A bacteria injection scheme for in situ bioaugmentation.

This article presents an innovative design for inoculating the desired organisms to stratified geological layers at desired rates during in-situ bioaugmentation. The new delivery system consists of intermittent porous tubes connected in series with impermeable polyethylene tubes that run horizontally in each stratified layer of a contaminated aquifer. A bioaugmentation test using the new delivery system was conducted to inject an enriched culture of Escherichia coli (E. coli). Results of the test indicated that the distribution of E. coli through each porous tube was fairly uniform. A mathematical model previously developed to calculate the distribution of water flow through each porous tube was modified to calculate the distribution of E. coli. Geological layers often have different hydraulic conductivities. By controlling the permeability and the length of porous tubes placed in stratified layers, the new design provides a means to selectively deliver aqueous bacteria to various layers at desired rates according to aquifer heterogeneity.

A nutrient injection scheme for in situ bio-remediation.

Geological layers often have different hydraulic conductivities. This paper presents an innovative design for delivering aqueous substrates and nutrients to various stratified layers at desired rates during in-situ bio-stimulation. The new delivery system consists of intermittent porous tubes connected in series with impermeable polyethylene tubes that run horizontally in each stratified layer of a contaminated aquifer. Results of the tracer test indicated that the distribution of tritium through each porous tube was fairly uniform. A mathematical model was also developed to calculate the distribution of water flow through each porous tube. By controlling the permeability and the length of porous tubes placed in stratified layers, the new design provides a means to selectively deliver nutrients to various layers at desired rates according to aquifer heterogeneity.

Study of Water Relative Permeability in Fractures Using Well Tests and Radon: Gas Bubbles Effect.

Few published data are available for two-phase flow in fractures from field studies. All measurements of relative permeability reported in the literature were done in laboratory-scale. The in situ water saturations are normally not known for multiphase flow in natural fractures; therefore, the direct measurements of relative permeability are difficult in field-scale. With the help of a case study before and after the 2008 Mw 5.4 Antung earthquake, groundwater radon was used as a tracer to determine the gas and water saturations in a small naturally fractured aquifer. Well tests were also conducted to estimate aquifer transmissivity before and after the 2008 Antung earthquake. Anomalous declines in both groundwater radon concentration and transmissivity were observed precursory to the 2008 Antung earthquake. Both declines are two precursory phenomena having a common effect of gas bubbles. Using the data from well tests and radon tracer, one data point of water relative permeability can be obtained for in situ fractures. This data point reveals strong phase interference between water and gas bubbles for multiphase flow in natural fractures. Both the data of well tests and radon tracer are essential to gain an improved understanding of mass transfer behavior of groundwater-dissolved gases between water and gas phases.

Validation of the Chinese Version of Dementia Quality of Life Measure - Proxy in Care Home Residents with Dementia.

OBJECTIVE: To evaluate psychometric properties of the Chinese version of Dementia Quality of Life Measure - Proxy (C-DEMQoL-Proxy). METHODS: Care home residents aged ≥60 years who were diagnosed with dementia or demonstrated impairment in cognition were recruited from four care facilities in Hong Kong. Caregivers of these participants were also invited to participate. The original DEMQoL-Proxy was translated into Chinese (Cantonese) by a trained translator. The forward-translated version was reviewed by an expert panel of six experienced healthcare professionals. Revisions were made based on comments. The instrument was back-translated to English to check whether further changes were necessary. Demographic data (age, sex, type and severity of dementia, and Mini-Mental State Examination [MMSE] score) were collected from medical records of participants with dementia. Caregivers were interviewed by an occupational therapist or personnel supervised by the occupational therapist using the C-DEMQoL-Proxy and the Chinese version of Quality of Life-Alzheimer's Disease-Proxy (C-QoL-AD-Proxy). Acceptability, reliability, and validity of the C-DEMQoL-Proxy were evaluated using standard psychometric methods. RESULTS: 90 individuals (82.2% women) with dementia aged 72 to 102 years were included. Their diagnosis included Alzheimer's disease (23.3%), vascular dementia (15.6%), mixed and other types of dementias (51.1%), and missing (10%). Severity was mild in 12.2%, moderate in 62.2%, and severe in 25.6%. The mean MMSE score was 12.0 ± 4.9. 20% of the caregivers were family members and the rest were professional carers. The C-DEMQoL-Proxy had good acceptability, with no floor or ceiling effects or missing data. It had good internal consistency (Cronbach alpha = 0.91) and test-retest reliability (intraclass correlation coefficients = 0.83). It was mildly correlated with C-QoL-AD-Proxy (r = 0.29, p < 0.01). Age and sex were not correlated with C-DEMQoL-Proxy scores. C-DEMQoL-Proxy scores were not significantly different between dementia types, severity levels, or between those with higher or lower MMSE scores. CONCLUSION: The C-DEMQoL-Proxy is a valid and reliable instrument to assess health-related quality of life in individuals with dementia.

