Biotic and abiotic regulation of resting spore formation in vivo of obligate aphid pathogen Pandora nouryi: Modeling analysis and biological implication

Entomophthoralean fungus Pandora nouryi is an obligate aphid pathogen that enables to produce resting spores (azygospores) for surviving host absence. To explore possible mechanisms involved in the regulation of resting spore formation in vivo, host cohorts consisting of 40-60 nymphs of green peach aphid Myzus persicae produced within 24 h on cabbage leaf discs in petri dishes were exposed to spore showers of P. nouryi at the concentrations (C) from a very few to nearly 2000 conidia/mm² and then reared for 7-11 days at the regimes of 10-25 °C (T) and 8-16 h daylight (H_L) or ambient (17.5 ± 3.1 °C, 13:11 L:D). Aphid mortalities observed from 35-83 cohorts (showered separately) at each regime showed typical sigmoid trend and fit well a general logistic equation (0.79 ? r² ? 0.88), yielding similar LC₅₀ estimates of 1.7-6.1 conidia/mm². The proportions (P) of cadavers forming resting spores in the cohorts also fit the same equation (0.73 ? r² ? 0.85) at all tested regimes except at 10 °C, a low temperature for the host-pathogen interaction. This indicates the dependence of resting spore formation on the spore concentration. The effects of T and H_L on P over C were well elucidated by the fitted modified logistic equations P = 0.578/{1 + exp[1.710 − (0.136 − 0.0053T)C]} and P = 0.534/{1 + exp[1.639 + (0.034 − 0.0053H_L)C]} (both r² = 0.79). Our results highlight that the resting spore formation in vivo of P. nouryi is regulated primarily by the concentration of host-infecting conidia discharged from cadavers and facilitated by lower temperature and longer daylight.     

Entomophthora muscae resting spore formation in vivo in the host Delia radicum

The formation in vivo of Entomophthora muscae resting spores was investigated in the host, Delia radicum (cabbage root fly), by analysis of field data on the seasonal occurrence of E. muscae resting spores over 4 years. E. muscae resting spores in D. radicum were spherical with an average diameter of 39.4 microm, and the average numbers produced were estimated at 5.7 x 10(4) resting spores/female cadaver. Resting spores were found only after midsummer in D. radicum and almost exclusively in females. The proportion of infected females with resting spores was negatively correlated with average weekly day length after midsummer. We did not detect any significant year effect; thus, the results support the hypothesis that the photoperiod is the most important abiotic factor controlling E. muscae resting spore formation in D. radicum.     

Sporulation,storage and infectivity of obligate aphid pathogen Pandora nouryi grown on novel granules of broomcorn millet and polymer gel

Aims: Producing granular cultures of obligate aphid pathogen Pandora nouryi for improved sporulation and storage. Methods and Results: Small millet–gel granules were made of the mixtures of 80–95% millet powder with 5–20% polymer gel (polyacrylamide, polyacrylate or acrylate‐acrylamide copolymer) and inoculated with mycelia at 30 mg biomass g^?1 dry granules plus 87·5% water, followed by static incubation at 20°C for 4–12 days. The fungus grew well on 12 preparations but best on that including 10% copolymer. An 8‐day culture of this preparation discharged maximally 58·5 × 10⁴ conidia mg^?1 granule at 100% RH and was capable of ejecting conidia at the nonsaturated regimes of 86–97% RH. During storage at 6°C, granular cultures with >85% water content had twofold longevity (120 days) and half‐decline period (34–36 days) of those stored at room temperature. The steadily high water content preserved the cultures better than that decreasing at 6°C. However, conidia from 70‐day‐stored granules were less infective to Myzus persicae nymphs than those from fresh ones based on their LC₅₀s. Conclusions: The millet–gel granules had higher sporulation capacity than reported Pandora cultures and a capability of spore discharge at nonsaturated humidity. Significance and Impact of the Study: The granular cultures are more useful for aphid control.     

