ICE1 demethylation drives the range expansion of a plant invader through cold tolerance divergence.

Cold tolerance adaption is a crucial determinant for the establishment and expansion of invasive alien plants into new cold environments; however, its evolutionary mechanism is poorly understood. Crofton weed (Ageratina adenophora), a highly invasive alien plant, is continuously spreading across subtropical areas in China, north-eastward from the first colonized south-western tropical regions, through cold tolerance evolution. Close relations between the cold tolerance levels of 34 populations, represented by 147 accessions, and the latitude, extreme lowest temperature, coldest month average temperature, and invasion period have provided direct insight into its cold tolerance divergence. A comparative study of the CBF pathway, associated with the cold tolerance enhancement of cold-susceptible CBF1-transgenic plant, among four geographically distinct crofton weed populations revealed that the CBF pathway plays a key role in the observed cold tolerance divergence. Four epialleles of the cold response regulator ICE1 ranged from 66 to 50 methylated cytosines, representing a 4.4% to 3.3% methylation rate and significantly corresponding to the lowest to highest cold tolerance levels among these different populations. The significant negative relation between the transcription levels of the primary CBF pathway members, except for CBF2, and the methylation levels among the four populations firstly demonstrates that the demethylation-upregulated transcription level of CBF pathway is responsible for this evolution. These facts, combined with the cold tolerance variation and methylation found among three native and two other introduced populations, indicate that the ICE1-demethylated upregulation of cold tolerance may be the underlying evolutionary mechanism allowing crofton weed to expand northward in China.

First Report of Diplodia Cane Dieback of Grapevine in Bolivia.

Grape (Vitis vinifera L.) is an important commercial crop in the temperate regions of Bolivia where it has been grown for hundreds of years. In October of 2001, diseased canes of grape (cv. Muscat of Alexandria) were collected in a vineyard in Yotala, Department of Chuquisaca in southern Bolivia. In this planting of more than 1,000 plants, more than 75% were exhibiting cane dieback symptoms and many were dead or dying. No disease was observed on grape berries. Symptoms of the disease were similar to those reported for Diplodia cane dieback (1). Cankers ranging from 2 to 10 cm long and 0.5 to 3 cm wide were observed. When diseased canes were placed in a moist chamber, conidia oozed from pycnidia in black cirri. Immature conidia were hyaline and one-celled, but mature conidia were dark brown (20 to 30 × 10 to 15 µm) with one median septum and longitudinal striations. The pathogen was tentatively identified as Lasiodiplodia theobromae (Pat.) Griffon & Maubl. (synonyms Diplodia natalensis Pole-Evans and Botryodiplodia theobromae Pat.), teleomorph Botryosphaeria rhodina (Cooke) Arx) (2). Fungi were isolated from cankers on diseased canes by surface disinfestation in 0.25% NaOCl for 5 min and placing small pieces of tissue on 2% water agar or potato dextrose agar (PDA). L. theobromae was isolated from these tissues. Koch's postulates were fulfilled by inoculating grape berries and canes with the pathogen. Five grape berries were surface disinfested and inoculated by wounding with a sterile scalpel and inserting a piece of fungal mycelium on PDA in the wounded sites. The same number of control berries was similarly treated with sterile PDA. Inoculated and control berries were placed in plastic, moist chambers in the laboratory at ambient temperature (15 to 28°C) in the dark. Five canes on two potted plants were inoculated with the same isolate of the pathogen in a similar manner as the berries. The inoculated and control sites on canes were wrapped with masking tape. Plants were placed in a moist chamber for 5 days. After 8 days, inoculated berries were rotting and the inoculated sites were covered with grayish mycelium. Within 12 days, cankers as much as 3 cm long developed on the inoculated canes, and in some lesions, black pycnidia were observed. No lesions developed in the wounded control canes. The pathogen was reisolated from inoculated berries and canes, but not from control berries or canes. The teleomorph was not observed on any naturally infected canes or on those inoculated with the anamorph. The pathogen was identified as L. theobromae based on symptoms (1), cultural and morphological characteristics (2), and pathogenicity tests. The disease poses a potential threat to the cultivation of grapevine in southern Bolivia. To our knowledge, this is the first report of Diplodia cane dieback of grapevine in Bolivia. References: R. C. Pearson and A. C. Goheen. Compendium of Grape Diseases. The American Phytopathological Society, St. Paul, MN, 1988. (2). E. Punithalingam. No. 519 in: Descriptions of Pathogenic Fungi and Bacteria. CMI, Kew, Surrey, England, 1976.