A novel HLA-D/DR-like antigen specific for human B lymphoid cells. Biochemical evidence for similarity to but nonidentity with known HLA-D/DR antigens.

The polymorphic human B cell-specific antigen, 33.1, detected by a murine monoclonal antibody, was compared by genetics and structural analysis with known human Ia antigens from a panel of DR homozygous Epstein-Barr virus-transformed B lymphoblastoid cell lines. Cells homozygous for DR 1, 2, 4, 5, and w6 were positive, while cells that are DR3,3 or DR7,7 usually failed to express this antigen. Mutant DR null, DC/MB-positive cells were 33.1 positive while DR null, DC/MB-negative cells failed to express this antigen, suggesting the segregation of 33.1 with the DC antigen. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis revealed that 33.1 alpha and beta chains were of lower molecular weights than the DR alpha and beta chains isolated from the same cell line. Partial N-terminal amino acid sequence analyses were carried out for the heavy and light chains of the 33.1 antigen radiolabeled with [3H] phenylalanine. The results of these analyses, in conjunction with previous data on tissue distribution, indicate that the 33.1 antigen is a non-DR but Ia-like antigen closely related to the previously defined I-A homologues, DC and DS.

Complications impaired endothelial progenitor cell function in Type 2 diabetic patients with or without critical leg ischaemia: implication for impaired neovascularization in diabetes.

AIMS: This study tested the hypothesis that migratory function of endothelial progenitor cells (EPCs) is impaired in Type 2 diabetic patients with or without critical leg ischaemia. METHODS: Seventy-four patients were classified into four groups: Type 2 diabetic (n = 21) and non-diabetic patients (n = 10) with critical leg ischaemia and Type 2 diabetic patients without lower extremity vascular disease (n = 30) and healthy subjects (n = 13). The number and functional activity of circulating and cultured EPCs were determined. RESULTS: The migratory function of cultured EPCs was significantly impaired in diabetic patients without (median, 48, interquartile range, 46, 49 count/view/well) and with (median, 51, interquartile range, 46, 60 count/view/well) critical leg ischaemia and non-diabetic patients with critical leg ischaemia (median, 49, interquartile range, 47, 55 count/view/well) compared with healthy subjects (median, 63, interquartile range, 57, 65 count/view/well) (P < 0.0001). The number of circulating EPCs was lower in Type 2 diabetic patients without lower extremity vascular disease (median, 3500, interquartile range, 1600, 6600/10(6) cytometric events) than Type 2 diabetic patients with critical leg ischaemia (median, 5300, interquartile range, 2400, 11,100/10(6) cytometric events), non-diabetic patients with critical leg ischaemia (median, 5550, interquartile range, 2000, 32,100/10(6) cytometric events) and healthy subjects (median, 5400, interquartile range, 2700, 8700/10(6) cytometric events) (P = 0.413). CONCLUSIONS: The migratory function of EPCs is impaired in patients with Type 2 diabetes, even in those without critical leg ischaemia. These findings present an important new insight into the pathogenesis of impaired neovascularization and critical limb ischaemia in diabetic patients and provide avenues of future clinical study.

