Antagonistic effects of the endophytic fungus Meira geulakonigii on the citrus rust mite Phyllocoptruta oleivora

AIMS: The fungus Meira geulakonigii has been shown to reduce populations of citrus rust mite (CRM; Phyllocoptruta oleivora) on citrus leaves and fruits, in both the field and laboratory. However, attempts to isolate the fungus from leaves and fruits have been unsuccessful. The aims of this study were therefore to determine whether M. geulakonigii is a citrus endophyte, and to assess possible mechanisms involved in its mite-antagonist activity. METHODS AND RESULTS: A quantitative real-time PCR and regular PCR approaches were developed to detect M. geulakonigii in both the field and laboratory. The fungus was detected throughout. Different methods revealed that M. geulakonigii is an endophyte, which colonizes both the peel of grapefruits. Applications of conidia protected the grapefruits against CRM, and fungal secretions extracted from growth media caused 100% CRM mortality. CONCLUSIONS: Meira geulakonigii is a beneficial endophyte of grapefruits that colonizes the fruit's peel, and protects it from CRM. SIGNIFICANCE AND IMPACT OF THE STUDY: Findings from this study demonstrate the endophytic nature of M. geulakonigii in its interaction with grapefruits. In addition, a molecular approach was developed to specifically detect the fungus inside the grapefruit peel. This approach can be used to assess the natural occurrence of M. geulakonigii in grapefruit.     

毛竹上两株不同菌落形态的格氏梅拉菌Meira geulakonigii菌株的分离和特征记述

格氏梅拉菌Meira geulakonigii最早是在柑桔锈螨病的枯枝中发现的,它是一种担子菌类酵母样无性态真菌,归属于黑粉菌纲外担子菌亚纲。目前仅了解这种真菌在植物中的生长状态,关于它的宿主范围、区域分布和生长特性依然知之甚少。首次从竹子中分离了两株新的Meira geulakonigii菌株(PM1和PM2),并且更深一步地描述了他们的生理特性,尤其着重表述了菌丝体的独特生长现象。与先前报导的两株M.geulakonigii菌株PFS004和AS004相比较,PM1和PM2在分子系统上与AS004更相近,而生理现象与PFS004更相似。这两株菌株的成功分离可能有助于毛竹中Meira真菌菌丝体形成和功能的进一步研究。     

The Biology of the Acaropathogenic Fungus Hirsutella kirchneri

The acaropathogenic fungus Hirsutella kirchneri (Rostrup) Minter , Brady and Hall grew best and produced most mycelia on a medium containing yeast extract , dextrose and agar; conidial production , however , was maximal or potato dextrose agar (PDA) . The best growth on both media was at 25 C and conidial germination was high within a wide range of temperatures (10 - 35 C) . Colony growth , mycelial mass production and conidial yield were best under alternating dark and light regimes . Maximal germination occurred under dark conditions . When grown in continuous light the fungus produced synnemata (compacted conidiophores) which remained viable for 22 weeks . Of six species of phytophagous mites assayed , three spider mites and a rust mite became infected by the fungus , as did , to a limited degree , a parasitic mite . Another four mites , including two pests , a scavenger and a predator , were unaffected . The fungus grew on and sporulated from heat - killed cadavers of a dead mealybug (Homoptera) . Conidial germination and penetration into live mites , under saturation condi tions , were affected little by temperatures . Intra - host growth was temperature dependent , with mite death beginning on the second or third day post - infection . Maximal sporulation from infected mites took place at 25 C . Mortality was quickest at 25 C when mites were held under saturation conditions , but some death also occurred at lower relative humidities . These data are discussed with a view to using H. kirchneri in the biological control of plant mites .     

Assaying three new fungi against citrus mites in the laboratory,and a field trial

The Basidiomycotine fungi Meira geulakonigii, Meira argovae and Acaromyces ingoldii were assayed in the laboratory against five species of herbivorous mites: Phyllocoptruta oleivora (Eriophyidae), Panonychus citri, Eutetranychus orientalis, Tetranychus urticae and Tetranychus cinnabarinus (all four Tetranychidae). All fungi caused significantly high mortality rates (as compared to controls) after 14 days, some after 1 week. Phyllocoptruta oleivora was the most susceptible, showing >80% mortality even after 1 week. In a field trial, grapefruits sprayed either once a month or once a season with M. geulakonigii had significantly fewer P. oleivora and less damage than unsprayed fruit. These results suggest that M. geulakonigii may protect grapefruits against the injurious P. oleivora.     

