Canine assistants for conservationists.
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Biomedical subjects
Publications and source records attributed to J E Maldonado.
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We examined cytochrome b sequence variation in 251 ornate shrews (Sorex ornatus) from 20 localities distributed throughout their geographical range. Additionally, vagrant (S. vagrans) and montane (S. monticolus) shrews from four localities were used as outgroups. We found 24 haplotypes in ornate shrews from California (USA) and Baja California (Mexico) that differed by 1-31 substitutions in 392 bp of mitochondrial DNA (mtDNA) sequence. In a subset of individuals, we sequenced 699 bp of cytochrome b to better resolve the phylogeographic relationships of populations. The ornate shrew is phylogeographically structured into three haplotype clades representing southern, central and northern localities. Analysis of allozyme variation reveals a similar pattern of variation. Several other small California vertebrates have a similar tripartite pattern of genetic subdivision. We suggest that topographic barriers and expansion and contraction of wetland habitats in the central valley during Pleistocene glacial cycles account for these patterns of genetic variation. Remarkably, the northern ornate shrew clade is phylogenetically clustered with another species of shrew suggesting that it may be a unique lowland form of the vagrant shrew that evolved in parallel to their southern California counterparts.
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The Pampas deer (Ozotoceros bezoarticus L. 1758) is the most endangered neotropical cervid, and in the past occupied a wide range of open habitats including grassland, pampas, savanna, and cerrado (Brazil) from 5 degrees to 41 degrees S. To better understand the effect of habitat fragmentation on gene flow and genetic variation, and to uncover genetic units for conservation, we examined DNA sequences from the mitochondrial control region of 54 individuals from six localities distributed throughout the present geographical range of the Pampas deer. Our results suggest that the control region of the Pampas deer is one of the most polymorphic of any mammal. This remarkably high variability probably reflects large historic population sizes of millions of individuals in contrast to numbers of fewer than 80,000 today. Gene flow between populations is generally close to one migrant per generation and, with the exception of two populations from Argentina, all populations are significantly differentiated. The degree of gene flow was correlated with geographical distance between populations, a result consistent with limited dispersal being the primary determinant of genetic differentiation between populations. The molecular genetic results provide a mandate for habitat restoration and reintroduction of Pampas deer so that levels of genetic variation can be preserved and historic patterns of abundance can be reconstructed. However, the source of individuals for reintroduction generally should be from populations geographically closest to those now in danger of extinction.
Mitochondrial DNA control region sequences were analyzed from 162 wolves at 27 localities worldwide and from 140 domestic dogs representing 67 breeds. Sequences from both dogs and wolves showed considerable diversity and supported the hypothesis that wolves were the ancestors of dogs. Most dog sequences belonged to a divergent monophyletic clade sharing no sequences with wolves. The sequence divergence within this clade suggested that dogs originated more than 100,000 years before the present. Associations of dog haplotypes with other wolf lineages indicated episodes of admixture between wolves and dogs. Repeated genetic exchange between dog and wolf populations may have been an important source of variation for artificial selection.
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The bone marrow erythrocytic precursors of 12 patients with refractory anemia (preleukemia) or myelomonocytic leukemia were studied by transmission electron microscopy. The results were tabulated in a semiquantitative manner and a comparison was established between the two main diagnostic groups. The following results are reported. 1. Similar nuclear and cytoplasmic abnormalities of the normoblasts were observed in preleukemia and leukemia. 2. A nuclear lesion consisting of nuclear clefts and blebs was demonstrated in at least some of the normoblasts in all of the patients. Although not specific, this finding appears to be a new contribution in the field of preleukemia and myelomonocytic leukemia. 3. Iron overload, including the presence of pathologic sideroblasts, is common to both preleukemia and leukemia.
