Potato Cultivar, Pathogen Isolate and Antagonist Cultivation Medium Influence the Efficacy and Ranking of Bacterial Antagonists of Fusarium Dry Rot

The process of selecting biological control agents for further development frequently does not involve conducting bioassays of strain effectiveness on a range of pathogen isolates or host cultivars. Additionally, though previous studies have demonstrated that the medium used to produce biomass of an antagonist can alter its efficacy, this factor is also rarely considered when selecting for the most effective antagonist. Host cultivar, pathogen isolate, and the cultivation medium used to produce the antagonists' biomass were examined as factors of potential importance for assessing the relative effectiveness of bacterial biocontrol strains accurately. Five bacterial antagonists that control Fusarium dry rot on stored potato tubers were assayed for effectiveness against 10 isolates of Gibberella pulicaris . All antagonists reduced disease severity (35-81%) regardless of the specific assays conducted. However, when the antagonists' biomass were produced on two media that differed both in nutrient composition and phase, the efficacy ranking of antagonist Enterobacter sp. S11:P:08 varied from first to fourth most effective. For the antagonists studied, the phase of a nutritionally identical medium had little impact on the efficacy ranking of the five antagonists. Four of the five antagonists had efficacy rankings that ranged from first to last depending on the isolate of the pathogen used to conduct the bioassay. The cultivar of the host also caused variations in the efficacy ranking of the antagonists. These results indicate that bioassays should be conducted using a range of liquid culture production media, pathogen isolates, and host cultivars in order to choose an antagonist that has the highest likelihood for commercial development as an effective biological control product.     

Influence of antagonist, host fruit and pathogen on the biological control of postharvest fungal diseases by yeasts

The yeasts Rhodotorula glutinis (LS-11), Cryptococcus laurentii (LS-28), Candida famata (21-D) and Pichia guilliermondii (29-A) and the yeast-like fungus Aureobasidium pullulans (LS-30), previously selected and characterized for mechanisms of action and antagonistic activity against postharvest pathogens in small and large-scale experiments, were used in this study in order to assess interrelationships among the main factors (antagonist, host fruit and fungal pathogen) involved in biological control of postharvest diseases. The antagonists were evaluated for their inhibitory activity (IA) against six common postharvest fungal pathogens on six different host fruits. Artificially wounded fruits were first inoculated with the antagonist and 2 h later with the pathogen; subsequently they were kept at 20°C for 4–6 days. The IA of each antagonist was evaluated and data were submitted to factorial analysis of variance. The populations of antagonists were also monitored on wounded and unwounded fruits kept at 20°C for 7 days. Each factor examined (antagonist, host fruit and fungal pathogen) as well as their interactions significantly affected the IA. However, among the antagonists, isolates LS-28 and LS-30 were only slightly affected by both host and pathogen, showing a wide range of activity, whereas isolate LS-11 had a variable IA. All the antagonists rapidly colonized the wounds, while their population remained substantially unchanged on unwounded fruits. These results suggest that in order to select yeasts with a broad spectrum of action, more suitable for commercial development, it would be advantageous to perform preliminary assays against several pathogens and in particular on different fruit species. Received 23 February 1999/ Accepted in revised form 09 July 1999     

Parameter estimation and sensitivity analysis in an agent-based model of Leishmania major infection

Computer models of disease take a systems biology approach toward understanding host-pathogen interactions. In particular, data driven computer model calibration is the basis for inference of immunological and pathogen parameters, assessment of model validity, and comparison between alternative models of immune or pathogen behavior. In this paper we describe the calibration and analysis of an agent-based model of Leishmania major infection. A model of macrophage loss following uptake of necrotic tissue is proposed to explain macrophage depletion following peak infection. Using Gaussian processes to approximate the computer code, we perform a sensitivity analysis to identify important parameters and to characterize their influence on the simulated infection. The analysis indicates that increasing growth rate can favor or suppress pathogen loads, depending on the infection stage and the pathogen's ability to avoid detection. Subsequent calibration of the model against previously published biological observations suggests that L. major has a relatively slow growth rate and can replicate for an extended period of time before damaging the host cell.     