Visceral leishmaniasis in Costa Rica: first case report.

We describe a 15-month-old eutrophic immunocompetent male who presented with fever, hepatosplenomegaly, pancytopenia, and hypergammaglobulinemia. Leishmania amastigotes were identified in spleen and bone marrow specimens. In addition, tissue culture, animal inoculation, and isoenzyme analysis identified the parasite as Leishmania donovani infantum or Leishmania donovani chagasi. The infant was successfully treated with an antimonial drug. These findings represent the first case of visceral leishmaniasis reported in Costa Rica.

Correlation of rhodamine 123 efflux by neoplastic plasma cells with clinical and biological characteristics of multiple myeloma.

Although a variable proportion of multiple myeloma patients can achieve response with conventional chemotherapy, residual tumor cells, which are refractory, finally reemerge leading to disease progression. The expression of the multidrug resistance protein (MDR1) has been one of the most extensively explored mechanisms of drug resistance and has been related to a poor response to chemotherapy in several human tumors. Nevertheless, a careful analysis of the literature on MDR1 expression in multiple myeloma (MM) shows the existence of disturbing discrepancies as regards both the incidence of MDR1 over-expression and its clinical value. A prerequisite for the assessment of MDR1 in tumor cells should be the identification of the neoplastic cells present in the sample. This is particularly important in MM, where the percentage of tumor cells in bone marrow (BM) is relatively low. In the present study we have analyzed the functional expression of MDR1 in BM plasma cells (PC), from a group of 40 untreated MM patients. For that purpose, the rhodamine 123 efflux assay was used in combination with specific staining for plasma cells (CD38 strong+). The mean fluorescence channel (MFC) of rhodamine 123 in myelomatous PC from MM patients was 311 and 110 after incubating cells with this fluorochrome for 15 and 60 min, respectively. The median percentage of rhodamine 123 elimination by BM PC was of 61% (range: 0.29 to 88%). Upon analyzing the relationship between the ability of myelomatous PC to eliminate rhodamine 123 and other clinical and biological disease characteristics we found that, within the group of patients displaying high MDR1 expression (>61% rhodamine efflux), there was a higher incidence of cases with bone disease (P = 0.014) and advanced clinical stages (P = 0.031), greater calcium (P = 0.007) and creatinine serum levels (P = 0.061), and lower levels of albumin in serum (P = 0.015). All these parameters are usually associated with a poor prognosis. When we analyzed the possible relationship between the ability of BM PC to eliminate rhodamine 123 and the presence of numerical chromosome abnormalities we observed that a low MDR1 expression was related to a higher incidence of trisomies of chromosomes 6 and 17, although these differences did not reach statistical significance (P = 0.06). In spite of these associations, from the prognostic point of view, MDR1 expression did not correlate with other relevant prognostic factors, response to treatment (P = 0.38) or overall survival (P = 0.12).

Prognostic value of immunophenotypic detection of minimal residual disease in acute lymphoblastic leukemia.

PURPOSE: The identification of immunophenotypic aberrancies through multiparametric flow cytometry makes the differentiation between normal and leukemic cells relatively simple and quick, and is therefore an attractive method for the investigation of minimal residual disease (MRD). In this report, we have analyzed the impact on relapse and relapse-free survival (RFS) of detecting immunophenotypical aberrant cells in acute lymphoblastic leukemia (ALL) patients in cytomorphologic complete remission (CR). MATERIALS AND METHODS: Two hundred eleven bone marrow (BM) samples from 53 consecutive ALL (37 precursor B-ALL and 16 T-ALL) patients were analyzed. The only selection criteria were to have at least one aberrant immunophenotypic feature at diagosis and to have achieved cytomorphologic CR after induction therapy. For MRD detection, all follow-up samples were analyzed with triple labelings using a two-step acquisition procedure, in which 106 cells were screened for the possible persistence of residual leukemic cells with the same phenotypic aberrancy as that identified diagnosis. RESULTS: Patients who displayed a gradual increase in MRD levels showed a higher relapse rate (90% v22%; P < .00001) and shorter median RFS (12 months v not reached; P < .0001) than those with stable or decreasing MRD levels. This adverse prognostic influence also was observed when children and adults, as well as B-ALL and T-ALL patients, were analyzed separately. An MRD level > or = or greater than 10(-3) discriminated two risk groups of ALL patients with significantly different relapse rates and RFS at all treatment phases (end of induction, consolidation, maintenance, and out of treatment). CONCLUSION: Multiparametric flow cytometry of MRD in ALL patients is a valuable tool for relapse prediction and for the identification of a cohort of patients with very poor prognosis.

