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This study evaluated the impact of Aporrectodea molleri's coelomic fluid-associated bacteria (CFB) on Zea mays L. growth and soil biochemical characteristics under abiotic stress conditions, including alkaline soil (pH = 8) and nitrogen (N), phosphate (P), and potassium (K) deficit. Compared to maize cultivated in uninoculated soil, the effect of CFB on boosting plant growth under abiotic stress was notably exceptional. Different CFB treatments increased significantly root and shoot length by 50% and 21%, respectively. Furthermore, the presence of isolates in soil resulted in a significant increase in plant fresh and dry weights (of up to 113% and 91% for roots, and up to 173% and 44% for shoots), leaf surface (78%), and steam diameter (107%). Overall, soil inoculation with CFB significantly (P 0.05) enhanced chlorophyll and water content in the plant compared to the untreated soil. Despite the soil's alkaline condition, CFB drastically boosted soil quality by increasing nutrient availability (up to 30 ppm for N, 2 ppm for P, and 60 ppm for K) and enzyme activity (up to 1.14 μg p-NP h

Earthworms are considered as a rich microhabitat for the growth and proliferation of diverse soil microorganisms. Hence, earthworms' associated bacteria attracted interest due to their high metabolic profiles and benefits to soil fertility and plant growth. In this study, we aimed to isolate for the first-time aerobic bacteria present in the chloragogenous tissue of the earthworm Apporectodea molleri and test their Plant Growth-Promoting abilities and their resistance to heavy metals (Mn, Zn, Cu, Cd, and Ni). The 16S rRNA gene sequencing revealed the affiliation of the fifteen isolates to six main bacterial genera: Enterobacter, Citrobacter, Aeromonas, Pseudomonas, Bacillus, Terribacillus. These strains displayed different plant growth promoting traits (e.g., indole-3-acetic acid IAA, siderophores, nitrogen fixation, phosphate, and potassium solubilization), in addition, they were able to resist differently to heavy metals. Bacillus strains were most effective as three strains, namely B. subtilis strain TC34; B. circulans strain TC7 and Bacillus sp. strain TC10, were positive to all PGP traits and resisted to all heavy metals. This study illustrates the potential of bacteria from the chloragogenous tissue to exhibit multiple properties, which can be related to the functional feature of this tissue to stock metabolites and neutralize toxic elements.

Earthworms are ecological engineers that can contribute to the displacement of biological control agents such as the entomopathogenic nematodes (EPNs) and fungi (EPF). However, a previous study showed that the presence of cutaneous excreta (CEx) and feeding behavior of the earthworm species Eisenia fetida (Haplotaxida: Lumbricidae) compromise the biocontrol efficacy of certain EPN species by reducing, for example, their reproductive capability. Whether this phenomenon is a general pattern for the interaction of earthworms-entomopathogens is still unknown. We hypothesized that diverse earthworm species might differentially affect EPN and EPF infectivity and reproductive capability. Here we investigated the interaction of different earthworm species (Eisenia fetida, Lumbricus terrestris, and Perionyx excavatus) (Haplotaxida) and EPN species (Steinernema feltiae, S. riojaense, and Heterorhabditis bacteriophora) (Rhabditida) or EPF species (Beauveria bassiana and Metarhizium anisopliae) (Hypocreales), in two independent experiments. First, we evaluated the application of each entomopathogen combined with earthworms or their CEx in autoclaved soil. Hereafter, we studied the impact of the earthworms' CEx on entomopathogens applied at two different concentrations in autoclaved sand. Overall, we found that the effect of earthworms on entomopathogens was species-specific. For example, E. fetida reduced the virulence of S. feltiae, resulted in neutral effects for S. riojaense, and increased H. bacteriophora virulence. However, the earthworm P. excavates increased the virulence of S. feltiae, reduced the activity of H. bacteriophora, at least at specific timings, while S. riojaense remained unaffected. Finally, none of the EPN species were affected by the presence of L. terrestris. Also, the exposure to earthworm CEx resulted in a positive, negative or neutral effect on the virulence and reproduction capability depending on the earthworm-EPN species interaction. Concerning EPF, the impact of earthworms was also differential among species. Thus, E. fetida was detrimental to M. anisopliae and B. bassiana after eight days post-exposure, whereas Lumbricus terrestris resulted only detrimental to B. bassiana. In addition, most of the CEx treatments of both earthworm species decreased B. bassiana virulence and growth. However, the EPF M. anisopliae was unaffected when exposed to L. terrestris CEx, while the exposure to E. fetida CEx produced contrasting results. We conclude that earthworms and their CEx can have positive, deleterious, or neutral impacts on entomopathogens that often coinhabit soils, and that we must consider the species specificity of these interactions for mutual uses in biological control programs. Additional studies are needed to verify these interactions under natural conditions., MC is supported by a Moroccan scholarship of the Ministry of National Education, Vocational Training, Higher Education and Scientific Research, and the travel assistance associated with the grant CSIC ICOOP + 2018 grant (COOPA20231). RBP is financed with an FPI contract (CAR-2018) by the Department of Economic Development and Innovation of The Government of La Rioja (Spain). IVD is supported with an FPI-UR fellowship by the Universidad de La Rioja (Spain). RCH is awarded by Ramon y Cajal contract award (RYC-2016-19939) of the Government of Spain. This study is supported by CSIC I-COOP + 2018 grant (COOPA20231).