Susceptibility to an inoculum of infectious hypodermal and haematopoietic necrosis virus (IHHNV) in three batches of whiteleg shrimp Litopenaeus vannamei (Boone, 1931)

The present study evaluated the susceptibility of three different batches of whiteleg shrimp Litopenaeus vannamei from Mexico to an inoculum of infectious hypodermal and haematopoietic necrosis virus (IHHNV). Each of the three shrimp batches came from a different hatchery. Because of their origin, it was possible that the genetic makeup of these batches was different among each other. The three batches tested showed differences in IHHNV susceptibility. Here, susceptibility is defined as the capacity of the host to become infected, and it can be measured by the infectivity titer. Susceptibility to IHHNV was observed in decreasing order in shrimp from batch 1 (hatchery from El Rosario, Sinaloa), batch 3 (hatchery from Nayarit) and batch 2 (hatchery from El Walamo, Sinaloa), respectively. The largest susceptibility difference between batches was 5012 times, and that between early and late juveniles from the same batch was 25 times. These results indicate that within a species, susceptibility to a pathogen such as IHHNV can have large differences. Susceptibility to pathogens is an important trait to consider before performing studies on pathogenesis. It may influence virological parameters such as speed of replication, pathogenicity and virus titer. In order to evaluate the potential use of IHHNV as a natural control agent against white spot syndrome virus (WSSV), it is necessary to know host susceptibility and the kinetics of IHHNV infection. These features can help to determine the conditions in which IHHNV could be used as antagonist in a WSSV infection.     

Periodic host absence can select for higher or lower parasite transmission rates

This paper explores the effect of discontinuous periodic host absence on the evolution of pathogen transmission rates by using R ₀ maximisation techniques. The physiological consequence of an increased transmission rate can be either an increased virulence, i.e. there is a transmission-virulence trade-off or ii) a reduced between season survival, i.e. there is a transmission-survival trade-off. The results reveal that the type of trade-off determines the direction of selection, with relatively longer periods of host absence selecting for higher transmission rates in the presence of a trade-off between transmission and virulence but lower transmission rates in the presence of a trade-off between transmission and between season survival. The fact that for the transmission-virulence trade-off both trade-off parameters operate during host presence whereas for the transmission-survival trade-off one operates during host presence (transmission) and the other (survival) during the period of host absence is the main cause for this difference in selection direction. Moreover, the period of host absence seems to be the key determinant of the pathogen’s transmission rate. Comparing plant patho-systems with contrasting biological features suggests that airborne plant pathogens respond differently to longer periods of host absence than soil-borne plant pathogens.     

Effect of bottlegourd seedcoating with antagonists on seedlings,quantum of the pathogen inside the seedlings and population of the soil againstFusarium oxysporum

Bottlegourd seedlings raised from antagonists coated seeds did not show any wilting symptoms and remained completely healthy. The pathogen quantum inside the host tissue was found least. The pathogen propagules per gram of host tissue were restricted and the soil population of theFusarium oxysporum was reduced very much in antagonists introduced soil.     

Screening of antagonistic bacteria for biological control of nursery wilt of black pepper (Piper nigrum)

Bacterial antagonists of Phytophthora capsici were isolated from underground shoot portions of rooted cuttings of black pepper. Initially isolates were screened by dual culture on potato dextrose agar and carrot agar. Further, a screening was done on black pepper shoots for supression of lesion caused by the pathogen. Most of the antagonists showed varying levels of antagonism in the dual culture and the shoot assay. Isolate PN-026, showing the highest suppression of lesion development in the shoot assay was found to be the most efficient antagonist in reducing Phytophthora capsici induced nursery wilt of black pepper. This screening involving the host, pathogen, and the antagonist, performed on black pepper shoot (the planting material for this vegetatively propagated crop), could be used as a rapid and reliable method for the isolation of efficient bacterial antagonists of P. capsici.     

