The nematodes C. elegans and P. pacificus populate diverse habitats and display distinct patterns of behavior. To understand how their nervous systems have diverged, we undertook a detailed examination of the neuroanatomy of the chemosensory system of P. pacificus. Using independent features such as cell body position, axon projections and lipophilic dye uptake, we have assigned homologies between the amphid neurons, their first-layer interneurons, and several internal receptor neurons of P. pacificus and C. elegans. We found that neuronal number and soma position are highly conserved. However, the morphological elaborations of several amphid cilia are different between them, most notably in the absence of ‘winged’ cilia morphology in P. pacificus. We established a synaptic wiring diagram of amphid sensory neurons and amphid interneurons in P. pacificus and found striking patterns of conservation and divergence in connectivity relative to C. elegans, but very little changes in relative neighborhood of neuronal processes. These findings demonstrate the existence of several constraints in patterning the nervous system and suggest that major substrates for evolutionary novelty lie in the alterations of dendritic structures and synaptic connectivity.

The nematodes C. elegans and P. pacificus populate diverse habitats and display distinct patterns of behavior. To understand how their nervous systems have diverged, we undertook a detailed examination of the neuroanatomy of the chemosensory system of P. pacificus. Using independent features such as cell body position, axon projections and lipophilic dye uptake, we have assigned homologies between the amphid neurons, their first-layer interneurons, and several internal receptor neurons of P. pacificus and C. elegans. We found that neuronal number and soma position are highly conserved. However, the morphological elaborations of several amphid cilia are different between them, most notably in the absence of 'winged' cilia morphology in P. pacificus. We established a synaptic wiring diagram of amphid sensory neurons and amphid interneurons in P. pacificus and found striking patterns of conservation and divergence in connectivity relative to C. elegans, but very little changes in relative neighborhood of neuronal processes. Impact StatementThe substrate for evolutionary divergence does not lie in changes in neuronal cell number or targeting, but rather in sensory perception and synaptic partner choice within invariant, prepatterned neuronal processes.

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Developmental and behavioral plasticity allow animals to prioritize alternative genetic programs during fluctuating environments. Behavioral remodeling may be acute in animals that interact with host organisms, since reproductive adults and the developmentally arrested larvae often have different ethological needs for chemical stimuli. To understand the genes that coordinate development and host-seeking behavior, we used the entomophilic nematode Pristionchus pacificus to characterize dauer-constitutive mutants (Daf-c) that inappropriately enter developmental diapause to become dauer larvae. We found two Daf-c loci with dauer-constitutive and cuticle exsheathment phenotypes that can be rescued by the feeding of Δ7-dafachronic acid, and that are dependent on the conserved canonical steroid hormone receptor Ppa-DAF-12. Specifically at one locus, deletions in the sole HydroxySteroid Dehydrogenase (HSD) in P. pacificus resulted in Daf-c phenotypes. Ppa-hsd-2 is expressed in the canal neurons (CAN) and excretory cells whose homologous cells in C. elegans are not known to be involved in the dauer decision. While in wildtype only dauer larvae are attracted to host odors, hsd-2 mutant adults show enhanced attraction to the host beetle pheromone, along with ectopic activation of a marker for putative olfactory neurons, Ppa-odr-3. Surprisingly, this enhanced odor attraction acts independently of the Δ7-DA/DAF-12 module, suggesting that Ppa-HSD-2 may be responsible for several steroid hormone products involved in coordinating the dauer decision and host-seeking behavior in P. pacificus.

1Developmental and behavioral plasticity allow animals to prioritize alternative 2 genetic programs during fluctuating environments. Behavioral remodeling may be 3 acute in animals that interact with host organisms, since reproductive adults and 4 the developmentally arrested larvae often have different ethological needs for 5 chemical stimuli. To understand the genes that coordinate development and 6 behavior, we used the nematode model Pristionchus pacificus to characterize 7 mutants that inappropriately enter developmental diapause to become dauer 8 larvae (Daf-c). We found several key olfactory differences between P. pacificus 9 and C. elegans Daf-c dauers. In addition, the two P. pacificus Daf-c alleles disrupt 10 steroid synthesis required for proper regulation of the conserved canonical steroid 11 hormone receptor DAF-12, whose dauer-constitutive and cuticle exsheathment 12 phenotypes can be rescued by the feeding of ∆7-dafachronic acid. One allele, 13 csu60, has a deletion in the sole HydroxySteroid Dehydrogenase (HSD) in P. 14 pacificus. Both hsd-2(csu60) adults and dauers show enhanced attraction to a 15 beetle pheromone, possibly due to the heterochronic activation of dauer-specific 16 neuronal development in the adults. Surprisingly, this enhanced odor attraction 17 acts independently of daf-12, revealing unexpected targets of steroid hormones 18 regulating ecdysis and olfaction in P. pacificus. analysis in diverse species of nematodes as a result of comparative studies in non-52Caenorhabditis species [8][9][10]. 53The commitment to dauer entry requires external as well as internal factors. 54Multiple parallel-acting signaling factors in the dauer regulation pathway have been 55 characterized in C. elegans, such as the recognition of pheromones by G-protein-56 coupled receptors in the chemosensory amphid neurons and signal transduction 57 to the DAF-11/guanylyl cyclase pathway [11]. Favorable conditions also lead to 58 signaling in the DAF-7/TGF-ß and DAF-2/insulin-like pathways [11][12][13][14]. This 59 signaling facilitates the production of ∆4-and ∆7-dafachronic acid from the 60 cholesterol precursor, which act as ligands that bind to the highly conserved 61 nuclear hormone receptor DAF-12 and prevent DAF-12 from repressing genes 62 necessary for development to reproductive adulthood [11,[15][16][17]. Less favorable 63 conditions produce pheromones that reduce signaling in these pathways, 64 subsequently shifting DAF-12 from its inactive ligand-bound form to its active, 65ligand-free form that specifies dauer development. Despite being a non-feeding 66 and stress-resistant stage, the DL are actively motile and receptive to external 67 chemosensory and mechanical cues. For both parasitic and free-living nematodes, 68 DL share the ability to survive harsh conditions but remain receptive to 69 semiochemicals from their environment. 70The commitment to dauer also prompts the rewiring of the sensory nervous 71 system in C. elegans [6,18,19], but the molecular basis of this process remains 72 poorly unde...