Bacterial adaptation to host association

Microorganisms are commonly found living in multicellular organisms. Taxonomically, individual symbionts and communities of microbes, microbiotas, of a variety of animals and plants have been well-described and it has become clear that they can have profound effects on their hosts' biology. The bacterial side of the association, however, has received comparatively little attention. In particular how microbes adjust to life with a host, and what drives the emergence and maintenance of microbe-host associations remains understudied. In this thesis, my main objective is therefore to improve our understanding of the traits and life history strategies that facilitate microbe-host associations. To that end I combine experimental study of bacterial isolates from the natural microbiota of the nematode Caenorhabditis elegans as a model system with predictions from mathematical models. In the first part of the thesis I focus on the functional ecology of the natural C. elegans microbiota. Integrating whole genome sequencing of bacterial isolates, metabolic modeling, and experiments, we found that the microbiota can synthesize all amino acids and vitamins essential to the worm. Further, we demonstrate that their predicted traits and metabolic repertoires lead to distinct life history strategies, shape interactions within the microbiota as well as worm colonization and worm population growth. The second part of the thesis addresses the evolution of bacteria in host-association. The first step is the development of the concept of a biphasic life cycle as an evolutionary intermediate between a free-living and a host-associated lifestyle. At this transition, matrix population models predict that bacteria can optimize fitness either by increasing replication rates or by modulating migration rates between host and environment. Experimentally evolving the microbiota isolate Pseudomonas lurida MYb11 in a biphasic life cycle with C. elegans, we find that a key step to associating is to simply stick to the host. Specifically, we observe the evolution of wrinkly colony types that show improved persistence in worms and a greater ability to form biofilms than the ancestral MYb11. This goes hand in hand with genetic mutations in regulators of the universal second messenger cyclic di-GMP, previously linked to the life history transition from motile to sessile and stress-tolerant. These findings highlight that bacteria can adapt to a host that they encounter periodically during a biphasic life cycle. Finally, I extend the focus to a two-member microbiota of MYb11 and Ochrobactrum vermis MYb71 and study interactions between these species and differences in their life history strategies when co-associating in C. elegans. This confirms that interactions within the microbiota are context-dependent and shift along the stages of the biphasic life cycle. Further, it demonstrates that different life history strategies co-exist in the microbiota, with the one centered on stress-tolerance being the most advantageous. Overall, this thesis emphasizes the importance of bacterial traits and life history to shaping life in association with a host. It thereby provides fundamental insights into the driving forces and possibly origin of microbe-host associations

DNA barcodes reveal a new host record for Carcelia atricosta Herting (Diptera Tachinidae) in Italy

The parasitoid-host association between Orgyia antiqua (Linnaeus) (Lepidoptera: Lymantriidae) and Carcelia atricosta Herting (Diptera: Tachinidae) is recorded here for the first time in Italy. A single caterpillar of O. antiqua was collected in Northern Italy (Arzergrande, Padua, Veneto Region) in June 2015. After the specimen died, a single tachinid larva emerged and pupariated. The emerged parasitoid was identified using DNA barcoding, with DNA extracted from the tachinid pupa. This is the first distributional record of C. atricosta in Northern Italy and the second for Italy, only two other specimens having being recorded previously (Abruzzo Region, Central Italy)

Aphidiinae (Hymenoptera, Braconidae) aphid parasitoids of Malta : review and key to species

This paper brings an annotated list of the 16 aphid parasitoids detected up to 2015 in Malta. All the species were reared from identified aphid-plant associations. An illustrated key to the identification of the recorded species from Malta is provided. Taxonomy and peculiarities of the individual taxa are discussed and research outlines are also presented. Two species, Aphidius absinthii and Trioxys pallidus are newly recorded for the Maltese fauna, in association with Uroleucon inulae and Hoplocallis picta, respectively. The first host association has never been reported so far. The documented aphid fauna of the Maltese Islands indicates the probable existence of other species of aphid parasitoids that may be present in the archipelago, especially those already known in nearby Mediterranean territories.peer-reviewe

Molecular diversity of arbuscular mycorrhizal fungi and patterns of host association over time and space in a tropical forest