Indirect nontarget effects of host-specific biological control agents: Implications for biological control

Classical biological control of weeds currently operates under the assumption that biological control agents are safe (i.e., low risk) if they do not directly attack nontarget species. However, recent studies indicate that even highly host-specific biological control agents can impact nontarget species through indirect effects. This finding has profound implications for biological control. To better understand the causes of these interactions and their implications, we evaluate recent case studies of indirect nontarget effects of biological control agents in the context of theoretical work in community ecology. We find that although particular indirect nontarget effects are extremely difficult to predict, all indirect nontarget effects of host specific biological control agents derive from the nature and strength of the interaction between the biological control agent and the pest. Additionally, recent theoretical work suggests that the degree of impact of a biological control agent on nontarget species is proportional to the agent’s abundance, which will be highest for moderately successful control agents. Therefore, the key to safeguarding against indirect nontarget effects of host-specific biological control agents is to ensure the biological control agents are not only host specific, but also efficacious. Biological control agents that greatly reduce their target species while remaining host-specific will reduce their own populations through density-dependent feedbacks that minimize risks to nontarget species.     

Host persistence or extinction from emerging infectious disease: insights from white-nose syndrome in endemic and invading regions

Predicting species'' fates following the introduction of a novel pathogen is a significant and growing problem in conservation. Comparing disease dynamics between introduced and endemic regions can offer insight into which naive hosts will persist or go extinct, with disease acting as a filter on host communities. We examined four hypothesized mechanisms for host–pathogen persistence by comparing host infection patterns and environmental reservoirs for Pseudogymnoascus destructans (the causative agent of white-nose syndrome) in Asia, an endemic region, and North America, where the pathogen has recently invaded. Although colony sizes of bats and hibernacula temperatures were very similar, both infection prevalence and fungal loads were much lower on bats and in the environment in Asia than North America. These results indicate that transmission intensity and pathogen growth are lower in Asia, likely due to higher host resistance to pathogen growth in this endemic region, and not due to host tolerance, lower transmission due to smaller populations, or lower environmentally driven pathogen growth rate. Disease filtering also appears to be favouring initially resistant species in North America. More broadly, determining the mechanisms allowing species persistence in endemic regions can help identify species at greater risk of extinction in introduced regions, and determine the consequences for disease dynamics and host–pathogen coevolution.     

Potential of Lecanicillium spp. for management of insects, nematodes and plant diseases

Fungi in the genus Lecanicillium (formerly classified as the single species Verticillium lecanii) are important pathogens of insects and some have been developed as commercial biopesticides. Some isolates are also active against phytoparasitic nematodes or fungi. Lecanicillium spp. use both mechanical forces and hydrolytic enzymes to directly penetrate the insect integument and the cell wall of the fungal plant pathogen. In addition to mycoparasitism of the plant pathogen, the mode of action is linked to colonization of host plant tissues, triggering an induced systemic resistance. Recently it was demonstrated that development of Lecanicillium hybrids through protoplast fusion may result in strains that inherit parental attributes, thereby allowing development of hybrid strains with broader host range and other increased benefits, such as increased viability. Such hybrids have demonstrated increased virulence against aphids, whiteflies and the soybean cyst nematode. Three naturally occurring species of Lecanicillium, L. attenuatum, L. longisporum, and an isolate that could not be linked to any presently described species based on rDNA sequences have been shown to have potential to control aphids as well as suppress the growth and spore production of Sphaerotheca fuliginea, the causal agent of cucumber powdery mildew. These results suggest that strains of Lecanicillium spp. may have potential for development as a single microbial control agent effective against several plant diseases, pest insects and plant parasitic nematodes due to its antagonistic, parasitic and disease resistance inducing characteristics. However, to our knowledge, no Lecanicillium spp. have been developed for control of phytopathogens or phytoparasitic nematodes.     

