Subcellular degeneration of mycetomal endocytobionts in tsetse,Glossina morsitans morsitans, inoculated twice withEscherichia coliJura, Walter; Kaaya, Godwin
doi: 10.1007/BF01570880pmid: N/A
Specimens of mycetome, a portion of anterior midgut harboring intracellular bacterioids (endocytobionts), obtained from both untreated control female tsetse,Glossina morsitans morsitans, and those inoculated twice with strain D31 ofEscherichia coli, were processed for routine electron microscopy, and the endocytobionts were examined for structural alterations. In the controls, mycetocytes contained intact bacterioids with numerous, electron-dense ribosomal particles in the cytoplasm. FemaleG. m. morsitans subjected to two hemocoelic inoculations with the liveE. coli showed severe degeneration of the subcellular components of the endocytobionts characterized by advanced lysis and rarefaction. The observed endocytobiotic degeneration is attributed to effects of induced humoral antibacterial factors.
A novel, symbiotic bacterium isolated from marine shipworm secretes proteolytic activityGreene, Richard; Cotta, Michael; Griffin, Harold
doi: 10.1007/BF01570881pmid: N/A
Proteolytic activity was identified in cultures of the marine shipworm bacterium by monitoring of the release of acid-soluble azopeptides from azocasein. Activity was predominantly extracellular (>80%), and its production was coincident with logarithmic cell growth. The protease(s) appeared to be constitutive, since it was present even when the bacterium was grown under nitrogen-fixing conditions. However, activity was stimulated up to 8.6-fold by the addition of complex nitrogen (casein, amino acids) to the growth medium. Maximum activity was observed at 40°C and between pH 6.5 and 9.0. Relatively low concentrations (0.1 mM) of phenylmethylsulfonyl fluoride (PMSF) abolished activity, indicative of a serine protease(s).
ELISA and dot-blot detection of flavescence dorée-MLO in individual leafhopper vectors during latency and inoculative stateBoudon-Padieu, Elisabeth; Larrue, Jean; Caudwell, Antoine
doi: 10.1007/BF01570882pmid: N/A
Flavescence dorée, an MLO (mycoplasma-like organism) disease of grapevine, is vectored in the field by the leafhopperScaphoïdeus littoralis, and in the laboratory fromVicia faba toVicia faba by the leafhopperEuscelidius variegatus. Antibodies to partially purified MLOs from broadbeans orE. variegatus were raised in rabbits. ELISA allowed individual assay of leafhoppers almost without background to healthy leafhopper control. Positive ELISA reading was associated with biological transmission and demonstrated the infection in noninoculative leafhoppers, even during latency. Thus, ELISA detection was fast and more accurate than inoculative state. A first multiplication of antigen, leading to infectivity of leafhoppers, was followed by a fast and strong multiplication during the following weeks. Males were particularly affected. Differential acquisition between individuals was shown. Application toS. titanus naturally contaminated in the field allows survey of epidemic outbreak and search for other vector species.
Evaluation of a method to measure conjugal transfer of recombinant DNA in soil slurriesWalter, Michael; Porteous, L.; Seidler, Ramon
doi: 10.1007/BF01570883pmid: N/A
Release of recombinant microbes into the environment necessitates an evaluation of their ability to transfer genetic material. The present report evaluates a method to detect conjugal DNA plasmid transfer in soil slurries under various environmental conditions. DonorPseudomonas cepacia containing pR388::Tn1721 andP. cepacia recipient cultures were coincubated in soil slurries containing autoclaved or natural soil and treated with one or more of 14 experimental conditions. Conjugal mating frequency (transconjugants per initial donor) ranged from 4.8×10−1 to 1.9×10−7. Highest numbers of transconjugants, 1.5×107 colony forming units/ml soil slurry, were observed following incubation at 35°C with an enriched nutrient supplement added to the soil. Low numbers of transconjugants, 103 colony forming units/ml soil slurry, were observed when mating pairs were subjected to low nutrient or pH stress even though initial donor and recipient populations were maintained at high levels. This test system provides a simple way to estimate effects of changing environmental factors on plasmid transfer rates and on the survival of recombinant microorganisms. By use of soil collected from sites proposed to receive genetically engineered microorganisms, preliminary risk assessments can be obtained regarding the potential for gene transfer and microorganism survival with this soil slurry test system.
Tolerance of extremely halophilic archaebacteria towards bromideOren, Aharon; Bekhor, David
doi: 10.1007/BF01570884pmid: N/A
The tolerance of halophilic archaebacteria towards bromide was tested in view of the fact that bromide occurs in natural brines in concentrations of up to 66 mM. It was found that, while concentrations of up to 0.8–1M are tolerated well by all halobacterial types examined, great differences exist between species with respect to bromide tolerance. WhileHalobacterium (H. salinarium, H. halobium, andH. sodomense) andNatronobacterium species are only moderately tolerant,Haloarcula (H. vallismortis, “H. marismortui”), andHaloferax species (H. mediterranei, H. gibbonsii) tolerate higher concentrations.Haloferax volcanii proved extremely tolerant and showed growth in bromide media at very low chloride concentrations (below 50 mM). No correlation was found between bromide tolerance and the bromide concentration in the habitat from which the strains were isolated. Iodide proved much more toxic than bromide. Bromide-tolerant strains also proved relatively resistant to growth inhibition by iodide.
Detection of plasmid RP4 transfer in soil and rhizosphere, and the occurrence of homology to RP4 in soil bacteriaElsas, Jan; Nikkel, Marcel; Overbeek, Leo
doi: 10.1007/BF01570885pmid: N/A
Plasmid RP4 transfer between introduced pseudomonads was studied in non-rhizosphere and rhizosphere soil. The addition of nutrients to the non-rhizosphere soil stimulated plasmid transfers between introduced donor and recipient cells, and no transfer was detected in nonamended soil. Transfer was also detected in soil in a model rhizosphere, but not in corresponding non-rhizosphere soil. Colony hybridization with whole plasmid RP4 DNA as a probe was employed to detect transfers to indigenous organisms in soil. Although transfers to introduced recipient cells were easily detected in parallel controls, no indigenous organisms were identified that had received RP4. Background levels of soil organisms with the RP4 resistance pattern were considerable, and about 10% of these populations contained DNA sequences with homology to RP4. However, no plasmids could be detected in any of 20 isolates, nor was resistance transfer to aPseudomonas fluorescens recipient detected in filter matings.
Identification of surface proteins ofSpiroplasma citri with protease and antibody probesFletcher, Jacqueline; Wills, John; Denman, Sue
doi: 10.1007/BF01570886pmid: N/A
Spiroplasma citri, a helical, wall-less prokaryote, is an insect-borne phytopathogen. Though proteins having domains on the surface ofS. citri cells may be important in pathogenicity or transmissibility, only one surface protein, spiralin (29 KDa), has previously been identified. Intact cells of strain BR3 were treated with chymotrypsin, proteinase K, or trypsin, and the surviving proteins were analyzed by SDS-PAGE. Seven proteins, in addition to spiralin, were degraded, indicative of surface exposure of those polypeptides. Surface immunoprecipitation (SIP) was used to test accessibility of the proteins to anti-S. citri membrane serum, another indication of surface exposure. With unlabeled cells, five such proteins were identified. Four of these have sizes that correspond to those seen with protease treatments. When125I surfacelabeled spiroplasmas were used for SIP, twelve surface proteins were detected, eight of which correspond to bands identified by the other methods. A protein of 89 KDa in strain BR3 was not universally detected in otherS. citri strains and spiroplasma species.