Regional-scale impact of the weed biocontrol agent Mecinus janthinus on Dalmatian toadflax (Linaria dalmatica)

Dalmatian toadflax (Linaria dalmatica (L.) Mill.) is an important invasive plant on rangelands throughout western North America. In 1991, the stem-mining weevil, Mecinus janthinus Germar, was introduced into Canada from Europe as a classical biological control agent to reduce toadflax densities and improve rangelands, particularly in British Columbia. To determine if the program was a success at a regional level, this paper answers three key questions: (1) has M. janthinus spread throughout the study area, (2) is M. janthinus causing a decline in toadflax plant size or density at the regional scale, and (3) has the distribution of toadflax plants changed following M. janthinus introduction? These questions are answered by combining historical survey data and mensurative experimental data on plant and weevil densities across a 40,000 km² area in southern British Columbia. The results show that through a combination of intentional redistribution and natural dispersal weevils have spread throughout the study area. Stem densities at naturally colonized sites and historical release sites were equally low. Across weevil populations between 3 and 10 years old, weevil densities peaked in the eighth year, and there was a negative relationship between weevil density and stem length. Between 2000 and 2007, toadflax patches were found to both decrease in density and become more fragmented over time, with 15% of patches disappearing completely. These findings show that M. janthinus has had a significant negative impact on both the density and distribution of Dalmatian toadflax throughout a large part of its range in British Columbia.     

Microscopic observations of a compatible host/pathogen interaction between a potential biocontrol agent (Uromyces pencanus) and its target weed (Nassella neesiana)

Uromyces pencanus is a prospective biocontrol agent for Nassella neesiana in Australia and New Zealand. The infection process of U. pencanus urediniospores in leaves of its susceptible host N. neesiana was found to be similar to that of most other rust species. A pattern against which to compare nonhost reactions in host range experiments was achieved using a modification of Bruzzese and Hasan’s whole leaf clearing and staining technique. The staining of microtome leaf sections provided supplemental information about this pathosystem.     

Effects of gall damage by the introduced biocontrol agent Epiblema strenuana (Lep., Tortricidae) on the weed Parthenium hysterophorus (Asteraceae)

Effects of gall damage by the introduced moth Epiblema strenuana on different growth stages of the weed Parthenium hysterophorus was evaluated in a field cage using potted plants with no competition and in naturally regenerated populations with intraspecific competition. Gall damage at early stages of plant growth reduced the plant height, main stem height, flower production, leaf production, and shoot and root biomass. All galled, potted plants with no competition produced flowers irrespective of the growth stage at which the plants were affected by galling, but lesser than in ungalled plants. Gall induction during early growth stages in field plants experiencing competition prevented 30% of the plants reaching flowering. However, 6% of the field plants escaped from gall damage, as their main stems were less vigorous to sustain the development of galls. Flower production per unit total plant biomass was lower in galled plants than in ungalled plants, and the reduction was more intense when gall damage was initiated at early stages of plant growth. In potted plants with no competition, the number of galls increased with the plant vigour, as the gall insects preferred more vigorous plants. But in field plants there were no relationship between gall abundance and plant vigour, as intraspecific competition enhanced the negative effects of galling by reducing the vigour of the weed.     