Exudate from sporulating cultures of Hirsutella thompsonii inhibit oviposition by the two-spotted spider mite Tetranychus urticae

The acaricidal mycopathogen Hirsutella thompsonii has been found to secrete metabolites that are active against femaleTetranychus urticae. Specifically, the rose-colored exudate produced on sporulating cultures of Mexican HtM120I strain sterilized female spider mites in a dose-dependent fashion. Topical application of the exudate resulted in a 100% reduction in mite fecundity over the initial six days of experimentation. Depending upon the exudate dosage, mites partially recovered within 3 and 6 d post-treatment and produced a limited number of eggs. The spider mite active HtM120I exudate contained less detectable HtA toxin than the HtM120I broth filtrate, and it was innocuous when injected into the greater wax moth Galleria mellonella L. larvae. Broth filtrates of HtM120I cultures, although toxic to assayed G. mellonella larvae, did not inhibit mite oviposition to the degree or duration of the exudate preparations. These findings suggest that the factor responsible for suppressing oviposition in female spider mites is linked to the sporulation process and is distinct from the well-characterized HtA produced by vegetative cells. This revised version was published online in July 2006 with corrections to the Cover Date.     

Fungal Biocontrol of Acari

Mites and ticks are susceptible to pathogenic fungi, and there are opportunities to exploit these micro-organisms for biological control. We have collated records of 58 species of fungi infecting at least 73 species of Acari, either naturally or in experiments. Fungal pathogens have been reported to kill representatives of all three orders of the Actinotrichida (the Astigmata, Oribatida and Prostigmata) and the Ixodida and Mesostigmata in the Anactinotrichida. Most reports concern infections in the Prostigmata, particularly in the families Tetranychidae and Eriophyidae. Two species of Acari-specific pathogens - Hirsutella thompsonii and Neozygites floridana - are important natural regulators of pestiferous eriophyoid and tetranychid mites respectively. Research has been done to understand the factors leading to epizootics of these fungi and to conserve and enhance natural pest control. Hirsutella thompsonii was also developed as the commercial product Mycar for the control of eriophyoid mites on citrus, but was withdrawn from sale in the 1980s, despite some promising effects in the field. Beauveria bassiana , Metarhizium anisopliae, Paecilomyces farinosus, Paecilomyces fumosoroseus and Verticillium lecanii infect ixodid ticks in nature, and B. bassiana and M. anisopliae are being studied as biological control agents of cattle ticks in Africa and South America. Beauveria bassiana also has potential as a mycopesticide of the two-spotted spider mite, Tetranychus urticae . There is scope to develop fungal biocontrol agents against a range of acarine pests, both as stand-alone treatments and for use in integrated pest management. Further research is required to clarify the taxonomic status of fungal pathogens of Acari, to study their ecosystem function, and to develop efficient mass production systems for species of Hirsutella and Neozygites .     

Diseases of Mites

An overview is given of studies on diseases of mites. Knowledge of diseases of mites is still fragmentary but in recent years more attention has been paid to acaropathogens, often because of the economic importance of many mite species. Most research on mite pathogens concerns studies on fungal pathogens of eriophyoids and spider mites especially. These fungi often play an important role in the regulation of natural mite populations and are sometimes able to decimate populations of phytophagous mites. Studies are being conducted to develop some of these fungi as commercial acaricides.Virus diseases are known in only a few mites, namely, the citrus red mite and the European red mite. In both cases, non-occluded viruses play an important role in the regulation of mite populations in citrus and peach orchards, respectively, but application of these viruses as biological control agents does not seem feasible. A putative iridovirus has been observed in association with Varroa mites in moribund honeybee colonies. The virus is probably also pathogenic for honeybees and may be transmitted to them through this parasitic mite.Few bacteria have been reported as pathogens of the Acari but in recent years research has been concentrated on intracellular organisms such as Wolbachia that may cause distorted sex ratios in offspring and incompatibility between populations. The role of these organisms in natural populations of spider mites is in particular discussed. The effect of Bacillus thuringiensis on mites is also treated in this review, although its mode of action in arthropods is mainly due to the presence of toxins and it is, therefore, not considered to be a pathogen in the true sense of the word.Microsporidia have been observed in several mite species especially in oribatid mites, although other groups of mites may also be affected. In recent years, Microsporidia infections in Phytoseiidae have received considerable attention, as they are often found in mass rearings of beneficial arthropods. They affect the efficacy of these predators as biological control agent of insect and mite pests. Microsporidia do not seem to have potential for biological control of mites.     