Immunophenotypic analysis of CD19+ precursors in normal human adult bone marrow: implications for minimal residual disease detection.

BACKGROUND AND OBJECTIVE: Normal B-cell differentiation has been characterized extensively, but discrepancies persist regarding the exact sequence of antigen expression. Few systematic studies focusing on identification of the minor or undetectable B-cell subsets in normal human bone marrow (BM) which are frequently found in leukemic cells have been performed. Such studies could help to monitor minimal residual disease (MRD) in precursor-B-acute lymphoblastic leukemia (precursor-B-ALL). The aim of the present study was to analyze the sequence of antigen expression among normal human CD19+ B cells from adult BM. Our major goal was to identify infrequent and undetectable B-cell phenotypes that could be used for the detection of MRD in patients with precursor-B-ALL. DESIGN AND METHODS: Adult BM samples from a total of 33 healthy volunteers were analyzed using triple stainings, and measured by flow cytometry. A sensitive method based on the two-step acquisition procedure was used for the identification and characterization of cells present at very low frequencies. RESULTS: Five different subsets of CD19+ cells were identified in normal BM samples according to their degree of maturation: 1) CD19+/CD34+/CD10-/CD20-/CD22dlm+ (0.5 +/- 0.4% B cells); 2) CD19+/CD34-/CD10++/CD20-/CD22dlm+ (3.4 +/- 2.7%); 3) CD19+/CD34-/CD10+/CD20-/CD22dlm+ (3.5 +/- 2.2%); 4) CD19+/CD34-/CD10+/CD20+,++/CD22dlm+ (21 +/- 11%), and 5) CD19+/CD34-/CD10-/CD20++/CD22+ (73 +/- 19%). We observed that several B-cell phenotypes are frequent among precursor-B-ALL, but are infrequent or undetectable in normal human B cell differentiation. Accordingly, in all normal BM samples analyzed, less than 4 x 10(-5) cells co-expressed CD19 and CD117; CD20strong+/CD34+ and CD22strong+/CD34+ events were found at frequencies less than 5 x 10(-4), while CD20+/CD34+ phenotypes were found in less than 1 x 10(-3) BM cells. Although both CD19+/CD13+ and CD19+/CD33+ events were found at frequencies of up to 3 x 10(-3), they never formed a well-defined population of cells and therefore these latter phenotypic patterns could also be of use for MRD investigation in CD13+ and/or CD33+ precursor-B-ALL cases. INTERPRETATION AND CONCLUSIONS: Our results show that in adult BM normal B-cells display constant patterns of maturation as regards both their phenotypic characteristics and their relative distribution. Abnormalities in these patterns provide a potentially useful tool for monitoring MRD in precursor-B-ALL patients who achieve cytomorphologic complete remission.

Functional expression of MDR-1 in acute myeloid leukemia: correlation with the clinical-biological, immunophenotypical, and prognostic disease characteristics.