OBSERVATIONS ON CHYTRIDIACEOUS PARASITES OF PHANEROGAMS. XXVII. A FURTHER STUDY OF PHYSODERMA HYDROCOTYLIDIS VIÉGAS AND TEIXEIRA

A further study of Physoderma hydrocotylidis from California on its host Hydrocotyle ranunculoides gives further details on its structure and development. EM micrographs reveal that the notably thin wall of the resting spore consists of an outer, an intermediate, and an inner layer. The complete wall thickness from measurements of these EM preparations is only 0.5 μm, the thinnest of any known Physoderma resting spore. Such resting spores germinate readily in 2–5 h by the dehiscence of a broad cap and formation of a protruding endosporganium. Zoospores from the latter infect epidermal host cells and produce either the endobiotic, polycentric rhizoidal system with tenuous filaments, turbinate cells and eventually resting spores, or a hitherto unknown epibiotic, monocentric, rhizidiaceous sporangial stage. Zoospores from the latter may all bear colorless lipid globules or all faintly orange-colored ones.     

Effect of temperature,relative humidity and aphid developmental stage on the efficacy of the mycoinsecticide Mycotal® against Myzus persicae

The green peach aphid, Myzus persicae, is a major pest worldwide. An examination of the impact of temperature, relative humidity (RH) and developmental stages of M. persicae on the efficacy of the whitefly mycoinsecticide Mycotal®, based on Lecanicillium muscarium and the effects of infection on aphid fecundity was evaluated under controlled conditions. Although this fungus can be grown at a broad range of temperatures (15–30°C), the optimum temperature for control of M. persicae ranged between 20 and 30°C. L. muscarium had high efficacy as a microbial control agent against M. persicae between 55% and 90% RH. Total mortality of aphids treated with different spore dosages of L. muscarium varied according to the developmental stage: adults, fourth and third instar nymphs proved more susceptible than first instar nymphs. Although the fungus did not affect the rate of nymph production, the reproductive period of aphids significantly decreased with increasing the spore dosage. Thus, total fecundity of treated aphids was 22.6 ± 1.1 and 31.6 ± 2.4 offspring per adult at the medium (644 ± viable spore/mm²) and low (330 ± 40 viable spore/mm²) dosages, compared with 45.7 ± 4.3 offspring per untreated aphid. The results suggest that L. muscarium has the potential as a biological control agent of M. persicae.     

Importance of ambient saturation deficits in an epizootic of the fungus Neozygites floridana in cassava green mites (Mononychellus tanajoa)

The mite-pathogenic fungus Neozygites floridana Fisher (Entomophthorales: Neozygitaceae) is considered to have potential for the biological control of the cassava green mite, Mononychellus tanajoa (Bondar). However, its activity is sporadic and laboratory data suggest a strong dependence on night-time saturation deficits for transmission. We report on an epizootic of this fungus in a mite population in northeastern Brazil. During the epizootic, host populations appeared to be limited by a combination of the pathogen and a predatory mite Neoseiulus idaeus (Acari: Phytoseiidae). When temperatures increased, the epizootic finished and the host population began to grow. Abiotic conditions could not explain the variation in host mortality following pickup of infective propagules in this epizootic. However, night-time saturation did help to explain the variation in transmission from infective cadavers to newly killed hosts. This supports laboratory observations that horizontal transmission between hosts is determined mainly by saturation deficits, while the process of infection is little affected by abiotic conditions. A further field observation was the near-absence of resting spores in dead mites (ca. 0.1% of cadavers), suggesting that the pathogen population was unsuccessful in producing inoculum to infect future M. tanajoa populations. The implications are that this pathogen will only be effective as a biological control agent in periods of high relative humidity, and establishment in new areas may be limited by resting spore formation. This revised version was published online in July 2006 with corrections to the Cover Date.     

SPORE FORMATION IN THE LIFE CYCLES OF THE DIATOMS CHAETOCEROS DIADEMA AND LEPTOCYLINDRUS DANICUS1

A nitrogen limitation technique elicited the entire life cycle of the marine centric diatoms Chaetoceros diadema (Ehr.) Gran and Leptocylindrus danicus Cleve. In C. diadema the sexual cycle followed the same pattern as in the previously investigated C. didymus. Sexuality took place in narrow diameter cells, only at 2 and 5° C, and was seldom seen. Resting spore formation took place in cells of all sizes and at all temperatures at which the species grew vegetatively (2–15° C). The L. danicus life cycle is probably unique among diatoms. Nitrogen depletion induced sexuality in the entire culture at 10 and 15° C if the cell diameter was narrow (3–8 μm). Auxospore formation was followed by resting spore formation directly within the auxospore. In C. diadema, as in most centric diatoms, resting spores are not an obligate part of the life cycle, but they are in L. danicus. Resting spore formation is a versatile adaptive response in C. diadema, depending only on nitrogen depletion, although promoted by low temperatures. In L. danicus the linkage to the sexual process sharply limits conditions under which resting spores can form.     