Fluorometric microplate assay to measure glutathione S-transferase activity in insects and mites using monochlorobimane

Elevated levels of glutathione S-transferases (GSTs) play a major role as a mechanism of resistance to insecticides and acaricides in resistant pest insects and mites, respectively. Such compounds are either detoxicated directly via phase I metabolism or detoxicated by phase II metabolism of metabolites as formed by microsomal monooxygenases. Here we used monochlorobimane (MCB) as an artificial substrate and glutathione to determine total GST activity in equivalents of single pest insects and spider mites in a sensitive 96-well plate-based assay system by measuring the enzymatic conversion of MCB to its fluorescent bimane-glutathione adduct. The differentiation by their GST activity between several strains of the two-spotted spider mite, Tetranychus urticae (Acari: Tetranychidae), with different degrees of resistance to numerous acaricides was more sensitive with MCB compared to the commonly used substrate 1-chloro-2,4-dinitrobenzene (CDNB). Compared to an acaricide-susceptible reference strain, one field population of T. urticae showed a more than 10-fold higher GST activity measured with MCB, in contrast to a less than 2-fold higher activity when CDNB was used. Furthermore, we showed that GST activity can be sensitively assessed with MCB in homogenates of pest insects such as Heliothis virescens, Spodoptera frugiperda (Lepidoptera: Noctuidae), Plutella xylostella (Lepidoptera: Yponomeutidae), and Myzus persicae (Hemiptera: Aphididae).     

Resistance status of the carmine spider mite,Tetranychus cinnabarinus and the twospotted spider mite,Tetranychus urticae to selected acaricides on strawberries

The carmine spider mite, Tetranychus cinnabarinus (Boisduval) and the twospotted spider mite, Tetranychus urticae Koch, are serious pests of strawberries and many other horticultural crops. Control of these pests has been heavily dependent upon chemical acaricides. Objectives of this study were to determine the resistance status of these two pest species to commonly used acaricides on strawberries in a year‐round intensive horticultural production region. LC₉₀ of abamectin for adult carmine spider mites was 4% whereas that for adult twospotted spider mites was 24% of the top label rate. LC_90s of spiromesifen, etoxazole, hexythiazox and bifenazate were 0.5%, 0.5%, 1.4% and 83% of their respective highest label rates for carmine spider mite eggs, 0.7%, 2.7%, 12.1% and 347% of their respective highest label rates for the nymphs. LC_90s of spiromesifen, etoxazole, hexythiazox and bifenazate were 4.6%, 11.1%, 310% and 62% of their respective highest label rates for twospotted spider mite eggs, 3%, 13%, 432,214% and 15% of their respective highest label rates for the nymphs. Our results suggest that T. cinnabarinus have developed resistance to bifenazate and that the T. urticae have developed resistance to hexythiazox. These results strongly emphasize the need to develop resistance management strategies in the region.     

Diversity of acaropathogenic fungi in Poland and other European countries

The occurrence, species diversity and some aspects of taxonomical affinity and host selectivity of acaropathogenic fungi associated with phytophagous, saprotrophic and predacious mites in Poland and other European countries were investigated on wild and cultivated plants, in insect feeding sites under the bark and in decayed wood. From among 33 species of fungi affecting mites only five species of Entomophthorales were separated and the most numerous were Neozygites floridana mostly on Tetranychus urticae, N. abacaridis on a few eriophyid species, and Conidiobolus coronatus attacking gamasid mites most frequently of the genus Dendrolaelaps. The most frequent mite pathogens occurring in mite communities on plants and in wood infested by insects were of the genus Hirsutella. Until now 13 of their form-species have been recognized in these habitats, but only H. kirchneri, H. necatrix and H. thompsonii (including its variety synnematosa) can be treated as exclusive oligophagous pathogens of phytophagous mites, though their potential host range seems to embrace only selected eriophyid or tarsonemid mites. Taxonomical differentiation of fungal strains was based on close morphological observations and molecular analysis of ITS region sequences. Two new species of acaropathogenic fungi were described in these studies. Hirsutella danubiensis sp. nov. was found in the tetranychid T. urticae, whereas H. vandergeesti sp. nov. affected phytoseiid mites of the genera Amblyseius, Neoseiulus, Seiulus and Typhlodromus, and the tarsonemid Tarsonemus lacustris.     

