Distribution and altitudinal structuring of phlebotomine sand flies (Diptera: Psychodidae) in southern Anatolia, Turkey: their relation to human cutaneous leishmaniasis.

The two Old World genera, Phlebotomus and Sergentomyia, were both recorded in southern Anatolia in Turkey. Phlebotomus species predominated and comprised about 93% of the entire collection (3,172 specimens). Out of the sixteen species identified, two belonged to the genus Sergentomyia: S. dentata and S. theodori. The remaining fourteen species in the genus Phlebotomus were grouped under four subgenera including some species that are elsewhere known to act as vectors of human cutaneous leishmaniasis. Most of the Phlebotomus were P. tobbi (32.5%), but P. papatasi, P. transcaucasicus, P. halepensis, P. galilaeus, P. sergenti, P. syriacus, P. neglectus, P. simici, P. alexandri, P. similis, P jacusieli, P. perfiliewi, and P. brevis were also identified. There were two associations of sand fly fauna with altitudinal gradient; the first one at relatively higher altitudes and the second one at lower altitudes. The transition between these two assemblages was within the range of 800-1,000 m. It is likely that Adana and Hatay provinces are transitional areas between western and eastern Anatolia. Mountains do not appear to be important geographical barriers for sand fly distribution. We also found that the proven vector P. sergenti is a widely distributed species throughout southern Anatolia and this species, together with its closely related species P. similis, shows sympatry in Konya Province.     

Binding of linker histones to the core nucleosome

Binding of chicken erythrocyte linker histones H1/H5 to the core nucleosome has been studied. Histones H1/H5 bind very efficiently to the isolated core nucleosome in vitro. The binding of linker histones to the core nucleosome is associated with aggregation of the particles. Approximately one molecule of linker histone binds per core nucleosome in the aggregates, irrespective of the concentration of the linker histones and the salt used. Histone H5 shows greater binding affinity to the core nucleosome as compared to H1. The carboxyl-terminal fragment of the linker histones binds strongly to the core nucleosome while the binding of the central globular domain is weak. Each core nucleosome is capable of binding two molecules of carboxyl-terminal fragment of linker histone. The core nucleosome containing one molecule of carboxyl-terminal fragment of linker histone requires higher salt concentration for aggregation while the core nucleosome containing two molecules of carboxyl-terminal fragment of linker histone can self-associate even at lower salt concentrations. On the basis of these results we are proposing a novel mechanism for the condensation of chromatin by linker histones and other related phenomena.     

Development of a single probe for documentation of chimerism following bone marrow transplantation.

Although numerous genetic markers are available for studying chimerism after bone marrow transplantation (BMT), there remains a need for a practical and highly informative method that is applicable in the early posttransplantation period. Using DNA restriction-fragment-length polymorphisms (RFLPs), we have evaluated the feasibility of developing a single synthetic oligonucleotide probe to study post-BMT chimerism. We have thus tested three candidate probes, termed O-3315-32, O-3315-80, and O-AY-29, that are homologous to tandemly repetitive sequences. Our results demonstrated donor-specific and recipient-specific fragments in 11 of 11 HLA-matched sibling pairs tested using probes O-3315-32 and O-3315-80. When probe O-AY-29 was used, 14 of 17 sibling pairs showed both donor and recipient markers, one had only a recipient marker, and two were identical. We showed that each of the three synthetic probes was effective in documenting donor marrow engraftment, mixed hematopoietic chimerism, the patient's pre-BMT phenotype (by using cultured skin fibroblasts obtained after BMT), and the origin of the malignant hematopoietic cells (i.e., of donor or recipient origin) in patients who developed recurrent hematologic malignancy following BMT. Compared with the use of cloned genomic probes, there are several important advantages to the use of synthetic oligonucleotide probes in studying post-BMT chimerism. Synthetic probes have absolute hybridization specificity and can be designed to suit the purposes of an individual study, since they have adjustable specificity that can be altered by changes in the length of the probe and by changes in the hybridization temperature. A single synthetic probe analogous to several highly polymorphic loci can have a polymorphism information content sufficiently high so that all but a small percentage of BMT patients could be followed easily; for example, if a probe were complementary to three highly polymorphic unlinked loci, it would discriminate approximately 98% of sibling donor/recipient pairs. This would be accomplished using only one restriction-endonuclease digestion and only one gel electrophoresis. Since other genetic markers, e.g., red blood cell antigens, immunoglobulin allotypes, and chromosome analysis, are not uniformly informative and, in some cases, cannot be used in the early posttransplantation period, the use of synthetic oligonucleotide probes for analysis of DNA RFLP is emerging as the method of choice for studies of post-BMT chimerism. This method will allow for the development of new knowledge that has not been possible with previous methods.     