Role of serum sodium in assessing hospital mortality in cancer patients with spontaneous tumour lysis syndrome inducing acute uric acid nephropathy.

Spontaneous tumour lysis syndrome (STLS) inducing acute uric acid nephropathy, a rare and neglected disease, presents more insidiously than conventional post-treatment tumour lysis syndrome. Although STLS is a serious and potentially fatal complication in patients with neoplastic disorders, few investigations have addressed the relevance of clinical and laboratory features in assessing prognosis. A retrospective study was conducted, reviewing the records of all patients who developed acute renal failure (ARF) at Chang Gung memorial hospital between 1 July 1999 and 30 June 2003. STLS-induced acute uric acid nephropathy was identified in 12 of 1072 ARF patients (1.1%) during the study period. All patients had advanced stage tumours with large tumour burden, and 66.7% of cases had abdominal organ involvement. All 12 hyperuricemic patients became oliguric despite conservative therapy, and remained hyperuricemic (21.6 +/- 5.2 mg/dl) before dialysis therapy. Diuresis developed in eight patients (66.7%), with associated resolution of hyperuricemia, azotemia and metabolic derangements following dialysis initiation. Overall hospital mortality was 58.3%. Death in most patients was related to hyponatremia and hypoalbuminemia on admission. The serum sodium was found to have the best Youden index (0.86) and highest overall prediction accuracy (93%). Moreover, serum sodium and serum albumin for individual patients were significantly and positively correlated (r = 0.617, p = 0.032). This investigation confirms a grave prognosis for cancer patients with STLS inducing acute uric acid nephropathy. Hyponatremia and hypoalbuminemia on the first day of admission indicate poor prognosis in such patients.

Aristolochic acid nephropathy due to herbal drug intake manifested differently as Fanconi's syndrome and end-stage renal failure--a 7-year follow-up.

Aristolochic acid (AA) may reduce glomerular or proximal tubular function, or both. We report a married couple taking AA-containing herbal drugs. The man developed Fanconi's syndrome (FS) whereas his wife reached end-stage renal failure (ESRF). He was a 36-year-old alcoholic cirrhotic patient who had taken the Chinese herbal drugs for 6 years, presenting with muscle weakness and laboratory findings of FS; the renal pathological findings were compatible with the diagnosis of aristolochic acid nephropathy (AAN). His 38-year-old wife, who took a lower cumulative amount of the same herbal drug for a shorter duration, developed advanced renal failure and severe anemia with pathological findings of extensive tubular atrophy, interstitial fibrosis but spared glomeruli. AA-I was detected in one of the herbal drugs. The wife has been on hemodialysis for 7 years, but the husband is still at the stage of slowly progressive chronic renal failure and persistent FS. None of their 5 children ever took the herbal drug, and none had renal problems during follow-up. It is important to trace the history of herbal drug intake in all the family members because of the possibility of sharing of drugs within a family. In addition to the effect of cumulative doses of AAs and the potentially higher susceptibility of females to AAN, the roles of liver cirrhosis and related vasodilators in the protection of the renal interstitium from fibrosis are questions that warrant further study.

The Risk Factors of mTORi-Associated Posttransplant Proteinuria.

Treatment with mammalian target of rapamycin inhibitors (mTORi) has been associated with an increased incidence of proteinuria after kidney transplantation as compared to other immunosuppressive agents. Proteinuria after mTORi use may occur in different clinical conditions and the precise mechanism remains unclear. The objective of this study was to investigate the related risk factors for proteinuria after mTORi treatment in kidney transplant recipients. This retrospective observational study population consisted of kidney transplant recipients followed up in a medical center in Southern Taiwan from January 1999 to April 2016. The baseline characteristics and transplantation-related profiles were collected at the time of enrollment. We examined risk factors for mTORi-associated proteinuria using a multivariate logistic regression analysis. P < .05 was considered as statistically significant. Hyperlipidemia and obesity at the initiation of mTORi treatment were strong predictors for proteinuria. Earlier identification of these risk factors may assist physicians in deciding the best candidate for mTORi conversion in order to optimize transplantation outcomes.