The reproductive strategy of Argonauta nouryi (Cephalopoda: Argonautidae) in the Mexican South Pacific

ABSTRACT

Aspects of the reproductive biology of Argonauta nouryi Lorois, 1852 from the coast of Oaxaca, Mexico, were analysed. A total of 54 specimens with dorsal mantle length (ML) ranging from 6.2 to 41.1?mm were obtained from the stomach contents of five species of large predatory fishes from January 2016 to June 2018. The goals of this study were to collect data on reproductive indicators in order to evaluate the reproductive strategy of A. nouryi in the southeast Mexican Pacific and to use histological analysis to characterise the different ovarian phases based on oogenesis. Immature females (19.6?mm?ML) had sperm in their oviducal glands, indicating that female A. nouryi store sperm long after hectocotyli transmission by males. Potential fecundity was estimated as 31,000–75,026 eggs, with a batch fecundity of 2,000–8,000 eggs. Six distinct developmental stages were observed in the eggs suspended in the A. nouryi shell. Our data indicate that the A. nouryi reproductive strategy includes continuous low batch spawnings, mating early in life, and sperm storage in the spermatheca.     

Population dynamics of Myzus persicae on tobacco in Yunnan Province,China, before and after augmentative releases of Aphidius gifuensis

The population dynamics of Myzus persicae were investigated in the field in a year without releases of Aphidius gifuensis (1998–1999), in a year with A. gifuensis releases (2000–2001) and several years later (2005–2007). The results showed that both high mean population densities and damage rates did not differ between 1998 and 1999, but were significantly lower in 2000–2001 and 2005–2007. Moreover, farmers also reported the decrease of M. persicae populations and attributed the declines to augmentative releases of A. gifuensis in their own fields, indicating farmers’ recognition in the effectiveness of A. gifuensis for M. persicae control. In addition, compared with the historical data on pesticide use for M. persicae control, the number of insecticide applications and cost of M. persicae control was sustained at a low level in 2007 (several years after release of A. gifuensis). This suggests that the augmentative releases of A. gifuensis could be effective and sustainable in M. persicae control.     

努利虫疠霉感染桃蚜过程的扫描电镜观察

利用扫描电镜观察了努利虫疠霉Pandora nouryi (Remaudière & Hennebert) Humber初生分生孢子接种桃蚜Myzus persicae (Sulzer)后孢子萌发、入侵以及菌体突破虫体的整个侵染过程。结果表明：附着于虫体表面的初生分生孢子在3～5 h后即有60%以上的萌发率,萌发的孢子形成芽管或产生球形或棍棒状的附着胞;12 h后大部分孢子均已萌发,并成功入侵寄主虫体;接种60 h后,呈掌状分枝的假根首先从桃蚜胸部的腹面突破虫体长出体外,明显区别于新蚜虫疠霉Pandora neoaphidis (Remaudière & Hennebert) Humber假根突破虫体的位置;假囊状体不多见,且仅分布于蚜虫身体两侧,这可在一定程度上解释努利虫疠霉产孢对湿度条件要求较高的生物学现象。     

Evaluation of the reference genes for expression analysis using quantitative real‐time polymerase chain reaction in the green peach aphid,Myzus persicae

The green peach aphid, Myzus persicae Sulzer (Hemiptera, Aphididae), is an important cosmopolitan pest. Real time qRT‐PCR has been used for target gene expression analysis on M. persicae. Using real time qRT‐PCR, the expression levels are normalized on the basis of the reliable reference genes. However, to date, the stability of available reference genes has been insufficient. In this study, we evaluated nine candidate reference genes from M. persicae under diverse experimental conditions. The tested candidate genes were comprehensively ranked based on five alternative methods (RefFinder, geNorm, Normfinder, BestKeeper and the comparative ΔC_t method). 18s, Actin and ribosomal protein L27 (L27) were recommended as the most stable reference genes for M. persicae, whereas ribosomal protein L27 (L27) was found to be the least stable reference genes for abiotic studies (photoperiod, temperature and insecticide susceptibility). Our finding not only sheds light on establishing an accurate and reliable normalization of real time qRT‐PCR data in M. persicae but also lays a solid foundation for further studies of M. persicae involving RNA interference and functional gene research.     

