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Biomedical subjects

L Battisti

Publications and source records attributed to L Battisti.

At least 19 recordsLinked to original sources

Sequence divergence of the red and green visual pigments in great apes and humans.

We have determined the coding sequences of red and green visual pigment genes of the chimpanzee, gorilla, and orangutan. The deduced amino acid sequences of these pigments are highly homologous to the equivalent human pigments. None of the amino acid differences occurred at sites that were previously shown to influence pigment absorption characteristics. Therefore, we predict the spectra of red and green pigments of the apes to have wavelengths of maximum absorption that differ by < 2 nm from the equivalent human pigments and that color vision in these nonhuman primates will be very similar, if not identical, to that in humans. A total of 14 within-species polymorphisms (6 involving silent substitutions) were observed in the coding sequences of the red and green pigment genes of the great apes. Remarkably, the polymorphisms at 6 of these sites had been observed in human populations, suggesting that they predated the evolution of higher primates. Alleles at polymorphic sites were often shared between the red and green pigment genes. The average synonymous rate of divergence of red from green sequences was approximately 1/10th that estimated for other proteins of higher primates, indicating the involvement of gene conversion in generating these polymorphisms. The high degree of homology and juxtaposition of these two genes on the X chromosome has promoted unequal recombination and/or gene conversion that led to sequence homogenization. However, natural selection operated to maintain the degree of separation in peak absorbance between the red and green pigments that resulted in optimal chromatic discrimination. This represents a unique case of molecular coevolution between two homologous genes that functionally interact at the behavioral level.

Amino Acid Sequence

Treatment of chronic childhood immune thrombocytopenic purpura with intramuscular anti-D immunoglobulins.

Seven patients with chronic immune thrombocytopenic purpura (ITP) were treated with intramuscular anti-D (anti-D IgG) five times, on an alternate-day basis, or until a platelet count of 100 x 10(9)/l was achieved, and, subsequently, when necessary to maintain platelet counts above 50 x 10(9)/l. Five patients responded to therapy, two of whom entered long-term remission. Although signs of haemolysis were present in all patients, anaemia was never a problem. No patient developed haematomas at the site of injection. We suggest that intramuscular anti-D represents a safe and relatively inexpensive alternative to intravenous gamma globulins (IVGG) for children with severe chronic ITP.

Adolescent

Haplotype diversity in the human red and green opsin genes: evidence for frequent sequence exchange in exon 3.

We studied polymorphisms in the coding sequences of the human red and green opsin genes of 133 Caucasian males. Eleven polymorphic sites were discovered in the red opsin gene, seven of which were in exon 3, three in exon 4 and one in exon 5. Polymorphisms at 8 of these sites resulted in amino acid substitutions which generated a total of 18 unique red opsins in the population. The substitutions at three (S180A, I230T, and A233S) of the 8 sites involve hydroxyl-bearing to non-polar amino acid residues, and are therefore likely to alter spectral characteristics of the red pigment. Eight polymorphic sites were observed in the green opsin coding sequences, six of which were in exon 3, one in exon 2 and one in exon 5. Five of the eight involved amino acid substitutions which generated 15 unique green opsins in the population. Substitutions at two of these sites involve hydroxyl-bearing vs. non-polar residues. Six polymorphisms, all of which are located in exon 3, are shared between the red and green opsin genes, essentially making it difficult to assign this exon to either of these genes. Markers in exon 3 are in partial linkage disequilibrium with those in exons 4 and 5, whereas the latter two are in strong linkage disequilibrium with each other. Furthermore, markers in the 5' region of exon 3 are also in only partial (54%) disequilibrium with those in the 3' region. The above results strongly suggest a history of frequent gene conversion, mainly localized to exon 3, in the lineages leading to the human red and green opsin genes.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence

Selective expression of human X chromosome-linked green opsin genes.

