Hypercalcemia of malignancy: Cancer treatment can be a cause as well.

While the incidence of hypercalcemia of malignancy (HCM) is on the decline, it still occurs in up to 30% of patients with cancer. Immune checkpoint inhibitor (ICI)-related hypercalcemia is becoming increasingly recognized. We describe a case of cemiplimab-induced hypercalcemia in a patient with metastatic squamous cell carcinoma of the earlobe and discuss a management algorithm for HCM. Timely diagnosis and management of HCM is critical for optimal care and the prevention of complications.

Public Health System's Preparedness to Address Polycystic Ovarian Syndrome: A Rapid Assessment Survey of Health-care Providers in India.

BACKGROUND: Polycystic ovarian syndrome (PCOS) is one of the most common endocrine disorders largely affecting women of reproductive age group. OBJECTIVES: This study aimed to understand the Indian public health-care systems' preparedness in addressing PCOS. MATERIALS AND METHODS: A multicentric rapid assessment cross-sectional study was undertaken among 173 health-care providers serving across various public health-care facilities in India. This study was a component of a larger task force study that aimed to estimate the community-based prevalence of PCOS in India. Information on PCOS cases reported that knowledge about PCOS diagnosis, management practices, availability of diagnostic facilities, and drugs was explored. RESULTS: Irregular menstrual cycle was the most commonly reported PCOS symptom. Most of the health-care providers (HCPs) lacked correct knowledge about diagnostic criteria and investigation needed for the diagnosis of PCOS. Diagnostic facilities and drugs were inadequate. However, some facilities had access to investigations through public-private partnerships. Awareness programs on PCOS in the community were negligible, and PCOS cases were not documented. Training HCPs on PCOS along with the availability of specialists and strengthening diagnostic facilities were some major demands from the HCPs. CONCLUSION: Results suggest the need for training HCPs, strengthening infrastructure with good referral linkages, and adequate supply of drugs to help improve PCOS management at public health-care facilities in India. There is a need to develop national technical and operational guidelines to address PCOS using a multidisciplinary approach across all levels of care. Creating demand for services and advocating healthy lifestyles through community awareness can help early diagnosis and prevention of complications.

Complement Inhibitors in the Management of Complement-Mediated Hemolytic Uremic Syndrome and Paroxysmal Nocturnal Hemoglobinuria.

BACKGROUND: Complement-mediated HUS (CM-HUS) and paroxysmal nocturnal hemoglobinuria (PNH) are rare hematologic disorders that cause dysregulation and hyperactivation of the complement system. Historically, treatment of CM-HUS involved plasma exchange (PLEX), often with limited benefit and variable tolerance. Conversely, PNH was treated with supportive care or hemopoietic stem cell transplant. Within the last decade, monoclonal antibody therapies that block terminal complement pathway activation, have emerged as less invasive and more efficacious options for management of both disorders. This manuscript seeks to discuss a relevant clinical case of CM-HUS and the evolving landscape of complement inhibitor therapies for CM-HUS and PNH. AREAS OF UNCERTAINTY: Eculizumab, the first humanized anti-C5 monoclonal antibody, has been the standard of care in treating CM-HUS and PNH for over a decade. Although eculizumab has remained an effective agent, the variability in ease and frequency of administration has remained an obstacle for patients. The development of novel complement inhibitor therapies with longer half-lives, has allowed for changes in frequency and route of administration, thus improving patient QOL. However, there are limited prospective clinical trial data given disease rarity, and limited information on variable infusion frequency and length of treatment. THERAPEUTIC ADVANCES: Recently, there has been a push to formulate complement inhibitors that improve QOL while maintaining efficacy. Ravulizumab, a derivative of eculizumab, was developed to allow for less frequent administration, while remaining efficacious. In addition, the novel oral and subcutaneous therapies, danicopan and crovalimab, respectively, along with pegcetacoplan are currently undergoing active clinical trials, and poised to further reduce treatment burden. CONCLUSION: Complement inhibitor therapies have changed the treatment landscape for CM-HUS and PNH. With a significant emphasis on patient QOL, novel therapies continue to emerge and require an in-depth review of their appropriate use and efficacy in these rare disorders. CLINICAL CASE: A 47-year-old woman with hypertension and hyperlipidemia presented with shortness of breath and was found to have hypertensive emergency in the setting of acute renal failure. Her serum creatinine was 13.9 mg/dL; elevated from 1.43 mg/dL 2 years before. The differential diagnosis for her acute kidney injury (AKI) included infectious, autoimmune, and hematologic processes. Infectious work-up was negative. ADAMTS13 activity level was not low at 72.9%, ruling out thrombotic thrombocytopenic purpura (TTP). Patient underwent a renal biopsy, which revealed acute on chronic thrombotic microangiopathy (TMA). A trial of eculizumab was initiated with concurrent hemodialysis. The diagnosis of CM-HUS was later confirmed by a heterozygous mutation in complement factor I (CFI), resulting in increased membrane attack complex (MAC) cascade activation. The patient was maintained on biweekly eculizumab and was eventually transitioned to ravulizumab infusions as an outpatient. Her renal failure did not recover, and the patient remains on hemodialysis while awaiting kidney transplantation.