Observations on chytridiaceous parasites of Phanerogams

Summary A reexamination of Physoderma maculare Wallroth the type species of the genus, on Alisma, confirms Clinton's (1902) account of the production of an epibiotic stage from resting spore zoospores. The latter on mature host tissue may also give rise to the endobiotic stage which bears resting spores. On seedlings, however, resting spore zoospores produce only epibiotic sporangia. The fungus could not successfully infect Sagittaria, a closely related host.Contribution No. 1152     

Resting spore formation ofLeptocylindrus danicus (Bacillariophyceae) during short time-scale upwelling and its significance as predicted by a simple model

Resting spore formation during short time-scale upwelling and its significance were investigated in the field and by a simple theoretical model. Field observations of spore formation ofLeptocylindrus danicus were made off Izu Peninsula, Japan. A rapid increase in ratio of resting spore to vegetative cell numbers indicated thatL. danicus formed resting spores quickly as a response to nutrient depletion in the upwelled water, although only a very low number of resting spores was found in the upwelling. A simple model was constructed to investigate the possible advantages of spore formation during short time-scale upwelling. This showed that there is a critical time-scale for resting spore formation to be advantageous. The nutrient depletion period of the upwelling off Izu was shorter than the critical time-scale determined by the model. Rapid-sinking of resting spores may increase further the critical time-scale, unless spores return with upwelling water. For short time-scale upwelling, the vegetative cell may be better suited than the resting spore for enduring a short period of nutrient depletion. Contribution from Shimoda Marine Research Center, University of Tsukuba, No. 475.     

Probing behavior of aposymbiotic green peach aphid (Myzus persicae) on susceptible Solanum tuberosum and resistant Solanum stoloniferum plants

The green peach aphid, Myzus persicae Sulzer (Hemiptera: Aphididae) is one of the potato important pests; it is the most efficient vector of potato viruses. Myzus persicae harbors the endosymbiotic bacteria Buchnera aphidicola which supplements their diet. There is increasing evidence that B. aphidicola is involved in plant–aphid interactions and we previously demonstrated that B. aphidicola disruption (aposymbiosis) affected the probing behavior of M. persicae on radish plants, delaying host plant acceptance. In this work, we evaluated the effect of aposymbiosis on the probing behavior of M. persicae on 2 Solanum species with different compatibility with M. persicae, Solanum tuberosum (susceptible) and Solanum stoloniferum (resistant) with the electrical penetration graph technique (EPG). To disrupt B. aphidicola, rifampicin was administered to aphids through artificial diets. Aposymbiotic aphids, on both plant species, showed increased pathway activities, mechanical problems with the stylets, and delayed salivation in the phloem. The extended time in derailed stylet mechanics affected the occurrence of most other probing activities; it delayed the time to the first phloem phase and prevented ingestion from the phloem. The effect of aposymbiosis was more evident in the compatible interaction of M. persicae–S. tuberosum, than in the incompatible interaction with S. stoloniferum, which generated the M. persicae–S. tuberosum interaction to become incompatible. These results confirm that B. aphidicola is involved in the plant–aphid interaction in relation to plant acceptance, presumably through a role in stylets penetration in the plant.     

Identifying the abiotic and biotic drivers behind the elevational distribution shift of a parasitic plant

• Climate change will alter the biotic and abiotic environment and dissipate ecological barriers, reorganising maps of current distribution of parasites and their hosts. In this study, we analyse the population dynamics of the parasitic plant Viscum album subsp. austriacum and explore key biotic (host availability and seed dispersal) as well as abiotic (temperature) factors influencing elevational distribution.
• The study was conducted along an elevational gradient of a Mediterranean mountain, covering the distribution belts of three potential pine hosts: Pinus halepensis (1300–1500 m), P. nigra (1300–1900 m) and P. sylvestris var. nevadensis (1600–2000 m). Along this gradient, we measured multiple variables of mistletoe population (prevalence, abundance and demographic profile) and different factors that might define the current mistletoe distribution (host suitability and availability, temperature and seed dispersal services).
• We found a decline in mistletoe prevalence and abundance with increasing elevation, detecting larger values of both variables at lower elevations of the most suitable host (Pinus nigra). Pinus sylvestris var. nevadensis was a suboptimal but suitable host for the parasite at high elevations. Mistletoe found suitable temperatures and seed dispersal services all along the gradient, being able to recruit at any site.
• With warming temperatures, the presence of suitable vectors for parasite dispersion, and the presence of a sub‐optimal host (P. sylvestris var. nevadensis) at the mountain top, mistletoe currently has a window of opportunity to expand its present geographic distribution to the summits.