Earthworms are considered as a rich microhabitat for the growth and proliferation of diverse soil microorganisms. Hence, earthworms' associated bacteria attracted interest due to their high metabolic profiles and benefits to soil fertility and plant growth. In this study, we aimed to isolate for the first-time aerobic bacteria present in the chloragogenous tissue of the earthworm Apporectodea molleri and test their Plant Growth-Promoting abilities and their resistance to heavy metals (Mn, Zn, Cu, Cd, and Ni). The 16S rRNA gene sequencing revealed the affiliation of the fifteen isolates to six main bacterial genera: Enterobacter, Citrobacter, Aeromonas, Pseudomonas, Bacillus, Terribacillus. These strains displayed different plant growth promoting traits (e.g., indole-3-acetic acid IAA, siderophores, nitrogen fixation, phosphate, and potassium solubilization), in addition, they were able to resist differently to heavy metals. Bacillus strains were most effective as three strains, namely B. subtilis strain TC34; B. circulans strain TC7 and Bacillus sp. strain TC10, were positive to all PGP traits and resisted to all heavy metals. This study illustrates the potential of bacteria from the chloragogenous tissue to exhibit multiple properties, which can be related to the functional feature of this tissue to stock metabolites and neutralize toxic elements.

The effect on the bacterial growth of Aporrectodea molleri cutaneous excreta has given very satisfactory results. The cutaneous excreta were obtained using the electrical stimulation technique, recently developed by our laboratory. This technique made it possible to collect the excreta in sufficient quantities while ruling out any possible interference with the chemical constituents of the soil. The effectiveness of cutaneous excreta, tested as culture media at different concentrations on bacterial growth, was justified not only by the high growth rates but also by the concentrations used, which were several times lower than those of the conventional medium (nutrient agar). Indeed, the maximum growth rate recorded by E. coli and P. fluorescens is, respectively, 1.32 times and 2.99 times greater than that observed on the conventional medium. These higher levels are obtained at optimal excretion concentrations which are, respectively, 8 times and 133.33 times lower than the concentration of the conventional medium. The efficacy interval that delimited the excretion concentrations tested covers 8 successive concentrations ranging from 0.0075% to 1% for P. fluorescens, while for E. coli, this interval covers only three concentrations ranging from 0.06% at 0.25%. The growth rates of P. fluorescens are always higher than those of E. coli. The higher efficiency of excretions on the growth of P. fluorescens shows that this strain is more suited to use these excretions as a source of nutrients and to react positively to the stimulation of growth. These results, obtained in vitro, prove that, in the natural environment, the production of lubricating and viscous cutaneous excretions does not only have the role of facilitating the movement and digging of galleries by earthworms but also of enriching the soil in nutrients and growth factors with high added values for the activation and development of plant growth-promoting bacteria essential for soil fertility.