Slow Off-Rate Modified Aptamer (SOMAmer) Proteomic Analysis of Patient-Derived Malignant Glioma Identifies Distinct Cellular Proteomes.

Malignant gliomas derive from brain glial cells and represent >75% of primary brain tumors. This includes anaplastic astrocytoma (grade III; AS), the most common and fatal glioblastoma multiforme (grade IV; GBM), and oligodendroglioma (ODG). We have generated patient-derived AS, GBM, and ODG cell models to study disease mechanisms and test patient-centered therapeutic strategies. We have used an aptamer-based high-throughput SOMAscan® 1.3K assay to determine the proteomic profiles of 1307 different analytes. SOMAscan® proteomes of AS and GBM self-organized into closely adjacent proteomes which were clearly distinct from ODG proteomes. GBM self-organized into four proteomic clusters of which SOMAscan® cluster 4 proteome predicted a highly inter-connected proteomic network. Several up- and down-regulated proteins relevant to glioma were successfully validated in GBM cell isolates across different SOMAscan® clusters and in corresponding GBM tissues. Slow off-rate modified aptamer proteomics is an attractive analytical tool for rapid proteomic stratification of different malignant gliomas and identified cluster-specific SOMAscan® signatures and functionalities in patient GBM cells.

Financing health care in Bangladesh: Policy responses and challenges towards achieving universal health coverage.

Bangladesh has attained notable progress in most of the health indicators, but still, health system of the country is suffering badly from poor funding. Issues like burden of out-of-pocket expenditure, low per capita share in health, inadequate service facilities, and financial barriers in reducing malnutrition are being overlooked due to inadequacy and inappropriate utilization of allocated funds. We aimed to review the current status of health care spending in Bangladesh in response to national health policy (NHP) and determine the future challenges towards achieving universal health coverage (UHC). National health policy suggested a substantial increase in budgetary allocation for health care, although government health care expenditures in proportion to total public spending plummeted down from 6.2% to 4.04% in the past 8 years. Overall, 67% of the health care cost is being paid by people, whereas global standard is below 32%. Only one hospital bed is allocated per 1667 people, and 34% of total posts in health sector are vacant due to scarcity of funds. The country is experiencing demographic dividend with a concurrent rise of aged people, but there seems no financial protection schemes for the aged and working age populations. Such situation results in multiple obstacles in achieving financial risk protection as well as UHC. Policy makers must think effectively to develop and adapt systems in order to achieve UHC and ensure health for all.

Serendipitous identification of Pratylenchus curvicauda from the grainbelt of Western Australia.