Nucleoside triphosphate pyrophosphatase of rabbit matrix vesicles, a mechanism for the generation of inorganic pyrophosphate in epiphyseal cartilage

Inorganic pyrophosphate (PPi) may be important in the regulation of mineralisation but its origin in epiphyseal cartilage is ill-defined. Nucleoside triphosphate pyrophosphatase is one potential source, as this enzyme catalyses the formation of PPi from nucleoside triphosphates. This enzyme has been identified in matrix vesicles derived from rabbit epiphyseal cartilage and a method developed to measure the activity using ATP as substrate in intact matrix vesicles under relatively physiological conditions. The enzyme had a high affinity for ATP (Km less than 10 microM) and was also active towards GTP, CTP and UTP. Disruption of the matrix vesicle membrane by sonication failed to alter the activity. Treatment of sonicated matrix vesicles with Triton X-100 increased the activity which may indicate a direct effect of the detergent on the enzyme. Activity towards ATP was inhibited substantially by ADP and AMP and by another potential substrate beta,gamma-methyleneadenosine 5'-triphosphate. Dichloromethylene bisphosphonate, an analogue of the product PPi, inhibited the activity to a lesser extent. Two other potential substrates, NADP+ and thymidine 5'-monophosphate p-nitrophenyl ester were only weakly inhibitory as was 1-hydroxyethylidene 1,1-bisphosphonate. These results imply that nucleoside triphosphates are the substrates in vivo and the inhibitory effects of ADP and AMP suggest mechanisms whereby this activity could be regulated.     

Polygalacturonase activity and variation in ripening of papaya fruit with tissue depth and heat treatment

Effects of tissue position (viz. outer vs inner mesocarp) and heat treatment (48°C, 20 min) on variations in polygalacturonase (EC 3.2.1.15 and EC 3.2.1.67) activity and ripening of fruits of Carica papaya L. cv. Backcross Solo were investigated. Polygalacturonase activity increased during ripening concomitantly with an increase in tissue softness and soluble polyuronide level. Throughout ripening, inner mesocarp tissue was softer and contained higher polygalacturonase activity than outer mesocarp tissue. Titratable acidity as well as ß-galactosidase (EC 3.2.1.23) activity also increased during ripening; however, unlike polygalacturonase, their level or activity was lower in inner than in outer mesocarp. Ascorbic acid could partially account for the increase in titratable acidity during ripening but contributed very little to the differences in titratable acid levels between outer and inner mesocarp. Heat treatment had no effect on either fruit softness or titratable acidity, but it markedly reduced the increase in ascorbic acid and polygalacturonase activity during ripening. Ripening, as reflected by changes in tissue softness and polygalacturonase activity, progressed outwardly from the interior towards the exterior of the fruit. The effect of heat treatment in suppressing polygalacturonase activity was relatively greater in inner than in outer mesocarp, suggesting that sensitivity of the enzyme to heat treatment may vary with stage of ripeness of the tissue.     

