Respiratory infection remains a leading cause of morbidity and death in severely immunocompromised febrile neutropenic haematology patients, despite the introduction of numerous prophylactic strategies and advances in diagnosis and treatment. Prognosis is improved if an organism can be isolated and specific therapy commenced as soon as possible. Current practice in this population group is to commence empirical antibiotics and perform flexible bronchoscopy (FB) if temperature does not settle or after patients develop clinical or radiological features suggesting a respiratory source. This delay may result in a lower procedural diagnostic yield due to prior or prolonged anti-microbial treatment, and increased risk of respiratory compromise and procedural complications due to advanced respiratory infections. We hypothesised that proceeding to FB as early as possible after developing febrile neutropenia would improve treatment outcomes. With this randomised, prospective trial, we aim to further define the role of FB with reference to optimal timing of the procedure and its impact on diagnostic yield, future management and complication rate. We also aim to analyse the impact of proven infection on the cytokine profile of immunocompromised patients. Methods: Patients with acute leukaemia, allogeneic bone marrow transplantation or chronic lymphocytic leukaemia (CLL) being treated with Fludarabine/ Mabthera without an obvious non-respiratory source of infection were prospectively randomised into early bronchoscopy or conventional management groups at onset of febrile neutropenia. Bronchoalveolar lavage (BAL) fluid chemokine levels (IP-10, RANTES, MIG, IL-8, MCP-1) were measured using a human Chemokine cytometric bead array (CBA) kit. Results: Thirty-one episodes of febrile neutropenia in 29 patients were analysed; 17 conventional and 14 early. There was an increased yield in fungal growth in the early bronchoscopy group, which was not predicted by prior clinical or radiological changes. However, this had no impact on clinical management in the short-term due to the delayed growth. Overall diagnostic yield was not significantly different between the two groups. Procedural complication rate was negligible overall and there was no difference associated with either group. IP-10 and MIG were significantly lower in those patients who had a fungal pathogen isolated, compared with those study patients who did not (175.17 vs 1157.8, p=0.03, 30.33 vs 247.8, p=0.03 respectively). IP-10 levels were higher in the conventional than early group (1253.0 vs 261.14, p = 0.035) and the study population had higher MCP-1 (734 vs 2.83, p=0.006) and IL-8 levels (606.9 vs 14.25, p=0.00655) than normal controls. Those cases with fungal infection had higher mean MCP-1, RANTES and IL-8 levels than in normal controls (844.0 vs 2.83, p=0.007; 17.5 vs 2.1, p=0.03; 156.0 vs 14.25, p=0.004). Conclusions: Early bronchoscopy as a component of the septic screen in febrile neutropenic patients was feasible and safe. A significant difference in fungal yield was seen in the early bronchoscopy group compared to conventional methods, with a negligible complication rate, but this did not result in a change in immediate clinical management or outcomes. / Thesis (M.Clin.Sc.) - University of Adelaide, School of Medicine, 2009