A Pratylenchus species identified during a survey of Pratylenchus quasitereoides incidence at four locations of the grainbelt of Western Australia is described. Morphological and morphometric features indicated the previously undescribed morphotypes in nematode mixtures encountered were conspecific to P. curvicauda, and were clearly distinguishable from nine common Pratylenchus spp. Typical features of P. curvicauda were its body length (415-540 µm), which was curved to a c-shaped with a maximum body diameter of 20 µm, and the nature of its tail; 34 µm long, 2.8 µm wide at the anus and a typical ventrally arcuate with a round terminus. Sequenced for the first time, the sequences of the partial 18S-ITS1-5.8S-ITS2-partial 28S (80 clones, 14 individual nematodes) and the 28S-D3 (17 clones) regions of the rDNA of P. curvicauda had overall mean distances of 0.013 and 0.085, respectively. Phylogenetic analyses with sequences of both segments of the rDNA clearly showed the P. curvicauda isolates as monophyletic, distinct from ca 40 Pratylenchus species. Notably, it was distinct from Pratylenchus species present in Australia including P. quasitereoides and a Western Australia isolate of P. thornei. Further research into the biology of P. curvicauda is needed to facilitate development of strategies for its management, if it is an important pest.

Funds for treatment of hospitalized patients: evidence from Bangladesh.

This study was designed to explore sources of funds for health expenditure of patients if they are hospitalized. We have included 379 patients of 3 private and 7 public hospitals to estimate total expenditure. Of them, 229 (60.4%) were from public and 150 (39.6%) from private hospitals. Mean expenditure was Tk 60,613.3 and 8,262.7, and duration of hospital stay was 10.7 and 11.8 days in private and public hospitals respectively. More than half (55%) of the patients from middle class were treated in private hospitals. Of them, 278 (74.0%) were funded by themselves, 48 (12.8%) by loan with interest rate of 100% to 180%, 23 (6.1%) by loan without interest, 17 (4.5%) by losing their fixed asset, and 4 (1.1%) by begging in the street. Most of the patients did bear expenditure by themselves, followed by loan with high interest rate. 'Distress' selling of property was also a source. Middle-class patients could be comfortable with expenditure if they were in public hospitals.

Hemolytic Anemia and Pancytopenia Secondary to Vitamin B12 Deficiency: Evaluation and Clinical Significance.

Severe vitamin B12 deficiency presents a diagnostic challenge due to its diverse clinical manifestations, which can mimic serious hematologic disorders such as thrombotic thrombocytopenic purpura (TTP) or leukemia. The case we present here illustrates the unique characteristics of severe B12 deficiency, highlighting key differentiators from other conditions, including decreased reticulocyte counts and markedly elevated lactate dehydrogenase levels indicative of suppressed erythropoiesis. Advanced cobalamin deficiency affects all cell lines, leading to peripheral pancytopenia. Proposed mechanisms include fragile red blood cells prone to shearing, resulting in schistocyte formation, and hyperhomocysteinemia-induced oxidative stress exacerbating hemolysis. Prompt recognition and treatment with B12 replacement are critical, as illustrated by this case of hemolytic anemia and pancytopenia secondary to pernicious anemia, to prevent severe hematologic complications.

Kidney and Urinary Tract Involvement in Chronic Myelomonocytic Leukemia.

Chronic myelomonocytic leukemia (CMML) is a hematologic malignancy affecting the bone marrow and resulting in peripheral blood monocytosis. Kidney and urinary tract involvement is common and can present dramatically with life-threatening consequences. Kidney involvement can be the result of direct or indirect mechanisms, including prerenal azotemia, glomerular disease, tubulointerstitial involvement, and renovascular disorders. Urinary tract involvement, electrolyte and acid-base disorders, as well as nephrotoxicity from treatment of the disorder can also occur. Given this multifactorial pathogenesis involving several mechanisms concomitantly, nephrologists must exercise heightened awareness and maintain a low threshold for kidney biopsy. There is a pressing need for future research endeavors to elucidate and target the manifestations of CMML that involve the kidneys with the ultimate goal of augmenting overall prognosis and therapeutic outcomes.

Locking plate fixation of proximal humeral fractures with impaction of the fracture site to restore medial column support: a biomechanical study.