Guinea pig eosinophil granules contain a protein, the major basic protein (MBP), which accounts for more than half of the total granule protein, has a high content of arginine, and displays a remarkable tendency to form disulfide-linked aggregates. In this study we have purified a similar protein from human eosinophil granules and have compared the human MBP to the protein comprising the Charcot-Leyden crystal (CLC). Eosinophils from patients with various diseases were purified and disrupted, and the granule fraction was obtained. Examination of the granule fraction by transmission electron microscopy showed numerous typical eosinophil granules. Analyses of granule lysates by gel filtration and by polyacrylamide gel electrophoresis revealed the presence of peroxidase and MBP with properties similar to that previously found in guinea pig eosinophil granules. The human MBP had a molecular weight of 9,200, contained less than 1% carbohydrate, was rich in arginine, and readily formed disulfide-bonded aggregates. CLC were prepared from eosinophil-rich cell suspensions by homogenization in hypotonic saline. The supernates following centrifugation of cell debris spontaneously formed CLC. Analysis of CLC revealed the presence of a protein with a molecular weight of 13,000 containing 1.2% carbohydrate. The protein displayed a remarkable tendency to aggregate even in the presence of 0.2 M acetic acid. Human MBP and CLC protein differed in their molecular weights, carbohydrate compositions, and amino acid analyses. Mixtures of the MBP and the CLC protein yielded two bands in polyacrylamide gel electrophoresis. Neither eosinophil protein increased vascular permeability in the guinea pig skin or contracted the guinea pig ileum. The results indicate that the human MBP and the CLC are distinct substances with properties such that one cannot be derived from the other.
In vitro aggregation of the platelets from four patients with refractory anemia and two patients with acute myelomonocyctic leukemia revealed distinctive abnormalities. In five patients, there was deficient or minimal aggregation with adenosine diphosphate (ADP), epinephrine, or collagen and only one wave of aggregation could be elicited with ADP at any concentration. Ultrastructural studies revealed numerous isolated platelets, small aggregates with few platelet pseudopods, and the presence of a characteristic type of aggregate with heterogeneous platelet composition combining features of both the primary and the secondary waves of aggregation. These "mixed aggregates" were particularly abundant in the four patients who had refractory anemia and may constitute the structural basis of the single wave of aggregation observed.
We have previously reported on the ultrastructure of platelets in preleukemia and myelomonocytic leukemia. We referred to an unusual and distinct anomaly of the platelet granules found in 15 of 16 patients. In the present communication we wish to describe and illustrate the light microscopic appearance of giant anomalous granules. Close scrutiny of the platelet morphology and a search for the aforementioned platelet granulopathy are important in the evaluation of patients with myeloproliferative diseases. In this paper we describe and illustrate in more detail the ultrastructure and ultrastructural histochemistry of the abnormal granules. In those patients with the platelet granulopathy, we have conducted in vitro platelet aggregation studies and carried out an electron microscopic evaluation of the aggregates. At least some of the giant granules remained morphologically intact in advanced stages of the aggregation phenomenon, and thus they are probably composed of elements that were not released during aggregation.
In a combined scanning and transmission electron microscopic study of the red blood cell line in patients with refractory anemia (preleukemia) and myelomonocytic leukemia, certain elements with abundant, convoluted membranes that formed projections over the cell surface were observed. These cells were both normoblasts (nucleated) and reticulocytes (anucleated) and constituted what appeared to be a distinct subpopulation of the red blood cell line. It is possible that in these erythrocytes there is a nucleocytoplasmic asynchronism. We are not certain, however, that they are linked to the leukemic process.
Studies of in vitro platelet aggregation were done in five patients with refractory anemia and two with acute myelomonocytic leukemia. The macroscopic results as well as the general ultrastructural findings were reviewed in a companion paper. Electron microscopic analysis of changes in the individual platelets within aggregates revealed a striking heterogeneity, both in the degree of response of each platelet are in the ultrastructural characteristics of the platelet population. Many of the unaggregated platelets had reacted individually, resembling the platelets of patients with Glanzmann's thrombasthenia. There were other abnormalities suggesting the presence of surface defects, such as the presence of areas of obliteration of the interplatelet space (so-called tight connections). One of the most striking findings was a peculiar dissociation between the different components of the aggregation sequence.
