Phytobacter diazotrophicus from Intestine of Caenorhabditis elegans Confers Colonization-Resistance against Bacillus nematocida Using Flagellin (FliC) as an Inhibition Factor

Symbiotic microorganisms in the intestinal tract can influence the general fitness of their hosts and contribute to protecting them against invading pathogens. In this study, we obtained isolate Phytobacter diazotrophicus SCO41 from the gut of free-living nematode Caenorhabditis elegans that displayed strong colonization-resistance against invading biocontrol bacterium Bacillus nematocida B16. The colonization-resistance phenotype was found to be mediated by a 37-kDa extracellular protein that was identified as flagellin (FliC). With the help of genome information, the fliC gene was cloned and heterologously expressed in E. coli. It could be shown that the B. nematocida B16 grows in chains rather than in planktonic form in the presence of FliC. Scanning Electronic Microscopy results showed that protein FliC-treated B16 bacterial cells are thinner and longer than normal cells. Localization experiments confirmed that the protein FliC is localized in both the cytoplasm and the cell membrane of B16 strain, in the latter especially at the position of cell division. ZDOCK analysis showed that FliC could bind with serine/threonine protein kinase, membrane protein insertase YidC and redox membrane protein CydB. It was inferred that FliC interferes with cell division of B. nematocidal B16, therefore inhibiting its colonization of C. elegans intestines in vivo. The isolation of P. diazotrophicus as part of the gut microbiome of C. elegans not only provides interesting insights about the lifestyle of this nitrogen-fixing bacterium, but also reveals how the composition of the natural gut microbiota of nematodes can affect biological control efforts by protecting the host from its natural enemies.

On the role of dauer in the adaptation of nematodes to a parasitic lifestyle

Nematodes are presumably the most abundant Metazoa on Earth, and can even be found in some of the most hostile environments of our planet. Various types of hypobiosis evolved to adapt their life cycles to such harsh environmental conditions. The five most distal major clades of the phylum Nematoda (Clades 8–12), formerly referred to as the Secernentea, contain many economically relevant parasitic nematodes. In this group, a special type of hypobiosis, dauer, has evolved. The dauer signalling pathway, which culminates in the biosynthesis of dafachronic acid (DA), is intensively studied in the free-living nematode Caenorhabditis elegans, and it has been hypothesized that the dauer stage may have been a prerequisite for the evolution of a wide range of parasitic lifestyles among other nematode species. Biosynthesis of DA is not specific for hypobiosis, but if it results in exit of the hypobiotic state, it is one of the main criteria to define certain behaviour as dauer. Within Clades 9 and 10, the involvement of DA has been validated experimentally, and dauer is therefore generally accepted to occur in those clades. However, for other clades, such as Clade 12, this has hardly been explored. In this review, we provide clarity on the nomenclature associated with hypobiosis and dauer across different nematological subfields. We discuss evidence for dauer-like stages in Clades 8 to 12 and support this with a meta-analysis of available genomic data. Furthermore, we discuss indications for a simplified dauer signalling pathway in parasitic nematodes. Finally, we zoom in on the host cues that induce exit from the hypobiotic stage and introduce two hypotheses on how these signals might feed into the dauer signalling pathway for plant-parasitic nematodes. With this work, we contribute to the deeper understanding of the molecular mechanisms underlying hypobiosis in parasitic nematodes. Based on this, novel strategies for the control of parasitic nematodes can be developed.

New free-living nematode species and records (Chromadorea: Plectida and Desmodorida) from the edge and axis of Kermadec Trench, Southwest Pacific Ocean

One new nematode species is described and two new species records are provided from the edge (6,080 m depth) and axis (7,132 m) of Kermadec Trench, Southwest Pacific. Leptolaimus hadalis sp. nov. is characterised by medium body 587–741 μm long, labial region not offset from body contour, inconspicuous labial sensilla, amphid located 12–19 μm from anterior end, female without supplements, male with four tubular precloacal supplements (alveolar supplements absent), tubular supplements almost straight with dentate tip, arcuate spicules and weakly cuticularized dorsal gubernacular apophyses strongly bent distally. In a previously published ecological survey of Kermadec Trench, L. hadalis sp. nov. was the most abundant species in a core obtained at 8,079 m water depth and third most abundant species in a core obtained at 7,132 m, while only one individual was found at 6,096 m depth, and none at 9,175 m depth (Leduc & Rowden, 2018). Alaimella aff. cincta and Desmodora aff. pilosa are recorded for the first time from the Southwest Pacific region. Prior to the present study, Alaimella had only been recorded from coastal locations and from the Weddell sea to a depth of 2,000 m. The record of Desmodora aff. pilosa at 6,080 m depth is the deepest record of a Desmodora species to date, although unidentified Desmodora specimens have been found as deep as 6,300 m in the South Sandwich Trench. The morphology of the Kermadec Trench Alaimella aff. cincta and Desmodora aff. pilosa specimens bear a strong resemblance to their respective type populations from the Northern Hemisphere, but further morphological and molecular data are required to ascertain whether they in fact represent distinct species.