We have used molecular techniques to investigate the diversity and distribution of the arbuscular mycorrhizal (AM) fungi colonizing tree seedling roots in the tropical forest on Barro Colorado Island (BCI), Republic of Panama. In the first year, we sampled newly emergent seedlings of the understory treelet Faramea occidentalis and the canopy emergent Tetragastris panamensis, from mixed seedling carpets at each of two sites. The following year we sampled surviving seedlings from these cohorts. The roots of 48 plants were analysed using AM fungal-specific primers to amplify and clone partial small subunit (SSU) ribosomal RNA gene sequences. Over 1300 clones were screened for random fragment length polymorphism (RFLP) variation and 7% of these were sequenced. Compared with AM fungal communities sampled from temperate habitats using the same method, the overall diversity was high, with a total of 30 AM fungal types identified. Seventeen of these types have not been recorded previously, with the remainder being similar to types reported from temperate habitats. The tropical mycorrhizal population showed significant spatial heterogeneity and nonrandom associations with the different hosts. Moreover there was a strong shift in the mycorrhizal communities over time. AM fungal types that were dominant in the newly germinated seedlings were almost entirely replaced by previously rare types in the surviving seedlings the following year. The high diversity and huge variation detected across time points, sites and hosts, implies that the AM fungal types are ecologically distinct and thus may have the potential to influence recruitment and host composition in tropical forests

The regional economic impact of the 2009 New Zealand National Masters Hockey Tournament

This paper analyses the direct and subsequent flow-on expenditure resulting from the weeklong National Masters Hockey Tournament held in Hamilton, New Zealand in March 2009. During the tournament, information was collected on the expenditure from a sample of participating players and officials and from the host association, Waikato Hockey. The information was averaged over all participants and used in a regional economic model. The economic impact of the 2009 tournament was also compared to the outcome for the Hamilton tournament held previously in 2003. Total direct expenditure in the Waikato region from the 2009 tournament was approximately $2.5 million compared to$1.2 million in 2003. The 2009 expenditure led subsequently to an extra $1.13 million of value-added to the regional economy mainly in the hospitality and accommodation sectors

Natural Enemies of Cranberry Fruitworm, \u3ci\u3eAcrobasis Vaccinii\u3c/i\u3e, (Lepidoptera: Pyraudae) in Michigan Highbush Blueberries

A two-year study was conducted in Michigan highbush blueberries to determine the complex of parasitoids attacking cranberry fruitworm, Acrobasis vaccinii. Eight parasitoid species and one fungal pathogen were collected. Parasitism of collected hosts ranged from 6.6% to 28.1%. The more common larval parasitoid encountered was Campoletis patsuiketorum (Hymenoptera: Ichneumonidae). The more common parasitoid recovered from fruitworm hibernacula was Villa lateralis (Diptera: Bombyliidae). This study documented six unreported natural enemies of cranberry fruitworm, including C. patsuiketorum; V. lateralis; Diadegma compressum (Hymenoptera: Ichneumonidae); Compsilura concinnata (Diptera: Tachinidae); Memorilla pyste (Diptera: Tachinidae); an undescribed Microtypus species (Hymenoptera: Braconidae); and a fungal pathogen, Paecilomyces near farinosus. This is the first known host association for the undescribed Microtypus species, and increases the known parasitoid complex of cranberry fruitworm to 17 species

Evolutionary genetic aspects of host association in generalist ectoparasites

Despite the use of the host for dispersal by most parasite species, the extremely loose relationship typical between highly mobile hosts and generalist ectoparasites may lead to very different gene flow patterns between the two, leading in turn to different spatial genetic structure, and potentially different demographic history. I examined how similar gene flow patterns are between Cimex adjunctus, a generalist ectoparasite of bats present throughout North America, and two of its key bat hosts. I first analyzed the continent-scale genetic structure and demographic history of C. adjunctus and compared it to that of two of its hosts, the little brown myotis and the big brown bat, using microsatellite and mitochondrial data. Second, I compared spatial genetic structure of C. adjunctus with Cimex lectularius, or common bed bug, which associates with a broader range of host species. Third, I compared the effect of land cover on spatial genetic structure of C. adjunctus and of the big brown bat in the Great Lakes region. My results support the emerging hypothesis that generalist ectoparasites and their highly mobile hosts display weak, but positive, correlation in spatial genetic structure and demographic history. Generalist parasites associate with different hosts, which are, in some cases, evolutionarily divergent from each other. In such cases, it is not clear how hosts may affect adaptive genetic variation in the parasites. In the Cimex genus, parasite species associate with a range of hosts, including bats, humans, and swallows. I examined how hosts affect adaptive genetic variation in these generalist ectoparasites. I analyzed variation at two salivary protein genes, one coding for an apyrase and the other for a nitrophorin, in 10 species of Cimex. These proteins affect the way parasites feed on their hosts, by preventing clotting and vasoconstriction, and may experience selection depending on host ecology or physiology. I also analyzed allelic divergence at the same two genes in a single species, C. adjunctus, associated with several bat species in North America. My results suggest selection and adaptation to the host at genes coding for salivary proteins of blood-feeding ectoparasites across the Cimex genus, and also within C. adjunctus

Reproductive Compatibility among Populations and Host‐Associated Lineages of the Common Bed Bug (\u3cem\u3eCimex lectularius\u3c/em\u3e L.)