Platelets and megakaryocytes from 11 patients with the carcinoid syndrome have been studied by transmission electron microscopy. Cells fixed in phosphate-buffered glutaraldehyde are oval to discoid, with pseudopods, a dilated open-channel system, and a prominent dense tubular system as defined by peroxidase activity and alkaline bismuth stain. Atypical with hexagonal lattices and treaded substructures and large (diameter greater than 0.5 mum), phosphatase-positive, debris-containing vacuoles are four times more numerous than in normal platelets. Incubation of platelets in a 0.05% suspension of latex results in particle incorporation into phagosomes and the debris-containing vacuoles. Molybdate-dichromate stain reveals two classes of dense bodies, one of which (with a reticular core) is 20 times more numerous than in normal platelets. Bone marrow megakaryocytes lack both dense bodies and debris vacuoles analogous to those found in circulating platelets. These results suggest autophagy or endocytosis abnormalities and provide evidence for multiple types of dense bodies in carcinoid platelets.
Incubation of human platelets in plasma containing a suspension of latex particles for 1-90 min resulted in progressive accumulation of particles in the open-channel system, followed by localization of latex in electron-opaque vacuoles. After 60 min, acid phosphatase was localized within latex-containing vacuoles. The periodate-alkaline-bismuth reaction intensely stained external membranes and membranes of the open-channel system. Membranes of latex-containing organelles were not stained. Latex phagocytosis was independent of both anticoagulant choice and aspirin effects. Our results indicate that the platelet can act as a true phagocyte, and we suggest that the phagocytic process is chronologically similar to that reported for polymorphonuclear leukocytes.
Gel filtration of human platelet-rich plasma (PRP) on columns of Sepharose 2B removed at least 99.85% of the plasma proteins from platelets when a column 10 cm in height was used and a plasma volume 11 to 14% of the gel-bed volume was applied. ADP and ATP levels in gel-filtered platelets (GFP) were not significantly different from those in PRP. By transmission electron microscopy, GFP were indistinguishable from PRP. Gel filtration appears to be a highly satisfactory technique of separating platelets from plasma without modifying structure, function, or contents significantly. The roles of several crude protein fractions in platelet aggregation and aspirin's inhibition of aggregation were examined. Fraction I (mostly fibrinogen) enhanced collagen-induced aggregation of gel-filtered platelets; Fraction V (mostly albumin) was inhibitory. Fraction II (mostly gamma-globulin) or gelatin had no significant effect. Aspirin added to gel-filtered platelets inhibited aggregation by 80%. The addition of mixtures of plasma proteins containing albumin increased albumin's inhibitory effect. Incubation of gel-filtered platelets with aspirin labeled in the carboxyl position resulted in no uptake of the label. In contrast, incubation with acetyl-labeled aspirin was followed by uptake of more than 2 X 10(6) acetyl groups per platelet in 1 minute. Incubation for 30 minutes resulted in a five- to sixfold further increase in uptake of the label. Aspirin can acetylate platelets and inhibit aggregation directly. Plasma proteins, in particular albumin or a contaminant of the albumin fraction tested, enhance the inhibitory effect of aspirin on platelet aggregation.
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Five patients were seen at the Mayo Clinic over an 8-year period with the following complex of clinical and morphologic features; striking eosinophilia, cardiomyopathy, hepatosplenomegaly, and either a rapidly fatal or a prolonged, debilitating illness. In recent years, controversy has raged over the precise designation of this syndrome, with proposals ranging from eosinophilic leukemia to hypereosinophilic syndromes. To focus on the major target organ of the disease, we have favored the term endomyocardiopathy with eosinophilia. Experience with these five patients showed that (1) eosinophilia can persist for many years before symptoms appear; (2) progressive restrictive cardiac disease was the major cause of death and debility; (3) osmiophilic cytoplasmic inclusions are present in eosinophils of these patients and also in cells from other patients with marked eosinophilia; and (4) echocardiography may prove to be a useful noninvasive tool to diagnose and follow the progress of cardiac involvement. Although none of these patients was thought to have leukemia, intensive therapy with steroids or cytotoxic agents, or both, is considered necessary to control the progression of the disease.
The designation "pseudomyeloma'' is used to describe three patients who had a clinical picture that closely resembled multiple myeloma and was characterized by severe osteoporosis and a serum monoclonal paraprotein peak. The diagnosis of myeloma could not be made histologically, initally or after three, four, or ten years of observation. The protein abnormalities and the bone marrow picture remained stable. It is not known if this association has pathophysiologic importance or if it represents a chance phenomenon.