Adi Behar

Enterobacteria-mediated nitrogen fixation in natural populations of the fruit fly Ceratitis capitata

Nitrogen, although abundant in the atmosphere, is paradoxically a limited resource for multicellular organisms. In the Animalia, biological nitrogen fixation has solely been demonstrated in termites. We found that all individuals of field‐collected Mediterranean fruit flies (Ceratitis capitata) harbour large diazotrophic enterobacterial populations that express dinitrogen reductase in the gut. Moreover, nitrogen fixation was demonstrated in isolated guts and in live flies and may significantly contribute to the flys nitrogen intake. The presence of similar bacterial consortia in additional insect orders suggests that nitrogen fixation occurs in vast pools of terrestrial insects. On such a large scale, this phenomenon may have a considerable impact on the nitrogen cycle.

Bringing back the fruit into fruit fly–bacteria interactions

Female Mediterranean fruit flies (Ceratitis capitata) oviposit in fruits, within which the larvae develop. This development is associated with rapid deterioration of the fruit, and frequently with invasion by secondary pests. Most research on the associations between medflies and microorganisms has focused on the bacteria inhabiting the digestive system of the adult fly, while the role of the fruit in mediating, amplifying or regulating the fruit fly microflora has been largely neglected. In this study, we examine the hypothesis that the host fruit plays a role in perpetuating the fly‐associated bacterial community. Using direct and cultured‐based approaches, we show that this community is composed in its very large majority of diazotrophic and pectinolytic Enterobacteriaceae. Our data suggest that this fly‐associated enterobacterial community is vertically transmitted from the female parent to its offspring. During oviposition, bacteria are transferred to the fruit, establish and proliferate within it, causing its decay. These results show that the host fruit is indeed a central partner in the fruit fly–bacterial interaction as these transmitted bacteria are amplified by the fruit, and subsequently maintained throughout the flys life. This enterobacterial community may contribute to the flys nitrogen and carbon metabolism, affecting its development and ultimately, fitness.

Phylogenetic diversity of bacteria associated with the mucus of Red Sea corals.

Coral reefs are the most biodiverse and biologically productive of all marine ecosystems. Corals harbor diverse and abundant prokaryotic communities. However, little is known about the diversity of coral-associated bacterial communities. Mucus is a characteristic product of all corals, forming a coating over their polyps. The coral mucus is a rich substrate for microorganisms. Mucus was collected with a procedure using sterile cotton swabs that minimized contamination of the coral mucus by surrounding seawater. We used molecular techniques to characterize and compare the bacterial assemblages associated with the mucus of the solitary coral Fungia scutaria and the massive coral Platygyra lamellina from the Gulf of Eilat, northern Red Sea. The bacterial communities of the corals F. scutaria and P. lamellina were found to be diverse, with representatives within the Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria, Deltaproteobacteria and Epsilonproteobacteria, as well as the Actinobacteria, Cytophaga-Flavobacter/Flexibacter-Bacteroides group, Firmicutes, Planctomyces, and several unclassified bacteria. However, the total bacterial assemblage of these two corals was different. In contrast to the bacterial communities of corals analyzed in previous studies by culture-based and culture-independent approaches, we found that the bacterial clone libraries of the coral species included a substantial proportion of Actinobacteria. The current study further supports the finding that bacterial communities of coral mucus are diverse.

Rickettsia felis Infection in a Common Household Insect Pest, Liposcelis bostrychophila (Psocoptera: Liposcelidae)

ABSTRACT Many species of Rickettsia are well-known mammalian pathogens transmitted by blood-feeding arthropods. However, molecular surveys are continually uncovering novel Rickettsia species, often in unexpected hosts, including many arthropods that do not feed on blood. This study reports a systematic molecular characterization of a Rickettsia infecting the psocid Liposcelis bostrychophila (Psocoptera: Liposcelidae), a common and cosmopolitan household pest. Surprisingly, the psocid Rickettsia is shown to be Rickettsia felis, a human pathogen transmitted by fleas that causes serious morbidity and occasional mortality. The plasmid from the psocid R. felis was sequenced and was found to be virtually identical to the one in R. felis from fleas. As Liposcelis insects are often intimately associated with humans and other vertebrates, it is speculated that they acquired R. felis from fleas. Whether the R. felis in psocids causes disease in vertebrates is not known and warrants further study.

