019-0119-3.].

Surface effects on a photochromic spin-crossover iron(ii) molecular switch adsorbed on highly oriented pyrolytic graphite.

Thin films of an iron(ii) complex with a photochromic diarylethene-based ligand and featuring a spin-crossover behaviour have been grown by sublimation in ultra-high vacuum on highly oriented pyrolytic graphite and spectroscopically characterized through high-resolution X-ray and ultraviolet photoemission, as well as via X-ray absorption. Temperature-dependent studies demonstrated that the thermally induced spin-crossover is preserved at a sub-monolayer (0.7 ML) coverage. Although the photochromic ligand ad hoc integrated into the complex allows the photo-switching of the spin state of the complex at room temperature both in bulk and for a thick film on highly oriented pyrolytic graphite, this photomagnetic effect is not observed in sub-monolayer deposits. Ab initio calculations justify this behaviour as the result of specific adsorbate-substrate interactions leading to the stabilization of the photoinactive form of the diarylethene ligand over photoactive one on the surface.

Study of the Photoswitching of a Fe(II) Chiral Complex through Linear and Nonlinear Ultrafast Spectroscopy.

Photoswitching the physical properties of molecular systems opens large possibilities for driving materials far from equilibrium toward new states. Moreover, ultrashort pulses of light make it possible to induce and to record photoswitching on a very short time scale, opening the way to fascinating new functionalities. Among molecular materials, Fe(II) complexes exhibit an ultrafast spin-state transition during which the spin state of the complex switches from a low spin state (LS, S = 0) to a high spin state (HS, S = 2). The latter process is remarkable: It takes place within ∼100 fs with a quantum efficiency of ∼100%. Moreover, the spin-state switching induces an important shift of the broad metal-to-ligand absorption band of the complex, and it results in large modifications of the physical and chemical properties of the compounds. But because most of the Fe(II) complexes crystallize in centrosymmetric space groups, this prevents them from exhibiting piezoelectric, ferroelectric, as well as second-order nonlinear optical properties such as second-harmonic generation (SHG). This considerably limits their potential applications. We have recently synthesized [Fe(phen)3] [Δ-As2(tartrate)2] chiral complexes that crystallize in a noncentrosymmetric 32 space group. Hereafter, upon the excitation of a thin film of these complexes by a femtosecond laser pulse and performing simultaneously transient absorption (TRA) and time-resolved SHG (TRSH) measurements, we have recorded the ultrafast LS to HS switching. Whereas a single TRA measurement gives only partial information, we demonstrate that TRSH readily reveals the different mechanisms in play during the HS-to-LS state relaxation. Moreover, a simple model makes it possible to evaluate the relaxation times as well as the hyperpolarizabilities of the different excited states through which the system travels during the spin-state transition.