Toxicity of bifenazate and its principal active metabolite, diazene, to Tetranychus urticae and Panonychus citri and their relative toxicity to the predaceous mites, Phytoseiulus persimilis and Neoseiulus californicus

Bifenazate is a novel carbazate acaricide discovered by Uniroyal Chemical (now Chemtura Corporation) for the control of phytophagous mites infesting agricultural and ornamental crops. Its acaricidal activity and that of its principal active metabolite, diazene, were characterized. Bifenazate and diazene had high toxicity and specificity both orally and topically to all life stages of Tetranychus urticae and Panonychus citri. Acute poisoning was observed with no temperature dependency. No cross-resistance was found to mites resistant to several other classes of acaricides, such as tebufenpyrad, etoxazole, fenbutatin oxide and dicofol. Bifenazate remained effective for a long time with only about a 10% loss of efficacy on T. urticae after 1 month of application in the field. All stages of development of the predatory mites, Phytoseiulus persimilis and Neoseiulus californicus, survived treatment by both bifenazate and diazene. When adult females of the two predatory mite species were treated with either bifenazate or diazene, they showed a normal level of fecundity and predatory activity in the laboratory, effectively suppressing spider mite population growth. Even when the predators were fed spider mite eggs that had been treated previously with bifenazate, they survived. These findings indicate that bifenazate is a very useful acaricide giving high efficacy, long-lasting activity and excellent selectivity for spider mites. It is, therefore, concluded that bifenazate is an ideal compound for controlling these pest mites.     

Effects of chlorpyrifos and sulfur on spider mites (Acari: Tetranychidae) and their natural enemies

In many agricultural systems spider mites are believed to be induced pests, only reaching damaging densities after pesticides decimate predator populations. Wine grapes typically receive two types of pesticides, insecticides and fungicides. Chemicals in either class could impact spider mite densities both directly through spider mite mortality, and indirectly by negatively affecting natural enemies. The impact of a broad-spectrum insecticide (chlorpyrifos) and an inorganic fungicide (sulfur) on mites and their natural enemies was monitored in replicate open-field experiments conducted in an abandoned vineyard in Washington State. In both experiments, chemicals were applied within a 2 × 2 factorial design, allowing assessment of both main and interactive effects of the two chemicals. Following typical management practices on wine grapes in Washington State, we made a single insecticide application early in the season, but repeatedly applied sulfur throughout the season. In the absence of sulfur, chlorpyrifos application led to higher spider mite densities. The main effect of chlorpyrifos appeared to be indirect, perhaps mediated through mortality of generalist phytoseiid mites; generalists appeared to be unable to recover following even a single insecticide application, while there was no evidence for harmful effects of chlorpyrifos on specialist phytoseiid mites. Sulfur had direct suppressive effects on both pest and predatory mites, although in the second experiment the suppressive effect of sulfur on spider mites was weaker when chlorpyrifos was also applied. These field experiments suggest that a complex mix of direct and indirect effects of the two chemicals impacted spider mite population dynamics in our system.     

Mycopathogens of Mites in Poland - A Review

Studies on diseases of mites caused by entomopathogenic fungi have been undertaken in Poland about a half a century later than in West European countries. Nevertheless, during the last 30 years almost 40 species of arthropod pathogenic fungi have been identified, among them 23 species of Entomophthorales, 12 species of Hyphomycetales and four species of ectoparasitic Laboulbeniales. The most common are representatives of the genus Hirsutella , found both on phytophagous and parasitic or predaceous mites. Percentage infection by H. thompsonii in populations of some tarsonemid and eriophyid mites increases slowly from the end of spring reaching a maximum of 30-60% in August-September. Neozygites floridana is a common entomophthoralean species causing epizootics in spider mite populations; infection also peaks in late summer and autumn. The most numerous fungal taxa have been identified from the resting spores produced internally within the body of the host and show features of the genus Tarichium . From the total number of the taxa recorded within these studies, 22 were described as species new to science. Current research on invertebrate pathology in Poland includes many groups of mites and among their pathogens fungal diseases seem to be the most common.     