Control of Colletotrichum coccodes on Tomato by Grafting and Soil Amendments

Eight trials were carried out in 2011 and 2012 in Northern Italy to evaluate the efficacy of grafting, compost and biofumigation with Brassica carinata against Colletotrichum coccodes on tomato. Four trials were carried out in commercial farms, and four trials were carried out in plastic tunnels at an experimental centre. The rootstocks ‘Armstrong’, ‘Arnold’, ‘Beaufort’, ‘Big Power’, ‘Brigeor’, ‘Emperador’, ‘King Kong’, ‘Spirit’ and ‘Superpro V295’ were tested. Host plants included several tomato F1 hybrids: ‘Amantino’, ‘Arawak’, ‘CLX 37438’, ‘Cauralina’, ‘CU 8301’, ‘CU 8506’, ‘DRK 7021’, ‘E 34431’, ‘E 50070’, ‘EXP’, ‘Gotico’, ‘Ingrid’, ‘ISI 61401’, ‘ISI 61402’, ‘Profitto’, ‘Punente’, ‘Rugantino’ and ‘Tomahawk’. Tomato roots from the control plots were 34 to 87% diseased in both naturally and artificially infested soil. Among the nineteen commercial tomato hybrids tested, in the presence of a very high disease pressure in a naturally infested soil, ‘Rugantino’ was the least affected by C. coccodes, showing 32% infected roots. ‘Tomahawk’ grafted onto ‘Arnold’, ‘Armstrong’ and ‘Superpro V295’ was significantly less affected by C. coccodes, while ‘Arawak’ grafted onto ‘Armstrong’, ‘Arnold’, ‘Emperador’ and ‘Beaufort’ provided very good control of root rot in the different trials. Compost addition and biofumigation with Brassica pellets were also tested with and without grafting. Soil amendment with compost, in the case of the ‘Arawak’ and ‘Tomahawk’, resulted in a slightly improved disease control only on non‐grafted plants. When grafting and biofumigation were combined in a soil naturally infested with C. coccodes and Meloidogyne arenaria, biofumigation did not improve C. coccodes control in comparison with grafting alone. In a naturally infested soil, compost alone and combined with biofumigation improved disease control only on non‐grafted ‘Tomahawk’ plants. In general, grafting by itself provided very good results in terms of disease control, which were not significantly improved by combination with compost and/or biofumigation.     

Pectinase and cellulase enhance the control of Abutilon theophrasti by Colletotrichum coccodes

Inundative mycoherbicidal biocontrol agents are typically insufficiently virulent to be commercially competitive with herbicides in row crop agriculture, and require enhancement. Pectinase and cellulase are typically used by pathogens during infection. Thus, it was hypothesized that adding exogenous cell wall degrading enzymes might enhance fungal infection. Pectinase or cellulase was added to inocula of aqueous chopped mycelial suspensions of a strain of Colletotrichum coccodes for control of Abutilon theophrasti. Plants treated with 5.3×10⁶ C. coccodes propagules mL^-1 and 1.65 U mL^-1 pectinase had more rapid and complete disease development. Similar trend was achieved when 10 U mL^-1 of cellulase were added to 2.2×10⁶ C. coccodes propagules mL^-1. Adding pectinase or cellulase did not increase the host range of the wild-type fungus. The results suggest that there might be value to transforming biocontrol agents to overproduce these enzymes.     

Real-time PCR for detection and quantification of the biocontrol agent Trichoderma atroviride strain SC1 in soil

Trichoderma (Hypocreales, Ascomycota) is a widespread genus in nature and several Trichoderma species are used in industrial processes and as biocontrol agents against crop diseases. It is very important that the persistence and spread of microorganisms released on purpose into the environment are accurately monitored. Real-time PCR methods for genus/species/strain identification of microorganisms are currently being developed to overcome the difficulties of classical microbiological and enzymatic methods for monitoring these populations. The aim of the present study was to develop and validate a specific real-time PCR-based method for detecting Trichoderma atroviride SC1 in soil. We developed a primer and TaqMan probe set constructed on base mutations in an endochitinase gene. This tool is highly specific for the detection and quantification of the SC1 strain. The limits of detection and quantification calculated from the relative standard deviation were 6000 and 20,000 haploid genome copies per gram of soil. Together with the low throughput time associated with this procedure, which allows the evaluation of many soil samples within a short time period, these results suggest that this method could be successfully used to trace the fate of T. atroviride SC1 applied as an open-field biocontrol agent.     

Detection and differentiation of Colletotrichum gloeosporioides isolates using PCR

An oligonucleotide primer (CgInt), synthesised from the variable internally transcribed spacer (ITS) 1 region of ribosomal DNA (rDNA) of Collectotrichum gloeosporioides was used for PCR with primer ITS4 (from a conserved sequence of the rDNA) to amplify a 450-bp fragment from the 25 C. gloeosporioides isolates tested. This specific fragment was amplified from as little as 10 fg of fungal DNA. A similar sized fragment was amplified from DNA extracted from C. gloeosporioides-infected tomato tissue. RAPD analysis divided 39 C. gloeosporioides isolates into more than 12 groups linked to host source and geographic origin. Based on the results obtained, the potential of PCR for detection and differentiation of C. gloeosporioides is discussed.     