The human red and green photopigments are specified by genes on the long arm of the X chromosome (Xq28). In individuals with normal color vision, the locus was proposed to consist of a single red pigment gene upstream of one or more copies of green pigment genes. The presence of a single red pigment gene in the array was confirmed by demonstration of only one retinal mRNA transcript coding for the red opsin. In individuals with multiple green pigment genes, it is unknown whether all genes are expressed. We analyzed the sequence of red- and green-specific mRNA from retinas of individuals with multiple green pigment genes in comparison with the corresponding genomic DNA sequences. The data showed that only a single green pigment gene is expressed. We therefore suggest that a locus control-like element, already known to be located 3.8 kilobases upstream of the transcription initiation site of the red pigment gene, allows transcription of only a single copy of the green pigment genes, probably the most proximal copy. This finding provides an explanation for the not-infrequent presence of 5' green-red hybrid genes in individuals with normal color vision. Such hybrid genes are usually associated with defective color vision. We suggest that 5' green-red hybrid genes produce defective color vision only when their position in the gene array allows expression in the retinal cone cells.

Alleles

Specific oligosaccharide form of the Rhizobium meliloti exopolysaccharide promotes nodule invasion in alfalfa.

Rhizobium meliloti strain SU47 produces both high molecular weight (HMW) and low molecular weight (LMW) forms of an acidic exopolysaccharide, succinoglycan. Genetic studies have shown that succinoglycan is required for alfalfa root nodule invasion. We found that LMW succinoglycan, when applied exogenously to alfalfa roots, restored nodule invasion to exoA, exoB, exoF, and exoH mutants. Nodule initiation signals were not involved, since LMW succinoglycan from R. meliloti nodD1D2D3 and nodA mutants and from luteolin-induced wild-type cultures elicited effects similar to LMW succinoglycan from the uninduced wild-type strain. In contrast, LMW fractions from an exoA mutant, nonsuccinylated LMW succinoglycan, and HMW succinoglycan did not promote invasion, nor did LMW exopolysaccharides from R. leguminosarum bv. trifolii and Rhizobium sp. strain NGR234. LMW succinoglycan could be separated by anion-exchange chromatography into several distinct subfractions differing in repeating subunit multiplicities (monomer, trimer, and tetramer) and charge. When tested singly, only the most charged, tetrameric form was active. These results show that a specific oligosaccharide form of succinoglycan promotes nodule invasion in alfalfa. The implications for the mode of action of succinoglycan are discussed.

Chromatography, Ion Exchange

Heterologous exopolysaccharide production in Rhizobium sp. strain NGR234 and consequences for nodule development.

Rhizobium sp. strain NGR234 produces large amounts of acidic exopolysaccharide. Mutants that fail to synthesize this exopolysaccharide are also unable to nodulate the host plant Leucaena leucocephala. A hybrid strain of Rhizobium sp. strain NGR234 containing exo genes from Rhizobium meliloti was constructed. The background genetics and nod genes of Rhizobium sp. strain NGR234 are retained, but the cluster of genes involved in exopolysaccharide biosynthesis was deleted. These exo genes were replaced with genes required for the synthesis of succinoglycan exopolysaccharide from R. meliloti. As a result of the genetic manipulation, the ability of these hybrids to synthesize exopolysaccharide was restored, but the structure was that of succinoglycan and not that of Rhizobium sp. strain NGR234. The replacement genes were contained on a cosmid which encoded the entire known R. meliloti exo gene cluster, with the exception of exoB. Cosmids containing smaller portions of this exo gene cluster did not restore exopolysaccharide production. The presence of succinoglycan was indicated by staining with the fluorescent dye Calcofluor, proton nuclear magnetic resonance spectroscopy, and monosaccharide analysis. Although an NGR234 exoY mutant containing the R. meliloti exo genes produced multimers of the succinoglycan repeat unit, as does the wild-type R. meliloti, the deletion mutant of Rhizobium sp. strain NGR234 containing the R. meliloti exo genes produced only the monomer. The deletion mutant therefore appeared to lack a function that affects the multiplicity of succinoglycan produced in the Rhizobium sp. strain NGR234 background. Although these hybrid strains produced succinoglycan, they were still able to induce the development of an organized nodule structure on L. leucocephala. The resulting nodules did not fix nitrogen, but they did contain infection threads and bacteroids within plant cells. This clearly demonstrated that a heterologous acidic exopolysaccharide structure was sufficient to enable nodule development to proceed beyond the developmental barrier imposed on mutants of Rhizobium sp. strain NGR234 that are unable to synthesize any acidic exopolysaccharide.