AML patients with CEBPalpha mutations mostly retain identical mutant patterns but frequently change in allelic distribution at relapse: a comparative analysis on paired diagnosis and relapse samples.

The roles of CEBPalpha mutations and its cooperating mutations in the relapse of acute myeloid leukemia (AML) are not clear. CEBPalpha mutations were analyzed on 149 patients with de novo AML at both diagnosis and relapse. Twenty-two patients (14.8%) had the mutations at diagnosis, two patients had N-terminal nonsense mutations alone, one had homozygous inframe duplication at the bZIP domain, and 19 patients had both N-terminal and bZIP mutations. Twenty patients relapsed with identical mutant patterns, two lost CEBPalpha mutations and none acquired the mutations at relapse. Cloning analysis showed that the N-terminal and C-terminal mutations occurred on separate cloned alleles and also on the same alleles in most of the diagnosis and relapse samples. Losing one of the two or more mutations on the same allele or acquiring the other mutation on the allele original carrying single mutation were observed not infrequently in the paired samples analyzed. Seven patients with CEBPalpha mutations had cooperating mutations with FLT3/ITD, FLT3/TKD or N-ras but not K-ras mutations. Our study showed that 91% of de novo AML harboring CEBPalpha mutations at diagnosis retained the identical mutant patterns but frequently changed in the allelic distribution at relapse.

Characterization of fusion partner genes in 114 patients with de novo acute myeloid leukemia and MLL rearrangement.

The fusion transcripts of MLL rearrangement [MLL(+)] in acute myeloid leukemia (AML) and their clinicohematologic correlation have not be well characterized in the previous studies. We used Southern blot analysis to screen MLL(+) in de novo AML. Reverse transcriptase-polymerase chain reaction was used to detect the common MLL fusion transcripts. cDNA panhandle PCR was used to identify infrequent or unknown MLL partner genes. MLL(+) was identified in 114 (98 adults) of 988 AML patients. MLL fusion transcripts comprised of 63 partial tandem duplication of MLL (MLL-PTD), 14 MLL-AF9, 9 MLL-AF10, 9 MLL-ELL, 8 MLL-AF6, 4 MLL-ENL and one each of MLL-AF1, MLL-AF4, MLL-MSF, MLL-LCX, MLL-LARG, MLL-SEPT6 and MLL-CBL. The frequency of MLL-PTD was 7.1% in adults and 0.9% in children (P<0.001). 11q23 abnormalities were detected in 64% of MLL/t11q23 and in none of MLL-PTD by conventional cytogenetics. There were no differences in remission rate, event-free survival and overall survival between adult MLL-PTD and MLL/t11q23 groups. Adult patients had a significantly poorer outcome than children. The present study showed that cDNA panhandle PCR can identify all rare or novel MLL partner genes. MLL-PTD was rare in childhood AML. MLL(+) adults had a poor outcome with no difference in survival between MLL-PTD and MLL/t11q23 groups.

Semicontinuous microcosm study of aerobic cometabolism of trichloroethylene using toluene.