Phylogenetic, metabolic, and taxonomic diversities shape mediterranean fruit fly microbiotas during ontogeny.

ABSTRACT The Mediterranean fruit fly (medfly) (Ceratitis capitata) lays eggs in fruits, where larvae subsequently develop, causing large-scale agricultural damage. Within its digestive tract, the fly supports an extended bacterial community that is composed of multiple strains of a variety of enterobacterial species. Most of these bacteria appear to be functionally redundant, with most strains sustaining diazotrophy and/or pectinolysis. At least some of these bacteria were shown to be vertically inherited, but colonization, structural, and metabolic aspects of the communitys dynamics have not been investigated. We used fluorescent in situ hybridization, metabolic profiling, plate cultures, and pyrosequencing to show that an initial, egg-borne, diverse community expands throughout the flys life cycle. While keeping “core” diazotrophic and pectinolytic functions, it also harbors diverse and fluctuating populations that express varied metabolic capabilities. We suggest that the metabolic and compositional plasticity of the flys microbiota provides potential adaptive advantages to the medfly host and that its acquisition and dynamics are affected by mixed processes that include stochastic effects, host behavior, and molecular barriers.

Bacteria–diet interactions affect longevity in the medfly –Ceratitis capitata

Mediterranean fruit flies (Ceratitis capitata Wiedemann, Dipt.: Tephritidae) harbour a diverse community of bacteria in their digestive system. This microbiota may have important functions impacting on the fly’s fitness. Recently, we described the effect of eliminating intestinal bacteria on the reproductive success of C. capitata males and females. Here, we expand the view on the nature of fly–bacteria interactions by examining the effect of bacteria on male and female longevity. Antibiotics were used to suppress the gut bacterial community and mortality rates were compared between antibiotic‐treated and non‐treated flies when either nutritionally stressed (maintained on sugar) or provided with a full diet. These tests revealed that eliminating the gut bacterial population prolonged longevity, but only when flies were nutritionally stressed, indicating that the effect of bacteria on lifespan was diet dependent. Considering these results in light of other known effects of bacteria on fitness components of the fly demonstrates a cost‐benefit relationship between C. capitata and its gut microbiota.

The Mediterranean fruit fly and its bacteria – potential for improving sterile insect technique operations

Mediterranean fruit flies (Ceratitis capitata Wiedemann, Diptera: Tephritidae), harbour a diverse community of bacteria in their digestive system. Molecular and culture‐based techniques show that members of the Enterobacteriaceae form the dominant populations in the gut of the Mediterranean fruit fly. Among them, many are diazotrophs and actively fix nitrogen in vivo. Most prominent are Klebsiella spp., Enterobacter spp., Pectobacterium spp. Citrobacter freundii and Providencia stuartii. A marked shift in community composition was observed between different developmental stages: in larvae, the pectinolytic Pectobacterium were most abundant, suggesting that pectinolysis plays a role early during the fly’s life. Additionally, pseudomonads, some of which are known entomopathogens, constitute a minor, yet common and stable community in the C. capitata gut. Microbial communities in VIENNA 8 flies, a mass rearing genetic sexing strain, differ from wild flies, and irradiation further affects the microbial community. We found increased levels of the pathogenic species Pseudomonas in the industrially used strain. Furthermore, although members of the Enterobacteriaceae family remain the dominant bacteria group present in the fly’s gut, the levels of Klebsiella species decrease significantly in the days after irradiation. Eliminating the bacterial population in normal flies by using antibiotics affects measurable physiological and behavioural parameters related to fitness. Finally, we tested the hypothesis that inoculation of sterile flies with members of the original bacterial community results in enhanced competitiveness. We found that addition of the bacteria Klebsiella oxytoca to the post‐irradiation diet significantly improves sterile male performance.