     

Temperature checks the Red Queen? Resistance and virulence in a fluctuating environment

Numerous studies have revealed genetic variation in resistance and susceptibility in host–parasite interactions and therefore the potential for frequency‐dependent selection (Red Queen dynamics). Few studies, if any, have considered the abiotic environment as a mediating factor in these interactions. Using the pea aphid, Acyrthosiphon pisum, and its fungal pathogen, Erynia neoaphidis, as a model host–parasite system, we demonstrate how temperature can mediate the expression of genotypic variation for susceptibility and virulence. Whilst previous studies have revealed among‐clone variation in aphid resistance to this pathogen, we show that resistance rankings derived from assessments at one temperature, are not conserved across differing temperature regimes. We suggest that variation in environmental temperature, through its nonlinear impact on parasite virulence and host defence, may contribute to the general lack of evidence for frequency‐dependent selection in field systems.     

Effects of abiotic factors and biocontrol agents on chlamydospore formation in Fusarium graminearum and Fusarium sporotrichioides

Chlamydospores are vital asexual resting cells, which allow most of the Fusarium pathogenic strains to retain their longevity, thus ensuring survival of viable reproductive cells. This study suggested that both abiotic – extreme temperature and growth media, and biotic – antagonistic Bacillus amyloliquefaciens SMCD 518 and mycoparasititic Acremonium strictum SMCD 504 are natural stressors able to shift chlamydospores formation in Fusarium graminearum and F. sporotrichioides under in vitro conditions. In F. sporotrichioides, Minimal Conversion Media (MCM) with mannitol supplement induced high chlamydospore size, and chain abundance at optimal 21°C and extreme 37°C temperatures, respectively. F. graminearum showed low chlamydospore formation on MCM–mannitol, even when exposed to 37°C under prolonged 5 days incubation. Generally, F. sporotrichioides has higher chlamydospore abundance, longer chlamydospore chain, and production rapidity compared to F. graminearum in both abiotic and biotic treatments. However, biocontrol bacteria and mycoparasite posed minimal effects on chlamydospore formation, as compared to abiotic stressors, thus controlling the Fusaria but not triggering them to generate chlamydospores as protection shields.     

Factors affecting the relative abundance of two coexisting aphid species on sugar beet

1 The two most common species of aphid colonizing sugar beet Beta vulgaris L. are Myzus persicae (Sulzer) (Hemiptera: Aphididae) and Aphis fabae Scopoli (Hemiptera: Aphididae). 2 M. persicae colonizes sugar beet earlier than A. fabae but the population of the former also declines earlier. Despite similar numbers of each species migrating at the time of colonization, M. persicae is usually less abundant on the crop than A. fabae, suggesting differences between the species in their selection of, and performance on, sugar beet. 3 The intrinsic rate of increase of both species declines as sugar beet matures, however, at any given plant age the intrinsic rate of increase of A. fabae is one and a half times greater than that of M. persicae. This results in more rapid population growth and a later decline of the population. 4 Intraspecific competition appears to result in M. persicae becoming very restless, but there is no evidence for interspecific competition between the two species on this host. 5 A population growth model which takes account of the decline in host quality of sugar beet shows that the M. persicae population peaks 30 days before that of A. fabae, and, excluding differences in emigration rate, the maximum A. fabae population is 14 times greater than the maximum M. persicae population. These results are compared to field data.     

OBSERVATIONS ON CHYTRIDIACEOUS PARASITES OF PHANEROGAMS. XX. RESTING SPORE GERMINATION AND EPIBIOTIC STAGE OF PHYSODERMA BUTOMI SCHROETER

The development and morphology of Physoderma butomi Schroeter on Butomus umbellalus L., found in Michigan, is followed from germination of the resting spore through formation of epibiotic and endobiotic stages, to resting spore. No evidence for sexuality was found.