Carbohydrate Sequence

Involvement of Tn4430 in transfer of Bacillus anthracis plasmids mediated by Bacillus thuringiensis plasmid pXO12.

The self-transmissible plasmid pXO12 (112.5 kilobases [kb]), originally isolated from strain 4042A of Bacillus thuringiensis subsp. thuringiensis, codes for production of the insecticidal crystal protein (Cry+). The mechanism of pXO12-mediated plasmid transfer was investigated by monitoring the cotransfer of the tetracycline resistance plasmid pBC16 (4.2 kb) and the Bacillus anthracis toxin and capsule plasmids, pXO1 (168 kb) and pXO2 (85.6 kb), respectively. In matings of B. anthracis donors with B. anthracis and Bacillus cereus recipients, the number of Tcr transcipients ranged from 4.8 x 10(4) to 3.9 x 10(6)/ml (frequencies ranged from 1.6 x 10(-4) to 7.1 x 10(-2), and 0.3 to 0.4% of them simultaneously inherited pXO1 or pXO2. Physical analysis of the transferred plasmids suggested that pBC16 was transferred by the process of donation and that the large B. anthracis plasmids were transferred by the process of conduction. The transfer of pXO1 and pXO2 involved the transposition of Tn4430 from pXO12 onto these plasmids. DNA-DNA hybridization experiments demonstrated that Tn4430 was located on a 16.0-kb AvaI fragment of pXO12. Examination of Tra- and Cry- derivatives of pXO12 showed that this fragment also harbored information involved in crystal formation and was adjacent to a restriction fragment containing DNA sequences carrying information required for conjugal transfer.

Bacillus anthracis

Identification of self-transmissible plasmids in four Bacillus thuringiensis subspecies.

The transfer of plasmids by mating from four Bacillus thuringiensis subspecies to Bacillus anthracis and Bacillus cereus recipients was monitored by selecting transcipients which acquired plasmid pBC16 (Tcr). Transcipients also inherited a specific large plasmid from each B. thuringiensis donor at a high frequency along with a random array of smaller plasmids. The large plasmids (ca. 50 to 120 megadaltons), pXO13, pXO14, pXO15, and pXO16, originating from B. thuringiensis subsp. morrisoni, B. thuringiensis subsp. toumanoffi, B. thuringiensis subsp. alesti, and B. thuringiensis subsp. israelensis, respectively, were demonstrated to be responsible for plasmid mobilization. Transcipients containing any of the above plasmids had donor capability, while B. thuringiensis strains cured of each of them were not fertile, indicating that the plasmids confer conjugation functions. Confirmation that pXO13, pXO14, and pXO16 were self-transmissible was obtained by the isolation of fertile B. anthracis and B. cereus transcipients that contained only pBC16 and one of these plasmids. pXO14 was efficient in mobilizing the toxin and capsule plasmids, pXO1 and pXO2, respectively, from B. anthracis transcipients to plasmid-cured B. anthracis or B. cereus recipients. DNA-DNA hybridization experiments suggested that DNA homology exists among pXO13, pXO14, and the B. thuringiensis subsp. thuringiensis conjugative plasmids pXO11 and pXO12. Matings performed between strains which each contained the same conjugative plasmid demonstrated reduced efficiency of pBC16 transfer. However, in many instances when donor and recipient strains contained different conjugative plasmids, the efficiency of pBC16 transfer appeared to be enhanced.

Bacillus anthracis

Demonstration of a capsule plasmid in Bacillus anthracis.

Virulent and certain avirulent strains of Bacillus anthracis harbor a plasmid, designated pXO2, which is involved in the synthesis of capsules. Two classes of rough, noncapsulated (Cap-) variants were isolated from the capsule-producing (Cap+) Pasteur vaccine strains ATCC 6602 and ATCC 4229. One class was cured of pXO2, and the other class still carried it. Reversion to Cap+ was demonstrable only in rough variants which had retained pXO2. Proof that pXO2 is involved in capsule synthesis came from experiments in which the plasmid was transferred by CP-51-mediated transduction and by a mating system in which plasmid transfer is mediated by a Bacillus thuringiensis fertility plasmid, pXO12. Cells of Bacillus cereus and a previously noncapsulated (pXO2-) strain of B. anthracis produced capsules after the acquisition of pXO2.