Up till now, clinical data on the possible prognostic influence of multidrug resistance (MDR) in hematological malignancies have been inconsistent, probably due to technical pitfalls. Moreover, in most studies qualitative information on the presence/absence of MDR-1 expression has been used instead of quantitative results. In addition, results usually refer to the total BM population and not specifically to blast cells. In the present study we analyzed the expression of MDR-1 in a series of 50 newly diagnosed de novo AML using a double-staining technique: (a) monoclonal antibodies for the specific identification of blast cells and (b) the rhodamine-123 efflux assay, which allows a quantitative and calibrated measurement of MDR-1 function. Expression of MDR-1 was correlated with clinical, biological, and immunophenotypical disease characteristics. All patients were uniformly treated according to the AML 87/91 protocols of the Spanish Pethema Cooperative Group; the median age was 51 +/- 19 years and the FAB distribution was as follows: 2 M0, 9 M1, 9 M2, 12 M3, 11 M4, 5 M5, and 2 M6 cases. Upon grouping the 50 AML patients analyzed according to the level of rh123 elimination, it was observed that those cases with > or = 30% decrease in rh123 fluorescence displayed higher WBC counts (9 +/- 12 vs 37 +/- 73 x 10(9)/l, p = 0.02) and platelet numbers (94 +/- 11 vs 35 +/- 25 x 10(9)/l, p = 0.02), together with a higher incidence of extramedullary involvement (35% vs 24%, p = 0.02). Half of the patients (47%) displaying a low rh123 elimination (< 30%) showed M3 morphology, while among the 33 patients with a higher rate of rh123 elimination (> or = 30%), only four (12%) corresponded to the M3 morphological subtype (p = 0.0006). From the immunophenotypic point of view, a low rate of rh123 elimination was associated with a lower expression of HLADR antigen (p = 0.003) and a higher expression of CD117 (p = 0.01). Regarding the possible prognostic influence of MDR1 expression, we found that a high rate of rh123 elimination (> 30%) was associated with a tendency towards poor disease outcome, illustrated by both a lower complete remission rate with the first cycle of chemotherapy (36% vs 56%) and a lower median disease-free survival (22 months vs median DFS not reached), although differences did not reach statistical significance (p = 0.1 in both comparisons). This data shows that although MDR-1 can be a relevant parameter in the evaluation of AML patients, larger series of patients using appropriate techniques for specifically analyzing the MDR of blast cells will be necessary in order to establish the final clinical value of this parameter.

[The prognosis for elderly patients admitted to a coronary unit for myocardial infarct: the changes over a decade]. / Pronóstico de los ancianos ingresados en la unidad coronaria por infarto de miocardio: cambios en una década.

INTRODUCTION AND AIM: New therapies have been added that improve prognosis of patients with myocardial infarction. Our purpose was to know if elderly patients reach benefit from these therapeutic changes. METHODS: We have analyzed the clinical data of 227 patients older than 70 who were admitted in our coronary care unit: 78 admitted in 1980-81 and 14 admitted in 1990-91. RESULTS: Although differences were not significant, in 1990-91 there were more women (36% versus 28%), less smoking (24% versus 33%), more patients with previous infarction (19% versus 12%), more hypertension (53% versus 43%), less anterior infarcts (38% versus 45%) more non Q infarcts (11% versus 1%), and less bundle branch block (21% versus 31%). In 1990-91, pacemakers were used less often (13% versus 27%, p = 0.03), thrombolytic therapy was given to 16 patients (10.7%), and the mortality rate was a little inferior (22% versus 30%, not significant). Female sex, not being a smoker, Killip class and bundle branch block were significantly related to mortality. After a multivariate analysis in which these factors we included, the date of admission resulted an independent predictor of mortality, with an odds ratio of 1990-91 to 1980-81 of 0.43 (p = 0.039). CONCLUSIONS: The management of patients older than 70 with myocardial infarction has improved, with a significantly lesser risk of dead after multivariate analysis, despite that thrombolysis has been scarcely applied.

Characterization of aberrant phenotypes in acute myeloblastic leukemia.

The existence of leukemic-associated phenotypes has been suggested to be a valuable tool for the detection of minimal residual disease (MRD) in AML patients, as they would allow to distinguish leukemic blast cells from normal hematopoietic progenitors. The present study was designed to analyze in which proportion of AML patients the immunological detection of MRD is feasible, based on the presence of aberrant phenotypes that allow the distinction of leukemic from normal cells. For this purpose we have prospectively investigated the blast cells from 40 AML patients at diagnosis with a large panel of MoAb in double and triple staining combinations analyzed at flow cytometry, in order to detect aberrant phenotypes on blast cells (lineage infidelity, antigenic overexpression, and asynchronous antigenic expression, as well as aberrant light-scatter pattern). In the analysis of the 40 AML cases more than one blast cell subset, distinguished by its different antigenic expression, was detected in 85% of the patients: five different phenotypic blast cell subsets were observed in six cases, four in 13 patients, three subsets in three cases, and two in 12 patients; only six cases showed a homogeneous phenotypical blast cell population. Twenty-nine of the 40 AML cases analyzed (73%) showed the existence of at least one aberrant phenotype: in 15 cases the myeloid blast cells co-expressed lymphoid-associated antigens (CD2, CD5, CD7, and/or CD19)--lineage infidelity--; asynchronous antigen expression was detected in 25 patients (CD34+CD56+, CD34+CD11b+, CD34+CD14+, CD117+CD15+, CD33-CD13+, CD13-CD15+, HLADR + CD15 , HLADR-CD14+CD11b+ CD4+); seven cases displayed antigen overexpression (CD13, CD33, CD15, or CD14); and in 13 patients leukemic cells had an abnormal FSC/SSC distribution according to their phenotype. These results suggest that immunological methods for the detection of MRD based on the existence of aberrant phenotypes could be used in the majority of AML patients.

