Linda Baumann

Study of genetic relationships among marine species of the genera Beneckea and Photobacterium by means of in vitro DNA/DNA hybridization

AbstractStrains representative of species of the marine genera Beneckea and Photobacterium were used as reference standards in in vitro DNA/DNA competition experiments. Within a given species, strains were found to be related by over 80% competition. (Competition was defined as the amount of radioactive DNA displaced by heterologous DNA relative to the amount displaced by homologous DNA.) On the basis of interspecies competition values (expressed as averages), the following groupings could be made:1.“Photobacterium” fischeri was related to strain ATCC 15382 by a competition of 38% and was distinct from all the other strains tested (competition ≤11%).2.The genus Photobacterium consisted of 3 species, P. phosphoreum, P. leiognathi, and a newly designated species. P. angustum (composed of non-luminous strains). The latter species was found to be related to P. leiognathi and P. phosphoreum by 56 and 28% competition, respectively, while P. phosphoreum was related to P. leiognathi by 29%.3.In the genus Beneckea, 65% competition was detected between B. harveyi and B. campbellii as well as between B. parahaemolytica and B. alginolytica. These pairs of species were related to each other by 51–58% and to B. natriegens by 34–56% competition. A newly designated pathogenic species, B. vulnifica, appeared to have a low but significant relationship to all the above mentioned species of Beneckea.4.Two biotypes, related by 68% competition, were recognized in the species B. splendida. Similarly, B. pelagia was found to consist of 2 biotypes related by a competition of 67%. The competition values between these species were 38–40%.5.B. nereida, B. nigrapulchrituda, and “Vibrio” anguillarum had competition values ≤30% to each other as well as to other species of Beneckea.6.With Vibrio cholerae as the reference standard, V. albensis was found to be related by a competition of 82%, while V. proteus and V. metschnikovii had competition values of 22 and 12%, respectively. These results suggested that V. albensis should be synonymized with V. cholerae, while the latter two organisms should remain distinct from this species. V. cholerae as well as the other terrestrial organisms tested did not appear to be significantly related to any of the marine strains (competition values ≤27%). The speciation derived from the results of the DNA/DNA competition experiments was compared to previous speciation based on phenotypic similarities.

Reevaluation of the taxonomy ofVibrio, beneckea, andPhotobacterium: Abolition of the genusBeneckea

As a result of studies on the evolution of glutamine synthetase and superoxide dismutase, the genusBeneckea has been abolished and its constituent species, along withPhotobacterium fischeri andP. logei, assigned to the genusVibrio. The definitions ofVibrio andPhotobacterium have been modified accordingly.

Organization of the mitochondrial genomes of whiteflies, aphids, and psyllids (Hemiptera, Sternorrhyncha)

BackgroundWith some exceptions, mitochondria within the class Insecta have the same gene content, and generally, a similar gene order allowing the proposal of an ancestral gene order. The principal exceptions are several orders within the Hemipteroid assemblage including the order Thysanoptera, a sister group of the order Hemiptera. Within the Hemiptera, there are available a number of completely sequenced mitochondrial genomes that have a gene order similar to that of the proposed ancestor. None, however, are available from the suborder Sternorryncha that includes whiteflies, psyllids and aphids.ResultsWe have determined the complete nucleotide sequence of the mitochondrial genomes of six species of whiteflies, one psyllid and one aphid. Two species of whiteflies, one psyllid and one aphid have mitochondrial genomes with a gene order very similar to that of the proposed insect ancestor. The remaining four species of whiteflies had variations in the gene order. In all cases, there was the excision of a DNA fragment encoding for cytochrome oxidase subunit III(COIII)-tRNAgly-NADH dehydrogenase subunit 3(ND3)-tRNAala-tRNAarg-tRNAasn from the ancestral position between genes for ATP synthase subunit 6 and NADH dehydrogenase subunit 5. Based on the position in which all or part of this fragment was inserted, the mitochondria could be subdivided into four different gene arrangement types. PCR amplification spanning from COIII to genes outside the inserted region and sequence determination of the resulting fragments, indicated that different whitefly species could be placed into one of these arrangement types. A phylogenetic analysis of 19 whitefly species based on genes for mitochondrial cytochrome b, NADH dehydrogenase subunit 1, and 16S ribosomal DNA as well as cospeciating endosymbiont 16S and 23S ribosomal DNA indicated a clustering of species that corresponded to the gene arrangement types.ConclusionsIn whiteflies, the region of the mitochondrial genome consisting of genes encoding for COIII-tRNAgly-ND3-tRNAala-tRNAarg-tRNAasncan be transposed from its ancestral position to four different locations on the mitochondrial genome. Related species within clusters established by phylogenetic analysis of host and endosymbiont genes have the same mitochondrial gene arrangement indicating a transposition in the ancestor of these clusters.

