Xylella fastidiosa is a fastidious, xylem-limited bacterium that causes a range of economically important plant diseases. Here we report the complete genome sequence of X. fastidiosa clone 9a5c, which causes citrus variegated chlorosis--a serious disease of orange trees. The genome comprises a 52.7% GC-rich 2,679,305-base-pair (bp) circular chromosome and two plasmids of 51,158 bp and 1,285 bp. We can assign putative functions to 47% of the 2,904 predicted coding regions. Efficient metabolic functions are predicted, with sugars as the principal energy and carbon source, supporting existence in the nutrient-poor xylem sap. The mechanisms associated with pathogenicity and virulence involve toxins, antibiotics and ion sequestration systems, as well as bacterium-bacterium and bacterium-host interactions mediated by a range of proteins. Orthologues of some of these proteins have only been identified in animal and human pathogens; their presence in X. fastidiosa indicates that the molecular basis for bacterial pathogenicity is both conserved and independent of host. At least 83 genes are bacteriophage-derived and include virulence-associated genes from other bacteria, providing direct evidence of phage-mediated horizontal gene transfer.

To contribute to our understanding of the genome complexity of sugarcane, we undertook a large-scale expressed sequence tag (EST) program. More than 260,000 cDNA clones were partially sequenced from 26 standard cDNA libraries generated from different sugarcane tissues. After the processing of the sequences, 237,954 high-quality ESTs were identified. These ESTs were assembled into 43,141 putative transcripts. Of the assembled sequences, 35.6% presented no matches with existing sequences in public databases. A global analysis of the whole SUCEST data set indicated that 14,409 assembled sequences (33% of the total) contained at least one cDNA clone with a full-length insert. Annotation of the 43,141 assembled sequences associated almost 50% of the putative identified sugarcane genes with protein metabolism, cellular communication/signal transduction, bioenergetics, and stress responses. Inspection of the translated assembled sequences for conserved protein domains revealed 40,821 amino acid sequences with 1415 Pfam domains. Reassembling the consensus sequences of the 43,141 transcripts revealed a 22% redundancy in the first assembling. This indicated that possibly 33,620 unique genes had been identified and indicated that >90% of the sugarcane expressed genes were tagged

The larval click-beetle Pyrearinus termitilluminans elicits the phenomenon of luminous termite mounds in the central-west region of Brazil. The bioluminescence (BL) spectrum of this larva (lambda max = 534 nm) is one of the most blue-shifted reported among known luminescent Coleoptera. We have isolated mRNA from larval thoracic lanterns and constructed a cDNA library into a lambda ZAP II vector. An expression library was obtained after excision of the pBluescript plasmid. This library was screened by photodetection and one clone that emitted green BL (lambda max = 538 nm) was isolated. The 2.2 kb cDNA insert includes a 543 residue open reading frame showing 82% homology with the luciferase isoenzymes of Pyrophorus plagiophthalamus (Coleoptera: Elateridae). As expected, the region between residues 223 and 247 that contains the putative active site for BL color determination showed a higher degree of homology among click-beetle luciferases that elicit closer BL colors. The in vitro BL spectrum of recombinant P. termitilluminans luciferase also peaks at 538 nm and, as in the case of native enzyme, does not show any bathochromic shift upon decreasing the pH.

The larval click-beetle Pyrearinus termitilluminans elicits the phenomenon of luminous termite mounds in the central-west region of Brazil. The bioluminescence (BL) spectrum of this larva (lambda max = 534 nm) is one of the most blue-shifted reported among known luminescent Coleoptera. We have isolated mRNA from larval thoracic lanterns and constructed a cDNA library into a lambda ZAP II vector. An expression library was obtained after excision of the pBluescript plasmid. This library was screened by photodetection and one clone that emitted green BL (lambda max = 538 nm) was isolated. The 2.2 kb cDNA insert includes a 543 residue open reading frame showing 82% homology with the luciferase isoenzymes of Pyrophorus plagiophthalamus (Coleoptera: Elateridae). As expected, the region between residues 223 and 247 that contains the putative active site for BL color determination showed a higher degree of homology among click-beetle luciferases that elicit closer BL colors. The in vitro BL spectrum of recombinant P. termitilluminans luciferase also peaks at 538 nm and, as in the case of native enzyme, does not show any bathochromic shift upon decreasing the pH.

Late in the fourth larval instar, several regions of the Rhynchosciara amercana salivary gland chromosomes undergo "DNA puffing. " We have constructed a library ofcloned cDNAs synthesized from poly(A)+RNA isolated from salivary glands'during the period ofdevelopment when the DNA puffs are active. From this library we have studied clones representative of three genes active during this period but not active at earlier developmental periods of the gland. One of these genes is not amplified during the developmental process and encodes a 0.6-kilobase RNA molecule. The other two genes are located within the DNA-puffsites C3 and C8and.encode 1.25-kilobase and 1.95-kldobase RNA molecules, respectively. We estimate -from the quantitation -of transfer hybridization experiments that each of these genes undergoes 16-fold amplification during DNA puffing.Gene amplification in somatic cells was first detected by morphological criteria in the larval salivary glands offlies ofthe family Sciaridae. Several regions of the Rhynchosciara americana polytene chromosomes were found to show a type ofpuffing in which, after puff regression, there was more DNA in the bands involved compared with neighboring bands as indicated by Feulgen staining (1). This was later confirmed. both by spectrophotometric measurements (2) and by autoradiographical studies on the incorporation of [3H]thymidine (3). Subsequently, similar observations-were made on the salivary chromosomes of larvae from the genus Sciara (4-6).The DNA puffs, which appear in late fourth instar in-Rhynchosciara salivary glands, are involved in the production of messenger RNAs (7,8). These encode several peptides of the communal cocoon, which are needed in large amounts over a short period of time (9, 10). Based on morphological and physiological criteria, the fourth instar of R. americana larvae has been divided into six periods (11). The first indication of DNA puff formation is the appearance of a fast-green staining band between two orcein (+) bands at the chromosomal sites ofthese puffs early in period 4 (ref. 12; unpublished data). Amplification and puffformation are subsequently maximal at different times for each site and are also dependent on the position of the cell within the gland (13, 14). The largest puffs are found in region 2 of the B chromosome and in regions 3 and 8 of the C chromosome. The B2 puffis formed preferentially in the first 50 cells of the gland in period 5, whereas C3 attains its largest size in the middle and distal section of the gland in period 6. The C8 puff is similar in all regions and is also maximal in period 6. As the larvae progress from period 3 to period 4; there is a dramatic change in the pattern of RNA and protein synthesis. This consists of an inhibition of rRNA synthesis (15) and the synthesis of new poly(A)+RNA species (7,8). This is accompanied by inhibition of the synthesis of certain peptides and the synthesis jtg of unfractionated RNA from the salivary glands of periods 3 and 5 larvae, was fractionated by electrophores...