Peripheral blood marker of residual acute leukemia after hematopoietic cell transplantation using multi-plex digital droplet PCR.

Background: Relapse remains the primary cause of death after hematopoietic cell transplantation (HCT) for acute leukemia. The ability to identify minimal/measurable residual disease (MRD) via the blood could identify patients earlier when immunologic interventions may be more successful. We evaluated a new test that could quantify blood tumor mRNA as leukemia MRD surveillance using droplet digital PCR (ddPCR). Methods: The multiplex ddPCR assay was developed using tumor cell lines positive for the tumor associated antigens (TAA: WT1, PRAME, BIRC5), with homeostatic ABL1. On IRB-approved protocols, RNA was isolated from mononuclear cells from acute leukemia patients after HCT (n = 31 subjects; n = 91 specimens) and healthy donors (n = 20). ddPCR simultaneously quantitated mRNA expression of WT1, PRAME, BIRC5, and ABL1 and the TAA/ABL1 blood ratio was measured in patients with and without active leukemia after HCT. Results: Tumor cell lines confirmed quantitation of TAAs. In patients with active acute leukemia after HCT (MRD+ or relapse; n=19), the blood levels of WT1/ABL1, PRAME/ABL1, and BIRC5/ABL1 exceeded healthy donors (p<0.0001, p=0.0286, and p=0.0064 respectively). Active disease status was associated with TAA positivity (1+ TAA vs 0 TAA) with an odds ratio=10.67, (p=0.0070, 95% confidence interval 1.91 - 59.62). The area under the curve is 0.7544. Changes in ddPCR correlated with disease response captured on standard of care tests, accurately denoting positive or negative disease burden in 15/16 (95%). Of patients with MRD+ or relapsed leukemia after HCT, 84% were positive for at least one TAA/ABL1 in the peripheral blood. In summary, we have developed a new method for blood MRD monitoring of leukemia after HCT and present preliminary data that the TAA/ABL1 ratio may may serve as a novel surrogate biomarker for relapse of acute leukemia after HCT.

COVID-19 Cliff Notes: A COVID-19 Multidisciplinary Care Compendium.

As we pass the nearly 9 month mark of the coronavirus virus disease 2019 (COVID-19) pandemic in the United States, we sought to compile a brief multi-disciplinary compendium of COVID-19 information learned to date. COVID-19 is an active viral pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that confers high morbidity and mortality. COVID-19 has been associated with: pulmonary compromise and acute respiratory distress syndrome, thrombotic events, inflammation and cytokine, and post-infectious syndromes. Mitigation of these complications and expeditious therapy are a global urgency; this is brief summary of current data and management approaches synthesized from publications, experience, cross-disciplinary expertise (Figure 1).

The incidence, mortality and timing of Pneumocystis jiroveci pneumonia after hematopoietic cell transplantation: a CIBMTR analysis.

Pneumocystis jiroveci pneumonia (PJP) is associated with high morbidity and mortality after hematopoietic stem cell transplantation (HSCT). Little is known about PJP infections after HSCT because of the rarity of disease given routine prophylaxis. We report the results of a Center for International Blood and Marrow Transplant Research study evaluating the incidence, timing, prophylaxis agents, risk factors and mortality of PJP after autologous (auto) and allogeneic (allo) HSCT. Between 1995 and 2005, 0.63% allo recipients and 0.28% auto recipients of first HSCT developed PJP. Cases occurred as early as 30 days to beyond a year after allo HSCT. A nested case cohort analysis with supplemental data (n=68 allo cases, n=111 allo controls) revealed that risk factors for PJP infection included lymphopenia and mismatch after HSCT. After allo or auto HSCT, overall survival was significantly poorer among cases vs controls (P=0.0004). After controlling for significant variables, the proportional hazards model revealed that PJP cases were 6.87 times more likely to die vs matched controls (P<0.0001). We conclude PJP infection is rare after HSCT but is associated with high mortality. Factors associated with GVHD and with poor immune reconstitution are among the risk factors for PJP and suggest that protracted prophylaxis for PJP in high-risk HSCT recipients may improve outcomes.

Introducing the low-dose pill to Bangladesh; issues of continuation and failure.

In response to government plans to introduce a low-dose pill to the national family planning program of Bangladesh, a comparison of the performance of low-dose and standard-dose pills among a rural Bangladeshi population was conducted. Continuation rates were found to be better among users of the low-dose pill and there was no evidence that failure rates were higher. The relative risk (standard-dose vs. low-dose) over the first 30 months following adoption was 1.25 for first method continuation, and 1.29 for extended use failure. This paper, thus, provides evidence that low-dose pills may be a suitable method of contraception for rural Bangladeshi women.

Tubercular breast abscess--a diagnostic dilemma.

Tuberculosis affects many organs, however isolated tubercular breast lesion is rare. Clinically as well as radiologically it may mimic both pyogenic abscess and malignancy. One such case is being reported where a middle aged woman presented with painful, gradually increasing breast lump and was diagnosed clinically, radiologically, cytologically and even histologically as pyogenic abscess. Poor response to antibiotics raised the suspicion of malignancy for which repeat fine needle aspiration cytology was done. Well formed granulomas, necrosis and finally demonstration of acid fast bacilli established the diagnosis of tubercular abscess. No other organs were involved by tuberculosis in this case. Patient responded well to antitubercular drugs. Thus diagnostic challenge lies in the demonstration of acid fast bacilli in cases of equivocal morphology in routine cytology as well as histology. High clinical suspicion, poor response to antibiotics, suggestive radiological findings, cytology, histology and demonstration of acid fast bacilli -all contributes to the diagnosis of breast tuberculosis.