Free fatty acids from the weed,Polygonum orientale leaves for attraction of the potential biocontrol agent,Galerucella placida (Coleoptera: Chrysomelidae)

Extraction, thin layer chromatography and gas chromatography-mass spectrometry of leaf surface waxes of Polygonum orientale L. (Polygonaceae) weed revealed 11, 15 and 11 free fatty acids in young, mature and senescent stages. Oleic acid was the predominant in young leaves (5950 ± 111 µg); whereas palmitic acid was the predominant fatty acids, representing 4247.5 ± 23 and 6644 ± 110 µg in mature and senescent leaves, respectively. Both tridecanoic and heneicosanoic acids were not detected in young and senescent leaves, and myristic and heptadecanoic acids were not identified in young leaves; whereas lauric and nonadecanoic acids were not detected in senescent leaves. The free fatty acids from young, mature and senescent weed leaves, and the mixtures of synthetic fatty acids mimicking free fatty acids of three types of weed leaves attracted female Galerucella placida (Coleoptera: Chrysomelidae) at the minimal amounts of 2, 1 and 2 leaf equivalent free fatty acids, respectively, in Y-shaped glass tube olfactometer bioassays under laboratory conditions. Individual synthetic pentadecanoic, palmitoleic, stearic, nonadecanoic and docosanoic acids at 44.82, 9.91, 92.22, 18.33 and 15.88 µg, respectively, elicited attraction of the insect. A synthetic blend of 3.59, 7.89, 44.82, 9.91, 32.31, 18.33 and 15.88 µg of lauric, myristic, pentadecanoic, palmitoleic, heptadecanoic, nonadecanoic and docosanoic acids, respectively, indicated highest attraction of the insect.     

Attraction of the biocontrol agent,Lema praeusta,towards two Commelinaceae weed volatiles

Two Commelinaceae weeds, Commelina benghalensis L. and Murdannia nudiflora (L.) Brenan, are abundant in rice fields of India. Larvae and adults of Lema praeusta (Fab.) (Coleoptera: Chrysomelidae) voraciously consume these two weeds. Synthetic herbicides are applied to control both weeds, but applications of these substances have harmful effects in environment and beneficial organisms. So, it is necessary to use native biocontrol agent to control these weeds. Hence, an attempt has been made to find volatile organic compounds (VOCs) from both weeds causing attraction of L. praeusta. Behavioural responses of L. praeusta towards volatile blends from undamaged (UD), insect-damaged (ID: plants after 6 or 48 hr of continuous insect feeding) and mechanically damaged (MD) plants were conducted by Y-tube olfactometer bioassays. Benzyl alcohol was predominant in VOCs of UD plants and M. nudiflora after 48 hr of insect feeding. Benzyl alcohol and cuminaldehyde were both predominant in VOCs of C. benghalensis after 48 hr of insect feeding. Females were more attracted towards volatile blends from plants after 48 hr of insect feeding compared to undamaged plants. Females showed attraction towards a synthetic blend of seven compounds—7.28 µg (Z)-3-hexen-1-ol, 0.93 µg trans-isolimonene, 19.18 µg benzyl alcohol, 0.16 µg undecane, 1.07 µg 1-nonanol, 1.23 µg 1-undecanol and 0.47 µg 1-eicosene resembling the amounts released by C. benghalensis after 48 hr of insect feeding during 1 hr or a synthetic blend of six compounds—18.10 µg benzyl alcohol, 0.25 µg undecane, 0.56 µg 1-nonanol, 1.37 µg 1-undecanol, 0.35 µg 1-eicosene and 2.19 µg phytol resembling the amounts released by M. nudiflora after 48 hr of insect feeding during 1 hr. This study concludes that both blends could be used to attract the biocontrol agent during early vegetative period of these two weeds, which could lead to eradication of weeds from rice fields.     