Abstract Experiments in this thesis were conducted to investigate the possibility of bestowing lambs with increased resistance to gastrointestinal parasites through maternal protein and copper supplementation. Reproductive outcomes such as birth weight, haematological parameters and faecal egg counts were used as indices of possible foetal programming. This thesis involved 5 experiments. The first three experiments were done using rats as a preliminary study animal on account of their short generational intervals and high fecundity. The final two experiments involved Merino sheep. The first experiment in Chapter 4 investigated the most optimum larval dose to use in order to elicit a measurable immune response. Weaned offspring were infected with a rat nematode, Nippostrongylus brasiliensis and their response measured by faecal egg counts, parasite recovery from intestines at sacrifice, spleen weights and leucocyte numbers, especially manifested as eosinophilia. There was no significant difference in parasite rejection for rats infected with 1000 larvae/rat. When rats were infected with variable larval doses to determine the optimum dose rate, eosinophilia and spleen weight were significantly increased as dose rate increased from 500 L3 to 2,000 larvae. Based on these results, it was decided to use 1,000 larvae for each rat in Chapter 6. The experiment in Chapter 5 involved feeding diets with 5 graded concentrations of copper (Cu) ranging from deficient (1 ppm diet) to high (16 ppm diet). Rats were fed for 4 weeks before mating after which half of them were sacrificed to determine liver Cu concentrations and haematological parameters. The rest were mated and maintained on their respective Cu diets into the second trimester of pregnancy. Pregnant females were sacrificed on approximately gestational Day 10 to recover foetuses and determine the incidence of foetal defects, foetal Cu status as well as maternal liver copper status. It was determined that most morphological defects occurred for the 1 ppm foetuses and both 2 ppm and 4 ppm had similar incidences of brain enlargements. The 16 ppm copper diet was excessive evidenced by reduced liver iron status and erythrocyte counts to similar levels as for 2 ppm rats although it had no adverse effect on foetal development. Significant differences were found for liver Cu status, erythrocyte counts and spleen weights due to the copper diets. A deficient copper diet containing 1 ppm Cu (LC) and an adequate diet containing 8 ppm Cu (SC) were used for the last rat experiment in Chapter 6 which was funded by the Science and Innovation Award. The LC diets were fed for 4 weeks prior to mating. Rats were then fed LC throughout pregnancy, for the 1st trimester only or for the 1st and 2nd trimesters. Other pregnant females were fed the SC diet throughout pregnancy. Offspring were challenged with 1000 L3 N. brasiliensis and their immune responses measured. Copper deficiency at variable stages of prenatal development caused significant postnatal mortalities but had no effect on response to parasite resistance. However, significant parasite and sex effects were found for parameters such as spleen weight, eosinophilia and weight loss during infection. The foetal brain enlargement caused by the deficient 1 ppm Cu diet was determined to be reversible in vivo upon exposure to a normal 8 ppm Cu diet during gestation. Chapter 7 involved Merino ewes which were fed either a high protein diet (21%) or adequate protein diet (12%) during the first 2 trimesters of pregnancy. Production parameters measured included pregnancy weight gain, fleece yield, protein content in milk as well as birth weight of lambs but none were significantly different. After weaning, the lambs were experimentally infected with 10,000 Haemonchus contortus larvae. Barber’s pole worm is responsible for millions of dollars in production losses in the sheep industry. Responses measured were eosinophilia, faecal egg count, anaemia (PCV) and weight gain/loss during the infection period. No significant differences were found for any parameter tested except for a parasite effect on erythrocyte numbers and PCV. In Chapter 8 Merino ewes were used which were mildly Cu deficient due to grazing on pasture that was copper deficient. Control ewes were supplemented with copper oxide wire particles at mating and mid-pregnancy. The rest of the experiment was the same as for Chapter 7 in terms of Barber’s pole worm larval dose. There were no significant differences in birth weight, weaning weight or ewe fleece weights due to copper status. There were no differences in parasite resistance in the lambs due to maternal Cu status measured by live weights, eosinophil concentrations or faecal egg counts. In conclusion, foetal programming by maternal nutritional supplements for postnatal parasite resistance appears to be impossible. It may be that if a different organ was targeted, such as the spleen, the results would have been different. The thymus appears to be non-programmable during foetal development in rats and sheep. However, it was a worthwhile attempt at conferring resistance to parasites in lambs due to the urgency in combating the global problem of parasite resistance to anthelmintics and the resultant large economic losses that are experienced by the global sheep industry.

No description available.

No description available.

In patients with aplastic anaemia that respond to immunosuppressive therapy, quantitative, morphological and functional haematologic derangement have been reported. To explain these findings, abnormalities in the marrow stroma or the stem cell have been postulated. To define the relative contribution of each of the latter, the integrity of the bone marrow from sixteen patients that responded to anti-lymphocyte globulin and high dose methyl prednisolone was compared to normal individuals. Bone marrow mononuclear cells were divided into two fractions. From the first, stroma was cultured in aMEM containing 12.5% of both horse and foetal calf serum and 10-5 M hydrocortisone at 37° C in 5% CO2 in 90% humidity. The medium was changed weekly. Upon confluence, these stromal layers were studied morphologically and with cytospin preparations stained with Sudan black, 0 red oil, alkaline and acid phosphatases. The remainder was monocyte and lymphocyte depleted, CD 34+ progenitors were selected with paramagnetic beads and the population morphologically and immunophenotypically defined. To determine the functional status, control or patient CD 34+ progenitors, were suspended for two hours on normal or aplastic stroma for adherence to take place. The non-adhesive fraction was decanted by standardised washing and cultured for fourteen days in the presence of PHA-conditioned medium in the CFU-gm assay. Strama-adherent progenitors were covered with 0.3% agar and cultured for five days. Aggregates with more than twenty cells were scored (CFU-bl). The remaining CD 34+ cells were cultured in the mixed colony assay with combinations of recombinant cytokines belonging to the G protein super-family and the tyrosine kinase group in dose response studies. Light density cells from patients with treated aplasia contained significantly fewer CD 34+ cells than those present in the control suspensions (mean 0.65%, SD 0.35% vs 1.62%, SD 1.4%; p= 0.002). Normal and aplastic stroma became confluent at three and four weeks. There was no difference on the morphology or the cytochemical stains between the two groups. Functionally, aplastic bone marrow stroma supported CFU-bl formation no differently from normal layers. However, CD 34+ precursors from the patients cultured on control stroma resulted in significantly fewer CFU-bl (p= 0.0002,) and CFU-gm (p= 0.0009). This work provides original evidence supporting the reduced clonogenicity of the corresponding populations of CFU-bl from patients with aplasia is unrelated to attachment to the stroma, but intrinsic to the CD 34+ cells. Moreover, this study shows for the first time that exposure of these progenitors to growth factors belonging to the G protein and tyrosine kinase receptor families have defective responses, correctable only at supra physiological concentrations, while effects on combinations containing c-kit ligand, appear preserved. Following immunosuppressive therapy, the bone marrow is repopulated by a hypoproliferative progenitor cell population which responds suboptimally to physiological cytokine stimulation. This suggests that abnormal interactions between receptors and their ligands or alterations in the signal transduction for cell division by the cytokines belonging to the G superfamily lead to suboptimal growth.