Immunophenotype of c-kit cells in normal human bone marrow: implications for the detection of minimal residual disease in AML.

In the present study, seven normal human bone marrow samples from healthy volunteers have been analysed in order to investigate the immunophenotypic characteristics of the normal CD117+ cells and their utility for the detection of minimal residual disease in 71 acute myeloid leukaemia patients. Our results show that most of normal BM CD117+ cells coexpress the HLADR and the myeloid associated CD33 antigen. In addition, almost half of CD117+ cells are CD34+, these cells displaying a different FSC/SSC distribution when compared to the CD117+/CD34- cells. No CD117+/CD15+ and CD117+/CD10+ cells were detected and very few CD117+ cells (< 1 x 10(-3) expressing the HLADR-/CD34-, CD33+/HLADR- and CD34+/HLADR- phenotypes were found to be present in normal BM. In contrast, from the 71 AML patients analysed, 34 had CD117+/CD15+ blast cells and eight had the CD117+ phenotypes detected at low frequencies (< 1 x 10(-3)) in normal BM. In summary, the present study shows that the use of the CD117 antigen in different monoclonal antibodies combinations may be of great help for the detection of minimal residual disease in a high proportion of AML cases, especially in those patients displaying the CD117+/CD15+ phenotype, because cells coexpressing both antigens in normal BM, if present, are at very low frequencies.

[Age is the independent prognostic factor in acute myocardial infarct. The clinical course of infarct in the elderly patient]. / La edad es factor pronóstico independiente en el infarto agudo de miocardio. Curso clínico del infarto en el anciano.

We analyze the influence of age in the evolution of patients with acute myocardial infarction admitted to our Coronary Care Unit throughout two years (1990 and 1991). All 542 patients admitted during this period were classified in three groups: 299 less than 65 year old (group A), 170 between 65 and 74 year old (group B), and 73 with 75 year old or more (group C). Aged patients had a worse clinical condition, with significantly more previous heart failure, diabetes or hypertension, and the Killip's class was worse in group C than in group B, and worse in this than in group A (p = 0.00000). The mortality rate was 6.7% in group A, 12.9% in group B, and 31.5% group C (p = 0.00000). After a multivariate analysis, only three factors were significantly associated to prognosis: previous stroke, Killip's class, and group of age. Fibrinolytic therapy and coronary arteriography were less frequent with old people (p = 0.00000 and p = 0.00000 respectively). We conclude that age is an independent factor of prognosis during myocardial infarction. Old people have a worse clinical condition and the treatment is less aggressive than in young people.

Acute lymphoblastic leukemia (ALL): detection of minimal residual disease (MRD) at flow cytometry.

In the present study the usefulness of a method combining multiple staining direct immunofluorescence technique together with flow cytometry in order to predict relapse in ALL is analyzed in a group of 47 patients (11 T-ALL and 36 B-ALL). Results show that this method can be applied to at least two-thirds of all ALL patients being specially useful for the T-ALL cases (100% vs 56%) as this corresponding to the incidence of "aberrant" phenotypes. The detection of an increase in the percentage of bone marrow cells displaying "aberrant" phenotypes in two consecutive samples from the same patient is of great help on predicting relapse (sensitivity of 92% and specificity of 75%).