The susceptibility of tubers of potato cultivars to black dot (Colletotrichum coccodes (Walk.) Hughes)

In experiments with commercial seed of different cultivars at Rothamsted and Woburn, Bedfordshire in 1985 – 88 the severity of black dot on daughter tubers at harvest differed between cultivars. The disease was most severe on Desiree tubers. Amounts of disease were similar at both sites in 1986 – 88 but in 1985 it was more severe at Woburn than at Rothamsted. Disease-free seed of 12 (1987) or 15 (1988) cultivars were planted in experiments at Rothamsted (inoculated with Colletotrichum coccodes or not) and at Mepal, Cambridgeshire (not inoculated) and black dot assessed at harvest in October 1987 and in September and October 1988. There were significant differences in the amount of disease on different cultivars and the order of severity was similar at the two sites, on the two harvest dates in 1988 and in both years. Desiree, Maris Piper, Maris Peer and Record were amongst those cultivars severely affected whereas Cara, Pentland Crown and Romano were least affected. Skin discoloration caused by black dot was more noticeable on white-skinned than red-skinned cultivars and was severe on the Dutch cultivars Estima, Marfona, Santé and Wilja.     

Detection of the low-germination-rate resting oospores of Pythium myriotylum from soil by PCR

AIMS: To establish a sensitive and specific polymerase chain reaction (PCR)-based method for detecting Pythium myriotylum in soils. METHODS AND RESULTS: Oospores of P. myriotylum were separated from large soil particles by flotation in sucrose solution. The thick-walled oospores were disrupted by vortex with sea sand and its DNA was extracted by the Cetyl trimethyl Ammonium Bromide (CTAB) method. The recovered DNA was verified by PCR amplification of a 150-bp target sequence of P. myriotylum. Samples of 10 g of soil were assayed; thus, the detection limit by PCR-based method was 10 oospores per gram soil. The method was successfully applied for the detection of P. myriotylum in soils collected in March, prior to planting of ginger crops. CONCLUSIONS: A PCR-based method for detecting P. myriotylum from soil was achieved. SIGNIFICANCE AND IMPACT OF THE STUDY: The PCR method has allowed us to monitor the presence of P. myriotylum in soil prior planting season as a way of reducing or eliminating disease.     

Development of black dot disease (Colletotrichum coccodes (Wallr.) Hughes) and its effects on the growth and yield of potato plants

Seed tubers of cvs Désirée and Pentland Crown with different severities of black dot were planted in 1988 and 1989 at Rothamsted in fields in 4– or 7-course rotations, respectively. Tubers treated with prochloraz (1988) or imazalil (1989) were planted in some plots, and in others Colletotrichum coccodes inoculum was added to the soil at planting. In further experiments at Mepal, Cambridgeshire in 1989 and 1990 and at Rothamsted in 1990 on sites where potatoes had not been grown for more than 15 years, large amounts of inoculum were added to the soil around disease-free seed tubers of two (1989) or three (1990) cultivars at planting. In all experiments plants were sampled during the season and the effects of treatments on disease development, growth and yield were recorded. Disease on roots, stem bases and tubers was found early in the season and was more severe on Désirée than on Pentland Crown plants from fields in 4– or 7-course rotations. Severity increased throughout the season and with increasing amounts of disease on the seed tubers, especially with Desiree. Disease was also found on plants from disease-free tubers and was more severe in 1988 than 1989. At harvest, black dot on tubers was significantly more severe from severely affected than from disease-free seed, and was most severe where inoculum, especially large amounts, had been added at planting. Fungicide treatment decreased disease early in the season but had no effect on tuber infection at harvest. In 1989 the weight loss of seed tubers during sprouting increased with increasing amounts of black dot, but the disease had little effect on plant size through the season. At harvest the yield of ware tubers (>50 mm) decreased with severe disease but total tuber yields were not significantly affected. However, at harvest in 1988 severely affected seed yielded significantly less than healthy seed. Plants grown from mini-tubers were free from disease on sites where potatoes had not been grown for at least 15 years. Inoculum applied at planting caused severe disease on all cultivars in both years, whereas disease was slight on uninoculated plants. Inoculated plants senesced early at Mepal in 1990, but there were no significant differences in total tuber yield in any experiment. However, yields of ware tubers (>50 mm) were sometimes decreased and the total tuber number per plant increased.     