Validated Stability-indicating Reverse-phase Ultra-performance Liquid Chromatography Method for Simultaneous Determination of Sodium Methylparaben, Sodium Propylparaben and Ketorolac Tromethamine in Topical Dosage Forms.

A sensitive, fast, and stability-indicating isocratic reverse-phase ultra-performance liquid chromatography method was developed and validated for quantitative simultaneous determination of sodium methylparaben, sodium propylparaben and ketorolac tromethamine in topical dosage forms. Separation of all peaks was achieved by using acquity ethylene bridged hybrid C18 (50×2.1 mm, 1.7 µ) as stationary phase, mobile phase used was triethylamine buffer (pH 2.5):tetrahydrofuran:methanol (665:35:300, v/v/v) with isocratic mode at a flow rate of 0.40 ml/min. All component were detected at 252 nm with 10 min run time. The described method was found to be linear in the concentration range of 248-744 µg/ml for ketorolac tromethamine, 20.8-62.4 µg/ml for sodium methylparaben and 2.38-7.13 µg/ml for sodium propylparaben with correlation coefficients more than 0.999. Method was validated in terms of specificity, linearity, accuracy, precision, solution stability, filter equivalency, and robustness as per International Conference on Harmonization guideline. Formulation was exposed to the stress conditions of peroxide, acid, base, thermal, and photolytic degradation and proven all components were well separated in the presence of degradants.

The investigation of bacteriology of chronic suppurative otitis media in patients attending a tertiary care hospital with special emphasis on seasonal variation.

BACKGROUND: Chronic suppurative otitis media takes a lot of time in the hospital outdoors and a considerable amount of O.T. timings. Chronic suppurative otitis media may be either active chronic otitis media or a sequel of previous otitis media. OBJECTIVE: (a) To understand the bacteriology of Chronic suppurative otitis media (b) Their susceptibility patterns to commonly used antibiotics in this age of emerging resistance (c) the seasonal variation in the bacteriological pattern (d) For better empirical treatment of C.S.O.M where culture facilities are not available so that both intracranial and extra cranial complications can be avoided. MATERIALS AND METHODS: The secretions of 160 samples belonging to various age groups and of both sexes of clinically proven chronic suppurative otitis media with definitive exclusion and inclusion criteria were collected and cultured by aerobic and anaerobic methods. Drug sensitivity was done according to standard laboratory protocols. A month wise visit of the patients has been recorded to note any seasonal variations in the isolates. RESULTS: The most common aerobic organism is Pseudomonas Spp and most common anaerobes isolated being Bacteriodes species. Their susceptibility patterns and seasonal variations have been discussed. CONCLUSIONS: It is observed that chronic suppurative otitis media affects mainly younger group of population, mostly (86.8%) below 40 years of age. Majority of them (31.9%) belonged to, 10-19 years of age. The most common isolate being Pseudomonas Spp. (64.4%) followed by Staphylococcus aureus (33.8%). In the anaerobic group (1.8%) the Bacteriodes species is most prevalent. The most effective antibiotic in the aerobic isolates is Amikacin followed by Gentamicin and Cefotaxime where as for anaerobic isolates Cefoperazone sodium has better sensitivity. The isolation rates of both aerobic and anaerobic groups of organisms are more in the month of July to September, which is the monsoon season in this place. Isolation of Pseudomonas Spp. in Monsoon and post monsoon season may have been due to filling up of water bodies with rain water and infection with an omnipresent saprophytic organism like Pseudomonas Spp.

Effect of dextrans on cryopreservation of goat cauda epididymal spermatozoa using a chemically defined medium.

Experiments were carried out to investigate the cryoprotecting potential of dextrans (ranging from 10 to 2000 kDa) using a synthetic model system developed recently in this laboratory. Goat spermatozoa from the cauda epididymidis were extracted in a chemically defined medium (modified Ringer's solution) and assayed for motility using a phase-contrast microscope. The sperm cells were subjected to a freezing protocol in a computer-controlled biofreezer (cooling at 0.25 degrees C min(-1) to 5 degrees C; 5 degrees C min(-1) to -20 degrees C; 20 degrees C min(-1) to -100 degrees C) and plunged into liquid nitrogen. The frozen cells were thawed rapidly at 37 degrees C in a thermostatic waterbath. In the absence of dextran, all the spermatozoa lost their motility. The cryoprotecting efficacy of each dextran was found to be biphasic in nature. Initially, as the concentration of dextran was increased, the recovery of sperm motility also increased and reached an optimum value; however, with further increases in dextran concentration, the recovery of motility decreased sharply. Of all the sugar polymers tested, 10 kDa dextran showed the highest cryoprotecting efficacy, whereas the 2000 kDa sugar polymer was the least active. Dextrans of 10, 40, 73, 173, 252, 500 and 2000 kDa offered maximum cryorecovery of forward motility to the extent of approximately 23%, 21%, 19%, 18%, 16%, 15% and 8%, respectively. Optimum concentrations of these dextrans for cryoprotection of sperm motility were 8.42, 2.50, 1.09, 0.37, 0.31, 0.10 and 0.04 mmol l(-1), respectively. It thus appears that each dextran has a characteristic cryoprotection profile. The data show that the cryoprotecting efficacy and optimum cryoprotecting concentrations of dextrans are inversely related to their molecular mass. Dextran also served as a significant cryoprotectant in the presence of glycerol (0.87 mol l(-1)) and dimethyl sulphoxide (0.76 mol l(-1)), which are well known cryoprotectants; the action of the combined cryoprotectants was almost additive. The presence of glycerol or glycerol plus dimethyl sulphoxide caused a significant reduction (from 8.42 to 6.27 mmol l(-1)) in the optimum concentration of dextran. In the presence of the three cryoprotectants, recovery of sperm motility was as high as 58% (forward motility) and 60% (total motility).