BACKGROUND: Despite the advent of locking plate techniques, proximal humeral fracture fixation can fail due to varus collapse, especially in osteoporotic bone with medial cortex comminution. This study investigated the effect of restoring the integrity of the medial column by fracture impaction and shaft medialization with locking plate fixation. This construct was compared with a traditional locking plate construct under conditions of varus cyclical loading. MATERIALS AND METHODS: Proximal humeral fractures with medial comminution were simulated by performing wedge-shaped osteotomies at the surgical neck in cadaveric specimens and removing 1 cm of medial cortex. For each cadaver (n = 6), 1 humeral fracture was fixed with a traditional locking plate construct. The other was fixed with the locking plate construct plus fracture impaction and shaft medialization, resulting in medial column restoration. The humeral head was immobilized, and a repetitive, varus force was applied to the humeral shaft until construct collapse or until 25,000 cycles were completed. RESULTS: None of the constructs with fracture impaction collapsed, whereas 5 of 6 of the nonaugmented constructs collapsed before reaching 25,000 cycles (P = .008). Collapse of the 5 nonimpacted constructs that failed occurred after an average of 11,470 ± 3589 cycles. CONCLUSION: Fracture impaction increased the ability of the locking plate to withstand repetitive varus loading. This technique provides a construct biomechanically superior to locking plate fixation alone.

Estimating the cost of illness of non-alcoholic fatty liver disease in Bangladesh.

Background and Aim: Non-alcoholic fatty liver disease (NAFLD) is a growing concern, affecting about 45 million of the Bangladeshi population. There is a paucity of research on the economic burden of NAFLD. The study aims to estimate the cost of illness of NAFLD in Bangladesh. Methods: In this prospective, cross-sectional study, a total of 250 patients of NAFLD, non-alcoholic steatohepatitis (NASH), and NASH cirrhosis were included from public and private hospitals. Costs of hospitalization, physician fees, investigation costs, expenditures on medical procedures, drugs; and nonmedical costs such as transport expenses and other informal payments (tips) were estimated. Results: The overall cost per patient per evaluation was (16.90-46 942.00) USD. The cost in public and private hospitals was 384.76 and 1146.93 USD, respectively. The cost per patient of NAFLD was 157.91 (16.90-955.08) USD, and for NASH cirrhosis was 1783.80 (422.48-46 942) USD. The cost of illness increased to USD 281.18 for diabetics and 254.52 USD for hypertensive. If all the NAFLD patients are evaluated once in healthcare settings, the projected cost will be 7.11 billion USD. In NAFLD, cost for investigations, medicines, transportation, and consultation of physicians was 49.08%, 32.41%, 11.11%, and 6.67%, respectively. Conclusions: NAFLD is causing a huge economic burden to the healthcare system. The cost of illness is increased with NASH cirrhosis. Overall, this study provides valuable insights into the economic burden of NAFLD in Bangladesh and emphasizes the several ways of intervention to reduce the cost by preventive measures and accessible healthcare for affected individuals.

Do customers' perceptions of Islamic banking services predict satisfaction and word of mouth? Evidence from Islamic banks in Bangladesh.

This study aims to investigate the customers' perceptions of Islamic banking services and their impact on satisfaction and word of mouth (WOM) with others. This study designs the bootstrapping procedures using a partial least square method to test path coefficient results. Structured questionnaires were distributed among clients of Islamic banks in Dhaka city, where 377 responses were collected for data analysis. The findings revealed that there is a highly significant relationship between security and customers' perception. Ethical responsibility and religious value have a positive and significant impact on customers' perception whereas benefit has a negative significant impact on customers' perception. Findings from this study also indicated that customers' perceptions mediate the effect of ethical responsibility, religious value, benefit, and security on satisfaction. In addition, customers' satisfaction mediates the effect of customers' perception and WOM. These findings can promote managers of Islamic banks to build customer satisfaction and WOM with Islamic banking services, and attain competitive advantage that may lead Islamic banks to succeed in the competitive business. This study also provides new insights into customers' WOM with others about Islamic banking services. This knowledge could assist Islamic banks to understand the customers' perceptions that would increase satisfaction and in turn, contribute to WOM with others in determining where would be best to target marketing attention of Islamic banking services with limited resources.