Bacillus anthracis

Mating system for transfer of plasmids among Bacillus anthracis, Bacillus cereus, and Bacillus thuringiensis.

To facilitate the analysis of genetic determinants carried by large resident plasmids of Bacillus anthracis, a mating system was developed which promotes plasmid transfer among strains of B. anthracis, B. cereus, and B. thuringiensis. Transfer of the selectable tetracycline resistance plasmid pBC16 and other plasmids from B. thuringiensis to B. anthracis and B. cereus recipients occurred during mixed incubation in broth. Two plasmids, pXO11 and pXO12, found in B. thuringiensis were responsible for plasmid mobilization. B. anthracis and B. cereus transcipients inheriting either pXO11 or pXO12 were, in turn, effective donors. Transcipients harboring pXO12 were more efficient donors than those harboring pXO11; transfer frequencies ranged from 10(-4) to 10(-1) and from 10(-8) to 10(-5), respectively. Cell-to-cell contact was necessary for plasmid transfer, and the addition of DNase had no effect. The high frequencies of transfer, along with the fact that cell-free filtrates of donor cultures were ineffective, suggested that transfer was not phage mediated. B. anthracis and B. cereus transcipients which inherited pXO12 also acquired the ability to produce parasporal crystals (Cry+) resembling those produced by B. thuringiensis, indicating that pXO12 carries a gene(s) involved in crystal formation. Transcipients which inherited pXO11 were Cry-. This mating system provides an efficient method for interspecies transfer of a large range of Bacillus plasmids by a conjugation-like process.

Bacillus

Effect of Senokot on rat intestinal electrolyte transport. Evidence of Ca++ dependence.

The mechanism of action of Senokot, a widely used laxative, has not been established. Senokot was given orally to rats 8-14 h before intestinal water and electrolyte transport were studied. Senokot significantly decreased colonic and jejunal water absorption measured in vivo by the single-pass perfusion technique. The Senokot changes were not associated with changes in jejunal or colonic histology or adenylate cyclase activity or colonic cyclic adenosine monophosphate content. Senokot also altered active electrolyte transport in rat descending colon as measured by the Ussing chamber-voltage clamp technique. These changes consisted of an increase in short-circuit current and a decrease in active Na and Cl transport that was due to a decrease in the mucosal-to-serosal fluxes. The changes in active electrolyte transport were dependent on Ca++ in the serosal but not the mucosal bathing solution. In contrast, addition of 10(-4) M verapamil to the serosal surface did not alter the Senokot effect. In spite of a dependence on serosal Ca++, Senokot did not alter 45Ca++ entry across the colonic serosal surface. The phospholipase A2 inhibitor quinacrine (10(-4) M) also did not alter the effect of Senokot on colonic Na and Cl transport. Senokot alters active colonic Na and Cl transport via a presently unknown mechanism that is dependent on serosal Ca++.

Adenylyl Cyclases

Dantrolene and basal ileal sodium and chloride transport: involvement of calcium stores.

The effect of dantrolene on active ion transport in rabbit ileum was determined using the Ussing chamber short-circuiting technique. Dantrolene prevents the release of calcium from intracellular stores in skeletal muscle and was used to probe the role of intracellular calcium stores in intestinal ion transport. A saturated solution of dantrolene (approx 25 microM) decreased ileal short-circuit current and potential difference, increased conductance and mucosal-to-serosal and net Na and Cl fluxes, but did not alter serosal-to-mucosal Na and Cl fluxes. The dantrolene stimulation of active Na and Cl absorption was specific since it did not alter glucose-dependent Na absorption, transport changes caused by Ca2+ ionophore A23187, or the increase in short-circuit current caused by dibutyryl cAMP or theophylline. These effects were associated with an increase in total ileal calcium content and a decreased rate of 45Ca2+ efflux without any change in 45Ca2+ influx from the serosal or mucosal surfaces. These findings are consistent with an effect of dantrolene to stimulate active ileal Na and Cl absorption by a mechanism involving lowered cytosol Ca2+ levels and compatible with trapping calcium in intracellular stores. It thus appears as if intracellular calcium stores have an important role in the control of basal ion transport in the intestine.