A Transcription Factor DAF5 Functions in Haemonchus Contortus Development

Background: Daf5 (Dauer abnormal formation gene), located in the downstream of DAF-7 signalling pathway, mainly functions in dauer formation and reproductive processes in the free-living nematode Caenorhabditis elegans. Although its structure and function have been studied clearly in C. elegans, it was totally unknown in Haemonchus contortus, a socio-economically important parasitic nematode of gastric ruminants.Methods: Here, we identified and characterized a homologue of Daf5, Hcdaf5 and its inferred product (HcDAF5) in H. contortus. Using an integrated molecular approach, we studied the transcriptional profiles of Hcdaf5 and the anatomical expression of HcDAF5 in H. contortus. RNA interference (RNAi) was performed to explore its function in transition from the exsheathed third-stage larvae (xL3) to the fourth-stage larvae (L4) in vitro. Interaction of HcDAF5 and HcDAF3 (a co-SMAD) was also detected by bimolecular fluorescence complementation system (BiFc) in vitro.Results: Here, we showed that HcDAF5 is a member of the Sno/Ski superfamily. Hcdaf5 was transcribed in all developmental stages of H. contortus, with a significant up-regulation in L3. Immunohistochemistry localized native HcDAF5 to the reproductive organs, cuticle and intestine. RNAi revealed specific siRNAs (small interfering RNA) could retard the xL3 development. In addition, the interaction between HcDAF5 and HcDAF3 indicated the SDS box region of HcDAF5 is dispensable for the binding of HcDAF5 to HcDAF3 and the region in HcDAF3 that binds to HcDAF5 is MH2 domain.Conclusion: In summary, these findings show that Hcdaf5 functions in developmental processes of H. contortus, and this is the first characterization of daf-5 gene in parasitic nematodes.

DIVERSITY AND ABUNDANCE OF NEMATODES IN GUAVA (Psidium guajava L.) CULTIVATION IN LAMPUNG

Diversity and abundance of nematodes in guava (Psidium guajava L.) cultivation in Lampung. Crystal guava agroecosystem is inhabited by many species of plant parasitic nematodes. However, information regarding this topic was still limited. This study aimed to understand the species dominancy of nematodes in crystal guava cultivation in Lampung. Sampling was carried out in three locations of guava crystal plantations: Lampung Timur, Lampung Tengah, and Tanggamus. The laboratory analysis was done at the Plant Pest Science Laboratory and Agricultural Biotechnology Laboratory, Universitas Lampung. The study was conducted in December 2019 – July 2020. Nematodes were identified to the level of the genus. The Prominance value (PV) was used to assess the nematodes genus dominancy. The results showed that the nematodes inhabiting the crystal guava agro-ecosystem in Lampung was both plant parasitic and free-living nematodes. The plant parasitic nematodes were identified as Meloidogyne, Aphelenchus, Hemicriconemoides, Tylenchus, Aphelenchoides, and Xiphinema, while free-living nematodes was Rhabditis, Dorylaimine, Dorylaimus, and Mononchus. The dominant plant parasitic nematode was Meloidogyne and the dominant free-living nematode was Rhabditis. The abundance of Meloidogyne /300 mL of soil was 351.47 individuals in Lampung Timur, 124.27 individuals in Lampung Tengah, and 82.18 individuals in Tanggamus. The dominant free-living nematode in the three locations was Rhabditis.

A Zinc Metalloprotease nas-33 Is Required for Molting and Survival in Parasitic Nematode Haemonchus contortus

Molting is of great importance for the survival and development of nematodes. Nematode astacins (NAS), a large family of zinc metalloproteases, have been proposed as novel anthelmintic targets due to their multiple roles in biological processes of parasitic nematodes. In this study, we report a well conserved nas-33 gene in nematodes of clade V and elucidate how this gene is involved in the molting process of the free-living nematode Caenorhabditis elegans and the parasitic nematode Haemonchus contortus. A predominant transcription of nas-33 is detected in the larval stages of these worms, particularly in the molting process. Knockdown of this gene results in marked molecular changes of genes involved in cuticle synthesis and ecdysis, compromised shedding of the old cuticle, and reduced worm viability in H. contortus. The crucial role of nas-33 in molting is closely associated with a G protein beta subunit (GPB-1). Suppression of both nas-33 and gpb-1 blocks shedding of the old cuticle, compromises the connection between the cuticle and hypodermis, and leads to an increased number of sick and dead worms, indicating essentiality of this module in nematode development and survival. These findings reveal the functional role of nas-33 in nematode molting process and identify astacins as novel anthelmintic targets for parasitic nematodes of socioeconomic significance.