The role of dorsal root ganglia activation and brain-derived neurotrophic factor in multiple sclerosis.

Multiple sclerosis (MS) is characterized by focal destruction of the white matter of the brain and spinal cord. The exact mechanisms underlying the pathophysiology of the disease are unknown. Many studies have shown that MS is predominantly an autoimmune disease with an inflammatory phase followed by a demyelinating phase. Recent studies alongside current treatment strategies, including glatiramer acetate, have revealed a potential role for brain-derived neurotrophic factor (BDNF) in MS. However, the exact role of BDNF is not fully understood. We used the experimental autoimmune encephalomyelitis (EAE) model of MS in adolescent female Lewis rats to identify the role of BDNF in disease progression. Dorsal root ganglia (DRG) and spinal cords were harvested for protein and gene expression analysis every 3 days post-disease induction (pdi) up to 15 days. We show significant increases in BDNF protein and gene expression in the DRG of EAE animals at 12 dpi, which correlates with peak neurological disability. BDNF protein expression in the spinal cord was significantly increased at 12 dpi, and maintained at 15 dpi. However, there was no significant change in mRNA levels. We show evidence for the anterograde transport of BDNF protein from the DRG to the dorsal horn of the spinal cord via the dorsal roots. Increased levels of BDNF within the DRG and spinal cord in EAE may facilitate myelin repair and neuroprotection in the CNS. The anterograde transport of DRG-derived BDNF to the spinal cord may have potential implications in facilitating central myelin repair and neuroprotection.

Proximal humeral fracture fixation: locking plate construct ± intramedullary fibular allograft.

BACKGROUND: Locking plate constructs for proximal humeral fractures can fail due to varus collapse, especially in osteoporotic bone with medial cortex comminution. Augmentation, using a fibular allograft as an intramedullary bone peg, may strengthen fixation preventing varus collapse. This study investigated the ability of the augmented locking plate construct to withstand repetitive varus stresses relative to the nonaugmented construct. MATERIALS AND METHODS: Proximal humeral fractures with medial comminution were simulated by performing wedge-shaped osteotomies at the surgical neck in cadaveric specimens. For each cadaver (n = 8), 1 humeral fracture was fixated with the locking plate construct alone and the other with the locking plate construct plus ipsilateral fibular autograft augmentation. The humeral head was immobilized and a repetitive, medially directed load was applied to the humeral shaft until construct collapse or until 25000 cycles were completed. RESULTS: No augmented construct collapsed. In comparison, 6 of 8 nonaugmented constructs collapsed (P < .05). Collapse in the 6 nonaugmented constructs occurred after an average ±SD of 6604 ± 1984 cycles. Screw penetration of the articular surface was found in only 1 of the nonaugmented constructs. CONCLUSION: Fibular allograft augmentation increased the ability of the locking plate to withstand repetitive varus loading. Clinically, this may assist proximal humeral fracture fixation in osteoporotic bone with medial cortex comminution.

The Role of Zinc in Cardiovascular Disease.

Zinc is an essential trace element due to its role as a key part of human enzymatic activity. As a cofactor in metalloenzymes and metalloproteins, zinc participates in diverse biological functions, including gene transcription, translation, and replication, phagocytosis, and immunoglobulin and cytokine production. In this review, we will focus on the role of zinc in the cardiovascular system, including heart failure, vascular calcification, and myocardial infarction. We will further highlight the role of zinc in cardiovascular pathology in individuals with chronic kidney disease, and type II diabetes mellitus, groups uniquely at risk for cardiovascular morbidity and mortality.

Patient satisfaction level and its determinants after admission in public and private tertiary care hospitals in Bangladesh.

