An investigation into the recovery of ignitable liquid residues from entomological samples using solid-phase microextraction

University of Technology Sydney. Faculty of Science.The analysis of fire debris can indicate the presence of an ignitable liquid, but the volatility of these substances means that the likelihood of detecting them diminishes over time. It is proposed in this thesis that when a scene contains burnt human remains, entomological samples can be analysed for the detection of ignitable liquids, as an alternative to fire debris. It is hypothesised that a larva’s ability to invade areas protected from the external environment, such as the natural body openings of cadavers, and accumulate substances present in the tissue in which they are feeding, will extend the period in which ignitable liquids can be detected. In small-scale experiments conducted under controlled laboratory conditions, petrol and kerosene were detected in larvae of the blow fly Lucilia cuprina, (Wiedemann) (Diptera: Calliphoridae) that had been fed on meat burnt using these ignitable liquids. Four sample sets of meat, each with six replicates (24 meat samples in total) were prepared. The first and second sets were burnt using petrol and kerosene, respectively. The final two sets were control groups. Six larvae were collected daily from each of the 24 meat samples for a period of five days. Once the adults had emerged, six adults and six puparia were also collected from each meat sample. All of the entomological samples collected were analysed using solid-phase microextraction gas chromatography-mass spectrometry (SPME-GC-MS). It was found that larvae of the blow fly Lucilia cuprina can be used in a small-scale setting to detect both petrol and kerosene from burnt substrates for at least five days. Positive results for the ignitable liquids of interest were also obtained for a limited number of adult flies and puparia. Given these findings, further research was conducted using a more realistic experiment (conducted in duplicate) that mirrored a casework scenario more closely. Sample sets identical to those in the small-scale experiments were prepared using 24 piglets, each approximately 1.39 kg in weight, instead of the meat samples. These piglets were placed a minimum of 51 m apart at the Holsworthy Military Area in New South Wales, Australia, for three days. After this time, the piglets were transferred to a controlled laboratory. It was found during these fieldwork experiments that petrol and kerosene could be successfully detected in larvae for as long as eight days, and in the adult and puparia samples for at least one month. These findings confirm the significant advantage of using entomological samples as an alternative to fire debris, in that they extend the period available for sampling volatile ignitable liquids by at least one month. In particular, puparia can withstand changing climatic conditions, and unlike the larvae and adults, are immobile and hence could be found close to human remains even after considerable time has elapsed

Reconstructing tephra fall deposits via ensemble-based data assimilation techniques

In recent years, there has been a growing interest in ensemble approaches for modelling the atmospheric transport of volcanic aerosol, ash, and lapilli (tephra). The development of such techniques enables the exploration of novel methods for incorporating real observations into tephra dispersal models. However, traditional data assimilation algorithms, including ensemble Kalman filter (EnKF) methods, can yield suboptimal state estimates for positive-definite variables such as those related to volcanic aerosols and tephra deposits. This study proposes two new ensemble-based data assimilation techniques for semi-positive-definite variables with highly skewed uncertainty distributions, including aerosol concentrations and tephra deposit mass loading: the Gaussian with non-negative constraints (GNC) and gamma inverse-gamma (GIG) methods. The proposed methods are applied to reconstruct the tephra fallout deposit resulting from the 2015 Calbuco eruption using an ensemble of 256 runs performed with the FALL3D dispersal model. An assessment of the methodologies is conducted considering two independent datasets of deposit thickness measurements: an assimilation dataset and a validation dataset. Different evaluation metrics (e.g. RMSE, MBE, and SMAPE) are computed for the validation dataset, and the results are compared to two references: the ensemble prior mean and the EnKF analysis. Results show that the assimilation leads to a significant improvement over the first-guess results obtained from the simple ensemble forecast. The evidence from this study suggests that the GNC method was the most skilful approach and represents a promising alternative for assimilation of volcanic fallout data. The spatial distributions of the tephra fallout deposit thickness and volume according to the GNC analysis are in good agreement with estimations based on field measurements and isopach maps reported in previous studies. On the other hand, although it is an interesting approach, the GIG method failed to improve the EnKF analysis.</p

Exploiting Human NK Cells in Tumor Therapy

NK cells play an important role in the innate defenses against tumor growth and metastases. Human NK cell activation and function are regulated by an array of HLA class I-specific inhibitory receptors and activating receptors recognizing ligands expressed de novo on tumor or virus-infected cells. NK cells have been exploited in immunotherapy of cancer, including: (1) the in vivo infusion of IL-2 or IL-15, cytokines inducing activation and proliferation of NK cells that are frequently impaired in cancer patients. Nonetheless, the significant toxicity experienced, primarily with IL-2, limited their use except for combination therapies, e.g., IL-15 with checkpoint inhibitors; (2) the adoptive immunotherapy with cytokine-induced NK cells had effect on some melanoma metastases (lung), while other localizations were not affected; (3) a remarkable evolution of adoptive cell therapy is represented by NK cells engineered with CAR-targeting tumor antigens (CAR-NK). CAR-NK cells complement CAR-T cells as they do not cause GvHD and may be obtained from unrelated donors. Accordingly, CAR-NK cells may represent an \u201coff-the-shelf\u201d tool, readily available for effective tumor therapy; (4) the efficacy of adoptive cell therapy in cancer is also witnessed by the \u3b1\u3b2T cell- and B cell-depleted haploidentical HSC transplantation in which the infusion of donor NK cells and \u3b3\u3b4T cells, together with HSC, sharply reduces leukemia relapses and infections; (5) a true revolution in tumor therapy is the use of mAbs targeting checkpoint inhibitors including PD-1, CTLA-4, the HLA class I-specific KIR, and NKG2A. Since PD-1 is expressed not only by tumor-associated T cells but also by NK cells, its blocking might unleash NK cells playing a crucial effector role against HLA class I-deficient tumors that are undetectable by T cells

