Coexistence of novel gammaproteobacterial and Arsenophonus symbionts in the scale insect Greenisca brachypodii (Hemiptera, Coccomorpha: Eriococcidae)

Abstract

Scale insects are commonly associated with obligate, intracellular microorganisms which play important roles in complementing their hosts with essential nutrients. Here we characterized the symbiotic system of Greenisca brachypodii, a member of the family Eriococcidae. Histological and ultrastructural analyses have indicated that G. brachypodii is stably associated with coccoid and rod-shaped bacteria. Phylogenetic analyses have revealed that the coccoid bacteria represent a sister group to the secondary symbiont of the mealybug Melanococcus albizziae, whereas the rod-shaped symbionts are close relatives of Arsenophonus symbionts in insects - to our knowledge, this is the first report of the presence of Arsenophonus bacterium in scale insects. As a comparison of 16S and 23S rRNA genes sequences of the G. brachypodii coccoid symbiont with other gammaprotebacterial sequences showed only low similarity (∼90%), we propose the name 'Candidatus Kotejella greeniscae' for its tentative classification. Both symbionts are transovarially transmitted from one generation to the next. The infection takes place in the neck region of the ovariole. The bacteria migrate between follicular cells, as well as through the cytoplasm of those cells to the perivitelline space, where they form a characteristic 'symbiont ball'. Our findings provide evidence for a polyphyletic origin of symbionts of Eriococcidae.

Bacterial associates of Orthezia urticae, Matsucoccus pini, and Steingelia gorodetskia - scale insects of archaeoccoid families Ortheziidae, Matsucoccidae, and Steingeliidae (Hemiptera, Coccomorpha)

“…Since the “new diet” required the support of symbionts, the particular groups of scale insects acquired different symbionts. It should be stressed that recent molecular analyses which show that the symbionts of scale insects belong to different bacterial taxa (von Dohlen et al 2001; Thao et al 2002; Gruwell et al 2007, 2010, 2014; Kono et al 2008; Matsuura et al 2009; Ramirez-Puebla et al 2010; Gatehouse et al 2011; Dhami et al 2012; Rosenblueth et al 2012; Koga et al 2013a; Rosas-Pérez et al 2014; Michalik et al 2016, 2018; Szabo et al 2017; Szklarzewicz et al 2018) strongly support this hypothesis.…”

“…It is commonly known that different groups of scale insects are characterized by diverse methods of symbiont transmission (Buchner 1965; Szklarzewicz and Michalik 2017). In most scale insects, symbionts infect ovaries which contain oocytes during the advanced stage of vitellogenesis, e.g., in members of the family Pseudococcidae and Eriococcidae examined thus far, in Puto superbus (Putoidae), bacteria invade the anterior pole of the vitellogenic ovariole, whereas in Palaeococcus fuscipennis (Monophlebidae) and Porphyrophora polonica (Margarodidae) the posterior pole of the vitellogenic ovariole is infected (Buchner 1965, 1966; von Dohlen et al 2001; Szklarzewicz et al 2006, 2018; Michalik et al 2016, 2018). In Marchalina hellenica (Marchalinidae) and Puto albicans (Putoidae), bacteria infect the undifferentiated germ cells (= cystocytes) constituting the larval ovaries (Szklarzewicz et al 2010, 2013).…”

“…Previous studies based on paraffin technique have revealed that scale insects, as opposed to remaining Sternorrhyncha (aphids, whiteflies, psyllids), are characterized by an enormous diversity of symbiotic associates (reviewed in Walczuch 1932; Buchner 1965; Tremblay 1977). More recent ultrastructural and molecular analyses have confirmed that the symbioses of scale insects are much more diverse than those in the remaining Sternorrhyncha, with respect to the systematic affiliation of symbionts, distribution in the host insect body, and the mode of transmission from the mother to the progeny (Fukatsu and Nikoh 2000; von Dohlen et al 2001; Thao et al 2002; Szklarzewicz et al 2006, 2013, 2018; Niżnik and Szklarzewicz 2007; Kono et al 2008; Matsuura et al 2009; Gruwell et al 2010, 2012; Ramirez-Puebla et al 2010; Gatehouse et al 2011; McCutcheon and von Dohlen 2011; Vashishtha et al 2011; Dhami et al 2012; Rosenblueth et al 2012, 2018; Husnik et al 2013; Koga et al 2013a; Sabree et al 2013; Rosas-Pérez et al 2014; Michalik et al 2016, 2018; Szabo et al 2017). Several families of scale insects, e.g., Steingeliidae, Xylococcidae, Matsucoccidae, Kermesidae, Kuwaniidae, Dactylopiidae, were regarded as asymbiotic (Buchner 1965; Tremblay 1977); however, the results of recent ultrastructural or molecular studies have revealed that some of them, i.e., Steingeliidae, Dactylopiidae, Kermesidae, and Matsucoccidae, may harbor bacterial or yeast-like associates (Koteja et al 2003; Ramirez-Puebla et al 2010; Szklarzewicz et al 2014; Podsiadło et al 2018; Rosenblueth et al 2018).…”

A Symbiont’s Guide to the Germline

“…The nested symbiont cells are transported within bacteriocytes from the symbiont-housing organ (bacteriome) in the abdomen to the ovary. There, they are released from host cells, cluster around the connection between the oocyte and supporting cells (similar to scale insects [89] ), and are taken into the germline at this point [23] .…”

Molecular characterization, ultrastructure, and transovarial transmission of Tremblaya phenacola in six mealybugs of the Phenacoccinae subfamily (Insecta, Hemiptera, Coccomorpha)

“…Since recent genomic analyses indicate that, similarly to the co-primary symbionts in Auchenorrhyncha, the primary and secondary symbionts of Sternorrhyncha complement one another in synthesis of nutrients, they may be regarded as co-primary symbionts (Pérez-Brocal et al 2006; Gosalbes et al 2008; Luan et al 2015). In contrast to aphids, whiteflies, and psyllids, scale insects are characterized by a great diversity of symbionts (Fukatsu and Nikoh 2000; von Dohlen et al 2001; Thao et al 2002; Baumann and Baumann 2005; Szklarzewicz et al 2006, 2018; Gruwell et al 2007, 2010; Niżnik and Szklarzewicz 2007; Matsuura et al 2009; Rosenblueth et al 2012; Husnik and McCutcheon 2016; Michalik et al 2016; Michalik et al 2018a). They may be host to two types of microorganisms which may exist as co-primary symbionts (Husnik and McCutcheon 2016; Szabo et al 2017), or possess only one obligate symbiont (Gruwell et al 2010, 2012; Michalik et al 2016).…”