Mixed-chain phosphatidylcholine analogues modified in the choline moiety: preparation of isomerically pure phospholipids with bulky head groups and one acyl chain twice as long as the other

Diacylphosphatidylcholines were synthesized with widely different acyl chain lengths and bulky head groups. Lysophosphatidylcholine was acylated at room temperature within 6 h with a 10-fold molar excess of fatty acid anhydride in dry, alcohol-free chloroform in the presence of 1.2 equivalents of 4-pyrrolidinopyridine as a catalyst, affording the mixed-acid phosphatidylcholines with widely different chain lengths in more than 90% yield and with less than 1% acyl migration. The syntheses of isomerically pure 1-stearoyl-2-decanoyl- and 1-stearoyl-2-undecenoyl(delta 10)-sn-glycero-3-phosphocholines C(18:0)/C(10:0)-PC and C(18:0)/C(11:1 delta 10)-PC, respectively), followed by conversion to various head-group analogues, are illustrated here. The transition peak widths at half-height of the endotherms obtained by differential scanning calorimetry are consistent with very high isomeric purity. Phospholipase D from Streptomyces chromofuscus was used as a catalyst in the hydrolysis of C(18:0)/C(10:0-PC to give the corresponding phosphatidic acid in quantitative yield. The latter compound was condensed with 10 molar equivalents of various N,N,N-trialkylammonium alkanols (as their p-toluenesulfonate or tetraphenylborate salt) in the presence of trichloroacetonitrile in dry pyridine under nitrogen atmosphere to yield the C(18:0)/C(10:0) phospholipids bearing modified head groups, which were purified by flash chromatography.     

Oxacillin-hydrolysing beta-lactamases. A comparative analysis at nucleotide and amino acid sequence levels

We have extended the sequence of the OXA-2 beta-lactamase which together with S1 mapping has enabled us to identify the promoter site for this gene. This lies in a region that is found upstream from a variety of resistance genes on different plasmids; each gene appears to have been inserted at the same specific site and to be expressed from the same promoter. The ancestral plasmid thus appears to function as a natural expression vector. The sequence of the recombination site at the 5' end of the OXA-2 gene shows a marked similarity with the attP sequence of lambda. DNA-probe analysis confirmed that the OXA-2 and OXA-3 beta-lactamases are related, and indicated no similarity with other beta-lactamase genes. However, a comparison of amino acid sequences demonstrates that the OXA-2, OXA-1 and PSE-2 beta-lactamases show some similarities to the typical class A enzymes, especially in the central helical domain of the latter, which is largely responsible for forming the active site of the enzyme. The three oxacillinases also show marked amino acid sequence similarity with the product of a regulatory gene, blaR1, required for beta-lactamase induction in Bacillus licheniformis.     

Odontometric profile of a prehistoric southeastern Florida population

This study analyzes and compares the dental size variation and sexual dimorphism of the Highland Beach Mound people, a late southeastern prehistoric aboriginal Florida population (ca. 800-1200 AD) with a somewhat earlier midwestern group (ca. 200-800 AD) and an Archaic sample (ca. 4160-2558 BC). The lengths and breadths of maxillary and mandibular permanent teeth of 99 individuals were measured, and crown surface areas were calculated for the molars. Results indicated that there was little sexual dimorphism. Only the mesiodistal dimensions of the upper P2 and lower C were significantly different at the P less than 0.05 level. In terms of dental size, what appears to be a chronologically based reduction in most points of comparison was seen in all three groups, with the exception of the molars, which were larger in the Highland Beach sample than in the earlier Schultz Mound.     

Altered hepatic microsomal function and elevated protooncogene expression as residual effects in rats exposed to delta-9-tetrahydrocannabinol

The microsomal activation of the potent hepatocarcinogen aflatoxin B1 (AFB1) and the expression of selected protooncogenes were investigated in the livers of rats exposed to delta 9-tetrahydrocannabinol (THC). At equimolar levels of cytochrome P-450, the microsome-mediated binding of AFB1 to DNA was significantly lower (56% of the controls) in preparations from drug exposed rats. Hepatic expression of the c-k-ras protooncogene was 3-fold higher in THC exposed animals. These results suggest the possible occurrence of long lasting residual effects in the rats exposed to THC.     

Generation of small mutation in large genomic fragments by homologous recombination: description of the technique and examples of its use

We have developed a technique of homologous recombination in bacteria which allows the mutagenesis of large genomic fragments cloned in cosmids. The desired mutation is first introduced into a plasmid clone and is then transferred to the appropriate cosmid clone by the means of double antibiotic selection coupled with phenotypic selection. We describe three different types of construct made by this technique.