Introduction: Patient satisfaction is an important indicator of the quality of care provided by health care facilities. The objective of this study was to investigate the rate of satisfaction and its associated factors among the patients admitted to tertiary care hospitals in Bangladesh. Methods: This cross-sectional study was conducted in a public and two private tertiary care hospitals in Bangladesh in December 2019, including 923 consecutive patients admitted to medical and surgical departments. Face-to-face interview using a structured questionnaire was conducted to collect patient-level data. Logistic regression models were used to determine the factors associated with patients' satisfaction. Results: Patients' overall satisfaction level was 65% (51% in public and 75% in private hospitals) with a satisfaction rate of 63% in hospital staff courtesy, 56.5% in a hospital environment, 67% in physician care, 63% in general patient satisfaction, and 58% in patient's family care. Private hospitals (aOR 3.64, 95% CI 2.2-6.03), conservative management (aOR 3.34, 95% CI 2.10-5.33), shorter hospital stay (aOR 1.58, 95% CI 1.05-2.37) and perceived improvement after treatment (aOR 1.67, 95% CI 1.01-2.76) were associated with patients' satisfaction. In contrast, patients' accommodation on the floor (aOR 0.38, aOR 0.22-0.66) and high health care costs (aOR 0.97, 95% CI 0.95-0.99) were associated with patients' dissatisfaction with the in-patient service they received in both public and private hospitals. Conclusion: Almost two-thirds of the patients were satisfied with the inpatient service they received, though, the satisfaction rate was higher in private hospitals. Treatment modality, cost, and outcome, as well as hospital environment like accommodation, were associated with their satisfaction level.

Detection of Ramularia collo-cygni from barley in Australia using triplex quantitative and droplet digital PCR.

BACKGROUND: Ramularia leaf spot (RLS), caused by Ramularia collo-cygni, is an emerging threat to barley (Hordeum vulgare L.) production. RLS has been reported in Australia, however only minimal information is available regarding its detection and distribution. Due to initial asymptomatic growth in planta, slow growth in vitro and symptomatic similarities to net blotch and physiological leaf spots, detection of this pathogen can be challenging. Quantitative polymerase chain reaction (PCR)-based methods for R. collo-cygni-specific identification and detection have been described, however these assays have been demonstrated to lack specificity. False-positive detections may have serious implications, thus we aimed to design a robust R. collo-cygni-specific PCR method. RESULTS: Using the phylogenetically informative RNA polymerase II second largest subunit (rpb2) and translation elongation factor 1-alpha (tef1-α) genes, along with the tef1-α gene of H. vulgare, a triplex assay was developed for both quantitative and droplet digital PCR. The triplex assay detected R. collo-cygni DNA in barley leaves from New South Wales, South Australia, Tasmania, Victoria and Western Australia. No R. collo-cygni DNA was detected in barley seed grown in Western Australia. CONCLUSION: The presence of R. collo-cygni DNA has been confirmed in Australian barley crops, suggesting a distribution ranging across the southern barley growing regions of Australia. The R. collo-cygni-specific assay will be a valuable tool to assist with monitoring the distribution and impact of R. collo-cygni in Australia and other regions. © 2021 Society of Chemical Industry.

A Broken Heart: Cardiac Arrest As the Initial Presentation of Myasthenic Crisis.

Myasthenic crisis is a life-threatening condition commonly associated with respiratory failure and may present in unusual ways. However, there is paucity in the literature about the cardiac manifestations of myasthenia gravis. We present a case of a 61-year-old male who presented to the emergency room with upper respiratory infection symptoms who soon thereafter suffered sudden cardiac arrest. He was found to have shortened PR interval pre and post arrest onelectrocardiogram (EKG). Only past medical history, discovered post cardiac arrest, was myastenia gravis. All other causes of cardiac arrest were ruled out, and it was deemed to be due to a manifestation of myastenia gravis. The patient was treated with intravenous steroids and plasmapheresis with resolution of shortened PR interval. It is hypothesized that striatial muscle antibodies may trigger inflammation in cardiac muscle and cause conduction abnormalities. In addition, anti-Kv1.4 antibodies have been associated with EKG abnormalities, including QT prolongation and T-wave inversion. To our knowledge, we are the first to report myasthenic crisis manifesting with isolated cardiac arrest with pulseless electrical activity and a shortened PR interval.