A semicontinuous slurry-microcosm method was applied to mimic trichloroethylene (TCE) cometabolic biodegradation field results at the Que-Jen in-situ pilot study. The microcosm study confirmed the process of aerobic cometabolism of TCE using toluene as the primary substrate. Based on the nucleotide sequence of 16S rRNA genes, the toluene-oxidizing bacteria in microcosms were identified, i.e. Ralstonia sp. P-10 and Pseudomonasputida. The first-order constant of TCE-degradation rate was 0.5 day(-1) for both Ralstonia sp. P-10 and P.putida. The TCE cometabolic-biodegradation efficiency measured from the slurry microcosms was 46%, which appeared pessimistic compared to over 90% observed from the in-situ pilot study. The difference in the TCE cometabolic-biodegradation efficiency was likely due to the reactor configurations and the effective time duration of toluene presence in laboratory microcosms (1 days) versus in-situ pilot study (3 days). The results of microcosm experiments using different toluene-injection schedules supported the hypothesis. With a given amount of toluene injection, it is recommended to maximize the effective time duration of toluene presence in reactor design for TCE cometabolic degradation.

Decreased B cells and IgA-secreting B cells partially explain the high prevalence of IgA deficiency in dialysis patients.

AIMS: We previously reported 2 hemodialysis (HD) patients with recurrent infections and selective immunoglobulin A deficiency (IgAD). We further demonstrated that serum IgA levels were lower and the prevalence of IgAD was higher in uremic patients. The exact mechanisms of IgAD in uremic patients largely remained unclear. In some patients, it was caused by anti-IgA antibody neutralization and subsequent destruction. We performed the present study to survey if there is any defect in IgA production. MATERIALS AND METHODS: 288 patients were initially included for examination of serum immunoglobulins. 16 normal persons, 16 dialysis patients without IgAD, and 12 dialysis patients with IgAD were enrolled after the initial examination. Blood was drawn into heparinized tubes. WBC counts and lymphocyte percentage were examined by a CBC counter. Lymphocytes were separated by the Ficoll-Paque method. Flow cytometry was utilized to isolate the B cell and IgA-secreting B cell after staining with CD 19 phycoerythrin and FITC-conjugated rabbit anti-human IgA antibody. RESULTS: There is no significant difference between WBC counts or total lymphocyte counts of these 3 groups. However, we found a lower percentage of total lymphocyte counts in dialysis patients, either with or without IgAD. The total B cell numbers were lower in dialysis patients with IgAD. In addition, there were fewer IgA-secreting B cells in dialysis patients with IgAD. CONCLUSION: Decreased B cell and IgA-secreting B cell counts are seen in uremic patients with IgAD. This, in turn, indicates that there might be a defect of IgA production in some patients, rather than IgA destruction by anti-IgA antibodies as seen in some other patients. Further study is needed to investigate the mechanisms of decreased B cells and IgA-secreting B cells.

Endothelin-1 regulates parathyroid hormone expression of human parathyroid cells.

OBJECTIVE: Parathyroid cells synthesize and release endothelin-1 (ET-1). ET-1 displays an in vitro inhibitory effect on basal parathyroid hormone (PTH) secretion and also counteracts PTH hypersecretion stimulated by low calcium. Such effects are further demonstrated in vivo, independent of the changes in calcitonin. We propose that ET-1 may regulate the pathogenesis of uremic hyperparathyroidism. However, this was not directly demonstrated in human parathyroid glands. DESIGN: Hyperplastic parathyroid glands are obtained from the surgical operation for uremic hyperparathyroidism. Cells are isolated by enzyme digestion and treated with ET-1, and are assessed for PTH mRNA expression. PTH in the plasma and the medium is measured by a newly developed method to detect the whole PTH (1-84). PATIENTS: Uremic patients with severe secondary hyperparathyroidism and ultrasonography-proved hypertrophy of parathyroid glands received elective surgical approaches under general anesthesia. The resected glands were immediately taken to the laboratory for fresh isolation. MEASUREMENTS: Following ET-1 treatment, PTH mRNA expression is evaluated by RT-PCR method. ET-1 is detected with radioimmunoassay kit and PTH is measured by a new commercially available Duo PTH kit. RESULTS: ET-1 exhibited a dose-dependent inhibitory effect (from 10(-12) - 10(-7) M) on PTH mRNA expression of parathyroid cells, either in the basal or in the low-calcium-stimulated states. Release of PTH into the medium is also gradually inhibited by the increase in ET-1 concentrations. CONCLUSIONS: Our results demonstrate that ET-1 attenuates PTH mRNA expression in freshly isolated human parathyroid cells, and PTH release is also decreased. This result is consistent with our previously reported in vitro and in vivo experiments.