A comparative study was performed to evaluate the immunostimulatory effect of yeast and yeast-derived glucan in white prawn Fenneropenaeus indicus (sub-adults of ∼20 gm). Feed with a whole cell biomass of marine yeast Candida sake S165 (CSY) at a concentration of 10% (w/w) and another feed with 0.2% glucan of C. sake S165 (CSG) were used in the study. Fenneropenaeus indicus were fed with these diets for 40 days and subsequently challenged with the white spot syndrome virus (WSSV). Haematological parameters such as the total haemocyte count, phenoloxidase activity, superoxide anion (O2-) level, haemolymph peroxidase level and post-challenge survival against WSSV infection were determined to assess the immune status. In the present experiment, a higher immunity index and post-challenge survival were recorded in shrimps fed with the whole cell yeast diet. The better immunostimulatory performance of the whole cell yeast diet compared with the glucan diet could be attributed to the cellular constituents of yeast including the cell wall glucan, nucleotides, carotenoid pigments and vitamins. Here we observed that whole cell yeast performed better as an immunostimulant than the extracted cell wall glucans. Therefore, the use of yeast biomass in diets, rather than the yeast cell wall extract, glucan, would confer better protection against microbial infection besides reducing the cost of shrimp production.

COVID-19 is an infectious disease caused by the novel coronavirus. It presents as an acute respiratory illness, however, it also affects multiple other organ systems. One such unique manifestation is systemic coagulopathy involving arterial and venous systems. We present a 29-year-old woman with Hodgkin's lymphoma, who was diagnosed with COVID-19 infection prior to initiating chemotherapy. Two months after resolution of symptoms and testing negative for COVID-19, she presented with multiple acute thromboembolic complications of the infection, including bilateral jugular venous thrombosis, right atrial clot and arterial emboli in the brain resulting in cerebrovascular injury. These were thought to be delayed manifestations of the systemic coagulopathy secondary to infection. Also, some of these thromboembolic phenomena occurred while the patient was on anticoagulation, which emphasises the extensive hyperinflammatory state caused by the virus. This case highlights the importance of thromboprophylaxis especially in high-risk patients with this infection.

Non-producing variant of plasma cell disorders with circulating plasma cells is an aggressive variant of plasma cell dyscrasias with relatively poor outcomes. A 75-year-old man was admitted due to anaemia (90 g/L) and thrombocytopenia (9×10 9 /L). Comprehensive metabolic panel showed creatinine of 1.34 mg/dL, total protein of 6 g/dL, and corrected calcium was normal. Peripheral smear review showed 8% circulating atypical plasmacytoid cells. Bone marrow biopsy (BMB) confirmed plasma cell myeloma involving 90%-95% of bone marrow cellularity. Serum protein electrophoresis showed no monoclonal protein. Due to aggressive biology of non-producer variant and outcomes based on circulating plasma cells, he was started on VD-PACE (bortezomib, dexamethasone, cisplatin, doxorubicin, cyclophosphamide and etoposide) chemotherapy. BMB after cycle 1 chemotherapy showed no morphologic, immunophenotypic, or flow cytometric features of a plasma cell neoplasm. Given excellent treatment response cycle 2 was changed to VRD (bortezomib, lenalidomide and dexamethasone). Following two cycles of VRD, he underwent autologous haematopoietic cell transplantation. Day 80 BMB suggested stringent complete response.

No description available.

Acute Myeloid leukemia (AML) is an aggressive haematological malignancy that mainly affects the elderly. Relapse is common and is thought to be due to the presence of chemotherapy resistant leukemic stem cells (LSC). Within the CD34+ disease (>5% of the blast cells expressing CD34) , two subtypes have been identified; an LMPP/GMPlike expanded type and a MPP/CMP-like expanded type, the former is the most common, accounting for around 80% of CD34+ AML. Both the GMP-like and LMPPlike expanded populations show LSC activity. To improve our understanding of the disease and gain better insight in to how to develop treatments, the molecular basis of the disease needs to be investigated. I investigated miRNAs in the GMP/LMPP-like expanded AML. miRNAs are small non-coding RNAs involved in the regulation of mRNA. In recent years miRNAs have been shown to be implicated in many different diseases. To investigate the role miRNAs play in AML, miRNA expression was profiled in leukemic and normal bone marrow. Bioinformatic analysis was then used to examine the different miRNA expression profiles between normal and leukemic marrow. Our study showed that miRNAs are dysregulated in AML. miRNAs from the miR-17-92 and its paralogous cluster miR-106b-92 were amongst the miRNAs to be found down regulated in AML As had been seen previously at an mRNA level, on an miRNA level the LSC populations more closely resembled more mature progenitor populations than HSC and MPP populations, however the LSC populations did display an aberrant stem cell-like miRNA signature.