Community Structure of the Mediterranean Fruit Fly Microbiota: Seasonal and Spatial Sources of Variation

Fruit flies (Diptera: Tephritidae) maintain diverse yet stable communities of Enterobacteriaceae that reside in their digestive system. Focusing on one species of this family, Ceratitis capitata, the Mediterranean fruit fly (medfly), we applied the PCR-DGGE approach to study the influence of temporal and spatial variations on the composition and diversity of the enterobacterial community associated with the medfly. Our results revealed that the Enterobacteriaceae constituted not only dominant, but also constant and stable populations in the medflys gut. Nevertheless, changes in both time and space had an impact on the structure and diversity of these bacterial populations, and while some populations (e.g., Klebsiella spp.) were stable, others fluctuated. Whether the bacterial population fluctuated according to the ecological needs of the insect during different environmental conditions, or reflected opportunistic colonization abilities of some bacterial strains, remains to be determined.

Morphological and molecular characterization of a sexually reproducing colony of the booklouse Liposcelis bostrychophila (Psocodea: Liposcelididae) found in Arizona

The booklouse, Liposcelis bostrychophila, is a worldwide pest of stored products. For decades, only thelytokous parthenogenetic reproduction was documented in L. bostrychophila. Male L. bostrychophila were first found in Hawaii in 2002. In 2009, a sexual strain was found in Arizona. We examined the morphology of both males and females of the Arizona strain and compared the Arizona sexual strain with the Hawaii sexual strain and the parthenogenetic strains of L. bostrychophila. The sexual and parthenogenetic strains show some differences in eye morphology. To examine the relationship between sexual and asexual lineages, we sequenced the mitochondrial 12S and 16S ribosomal RNA genes of males and females from the Arizona strain. Phylogenetic analyses of L. bostrychophila individuals revealed that: 1) the sexually reproducing colony found in Arizona contains two closely related mitochondrial DNA haplotypes – one present in only females and the other in both males and females; and 2) the Arizona sexual strain was most closely related to a parthenogenetic strain in Illinois. We detected Rickettsia in all of the parthenogenetic individuals we checked but not in any Arizona sexual individuals. Further evidence is required to establish whether the presence of Rickettsia is linked to asexual reproduction in Liposcelis.

Tri-Party Underground Symbiosis between a Weevil, Bacteria and a Desert Plant

Inhabitants of arid ecosystems face severe nitrogen and water limitations. Inventive adaptations by organisms occupying such habitats are essential for survival. This study describes a tri-party symbiotic interaction between a plant (Salsola inermis), a beetle (Conorhynchus pistor), and a bacterium (Klebsiella pneumonia). The weevil survives by living within a mud structure affixed to the plant roots, thus benefiting from increased carbon and water, and refuge from predators and parasites. Active nitrogen-fixing bacteria harbored within the weevils gut mediate this interaction, by supplying nitrogen to the system, which eventually promotes seed development. We studied the correlation between the weevils existence and (i) root carbon and nitrogen content, (ii) soil water content and (iii) seed weight. Roots hosting weevils contained more nitrogen, heavier seeds and less carbon. In addition, water content was higher around the roots than in open spaces a short distance from the plant stem. Bacterial studies and nitrogen-fixation analyses, including molecular and chemical assays, indicated atmospheric nitrogen fixation in the larval stage and identified the bacterium. The coexistence of weevil and bacterial behavior coinciding with the plants life cycle was revealed here by a long period of field observations. Out of over 60,000 known weevils, this is the only report of a weevil living most of its life underground without harming plants. The unique tri-party interaction described herein shows the important ecological role of desert plant roots and provides an example of a sustainable consortium of living organisms coping with the challenging desert environment.