Eruption plumes extended more than 30 km in altitude in both phases of the Millennium eruption of Paektu (Changbaishan) volcano

The Millennium Eruption of Paektu volcano, on the border of China and North Korea, generated tephra deposits that extend >1000 km from the vent, making it one of the largest eruptions in historical times. Based on observed thicknesses and compositions of the deposits, the widespread tephra dispersal is attributed to two eruption phases fuelled by chemically distinct magmas that produced both pyroclastic flows and fallout deposits. We used an ensemble-based method with a dual step inversion, in combination with the FALL3D atmospheric tephra transport model, to constrain these two different phases. The volume of the two distinct phases has been calculated. The results indicate that about 3-16 km3 (with a best estimate of 7.2 km3) and 4-20 km3 (with a best estimate of 9.3 km3) of magma were erupted during the comendite and trachyte phases of the eruption, respectively. Eruption rates of up to 4 × 108 kg/s generated plumes that extended 30-40 km up into the stratosphere during each phase

NK cells and other innate lymphoid cells in hematopoietic stem cell transplantation

Natural killer (NK) cells play a major role in the T-cell depleted haploidentical hematopoietic stem cell transplantation (haplo-HSCT) to cure high-risk leukemias. NK cells belong to the expanding family of innate lymphoid cells (ILCs). At variance with NK cells, the other ILC populations (ILC1/2/3) are non-cytolytic, while they secrete different patterns of cytokines. ILCs provide host defenses against viruses, bacteria, and parasites, drive lymphoid organogenesis, and contribute to tissue remodeling. In haplo-HSCT patients, the extensive T-cell depletion is required to prevent graft-versus-host disease (GvHD) but increases risks of developing a wide range of life-threatening infections. However, these patients may rely on innate defenses that are reconstituted more rapidly than the adaptive ones. In this context, ILCs may represent important players in the early phases following transplantation. They may contribute to tissue homeostasis/remodeling and lymphoid tissue reconstitution. While the reconstitution of NK cell repertoire and its role in haplo-HSCT have been largely investigated, little information is available on ILCs. Of note, CD34+ cells isolated from different sources of HSC may differentiate in vitro toward various ILC subsets. Moreover, cytokines released from leukemia blasts (e.g., IL-1β) may alter the proportions of NK cells and ILC3, suggesting the possibility that leukemia may skew the ILC repertoire. Further studies are required to define the timing of ILC development and their potential protective role after HSCT

Killer Ig-like receptors (kirs). their role in nk cell modulation and developments leading to their clinical exploitation

Natural killer (NK) cells contribute to the first line of defense against viruses and to the control of tumor growth and metastasis spread. The discovery of HLA class I specific inhibitory receptors, primarily of killer Ig-like receptors (KIRs), and of activating receptors has been fundamental to unravel NK cell function and the molecular mechanisms of tumor cell killing. Stemmed from the seminal discoveries in early ‘90s, in which Alessandro Moretta was the major actor, an extraordinary amount of research on KIR specificity, genetics, polymorphism, and repertoire has followed. These basic notions on NK cells and their receptors have been successfully translated to clinical applications, primarily to the haploidentical hematopoietic stem cell transplantation to cure otherwise fatal leukemia in patients with no HLA compatible donors. The finding that NK cells may express the PD-1 inhibitory checkpoint, particularly in cancer patients, may allow understanding how anti-PD-1 therapy could function also in case of HLA class Ineg tumors, usually susceptible to NK-mediated killing. This, together with the synergy of therapeutic anti-checkpoint monoclonal antibodies, including those directed against NKG2A or KIRs, emerging in recent or ongoing studies, opened new solid perspectives in cancer therapy

Inhibitory 2B4 contributes to NK cell education and immunological derangements in XLP1 patients

X-linked lymphoproliferative disease 1 (XLP1) is an inherited immunodeficiency, caused by mutations in SH2D1A encoding Signaling Lymphocyte Activation Molecule (SLAM)-associated protein (SAP). In XLP1, 2B4, upon engagement with CD48, has inhibitory instead of activating function. This causes a selective inability of cytotoxic effectors to kill EBV-infected cells, with dramatic clinical sequelae. Here, we investigated the NK cell education in XLP1, upon characterization of killer Ig-like receptor (KIR)/KIR-L genotype and phenotypic repertoire of self-HLA class I specific inhibitory NK receptors (self-iNKRs). We also analyzed NK-cell cytotoxicity against CD48+ or CD48− KIR-ligand matched or autologous hematopoietic cells in XLP1 patients and healthy controls. XLP1 NK cells may show a defective phenotypic repertoire with substantial proportion of cells lacking self-iNKR. These NK cells are cytotoxic and the inhibitory 2B4/CD48 pathway plays a major role to prevent killing of CD48+ EBV-transformed B cells and M1 macrophages. Importantly, self-iNKR defective NK cells kill CD48− targets, such as mature DCs. Self-iNKR− NK cells in XLP1 patients are functional even in resting conditions, suggesting a role of the inhibitory 2B4/CD48 pathway in the education process during NK-cell maturation. Killing of autologous mature DC by self-iNKR defective XLP1 NK cells may impair adaptive responses, further exacerbating the patients’ immune defect