Earthworms enhance soil structure, the decomposition of organic matter and the dissemination of beneficial soil organisms such as the entomopathogenic nematodes (EPNs). Nevertheless, the effects of earthworm feeding behavior or cutaneous excreta (CEx) on the performance of EPNs as biological control agents is poorly understood. We hypothesized that the presence of earthworms or their excreta reduces EPN fitness, measured in terms of pathogenicity and reproductive success. In laboratory experiments we first evaluated the killing ability of EPNs against Galleria mellonella (Lepidoptera: Pyralidae) larvae when inoculated in autoclaved soil alone or in combination with the earthworm Eisenia fetida (Haplotaxida: Lumbricidae) or their excreta. We also evaluated EPN efficacy and reproduction when exposed to CEx derived from E. fetida at two nematode concentrations (1.5 and 10 IJs/cm2). For both experiments, we tested four steinernematids (Steinernema carpocapsae, S. feltiae, S. glaseri, and S. khuongi) and two heterorhabditids (Heterorhabditis bacteriophora and H. zealandica). The presence of earthworms or their excreta resulted in significant reduction of the larval mortality caused by some of the steinernematids at certain timings depending of the species (P < 0.015), while heterorhabditids were mainly not affected. Both S. feltiae and H. zealandica progeny production was significantly reduced (P < 0.01) when exposed to CEx. Hence, we showed that the presence of CEx might alter the biocontrol performance of certain EPN species, especially steinernematids bigger than 600 μm in size (S. feltiae, S. glaseri, and S. khoungi), by affecting their pathogenicity and reproductive success., The Government of Portugal (FCT), thanks to the “Starting Grant” associate funds (grant number IF/00552/2014/CP1234/CT0007), supported this work. RCH was awarded an Investigator Programme contract (grant number IF/00552/2014). Similarly, FABP and RBP were financed by the scientific assistantship fellowships associated with this grant (BI, UAlg-2016/004 and UAlg-2016/003, respectively, Universidade do Algarve, Portugal). MC was supported as visitor student at the Universidade do Algarve (Portugal) thanks to the international mobility program “Mare-Nostrum”. Currently, MC is supported by a Moroccan scholarship for the Ministry of National Education, Vocational Training, Higher Education and Scientific Research, RBP is financed by the Department of Economic Development and Innovation, Government of La Rioja (Spain) with a predoctoral contract (CAR-2018), and RCH is awarded by Ramon y Cajal contract award (RYC-2016-19939) from the Government of Spain. This study is linked to the CSIC i-coop+ 2018 grant (COOPA20231).

In vitro application of total gross extract of earthworms (Lumbricus terrestris) in diverse dilutions stimulates rhizogenesis in young bean plants (Phaseolus vulgaris). The observed effect is similar to that of indol acetic acid, a well-known growth enhancer in plants, used here as a control in various concentrations. Fragmentation of worm extract by column chromatography results in three groups of fractions. Only the polar group of fractions has a significant rhizogenous effect, which is, however, inferior to that observed in the presence of total gross extract of worms or of indol acetic acid. Gross extract analyses using thin layer chromatography, with appropriate chromatography systems and reagents, revealed that indol acetic acid is not present, but is probably replaced by other indol-derived substances that have a neutral to basic chromatographic behaviour. These presumed indol-derived substances are identified as methyl-tryptophane, serotonin, and hydroxy-indol acetic acid. Analyses using mass spectrometry combined with gas chromatography, following fragmentation and purification of the group of rhyzogenous fractions, have revealed the presence of hydroxy-indol carboxylic acid, which seems to take the form of several isomeres.[Journal translation]