Impact of transgenic Bacillus thuringiensis cotton on a non-target pest Tetranychus spp. in northern China

Field studies were carried out to evaluate the effect of two transgenic cotton varieties (SGK321 carrying Cry1A+CpTI and DP99B carrying Cry1Ac) and the conventional variety (shiyuan321‐parental line of SGK321) on spider mites, Tetranychus spp. from 2002 to 2004. In 2002, this experiment included three treatments: Bt cotton field (SGK321) treated with acaricides against spider mites, untreated non‐Bt cotton field (Shiyuan321), and untreated Bt cotton field (SGK321). In 2003–2004, there are four types of treatments after a new transgenic Bt cotton variety, DP99B (non‐chemical control), was added into the experiments. The results showed that there were no significant difference in densities of spider mites among Bt without acaricides and non‐Bt without acaricides cotton fields, nor was there a significant difference in damage of spider mites to cotton among these treatments (P > 0.1). However, there are significant differences (P < 0.05) in densities of spider mites and damage caused by spider mites between cotton fields with and without acaricides. Acaricide significantly reduced the densities of spider mites in Bt cotton (P < 0.05). These results suggest that Bt cotton has no effect on spider mites populations. However, spider mites have the potential for severe damage in Bt cotton fields. Acaricides are essential tools in controlling cotton spider mites in northern China.     

Argovin,a novel natural product secreted by the fungus Meira argovae,is antagonistic to mites

A metabolite of the fungus Meira argovae Boekhout, Scorzetti, Gerson & Sztejnberg (Exobasidiomycetidae) was assayed as an antagonist of mites. Separation of extracted fungal metabolites by reversed phase liquid chromatography (RPLC), with subsequent testing of the obtained fractions, allowed us to isolate a single mite‐antagonistic fraction (also active against a bacterium) that primarily includes one major component. This active compound (herein termed ‘argovin’) was identified by analyzing its spectral characteristics as 4,5‐dihydroxyindan‐1‐one, which has previously only been described as a product of chemical reactions. The growth rate of the fungus was higher at a neutral pH than at an acidic one. Meira argovae adjusts the pH of its media to values optimal for its colony growth and toxic secretions. RPLC‐cleaned argovin at 0.2 mg ml^?1 killed 100% of a population of the citrus rust mite, Phyllocoptruta oleivora (Ashmead) (Acari: Eriophyidae). This trait may be used to control citrus rust mites in the field, as well as for toxin production for industrial and pharmaceutical uses.     

Hirsutella thompsonii a fungal parasite of the citrus rust mitePhyllocoptruta oleivora in the Rio Grande Valley of Texas

Hirsutella thompsonii, a hyphomycetous fungus, was found attacking the citrus rust mitePhyllocoptruta oleivora for the first time in May, 1972, in the Lower Rio Grande Valley of Texas. Sharp reductions in mite populations occurred during this period. The fungus was isolated from mites collected from citrus fruits and leaves. The parasitic nature and the distribution of the pathogen are described.     

Modelling Fungal (Neozygites cf. Floridana) Epizootics in Local Populations of Cassava Green Mites (Mononychellus Tanajoa)