Metarhizium anisopliae lipolytic activity plays a pivotal role in Rhipicephalus (Boophilus) microplus infection

Lipases secreted by Metarhizium anisopliae, an important biological control agent, could potentially be involved in the host infection process. Here, we present the activity profile during the host infection process and the effect of lipase activity inhibitor ebelactone B on infection. The previous treatment of spores with lipase activity inhibitor, ebelactone B, completely inhibited lipolytic activity and prevented the infection of the Rhipicephalus (Boophilus) microplus host. The results herein presented prove, for the first time, the importance of lipase activity in M. anisopliae host infection process. The filamentous fungus Metarhizium anisopliae is one of the most important and studied biological agents for the control of several arthropod pests, including the cattle tick Rhipicephalus (Boophilus) microplus. Lipases secreted by M. anisopliae could potentially be involved in the host infection process. This work presents the activity profile during the host infection process and the effect of lipase activity inhibitor ebelactone B on infection. During the course of tick exposure to spores (6-120 h) lipase activity increased from 0.03 ± 0.00 U to 0.312 ± 0.068 U using rho NP palmitate as substrate. In zymograms, bands of lipase activity were detected in ticks treated with spores without inhibitor. The previous treatment of spores with lipase activity inhibitor, ebelactone B, completely inhibited lipolytic activity, at all times specified, and prevented the infection of the R. microplus host. Spores treated with the inhibitor did not germinate on the tick, although this effect was not observed in the culture medium. The results herein presented prove, for the first time, the importance of lipase activity in M. anisopliae host infection process.     

Beauveria bassiana (Balsamo) Vuillemin as an endophyte in tissue culture banana (Musa spp.)

Beauveria bassiana is considered a virulent pathogen against the banana weevil Cosmopolites sordidus. However, current field application techniques for effective control against this pest remain a limitation and an alternative method for effective field application needs to be investigated. Three screenhouse experiments were conducted to determine the ability of B. bassiana to form an endophytic relationship with tissue culture banana (Musa spp.) plants and to evaluate the plants for possible harmful effects resulting from this relationship. Three Ugandan strains of B. bassiana (G41, S204 and WA) were applied by dipping the roots and rhizome in a conidial suspension, by injecting a conidial suspension into the plant rhizome and by growing the plants in sterile soil mixed with B. bassiana-colonized rice substrate. Four weeks after inoculation, plant growth parameters were determined and plant tissue colonization assessed through re-isolation of B. bassiana. All B. bassiana strains were able to colonize banana plant roots, rhizomes and pseudostem bases. Dipping plants in a conidial suspension achieved the highest colonization with no negative effect on plant growth or survival. Beauveria bassiana strain G41 was the best colonizer (up to 68%, 79% and 41% in roots, rhizome and pseudostem base, respectively) when plants were dipped. This study demonstrated that, depending on strain and inoculation method, B. bassiana can form an endophytic relationship with tissue culture banana plants, causing no harmful effects and might provide an alternative method for biological control of C. sordidus.     

Modeling the impact of a biocontrol agent, Puccinia lagenophorae, on interactions between a crop, Daucus carota, and a weed, Senecio vulgaris

The impact of a biocontrol agent spreading from a point source on crop–weed interactions was modeled. The model encompassed: (i) severity of crop–weed competition as affected by a rust pathogen, (ii) velocity of spread of the rust pathogen, and (iii) density of infected plants introduced in the weed population as starting points (inoculum sources) for an epidemic. The model was parameterized for a study system encompassing the crop Daucus carota (carrot), the weed Senecio vulgaris (common groundsel), and its antagonist Puccinia lagenophorae. The parameters of (i) were estimated in a greenhouse study using a response-surface design. Estimates of the parameter of (ii) were obtained from the literature. The density of infected plants (iii) was varied to simulate crop loss as function of density. Simulations were run for various densities of the weed and various velocities of rust pathogen spread. The results of the simulations indicated a crop-loss ranging from 5 to 10% at levels of relatively weak D. carota–S. vulgaris competition. Density of inoculum sources and velocity of P. lagenophorae spread had only minor effects on crop loss. In contrast, density of inoculum sources and velocity of spread had major effects on crop loss at levels of intermediate (range of 10–35% loss) and severe competition (range of 30–70% loss). The results are discussed both with respect to biological control of S. vulgaris using P. lagenophorae as biocontrol agent and as a general model describing the impact of the spatial dynamics of a pathogen (natural enemy) on plant competition.     