Characterisation of ACP5 missense mutations encoding tartrate-resistant acid phosphatase associated with spondyloenchondrodysplasia.

Biallelic mutations in ACP5, encoding tartrate-resistant acid phosphatase (TRACP), have recently been identified to cause the inherited immuno-osseous disorder, spondyloenchondrodysplasia (SPENCD). This study was undertaken to characterize the eight reported missense mutations in ACP5 associated with SPENCD on TRACP expression. ACP5 mutant genes were synthesized, transfected into human embryonic kidney (HEK-293) cells and stably expressing cell lines were established. TRACP expression was assessed by cytochemical and immuno-cytochemical staining with a panel of monoclonal antibodies. Analysis of wild (WT) type and eight mutant stable cell lines indicated that all mutants lacked stainable enzyme activity. All ACP5 mutant constructs were translated into intact proteins by HEK-293 cells. The mutant TRACP proteins displayed variable immune reactivity patterns, and all drastically reduced enzymatic activity, revealing that there is no gross inhibition of TRACP biosynthesis by the mutations. But they likely interfere with folding thereby impairing enzyme function. TRACP exists as two isoforms. TRACP 5a is a less active monomeric enzyme (35kD), with the intact loop peptide and TRACP 5b is proteolytically cleaved highly active enzyme encompassing two subunits (23 kD and 16 kD) held together by disulfide bonds. None of the mutant proteins were proteolytically processed into isoform 5b intracellularly, and only three mutants were secreted in significant amounts into the culture medium as intact isoform 5a-like proteins. Analysis of antibody reactivity patterns revealed that T89I and M264K mutant proteins retained some native conformation, whereas all others were in "denatured" or "unfolded" forms. Western blot analysis with intracellular and secreted TRACP proteins also revealed similar observations indicating that mutant T89I is amply secreted as inactive protein. All mutant proteins were attacked by Endo-H sensitive glycans and none could be activated by proteolytic cleavage in vitro. In conclusion, determining the structure-function relationship of the SPENCD mutations in TRACP will expand our understanding of basic mechanisms underlying immune responsiveness and its involvement in dysregulated bone metabolism.

Application of Loop-Mediated Isothermal Amplification in an Early Warning System for Epidemics of an Externally Sourced Plant Virus.

Restricting Turnip yellows virus (TuYV) spread in canola (Brassica napus) crops often relies upon the application of systemic insecticides to protect young vulnerable plants from wide-scale green-peach aphid (GPA; Myzus persicae) colonization and subsequent virus infection. For these to be applied at the optimal time to ensure they prevent epidemics, growers would need to be forewarned of incoming viruliferous aphid migration and colonization. This study was conducted to field validate a loop-mediated isothermal amplification (LAMP) protocol designed to detect TuYV in aphids caught on traps and develop an early warning system for TuYV epidemics. Double-sided yellow sticky traps were deployed at 30 sites sown with canola over a two-year period in the south-west Australian grainbelt. Using LAMP, the percentage (%) of trap sides with TuYV-carrying aphids was measured and related to TuYV infection incidence in the adjacent crop. When TuYV was detected in aphids on >30% trap sides in a six-week period from pre-emergence to GS15 (five-leaf stage), TuYV reached >60% crop incidence by GS30 (beginning of stem elongation). Whereas, TuYV detection in aphids on ≤15% trap sides during this period was associated with ≤6% TuYV incidence by GS30. Furthermore, when large numbers of aphids, including GPA, were caught during this period but no TuYV was detected in them, minimal TuYV spread (≤5%) occurred in the crop by GS30. Therefore, the LAMP TuYV protocol can be used in an early warning system for TuYV epidemics by providing detection of initial viruliferous aphid migration into a canola crop before they establish colonies throughout the crop and spread virus. This would enable proactive, non-prophylactic, and thereby more effective systemic insecticide applications to minimize seed yield and quality losses due to early season TuYV infection.