The eubacterial endosymbionts of whiteflies (homoptera: Aleyrodoidea) constitute a lineage distinct from the endosymbionts of aphids and mealybugs

Whiteflies (superfamily Aleyrodoidea) contain eubacterial endosymbionts localized within host cells known as mycetocytes. Sequence analysis of the genes for the 16S rRNA of the endosymbionts ofBemisia tabaci, Siphoninus phillyreae, andTrialeurodes vaporariorum indicates that these organisms are closely related and constitute a distinct lineage within the γ-subdivision of theProteobacteria. The endosymbionts of whiteflies are unrelated to the endosymbionts of aphids and mealybugs, which are in two separate lineages.

Levels of Buchnera aphidicola Chaperonin GroEL During Growth of the Aphid Schizaphis graminum

Abstract.Buchnera aphidicola is the prokaryotic, intracellular symbiont found in the aphid Schizaphis graminum. Using an immunological approach, we have quantitated the amount of the B. aphidicola chaperonin, GroEL, present in aphid cell-free extracts during the growth cycle of S. graminum at 23°C. Our results indicate that the increase in GroEL approximately follows the increase in aphid weight and endosymbiont number for the first 12 days after birth of the aphid. A 9-day-old aphid contains 1.6 × 105 molecules of GroEL per μm3 of cell volume. This number is similar to that found in Escherichia coli growing at 46°C, close to its maximal growth temperature, and a condition at which there is a major increase in the levels of chaperonins and other stress proteins. It is estimated that at 23°C, 10% of the B. aphidicola protein is GroEL. When S. graminum grown at 23°C was shifted to 33°C for 1 day and subsequently to 23°C, there was no change in the level of GroEL or the rate of growth. It is possible that the high level of GroEL in the endosymbiont masked an increase in the protein owing to a heat shock response.

Secondary endosymbionts of psyllids have been acquired multiple times

Previous studies have established that psyllids (Hemiptera, Psylloidea) contain primary endosymbionts, designated as Carsonella ruddii, which cospeciate with the psyllid host. This association appears to be the consequence of a single infection of a psyllid ancestor with a bacterium. Some psyllids may have additional secondary (S-) endosymbionts. We have cloned and sequenced the 16S–23S ribosomal RNA genes of seven representative psyllid S-endosymbionts. Comparison of the S-endosymbiont phylogenetic trees with those of C. ruddii indicates a lack of congruence, a finding consistent with multiple infections of psyllids with different precursors of the S-endosymbionts and/or possible horizontal transmission. Additional comparisons indicate that the S-endosymbionts are related to members of the Enterobacteriaceae as well as to several other endosymbionts and insect-associated bacteria.

The Evolution and Genetics of Aphid Endosymbionts

O ne of the striking attribute.s of prokaryotes (boch Bacteria and Archaea) is the dlversity of thei! catabolic pathways and ehe ir biosynthetic capabilities (Brock er al. 1994). Many prnkaryores are able co L1se unusual substrates for growth and synthesize a11 of the constituenes of cells from relative!y simple compounds. In contrast, many eukaryotcs lack such capa bi litics and ha ve developed elose associations in which they take advantage of thc metabolie versatility of prokaryotes. In some of these associations, the organisms live in elose contace bm remain separate. In orher cases, called endosymbioses, the prokaryote i5 scqucstered \vithin the eukaryotic eell. The c1assicaJ eompilation of endosymbiotic assoClations is that of suchner (1965). Inseets are particularly prone ta endosymbiotic associatiom .. Such associ~tions are widespread among members of the orders Homoptera (a phids, whiteflies, mealybugs, psylIid!>, and cicadas), Blattaria (cock-