Location of pathogenicity genes on dispensable chromosomes inNectria haematococca MPVI

Nectria haematococca MPVI can be found in many different biological habitats but has been most studied as a pathogen of pea (Pisum sativum). Genetic analyses of isolates obtained from a variety of biological sources has indicated that a number of genes control pathogenicity on pea but that one important PEa Pathogenicity (PEP) gene isPDA, which confers the ability to detoxify the pea phytoalexin pisatin. In these studies, all naturally occurring isolates that lackedPDA (i.e. Pda^– isolates) and all Pda^– progeny were essentially non-pathogenic on pea. However, we have demonstrated recently that Pda^– mutants created by transformation-mediated gene disruptions, while having a modest reduction in virulence, and more virulent than any naturally occurring Pda^– isolates. In addition we know thatPDA genes are on dispensable (DS) chromosomes in this fungus. We believed that the gene disruption mutants have allowed the detection of otherPEP genes that are present on the DS chomosomes along withPDA and that naturally occuring Pda^– isolates usually lack this DS chromosome. This would explain why naturally occurring Pda^– isolates are always low in virulence. We propose that the DS chromosomes in fungi are analogous to bacterial plasmids which allow those microorganisms to colonise different habitats, i.e. the DS chromosomes ofNectria haematococca contain genes that allow individual isolates of this broad host range pathogen to occupy different biological niches.     

Sensitivity of a digoxigenin-labelled DNA probe in detecting Mikrocytos mackini, causative agent of Denman Island disease (mikrocytosis), in oysters

The protistan parasite Mikrocytos mackini, causative agent of Denman Island disease (mikrocytosis), induces mortality and reduces marketability in the Pacific oyster, Crassostrea gigas, in British Columbia, Canada. This parasite is a pathogen of international concern because it infects a range of oyster species, and because its life cycle and mode of transmission are unknown. A digoxigenin-labelled DNA probe in situ hybridisation technique (DIG-ISH) was developed, and its detection sensitivity was compared to standard histological sections stained with haematoxylin and eosin stain (H&E-histo). In H&E-histo preparations, the detection of M. mackini was certain only when the parasite occurred within the vesicular connective tissue of adult oysters. However, the DIG-ISH technique clearly demonstrated the presence of infection in all other host tissues as well as in juvenile oysters with poorly developed vesicular connective tissue. The probe hybridised strongly to M. mackini, did not hybridise to oyster tissues or with the other shellfish parasites tested, and was more sensitive for detecting infections when compared to H&E-histo.     

Host Status of Endophyte-Infected and Noninfected Tall Fescue Grass to Meloidogyne spp

Tall fescue grass cultivars with or without endophytes were evaluated for their susceptibility to Meloidogyne incognita in the greenhouse. Tall fescue cultivars evaluated included, i) wild-type Jesup (E+, ergot-producing endophyte present), ii) endophyte-free Jesup (E-, no endophyte present), iii) Jesup (Max-Q, non-ergot producing endophyte) and iv) Georgia 5 (E+). Peach was included as the control. Peach supported greater (P ≤ 0.05) reproduction of M. incognita than all tall fescue cultivars. Differences in reproduction were not detected among the tall fescue cultivars and all cultivars were rated as either poor or nonhosts for M. incognita. Suppression of M. incognita reproduction was not influenced by endophyte status. In two other greenhouse experiments, host susceptibility of tall fescue grasses to two M. incognita isolates (BY-peach isolate and GA-peach isolate) did not appear to be related to fungal endophyte strain [i.e., Jesup (Max-Q; nontoxic endophyte strain) vs. Bulldog 51 (toxic endophyte strain)]. Host status of tall fescue varied with species of root-knot nematode. Jesup (Max-Q) was rated as a nonhost for M. incognita (BY-peach isolate and GA-peach isolate) and M. hapla, a poor host for M. javanica and a good host for M. arenaria. Bulldog 51 tall fescue was also a good host for M. arenaria and M. javanica, but not M. incognita. Jesup (Max-Q) tall fescue may have potential as a preplant control strategy for M. incognita and M. hapla in southeastern and northeastern United States, respectively.     