As populations differentiate across geographic or host‐association barriers, interpopulation fertility is often a measure of the extent of incipient speciation. The bed bug, Cimex lectularius L., was recently found to form two host‐associated lineages within Europe: one found with humans (human‐associated, HA) and the other found with bats (bat‐associated, BA). No unequivocal evidence of contemporary gene flow between these lineages has been found; however, it is unclear whether this is due to an inability to produce viable “hybrid” offspring. To address this question and determine the extent of compatibility between host‐associated lineages, we set up mating crosses among populations of bed bugs based on both their host association (human—HA vs. bat—BA) and geographic origin (North America vs. Europe). Within‐population fecundity was significantly higher for all HA populations (\u3e 1.7 eggs/day) than for BA populations (\u3c 1 egg/day). However, all within‐population crosses, regardless of host association, had \u3e 92% egg hatch rates. Contrary to previous reports, in all interlineage crosses, successful matings occurred, fertile eggs were oviposited, and the F1 “hybrid” generation was found to be reproductively viable. In addition, we evaluated interpopulation genetic variation in Wolbachia among host‐associated lineages. We did not find any clear patterns related to host association, nor did we observe a homogenization of Wolbachia lineages across populations that might explain a breakdown of reproductive incompatibility. These results indicate that while the HA and BA populations of C. lectularius represent genetically differentiated host‐associated lineages, possibly undergoing sympatric speciation, this is in its incipient stage as they remain reproductively compatible. Other behavioral, physiological, and/or ecological factors likely maintain host‐associated differentiation

Bacteriophage-Host Association in the Phytoplasma Insect Vector Euscelidius variegatus

Insect vectors transmit viruses and bacteria that can cause severe diseases in plants and economic losses due to a decrease in crop production. Insect vectors, like all other organisms, are colonized by a community of various microorganisms, which can influence their physiology, ecology, evolution, and also their competence as vectors. The important ecological meaning of bacteriophages in various ecosystems and their role in microbial communities has emerged in the past decade. However, only a few phages have been described so far in insect microbiomes. The leafhopper Euscelidius variegatus is a laboratory vector of the phytoplasma causing Flavescence dorée, a severe grapevine disease that threatens viticulture in Europe. Here, the presence of a temperate bacteriophage in E. variegatus (named Euscelidius variegatus phage 1, EVP-1) was revealed through both insect transcriptome analyses and electron microscopic observations. The bacterial host was isolated in axenic culture and identified as the bacterial endosymbiont of E. variegatus (BEV), recently assigned to the genus Candidatus Symbiopectobacterium. BEV harbors multiple prophages that become active in culture, suggesting that different environments can trigger different mechanisms, finely regulating the interactions among phages. Understanding the complex relationships within insect vector microbiomes may help in revealing possible microbe influences on pathogen transmission, and it is a crucial step toward innovative sustainable strategies for disease management in agriculture

Host association of cryptosporidium parvum populations infecting domestic ruminants in Spain

A stock of 148 Cryptosporidium parvum DNA extracts from lambs and goat kids selected from a previous study examining the occurrence of Cryptosporidium species and GP60 subtypes in diarrheic lambs and goat kids in northeastern Spain was further characterized by a multilocus fragment typing approach with six mini- and microsatellite loci. Various degrees of polymorphism were seen at all but the MS5 locus, although all markers exhibited two major alleles accounting for more than 75% of isolates. A total of 56 multilocus subtypes (MLTs) from lambs (48 MLTs) and goat kids (11 MLTs) were identified. Individual isolates with mixed MLTs were detected on more than 25% of the farms, but most MLTs (33) were distinctive for individual farms, revealing the endemicity of cryptosporidial infections on sheep and goat farms. Comparison with a previous study in calves in northern Spain using the same six-locus subtyping scheme showed the presence of host-associated alleles, differences in the identity of major alleles, and very little overlap in MLTs between C. parvum isolates from lambs and those from calves (1 MLT) or isolates from lambs and those from goat kids (3 MLTs). The Hunter-Gaston index of the multilocus technique was 0.976 (95% confidence interval [CI], 0.970 to 0.982), which supports its high discriminatory power for strain typing and epidemiological tracking. Population analyses revealed the presence of two host-associated subpopulations showing epidemic clonality among the C. parvum isolates infecting calves and lambs/goat kids, respectively, although evidence of genetic flow between the two subpopulations was also detected