A rapid and simple micro-assay to assess catalase activity in individual mosquito tissues.

Oxidative stress generated as a normal byproduct of aerobic metabolism is minimized by the enzyme catalase (CAT; EC 1.11.1.

Preparation of Infectious Wolbachia from a Mosquito Cell Line.

Eventual genetic engineering of Wolbachia will require maximizing recovery of infectious bacteria, maintaining Wolbachia in a viable state for efficient manipulation, and reinfection of host cells for propagation and expansion of recombinant progeny. Challenges to manipulating Wolbachia arise from its obligate intracellular lifestyle and inability to divide outside a host cell, requiring modifications of standard bacteriological methods. The Aedes albopictus C7-10 cell line has proven to be a good recipient for the Wolbachia supergroup B strain, wStri, from the planthopper Laodelphax striatellus; the persistently infected C/wStri1 population provides a source of wStri inoculum that can be used systematically to explore conditions that increase yields of infectious material from input Wolbachia and identify conditions conducive to Wolbachia replication.

Wolbachia: Advancing into a Second Century.

Wolbachia pipientis had its scientific debut nearly a century ago and has recently emerged as a target for therapeutic treatment of filarial infections and an attractive tool for control of arthropod pests. Wolbachia was known as a biological entity before DNA was recognized as the molecule that carries the genetic information on which life depends, and before arthropods and nematodes were grouped in the Ecdysozoa. Today, some investigators consider Wolbachia the most abundant endosymbiont on earth, given the numbers of its hosts and its diverse mutualistic, commensal, and parasitic roles in their life histories.

Paraquat is toxic to the soil-dwelling arthropod, Folsomia candida (Collembola: Isotomidae), and has potential effects on its Wolbachia endosymbiont.

The springtail, Folsomia candida, is a soil arthropod commonly used to evaluate environmental toxins. Conflicting data on the toxicity of the herbicide paraquat prompted re-evaluation of its effects on F. candida survival and reproduction.

Mitotically inactivated mosquito cells support robust Wolbachia infection and replication.

Wolbachia is an obligate intracellular bacterium that infects many species of insects, and has been of particular interest in recent efforts to reduce disease transmission by mosquitoes. Two aspects of Wolbachia biology underlie its applications for insect control: first, the bacterium behaves as a natural gene drive agent and, second, when introduced into mosquitoes that do not harbor Wolbachia in nature, infection reduces survival of pathogens. These properties support efforts to explore the basic biology of Wolbachia in insect cell lines, which can produce sufficient infectious material for microbiological studies and microinjection into novel hosts.

From Mosquito Ovaries to Ecdysone; from Ecdysone to : One Woman's Career in Insect Biology.

In anautogenous mosquitoes, synchronous development of terminal ovarian follicles after a blood meal provides an important model for studies on insect reproduction. Removal and implantation of ovaries, in vitro culture of dissected tissues and immunological assays for vitellogenin synthesis by the fat body showed that the (L.) (Diptera, Culicidae) mosquito ovary produces a factor essential for egg production.

Muramidase, nuclease, or hypothetical protein genes intervene between paired genes encoding DNA packaging terminase and portal proteins in Wolbachia phages and prophages.

Genomes of the obligate intracellular alpha proteobacterium Wolbachia pipientis often encode prophage-like regions, and in a few cases, purified particles have been recovered. Because the structure of a conserved WO phage genome has been difficult to establish, we examined paired terminase and portal genes in Wolbachia phages and prophages, relative to those encoded by the gene transfer agent RcGTA from the free-living alpha proteobacterium Rhodobacter capsulatus. Terminase and portal proteins from Wolbachia have higher similarity to orthologs encoded by RcGTA than to orthologs encoded by bacteriophage lambda.

Growth and Maintenance of in Insect Cell Lines.

The obligate intracellular microbe, (Rickettsiales; Anaplasmataceae), is a Gram-negative member of the alpha proteobacteria that infects arthropods and filarial worms. Although closely related to the genera and , which include pathogens of humans, is uniquely associated with invertebrate hosts in the clade Ecdysozoa. Originally described in mosquitoes, is currently represented by 17 supergroups and is believed to occur in half of all insect species.

DNA recombination and repair in Wolbachia: RecA and related proteins.

Wolbachia is an obligate intracellular bacterium that has undergone extensive genomic streamlining in its arthropod and nematode hosts. Because the gene encoding the bacterial DNA recombination/repair protein RecA is not essential in Escherichia coli, abundant expression of this protein in a mosquito cell line persistently infected with Wolbachia strain wStri was unexpected. However, RecA's role in the lytic cycle of bacteriophage lambda provides an explanation for retention of recA in strains known to encode lambda-like WO prophages.

Computational evidence for antitoxins associated with RelE/ParE, RatA, Fic, and AbiEii-family toxins in Wolbachia genomes.

Wolbachia is an obligate intracellular Gram-negative alpha-proteobacterium that has diverse effects on reproduction of arthropod hosts, including cytoplasmic incompatibility, male killing, feminization, and parthenogenesis. Some of these effects have important potential for control of insect pests, including mosquitoes that vector pathogens of humans. In mosquitoes, and in most other arthropods, elimination of Wolbachia by antibiotic treatment has no effect on host survival and reverses the Wolbachia-associated phenotype.

Conditions facilitating infection of mosquito cell lines with Wolbachia, an obligate intracellular bacterium.

Factors that influence establishment of Wolbachia, an obligate intracellular bacterium, in novel insect hosts or uninfected insect cell lines are poorly understood. Infectivity of Wolbachia strain wStr was correlated with flow cytometric profiles to define optimal conditions for harvesting an infectious inoculum. Wolbachia recovered from the cell culture supernatant after gentle pipetting of infected cells represented about 1% of the total bacterial population and were more infectious than Wolbachia that remained associated with intact cells and/or membranes after low-speed centrifugation.

