Comparative tissue expression of American lobster (Homarus americanus) immune genes during bacterial and scuticociliate challenge.

The American lobster (Homarus americanus) fishery is the most economically significant fishery in Canada; although comparatively little is known about the lobsters' response to pathogenic challenge. This is the first study to investigate the expression of immune genes in tissues outside of the lobster hepatopancreas in response to challenges by the Gram-positive bacteria, Aerococcus viridans var. homari or the scuticociliate parasite, Anophryoides haemophila. The hepatopancreas has been regarded as the major humoral immune organ in crustaceans, but the contribution of other organs and tissues to the molecular immune response has largely been overlooked. This study used RT-qPCR to monitor the gene expression of several immune genes including three anti-lipopolysaccharide isoforms (ALF) Homame ALF-B1, Homame ALF-C1 and ALFHa-1, acute phase serum amyloid protein A (SAA), as well as thioredoxin and hexokinase, in antennal gland and gill tissues. Our findings indicate that the gene expression of the SAA and all ALF isoforms in the antennal gland and gill tissues increased in response to pathogenic challenge. However, there was differential expression of individual ALF isoforms that were dependent on both the tissue, and the pathogen used in the challenge. The gene expression changes of several immune genes were found to be higher in the antennal gland than have been previously reported for the hepatopancreas. This study demonstrates that increased immune gene expression from the gill and antennal gland over the course of pathogen induced disease contributes to the immune response of H. americanus.

Cultivation reveals physiological diversity among defensive 'Streptomyces philanthi' symbionts of beewolf digger wasps (Hymenoptera, Crabronidae).

BACKGROUND: 'Candidatus Streptomyces philanthi' is a monophyletic clade of formerly uncultured bacterial symbionts in solitary digger wasps of the genera Philanthus, Philanthinus and Trachypus (Hymenoptera, Crabronidae). These bacteria grow in female-specific antennal reservoirs and - after transmission to the cocoon - produce antibiotics protecting the host larvae from fungal infection. However, the symbionts' refractoriness to cultivation has thus far hampered detailed in vitro studies on their physiology and on the evolutionary changes in metabolic versatility in response to the host environment. RESULTS: Here we isolated in axenic culture 22 'Streptomyces philanthi' biovars from different host species. Sequencing of gyrB revealed no heterogeneity among isolates within host individuals, suggesting low levels of (micro)diversity or even clonality of the symbionts in individual beewolf antennae. Surprisingly, however, isolates from different host species differed strongly in their physiology. All biovars from the Eurasian/African Philanthus and the South American Trachypus host species had high nutritional demands and were susceptible to most antibiotics tested, suggesting a tight association with the hosts. By contrast, biovars isolated from the genus Philanthinus and the monophyletic North American Philanthus clade were metabolically versatile and showed broad antibiotic resistance. Concordantly, recent horizontal symbiont transfer events - reflected in different symbiont strains infecting the same host species - have been described only among North American Philanthus species, altogether indicative of facultative symbionts potentially capable of a free-living lifestyle. Phylogenetic analyses reveal a strong correlation between symbiont metabolic versatility and host phylogeny, suggesting that the host environment differentially affects the symbionts' evolutionary fate. Although opportunistic bacteria were occasionally isolated from the antennae of different host species, only filamentous Actinobacteria (genera Streptomyces, Amycolatopsis and Nocardia) could replace 'S. philanthi' in the antennal gland reservoirs. CONCLUSION: Our results indicate that closely related bacteria from a monophyletic clade of symbionts can experience very different evolutionary trajectories in response to the symbiotic lifestyle, which is reflected in different degrees of metabolic versatility and host-dependency. We propose that the host-provided environment could be an important factor in shaping the degenerative metabolic evolution in the symbionts and deciding whether they evolve into obligate symbionts or remain facultative and capable of a host-independent lifestyle.

Refining the roots of the beewolf-Streptomyces symbiosis: antennal symbionts in the rare genus Philanthinus (Hymenoptera, Crabronidae).

Insects engage in symbiotic associations with a large diversity of beneficial microorganisms. While the majority of well-studied symbioses have a nutritional basis, several cases are known in which bacteria protect their host from pathogen infestation. Solitary wasps of the genera Philanthus and Trachypus (beewolves; Hymenoptera, Crabronidae) cultivate the actinomycete "Candidatus Streptomyces philanthi" in specialized antennal gland reservoirs. The symbionts are transferred to the larval cocoon, where they provide protection against pathogenic fungi by producing at least nine different antibiotics. Here we investigated the closest relatives of Philanthus and Trachypus, the rare genus Philanthinus, for the presence of antennal gland reservoirs and symbiotic streptomycetes. Molecular analyses identified "Ca. Streptomyces philanthi" in reservoirs of Philanthinus quattuordecimpunctatus. Phylogenies based on the 16S rRNA gene suggest that P. quattuordecimpunctatus may have acquired "Ca. Streptomyces philanthi" by horizontal transfer from other beewolf species. In histological sections and three-dimensional reconstructions, the antennal gland reservoirs were found to occupy six antennal segments (as opposed to only five in Philanthus and Trachypus) and to be structurally less complex than those of the evolutionarily more derived genera of beewolves. The presence of "Ca. Streptomyces philanthi" in antennal glands of Philanthinus indicates that the symbiosis between beewolves and Streptomyces bacteria is much older than previously thought. It probably evolved along the branch leading to the monophyletic tribe Philanthini, as it seems to be confined to the genera Philanthus, Trachypus, and Philanthinus, which together comprise 172 described species of solitary wasps.