Biological performance of Diclidophlebia smithi (Hemiptera: Psyllidae), a potential biocontrol agent for the invasive weed Miconia calvescens

Diclidophlebia smithi Burckhardt, Morais and Picanço (Hemiptera: Psyllidae) is a promising biological control agent of Miconia calvescens DC. (Melastomataceae), a neotropical invasive weed in forest ecosystems in French Polynesia and Hawaii and a threat in Australia, where it was also introduced. A study on the reproductive performance of D. smithi under laboratory conditions through life expectancy and fertility tables is presented. Results indicated that this psyllid has a high reproductive capacity (R ₀>1 and r _m >0) and a short life cycle (46–47 days) and can have up to nine generations per year. The critical period of its life cycle is during the nymphal stage which is clearly inadequate for field introductions. The best age for introducing D. smithi against M. calvescens is the 4th day of the adult stage. D. smithi is easily mass-reared and has a short life cycle and a high reproductive capacity, which are desirable characteristics for a biological control agent.     

Comparison of the effect of antibiotic substances on sclerotia of Mycosphaerella ligulicola and Colletotrichum coccodes

Summary Sclerotia ofColletotrichum coccodes tolerated much higher concentrations of actidione in agar than did sclerotia ofMycosphaerella ligulicola. With increase in concentration of the antibiotic sclerotia of both species took longer to germinate. Increased resistance of both species to actidione developed after growth of a single generation on media containing the antibiotic. Sclerotia ofC. coccodes survived 5 days immersion in a bacterial culture filtrate whereas scleroia ofM. ligulicola ceased to be viable after a similar period.Sclerotia ofC. coccodes andM. ligulicola exhibited strand and loose types of formation respectively. The degree of resistance of these sclerotia to antibiotic substances was correlated with both longevity in soil and type of formation, but, in general, there is unlikely to be a relationship between structure of the sclerotium and longevity.     

Detection and quantification of phnE gene from oil-contaminated soil samples by competitive quantitative PCR

The phnE gene encoding catechol 2,3-dioxygenase belonging to the meta-cleavage pathway was selected as the marker gene and was detected and quantified from soil samples by competitive quantitative PCR. A PCR primer pair was designed based on the phnE gene to amplify the target DNA bands and competitor DNA bands. The phnE gene was detected in two samples of three. In samples S1 and S2, the phnE gene copy number was 6.2×10⁷/g soil and 5.8×10⁷/g soil, respectively. But no phnE gene was detected in sample S3. The target DNA bands were extracted and expressed. The results confirmed that the target DNA bands were the native phnE genes.     

Secondary metabolites from species of the biocontrol agent Trichoderma

Trichoderma species are free-living fungi that are highly interactive in root, soil and foliar environments and have been used successfully in field trials to control many crop pathogens. Structural and biological studies of the metabolites isolated from Trichoderma species are reviewed. This review, encompassing all the literature in this field up to the present and in which 269 references are cited, also includes a detailed study of the biological activity of the metabolites, especially the role of these metabolites in biological control mechanisms. Some aspects of the biosynthesis of these metabolites and related compounds are likewise discussed.     

Impact of defoliation by the biocontrol agentZygogramma bicolorataon the weed Partheniumhysterophorus in Australia

The leaf-feeding beetle Zygogrammabicolorata Pallister was introduced from Mexico intoAustralia in 1980 as a biocontrol agent for the weedParthenium hysterophorus L. (Asteraceae). Z. bicolorata became abundant in 1990, and since 1992there has been regular outbreaks resulting in thedefoliation of the weed in central Queensland. In thisstudy we evaluated the impact of defoliation by Z. bicolorata on P. hysterophorus from 1996 to1998. Z. bicolorata caused 91–100% defoliationresulting in reductions in weed density by 32–93%,plant height by 18–65%, plant biomass by 55–89%,flower production by 75–100%, soil seed-bank by13–86% and seedling emergence in the following seasonby 73–90%. At sites with continued outbreaks ofZ. bicolorata, it is expected that the existing soilseed-bank will be minimised, resulting in reduceddensity of parthenium in 6 to 7 years.