Neonicotinoid insecticide hydrolysis and photolysis: Rates and residual toxicity.

Neonicotinoid insecticides are the most widely used class of insecticides worldwide. Concern has grown over their widespread environmental presence and potential unintended adverse effects. The present study examined hydrolysis and photolysis reaction rates of neonicotinoids and assessed any residual toxicity of reaction products.

Strain-specific response to ampicillin in Wolbachia-infected mosquito cell lines.

Wolbachia pipientis (Rickettsiales; Anaplasmataceae) is an obligate intracellular alpha proteobacterium that occurs in arthropods and filarial worms. Some strains of Wolbachia can be maintained as persistent infections in insect cell lines. C/wStr1 cells from the mosquito Aedes albopictus maintain a robust infection with Wolbachia strain wStr, originally isolated from the planthopper, Laodelphax striatellus.

The Wolbachia WO bacteriophage proteome in the Aedes albopictus C/wStr1 cell line: evidence for lytic activity?

Wolbachia pipientis (Rickettsiales), an obligate intracellular alphaproteobacterium in insects, manipulates host reproduction to maximize invasion of uninfected insect populations. Modification of host population structure has potential applications for control of pest species, particularly if Wolbachia can be maintained, manipulated, and genetically engineered in vitro. Although Wolbachia maintains an obligate mutualism with genome stability in nematodes, arthropods can be co-infected with distinct Wolbachia strains, and horizontal gene transfer between strains is potentially mediated by WO phages encoded within Wolbachia genomes.

Mosaic composition of ribA and wspB genes flanking the virB8-D4 operon in the Wolbachia supergroup B-strain, wStr.

The obligate intracellular bacterium, Wolbachia pipientis (Rickettsiales), is a widespread, vertically transmitted endosymbiont of filarial nematodes and arthropods. In insects, Wolbachia modifies reproduction, and in mosquitoes, infection interferes with replication of arboviruses, bacteria and plasmodia. Development of Wolbachia as a tool to control pest insects will be facilitated by an understanding of molecular events that underlie genetic exchange between Wolbachia strains.

Effects of mimosine on Wolbachia in mosquito cells: cell cycle suppression reduces bacterial abundance.

The plant allelochemical L-mimosine (β-[N-(3-hydroxy-4-pyridone)]-α-aminopropionic acid; leucenol) resembles the nonessential amino acid, tyrosine. Because the obligate intracellular alphaproteobacterium, Wolbachia pipientis, metabolizes amino acids derived from host cells, the effects of mimosine on infected and uninfected mosquito cells were investigated. The EC50 for mimosine was 6-7 μM with Aedes albopictus C7-10 and C/wStr cell lines, and was not influenced by infection status.

Flow cytometric evaluation of the intracellular bacterium, Wolbachia pipientis, in mosquito cells.

Conditions for flow cytometric evaluation of the intracellular bacterium, Wolbachia pipientis, in infected mosquito cells are described. This approach will streamline investigation of Wolbachia's interactions with host cells and facilitate identification of culture conditions that select for Wolbachia-infected cells.

Proteomic profiling of a robust Wolbachia infection in an Aedes albopictus mosquito cell line.

Wolbachia pipientis, a widespread vertically transmitted intracellular bacterium, provides a tool for insect control through manipulation of host-microbe interactions. We report proteomic characterization of wStr, a Wolbachia strain associated with a strong cytoplasmic incompatibility phenotype in its native host, Laodelphax striatellus. In the Aedes albopictus C/wStr1 mosquito cell line, wStr maintains a robust, persistent infection.

Depletion of host cell riboflavin reduces Wolbachia levels in cultured mosquito cells.

Wolbachia is an obligate intracellular alphaproteobacterium that occurs in arthropod and nematode hosts. Wolbachia presumably provides a fitness benefit to its hosts, but the basis for its retention and spread in host populations remains unclear. Wolbachia genomes retain biosynthetic pathways for some vitamins, and the possibility that these vitamins benefit host cells provides a potential means of selecting for Wolbachia-infected cell lines.

Detection of the Wolbachia protein WPIP0282 in mosquito spermathecae: implications for cytoplasmic incompatibility.

Cytoplasmic incompatibility (CI) is a conditional sterility induced by the bacterium Wolbachia pipientis that infects reproductive tissues in many arthropods. Although CI provides a potential tool to control insect vectors of arthropod-borne diseases, the molecular basis for CI induction is unknown. We hypothesized that a Wolbachia-encoded, CI-inducing factor would be enriched in sperm recovered from spermathecae of female mosquitoes.

The oxidizing agent, paraquat, is more toxic to Wolbachia than to mosquito host cells.

Cultured cells provide an important in vitro system for examining metabolic interactions between the intracellular bacterium, Wolbachia pipientis, and its insect hosts. To test whether Wolbachia-associated changes in antioxidant activities could provide a tool to select for infected cells, we tested the effects of paraquat (PQ) on Aedes albopictus mosquito cells. Like mammalian cells, mosquito cells tolerate PQ over a wide range of concentrations, and for considerable lengths of time, depending on cell density at the time of treatment.

Detection of the Wolbachia-encoded DNA binding protein, HU beta, in mosquito gonads.

Wolbachia are obligate intracellular bacteria that cause cytoplasmic incompatibility in mosquitoes. In an incompatible cross, eggs of uninfected females fail to hatch when fertilized by sperm from infected males. We used polyacrylamide gel electrophoresis and tandem mass spectrometry to identify Wolbachia proteins in infected mosquito gonads.