Tipos de circulaçäo e predominância das artérias coronárias em coraçöes de brasileiros / Types of circulation and predominance of the coronary arteries in brazilian hearts

O conhecimento anatômico do tipo de circulaçäo, com a identificaçäo da dominância das artérias coronárias na irrigaçäo do coraçäo, apresenta grande interesse clínico e cirúrgico, devido a que variaçöes nessa irrigaçäo ocasionam diferentes graus de gravidade em casos de obstruçäo. Foram estudados 50 coraçöes retirados de indivíduos adultos, de ambos os sexos e diferentes raças, nos quais as artérias coronárias foram dissecadas, visando identificar o tipo de circulaçäo. Realizamos, ainda, estudo morfométrico em coraçöes cujo peso médio foi 291 gramas e altura ventricular média de 97 mm. Em 72 por cento dos coraçöes estudados havia dominância da direita, 16 por cento circulaçäo balanceada e 12 por cento dominância da esquerda. Identificamos o número de ramos que ultrapassam a crux cordis, sendo o mínimo de um ramo e o máximo de cinco ramos, com valor médio de 2,2 nos casos de dominância da direita e em apenas dois coraçöes um ramo (em cada um) nos 8 de dominância da esquerda. Em 50 por cento dos coraçöes estudados o ramo interventricular anterior ultrapassa o ápice cardíaco, atingindo a sua face diafragmática.

Tipos de circulaçäo e predominância das artérias coronárias em coraçöes de brasileiros / Circulation types and predominance of the coronary arteries in brazilian hearts

O conhecimento anatômico do tipo de circulaçao, com a identificaçao da dominância das artérias coronárias na irrigaçao do coraçao, apresenta grande interesse clínico e cirúrgico, devido a que variaçoes nessa irrigaçao ocasionam diferentes graus de gravidade em casos de obstruçao. Foram estudados 50 coraçoes retirados de indivíduos adultos, de ambos os sexos e diferentes raças, nos quais as artérias coronárias foram dissecadas, visando identificar o tipo de circulaçao. Realizamos, ainda, estudo morfométrico em coraçoes cujo peso médio foi 291 gramas e altura ventricular média de 97 mm. Em 72 por cento dos coraçoes estudados havia dominância da direita, 16 por cento circulaçao balanceada e 12 por cento dominância da esquerda. Identificamos o número de ramos que ultrapassam a crux cordis, sendo o mínimo de um ramo e o máximo de cinco ramos, com valor médio de 2,2 nos casos de dominância da direita e em apenas dois coraçoes um ramo (em cada um) nos 8 de dominância da esquerda. Em 5O por cento dos coraçoes estudados o ramo interventricular anterior ultrapassa o ápice cardíaco, atingindo a sua face diafragmática.

Situacion actual de las exploraciones vasculares / Current situation of vascular exploration

La exploracion vascular con metodos no invasivos atraviesa una etapa de continuos avances en la actualidad, haciendo raramente necesaria la realizacion de arterio o angiografia

Sibship with 17-ketosteroid reductase (17-KSR) deficiency and hypothyroidism. Lack of linkage of histocompatibility leucocyte antigen and 17-KSR loci.

A family with nine children, three with male pseudohermaphroditism due to testicular deficiency of 17-ketosteroid reductase activity (17-KSR) and four with congenital hypothyroidism is presented. The three subjects with 17-KSR deficiency were raised as females until puberty, at which time they assumed a male gender role. Only one developed gynecomastia. Laparotomy on one of the three patients revealed normal epididymi and vas deferens with absence of Mullerian structures. Testicular biopsy in all three showed Leydig cell hyperplasia, hyalinization of the tubular basement membrane, normal Sertoli cells and maturational arrest at the spermatogonial stage. The endocrine profile in peripheral blood revealed markedly increased plasma androstenedione concentrations but normal testosterone, dihydrotestosterone, progesterone, 17-hydroxyprogesterone, and dehydroepiandrosterone. The levels of estradiol and estrone and of LH and FSH were elevated. Genital skin fibroblasts from the three patients exhibited normal dihydrotestosterone-binding activity and 5 alpha-reductase activity. Congenital hypothyroidism affected one of the three siblings with male pseudohermaphroditism. All four hypothyroid patients had thyroid enlargement and significant titers of circulating antithyroglobulin but not antithyroid microsomal antibodies. Neither the locus for the 17-KSR enzyme nor that for congenital hypothyroidism were linked to the histocompatibility leucocyte antigen complex in this sibship. Transmission of the trait for both congenital hypothyroidism and 17-KSR deficiency appeared to be autosomal recessive.