Host-parasite local adaptation after experimental coevolution of Caenorhabditis elegans and its microparasite Bacillus thuringiensis

Coevolving hosts and parasites can adapt to their local antagonist. In studies on natural populations, the observation of local adaptation patterns is thus often taken as indirect evidence for coevolution. Based on this approach, coevolution was previously inferred from an overall pattern of either parasite or host local adaptation. Many studies, however, failed to detect such a pattern. One explanation is that the studied system was not subject to coevolution. Alternatively, coevolution occurred, but remained undetected because it took different routes in different populations. In some populations, it is the host that is locally adapted, whereas in others it is the parasite, leading to the absence of an overall local adaptation pattern. Here, we test for overall as well as population-specific patterns of local adaptation using experimentally coevolved populations of the nematode Caenorhabditis elegans and its bacterial microparasite Bacillus thuringiensis. Furthermore, we assessed the importance of random interaction effects using control populations that evolved in the absence of the respective antagonist. Our results demonstrate that experimental coevolution produces distinct local adaptation patterns in different replicate populations, including host, parasite or absence of local adaptation. Our study thus provides experimental evidence of the predictions of the geographical mosaic theory of coevolution, i.e. that the interaction between parasite and host varies across populations.     

Endophytic fungi associated with Fallopia japonica (Polygonaceae) in Japan and their interactions with Puccinia polygoni-amphibii var. tovariae, a candidate for classical biological control

Fallopia japonica (Polygonaceae), or Japanese knotweed, is now spreading globally, causing serious problems in Europe and North America in both natural and urban habitats. There is an urgent need for alternative management solutions, and classical biological control, using coevolved natural enemies found in the native range, is currently being investigated. Here, we isolated fungal endophytes from F. japonica in Japan, its natural habitat, to find endophytes that might increase the virulence of a coevolved rust pathogen, Puccinia polygoni-amphibii var. tovariae. A total of 1581 fungal endophytes were recovered from F. japonica and classified into 15 taxa. Five genera (Colletotrichum, Pestalotiopsis, Phoma, Phomopsis, and Alternaria) were dominant as endophytes in F. japonica. A greenhouse study of the dominant endophyte-pathogen interactions revealed three types of reactions: suppressive, synergistic, and neutral. In particular, one Phomopsis isolate--closely related to Diaporthe medusaea, based on ITS sequences--promoted the pathogenic aggressiveness of P. polygoni-amphibii var. tovariae and, therefore, this interaction is potentially useful to increase the effectiveness of the rust fungus as a biological control agent of F. japonica in its invasive range.     

Comparative genomic analysis of Ralstonia solanacearum reveals candidate genes for host specificity

Background

Ralstonia solanacearum is a vascular soil-borne plant pathogen with an unusually broad host range. This economically destructive and globally distributed bacterium has thousands of distinct lineages within a heterogeneous and taxonomically disputed species complex. Some lineages include highly host-adapted strains (ecotypes), such as the banana Moko disease-causing strains, the cold-tolerant potato brown rot strains (also known as R3bv2) and the recently emerged Not Pathogenic to Banana (NPB) strains.

Results

These distinct ecotypes offer a robust model to study host adaptation and the emergence of ecotypes because the polyphyletic Moko strains include lineages that are phylogenetically close to the monophyletic brown rot and NPB strains. Draft genomes of eight new strains belonging to these three model ecotypes were produced to complement the eleven publicly available R. solanacearum genomes. Using a suite of bioinformatics methods, we searched for genetic and evolutionary features that distinguish ecotypes and propose specific hypotheses concerning mechanisms of host adaptation in the R. solanacearum species complex. Genome-wide, few differences were identified, but gene loss events, non-synonymous polymorphisms, and horizontal gene transfer were identified among type III effectors and were associated with host range differences.

Conclusions

This extensive comparative genomics analysis uncovered relatively few divergent features among closely related strains with contrasting biological characteristics; however, several virulence factors were associated with the emergence of Moko, NPB and brown rot and could explain host adaptation.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1474-8) contains supplementary material, which is available to authorized users.