Lamivudine reverses severe acute hepatitis B and pancytopenia after renal transplantation: a case report.

Pancytopenia is rare after acute hepatitis B infection. The use of lamivudine in the treatment of acute hepatitis B-associated pancytopenia in renal transplant recipients has not been documented. Herein we reported a 21-year-old woman who was infected with acute hepatitis B 6 months after renal transplantation, a condition complicated by pancytopenia. Lamivudine reversed the acute hepatitis in 1 month and the pancytopenia after 3 months, without a change in renal function. Lamivudine was maintained for 2 years without a hepatitis flare-up after 4 years.

A mechanism for anomalous decline in radon precursory to an earthquake.

Mechanisms for interpreting anomalous decreases in radon in ground water prior to earthquakes are examined with the help of a case study to show that radon potentially is a sensitive tracer of strain changes in the crust preceding an earthquake. The 2003 Chengkung earthquake of magnitude (M) 6.8 on December 10, 2003, was the strongest earthquake near the Chengkung area in eastern Taiwan since 1951. The Antung radon-monitoring station was located 20 km from the epicenter. Approximately 65 d prior to the 2003 Chengkung earthquake, precursory changes in radon concentration in ground water were observed. Specifically, radon decreased from a background level of 780 pCi/L to a minimum of 330 pCi/L. The Antung hot spring is situated in a fractured block of tuffaceous sandstone surrounded by ductile mudstone. Given these geological conditions, we hypothesized that the dilation of brittle rock mass occurred at a rate faster than the recharge of pore water and gas saturation developed in newly created cracks preceding the earthquake. Radon partitioning into the gas phase may explain the anomalous decrease of radon precursory to the 2003 Chengkung earthquake. To support the hypothesis, vapor-liquid, two-phase radon-partitioning experiments were conducted at formation temperature (60 degrees C) using formation brine from the Antung hot spring. Experimental data indicated that the decrease in radon required a gas saturation of 10% developed in rock cracks. The observed decline in radon can be correlated with the increase in gas saturation and then with the volumetric strain change for a given fracture porosity.

Risk Factors Associated With Mammalian Target of Rapamycin Inhibitor Withdrawal in Kidney Transplant Recipients.

BACKGROUND: Mammalian target of rapamycin inhibitors (mTORi) play an essential role as novel immunosuppressive agents in kidney transplantation (KT). Treatment cessation usually occurs after adverse effects occur. We investigated the risk factors associated with withdrawal of mTORi in KT recipients and evaluated the outcomes related to the withdrawal. METHODS: The study enrolled KT recipients being followed up in a medical center in southern Taiwan from January 1999 through December 2014. RESULTS: Risk factors associated with mTORi withdrawal were initial proteinuria level, higher initial serum creatinine level posttransplantation, and history of glomerulonephritis as the primary etiology of renal failure. mTORi withdrawal was associated with increased risk of graft failure (hazard ratio [HR], 9.97 [95% confidence interval (CI), 1.03-96.8]; P = .047). Higher body mass index (HR, 11.2 [95% CI, 1.63-76.6]; P = .01) and tacrolimus usage (HR, 8.30 [95% CI, 1.14-60.7]; P = .037) were associated with increased risk of new-onset diabetes after transplantation in mTORi withdrawal groups. CONCLUSIONS: Proteinuria, poor graft function, and primary glomerulonephritis were associated with cessation of mTORi treatment. Earlier identification of these risk factors may prevent further adverse events and optimize transplantation outcomes after mTORi conversion.

Comprehensive mutational analysis of primary and relapse acute promyelocytic leukemia.