Description of Deleya gen. nov. Created to Accommodate the Marine Species Alcaligenes aestus, A. pacificus, A. cupidus, A. venustus, and Pseudomonas marina

Recent investigations of the evolutionary relationships of marine species assigned to the genera Alcaligenes (Alcaligenes aestus, Alcaligenes pacificus, Alcaligenes cupidus, and Alcaligenes venustus) and Pseudomonas (Pseudomonas marina) are reviewed. Data obtained by a number of different methods indicate that these five species are part of a single evolutionary lineage distinct from their close relatives, the fluorescent pseudomonads, and unrelated to the type species of Alcaligenes (Alcaligenes faecalis). Consequently, all five species are transferred to a new genus, Deleya, with Deleya aesta designated as the type species. The phenotype properties of D. aesta, Deleya pacifica, Deleya cupida, Deleya venusta, and Deleya marina are presented, together with the distinguishing traits of these organisms.

Phylogenetic evidence for two new insect-associated Chlamydia of the family Simkaniaceae.

On the basis of 16S–23S ribosomal DNA analyses, the whitefly Bemisia tabaci (Sternorrhyncha, Aleyrodidae) and the eriococcid Eriococcus spurius (Sternorrhyncha, Eriococcidae) were each found to harbor novel related chlamydial species within the family Simkaniaceae. The generic designation Fritschea gen. nov. is proposed to accommodate the two species, F. bemisiae sp. nov. and F. eriococci sp. nov. The finding of chlamydial 16S–23S ribosomal DNA in B. tabaci is consistent with a previous electron microscopy study which found that bacteriocytes of this species contain structures that we consider to resemble the elementary and reticulate bodies of chlamydia (Costa HS, Westcot DM, Ullman DE, Rosell R, Brown JK, Johnson MW. Protoplasma 189:194–202, 1995). The cloning and sequencing of a 16.6 kilobase DNA fragment from F. bemisiae indicated that it contains six genes encoding for proteins similar to those found in other species of chlamydia. These results extend the range of organisms that harbor chlamydia.

The Tryptophan Biosynthetic Pathway of Aphid Endosymbionts (Buchnera): Genetics and Evolution of Plasmid-Associated Anthranilate Synthase (trpEG) Within the Aphididae

The bacterial endosymbionts (Buchnera) from the aphidsRhopalosiphum padi, R. maidis, Schizaphis graminum, andAcyrthosiphon pisum contain the genes for anthranilate synthase (trpEG) on plasmids made up of one or more 3.6-kb units. Anthranilate synthase is the first as well as the rate-limiting enzyme in the tryptophan biosynthetic pathway. The amplification oftrpEG on plasmids may result in an increase of enzyme protein and overproduction of this essential amino acid, which is required by the aphid host. The nucleotide sequence oftrpEG from endosymbionts of different species of aphids is highly conserved, as is an approximately 500-bp upstream DNA segment which has the characteristics of an origin of replication. Phylogenetic analyses were performed usingtrpE andtrpG from the endosymbionts of these four aphids as well as from the endosymbiont ofSchlechtendalia chinensis, in whichtrpEG occurs on the chromosome. The resulting phylogeny was congruent with trees derived from sequences of two chromosome-located bacterial genes (part oftrpB and 16S ribosomal DNA). In turn, trees obtained from plasmid-borne and bacterial chromosome-borne sequences were congruent with the tree resulting from phylogenetic analysis of three aphid mitochondrial regions (portions of the small and large ribosomal DNA subunits, as well as cytochrome oxidase II). Congruence of trees based on genes from host mitochondria and from bacteria adds to previous support for exclusively vertical transmission of the endosymbionts within aphid lineages. Congruence with trees based on plasmid-borne genes supports the origin of the plasmid-bornetrpEG from the chromosomal genes of the same lineage and the absence of subsequent plasmid exchange among endosymbionts of different species of aphids.