Animals

Effect of dopamine and bromocriptine on rat ileal and colonic transport. Stimulation of absorption and reversal of cholera toxin-induced secretion.

Water and electrolyte transport were determined in rat ileum and colon using the single-pass perfusion technique. Intraperitoneal dopamine caused prompt stimulation of both ileal and colonic water absorption. The dopamine effect was mediated by both specific dopamine and alpha 2-adrenergic receptors. Haloperidol, a specific dopamine antagonist, and yohimbine, an alpha 2-adrenergic antagonist, inhibited the effect of dopamine in ileal absorption; both antagonists alone had no effect on basal water transport. Bromocriptine (intravenous and intraluminal) stimulated ileal and colonic water absorption, which was inhibited by haloperidol and yohimbine, and reversed cholera toxin-induced ileal secretion. Magnitude and time-courses of the increased water absorption in ileal loops, inoculated with saline, were the same as in loops, inoculated with saline, suggesting that bromocriptine acted to reverse cholera toxin-induced secretion by stimulating absorption. Bromocriptine had no effect on the cyclic adenosine monophosphate increase caused by cholera toxin. We conclude (a) dopamine stimulates water absorption in vivo in rat ileum and colon; (b) this dopamine effect is via specific dopamine and alpha 2-receptors; (c) bromocriptine stimulates water absorption in ileum and colon and also acts by dopamine and alpha 2-receptors; and (d) bromocriptine reverses cholera toxin-induced secretion.

Animals

Dopamine stimulation of active Na and Cl absorption in rabbit ileum: interaction with alpha 2-adrenergic and specific dopamine receptors.

The effects of dopamine on active intestinal ion transport have been evaluated. An epithelial sheet preparation of rabbit ileum was used in vitro with the Ussing chamber-voltage clamp technique. Dopamine, in the presence of 1 mM ascorbic acid, added to the serosal bathing solution caused a dose-dependent decrease in short-circuit current, with a half-maximal effect at 1.2 muM and maximal effect of -50 muA/cm(2) at 50 muM; dopamine decreased the potential difference, and increased the conductance and net Na and net Cl absorption. There was no effect on the residual ion flux. Dopamine did not alter the change in short-circuit current caused by mucosal glucose (10 mM) or serosal theophylline (10 mM). Mucosal dopamine had no effect. The effect of dopamine on short-circuit current was inhibited by the dopamine antagonists haloperidol and domperidone and the alpha(2)-adrenergic antagonist yohimbine; there was no effect of the alpha(1)-antagonist prazosin and the beta-antagonist propranolol. In addition, the alpha(2)-adrenergic agonist clonidine, but not the alpha(1)-agonist methoxamine caused a dose-dependent decrease in short-circuit current. The ileal effects of dopamine did not occur via conversion into norepinephrine or release of norepinephrine from the peripheral nerves since "peripheral sympathectomy" with 6-hydroxydopamine did not alter the dopamine-induced change in ileal short-circuit current. The dopamine effects were not associated with a change in basal ileal cyclic AMP content but were associated with a decrease in total ileal calcium content as measured by atomic absorption spectrometry and as estimated by (45)Ca(++) uptake. The decrease in calcium content could be attributed to a dopamine-induced decrease in (45)Ca(++) influx from the serosal surface. Because of the presence of dopamine in ileal mucosa and these effects on ileal electrolyte transport, it is possible that dopamine may be involved in the physiologic regulation of active intestinal electrolyte absorption.

Animals

[The isolated thyroid nodule: notes on our personal experience].

An analysis of 255 patients operated for isolated thyroid nodes is made. Of these, 81% were cold nodes and 19% hot. Carcinomas were present in 21 cases. One carcinoma appeared as a scintigraphically hot node. The frequency of carcinomas, in relation to the nodules observed, was roughly the same in the two sexes. There is parallelism in behaviour of the rate in the two sexes as regards cold nodes, while this is not true for hot nodes. The importance of some recent diagnostic techniques is briefly discussed.

Adolescent