Acute promyelocytic leukemia (APL) is a subtype of myeloid leukemia characterized by differentiation block at the promyelocyte stage. Besides the presence of chromosomal rearrangement t(15;17), leading to the formation of PML-RARA (promyelocytic leukemia-retinoic acid receptor alpha) fusion, other genetic alterations have also been implicated in APL. Here, we performed comprehensive mutational analysis of primary and relapse APL to identify somatic alterations, which cooperate with PML-RARA in the pathogenesis of APL. We explored the mutational landscape using whole-exome (n=12) and subsequent targeted sequencing of 398 genes in 153 primary and 69 relapse APL. Both primary and relapse APL harbored an average of eight non-silent somatic mutations per exome. We observed recurrent alterations of FLT3, WT1, NRAS and KRAS in the newly diagnosed APL, whereas mutations in other genes commonly mutated in myeloid leukemia were rarely detected. The molecular signature of APL relapse was characterized by emergence of frequent mutations in PML and RARA genes. Our sequencing data also demonstrates incidence of loss-of-function mutations in previously unidentified genes, ARID1B and ARID1A, both of which encode for key components of the SWI/SNF complex. We show that knockdown of ARID1B in APL cell line, NB4, results in large-scale activation of gene expression and reduced in vitro differentiation potential.

Pseudo-Gaucher cells in mycobacterial infection: a report of two cases.

This report describes two cases of mycobacterial infection with pseudo-Gaucher cells. Both patients had no clinical evidence of inherited Gaucher disease. The first case was a patient with AIDS and Mycobacterium avium intracellulare involving the lung, spleen, and bone marrow. The bone marrow aspirates showed many histiocytes with needle-like inclusions. Acid fast staining showed that these histiocytes contained acid fast bacilli. Bone marrow biopsies revealed granulomatous lesions with aggregates of foamy histiocytes. The second case was an alcoholic patient with Mycobacteriumkanasassi infection involving the lung and lymph nodes. The lymph node aspirates showed infiltration of the same cells with acid fast bacilli in the cytoplasm.

Acquisition of FLT3 or N-ras mutations is frequently associated with progression of myelodysplastic syndrome to acute myeloid leukemia.

The role of internal tandem duplication of fms-like tyrosine kinase 3 (FLT3/ITD), mutations at tyrosine kinase domain (FLT3/TKD) and N-ras mutations in the transformation of myelodysplastic syndrome (MDS) to AML was investigated in 82 MDS patients who later progressed to AML; 70 of them had paired marrow samples at diagnosis of MDS and AML available for comparative analysis. Five of the 82 patients had FLT3/ITD at presentation. Of the 70 paired samples, seven patients acquired FLT3/ITD during AML evolution. The incidence of FLT3/ITD at diagnosis of MDS was significantly lower than that at AML transformation (3/70 vs 10/70, P<0.001). FLT3/ITD(+) patients progressed to AML more rapidly than FLT3/ITD(-) patients (2.5+/-0.5 vs 11.9+/-1.5 months, P=0.114). FLT3/ITD(+) patients had a significantly shorter survival than FLT3/ITD(-) patients (5.6+/-1.3 vs 18.0+/-1.7 months, P=0.0008). After AML transformation, FLT3/ITD was also associated with an adverse prognosis. One patient had FLT3/TKD mutation (D835Y) at both MDS and AML stages. Additional three acquired FLT3/TKD (one each with D835 H, D835F and I836S) at AML transformation. Five of the 70 matched samples had N-ras mutation at diagnosis of MDS compared to 15 at AML transformation (P<0.001), one lost and 11 gained N-ras mutations at AML progression. Coexistence of FLT3/TKD and N-ras mutations was found in two AML samples. N-ras mutations had no prognostic impact either at the MDS or AML stage. Our results show that one-third of MDS patients acquire activating mutations of FLT3 or N-ras gene during AML evolution and FLT3/ITD predicts a poor outcome in MDS.