The fungus, Neozygitis cf. floridana is parasitic on the cassava green mite, Mononychellus tanajoa (Bondar) (Acari: Tetranychidae) in South America and may be considered for classical biological control of cassava green mites in Africa, where cassava is an important subsistence crop, cassava green mites are an imported pest and specific natural enemies are lacking. Spider mites generally have a viscous structure of local populations, a trait that would normally hamper the spread of a fungus that is transmitted by the contact of susceptible hosts with the halo of capilliconidia surrounding an infectious host. However, if infected mites search and settle to produce capilliconidia on sites where they are surrounded by susceptible mites before becoming infectious, then the conditions for maximal transmission in a viscous host population are met. Because the ratio between spider mites and the leaf area they occupy is constant, parasite-induced host searching behaviour leads to a constant per capita transmission rate. Hence, the transmission rate only depends on the number of infectious hosts. These assumptions on parasite-induced host search and constant host density lead to a simple, analytically tractable model that can be used to estimate the maximal capacity of the fungus to decimate local populations of the cassava green mite. By estimating the parameters of this model (host density, per capita transmission rate and duration of infected and infectious state) it was shown that the fungal pathogen can reduce the population growth of M. tanajoa, but cannot drive local mite populations to extinction. Only when the initial ratio of infectious to susceptible mites exceeds unity or the effective growth rate of the mite population is sufficiently reduced by other factors than the fungus (e.g. lower food quality of the host plant, dislodgement and death by rain and wind and predation), will the fungal pathogen be capable of decimating the cassava green mite population. Under realistic field conditions, where all of these growth-reducing factors are likely to operate, there may well be room for effective control by the parasitic fungus. This revised version was published online in November 2006 with corrections to the Cover Date.     

Biological, Chemical and Integrated Control of Two-spotted Spider Mite Tetranychus urticae on Dwarf Hops

Chemical, biological and integrated programmes for the control of two-spotted spider mite, Tetranychus urticae, were compared on dwarf hops in 1997 and 1998. In both years integrated control, which consisted of an application of the ovicidal acaricide clofentezine followed by a release of the predatory mite Phytoseiulus persimilis at 10 individuals per plant, was the most effective treatment. Similar numbers of spider mites were recorded on plots that were treated with P. persimilis only or with a single application of the acaricide tebufenpyrad, and plots with either of these treatments had higher levels of infestation than the plots under integrated control. The highest numbers of spider mites were found on the untreated plots. This study indicates that integrated control of T. urticae using clofentezine in conjunction with P. persimilis is likely to be more effective than an approach based on chemical or biological measures only. It is suggested that an integrated system would have the added benefit over a pesticide-only programme of reducing pressure on the pest for the selection of strains resistant to acaricides.     

Evaluation of Metarhizium anisopliae (Deuteromycota: Hyphomycetes) for control of broad mite Polyphagotarsonemus latus (Acari: Tarsonemidae) in mulberry

A study on 12 entomopathogenic fungi for controlling broad mite (Polyphagotarsonemus latus (Banks)) in mulberry found that Metarhizium anisopliae CKM-048 was the most virulent strain in controlling both larvae and adult broad mites at the concentration of 2 x 10(8) conidia/ml. There was no ovicidal effect when tested with broad mite eggs. Median lethal concentrations (LC(50)) of M. anisopliae in killing larvae and adults were 8.7 x 10(6) and 1.3 x 10(7 )conidia/ml, respectively. Median lethal times (LT(50)) of larvae and adults were 2.4 and 3.8 days, respectively, at the concentration of 2 x 10(8) conidia/ml. The fungus was found to produce protease and chitinase. Scanning electron microscope (SEM) studies were done to monitor the infection steps of the fungus on broad mites. A greenhouse test on mulberry trees revealed that M. anisopliae could reduce the broad mite population within 4 days after treatment. However, after 7 days, its efficacy was decreased significantly.     

Genetic structure of a phytophagous mite species affected by crop practices: the case of Tetranychus urticae in clementine mandarins

Tetranychus urticae Koch is a cosmopolitan mite considered as the most polyphagous species among spider mites. This mite is a key pest of clementine mandarins in Eastern Spain, where Spanish clementine production concentrates. Crop management practices can affect the population dynamics of this mite and, consequently, its impact on the orchard. Microsatellite markers were used to study mite population genetics from two commercial orchards which had been managed differently following Integrated Pest Management (IPM) or Organic Pest Management (OPM) schemes during four consecutive years. A multiplex system including 20 microsatellite loci was designed specifically and allowed an efficient and inexpensive genotyping of individual mites. We found that the IPM population had a stronger fluctuation of population structure and higher genetic diversity compared to OPM population. Thus, our study concludes that crop management has an impact on the population genetics of T. urticae which may be related to the alternation of some acaricides under IPM.