Isolation, Pure Culture, and Characterization of “ Candidatus Arsenophonus arthropodicus,” an Intracellular Secondary Endosymbiont from the Hippoboscid Louse Fly Pseudolynchia canariensis

Abstract

Members of the genus Arsenophonus comprise a large group of bacterial endosymbionts that are widely distributed in arthropods of medical, veterinary, and agricultural importance. At present, little is known about the role of these bacteria in arthropods, because few representatives have been isolated and cultured in the laboratory. In the current study, we describe the isolation and pure culture of an Arsenophonus endosymbiont from the hippoboscid louse fly Pseudolynchia canariensis. We propose provisional nomenclature for this bacterium in the genus Arsenophonus as "Candidatus Arsenophonus arthropodicus." Phylogenetic analyses indicate that "Candidatus Arsenophonus arthropodicus" is closely related to the Arsenophonus endosymbionts found in psyllids, whiteflies, aphids, and mealybugs. The pure culture of this endosymbiont offers new opportunities to examine the role of Arsenophonus in insects. To this end, we describe methods for the culture of "Candidatus Arsenophonus arthropodicus" in an insect cell line and the transformation of this bacterium with a broad-host-range plasmid.Many members of the class Insecta maintain mutualistic associations with one or more specialized symbiotic bacteria (2). Bacteria that participate in these associations are classified either as primary (P) or secondary (S) endosymbionts, because they often coexist in a single insect host. The P-endosymbionts are predicted to be ancient in origin because their phylogenies are concordant with those of their host insects over a substantial period of evolutionary time, indicating long-term coevolution. On the other hand, the S-endosymbionts are predicted to be recent in origin because their phylogenies show little or no concordance with their insect hosts, indicating recent acquisition.While the ancient P-endosymbionts are known to have defined mutualistic functions in their insect hosts, the role of the S-endosymbionts is not yet well understood. From an evolutionary standpoint it seems likely that S-endosymbionts have beneficial (mutualistic) roles in their insect hosts because they are maintained predominantly through a maternal (vertical) transmission strategy. Several recent studies have provided experimental evidence for a number of beneficial effects conferred by the S-endosymbionts of aphids, which recently received new nomenclature (20). These benefits include host plant specialization (17, 27), increased resistance to hymenopteran parasitoids (21, 22), and increased tolerance to heat stress (5, 19). In addition, there is evidence indicating that S-endosymbionts can provide some level of functional compensation for the loss of P-endosymbionts in a laboratory population of aphids (16). While these studies are both exciting and encouraging, the ability to perform experimentation in these systems would be greatly enhanced with the opportunity to genetically manipulate S-endosymbionts. The application of recombinant DNA technology would permit functional analysis of individual genes in the endosymbiont genomes, providing a platform to e...

Reductive genome evolution, host–symbiont co-speciation and uterine transmission of endosymbiotic bacteria in bat flies

“…in the Gammaproteobacteria (Aksoy, 1995). Meanwhile, diverse Arsenophonusrelated endosymbionts have been detected from louse flies and stereblid bat flies (Dale et al, 2006;Trowbridge et al, 2006;Nováková et al, 2009), which are certainly allied to but distinct from the clade of nycteribiid endosymbionts (Figure 2; Nováková et al, 2009). These patterns favor the hypothesis that the hippoboscoid endosymbionts are of multiple evolutionary origins and have experienced repeated acquisitions and/or replacements from different bacterial sources to meet their physiological requirement for their obligate blood-feeding lifestyle.…”

“…The complete genome sequences of Wigglesworthia and Sodalis have already been determined (Akman et al, 2002;Toh et al, 2006). By contrast, much less attention has been paid to the endosymbiotic systems of hippoboscids, streblids and nycteribiids (Dale et al, 2006;Trowbridge et al, 2006;Nováková et al, 2009), except for early histological descriptions (Roubaud, 1919;Zacharias, 1928;Aschner, 1931;Buchner, 1965).…”

Thorsellia anophelis is the dominant bacterium in a Kenyan population of adult Anopheles gambiae mosquitoes

“…Therefore, although the physiological adaptations we describe here help explain the association of T. anophelis with adult mosquitoes that have acquired a blood meal, other factors may be involved in their apparent predominance in this environment. Research on Arsenophonus also suggests that the basis of symbiosis with its host is probably not based on nutrition but on still unknown, but critical, function(s) (Dale et al, 2006). Another group has recently described an association between An.…”

“…Artificial culture has been achieved only on insect cell lines (Hypsa and Dale, 1997;Dale et al, 2006). Different species of Arsenophonus from different hosts share 499% 16S rRNA gene sequence similarity, suggesting recent host acquisition (Dale et al, 2006).…”

Evolutionary origin of insect– Wolbachia nutritional mutualism

“…S6). In the bat fly, the louse fly, the grasshopper, and the termites, the F Wolbachia infections are not fixed in the host populations, suggesting facultative nature of these host-symbiont associations (32)(33)(34)(35). These observations suggest that (i) the bedbug-wCle nutritional mutualism evolved independently of the nematodeWolbachia mutualism within the Wolbachia F supergroup, (ii) the origin of the bedbug-wCle mutualistic association is more recent than the origin of the nematode-Wolbachia mutualistic association, (iii) plausibly, an ancestor of the cimicid bugs acquired wCle from an unrelated arthropod host, (iv) in the donor arthropod, wCle was likely a facultative endosymbiotic associate, and (v) the establishment of the bedbug-wCle association presumably entailed an evolutionary transition from facultative symbiosis to obligate nutritional mutualism for the symbiont side.…”