Studies on resistance to dichlofluanid and other fungicides in Botrytis cinerea

Amongst 489 mass isolates of Botrytis cinerea obtained from protected crops in Humberside during 1989, 67.3% were resistant to benomyl and 45.6% to vinclozolin. Two isolates resistant to dichlofluanid were obtained with EC₅₀ values for mycelial growth on agar of 21.45 and 19.78 μg dichlofluanid/ml, respectively, compared with values of 1.55-2.56 μg/ml for sensitive isolates. EC₅₀ values for germ tube emergence and length were much lower than for mycelial growth. In population studies carried out on tomatoes in a polythene tunnel, resistance to dichlofluanid and to vinclozolin declined over a 12-week period in the absence of fungicides. Field isolates of B. cinerea resistant to dichlofluanid were also resistant to vinclozolin. but strains resistant to dichlofluanid and sensitive to vinclozolin were obtained during population studies. Examination of a number of features of dichlofluanid-resistant strains revealed a degree of lack of fitness as compared with sensitive strains. There is no indication that resistance to dichlofluanid poses a great threat to the control of grey mould in Humberside at present.     

Dicarboximide resistance in Botrytis cinerea in protected lettuce

In surveys of protected lettuce crops in the Humberside area of the UK during 1984-5, 47.3% of Botrytis cinerea isolates were resistant to dicarboximides and 25% to benomyl. Dicarboximide-resistant isolates obtained direct from the field had a moderate degree of resistance with ED₅₀ values for mycelial growth on agar of 1.0-2.4 μg/ml vinclozolin for all except two of 30 isolates.
Isolates which were resistant to vinclozolin were also resistant to iprodione, myclozolin and procymidone. Thirty dicarboximide-resistant isolates obtained direct from the field were capable of infecting cucumber cotyledons sprayed with field-strength Ronilan (250 μg/ml vinclozolin) whereas sensitive strains were not. Strains showing different degrees of resistance to dicarboximides developed after incubation of sensitive and resistant strains on agar supplemented with vinclozolin. Some of these strains had ED₅₀ values for mycelial growth on agar of more than 100 μg/ml vinclozolin. No highly resistant isolates were obtained direct from the field. Resistance to dicarboximides in field isolates was stable.     

Sensitivity of South African Ramulispora herpotrichoides isolates to carbendazim and ergosterol biosynthesis inhibitors

In a survey conducted during 1991–1992, single-spored isolates of the eyespot fungus from the Swartland area were characterized and tested for sensitivity to carbendazim and ergosterol inhibiting fungicides. The 100 isolates tested were all fast growing, even marginate, and designated as Ramulispora herpotrichoides . Fungal growth was completely inhibit on PDA amended with carbendazim (1 μg/ml), indicating that the local population of the fungus is still at baseline sensitivity to benzimidazoles. The mean concentration of prochloraz calculated to inhibited growth by 50% (IC₅₀ value) was 0.043 ± 0.029 μg/ml, which is comparable with the baseline sensitivity reported for European isolates. Of the 36 representative isolates screened against 2 μg/ml triadimenol, 44% were sensitive, while 36% were resistant. The triadimenol-resistant isolates were sensitive to propiconazole and flusilazole. However, four of the triadimenol-resistant isolates were also resistant to tebuconazole. These results indicate that South African isolates of R. herpotrichoides are sensitive towards carbendazim, prochloraz, propiconazole and flusilazole. They were found to differ, however, in sensitivity towards triadimenol and tebuconazole, where some isolates had an IC₅₀ value greater than 2 μg/ml.     

Multiple fungicide resistance to benzimidazoles, dicarboximides and diethofencarb in field isolates of Botrytis cinerea in Israel

During the period January-March 1989, 15 greenhouses at 12 sites in Israel were surveyed for the presence of fungicide-resistant strains of Botrytis cinerea , using a fungicide-amended Botrytis -selective medium. Resistance to benzimidazoles (Ben^R) and to dicarboximides (Dic^R) was frequent in most sites. Resistance to carbendazim + diethofencarb (Ben^R NPC^R) was found in all eight sites in which a mixture of these fungicides had been used against grey mould, but not in other sites. A new phenotype of multiple fungicide resistance was found among these isolates. The new phenotype, designated Ben^R Dic^R NPC^R, combines the three previously described characteristics of resistance to benzimidazole, dicarboximide and N -phenylcarbamate fungicides. It was found only in cucumber greenhouses that had been sprayed with the fungicide mixture carbendazim + diethofencarb against grey mould. Isolates of this phenotype were pathogenic in artificial inoculation of cucumber cotyledons treated with carbendazim, iprodione or carbendazim + diethofencarb.     

Incidence of benomyl and iprodione resistance in isolates of Botrytis cinerea in tomato crops in England and Wales in 1986

In 1986, mass-mycelial isolates of Botrytis cinerea from 67 tomato crops in England and Wales were examined for benomyl and iprodione resistance. Of the 706 isolates obtained, 62.7% were resistant to benomyl at 2μg/ml and 43.2% were resistant to iprodione at 2 μg/ml. Iprodione resistance persisted in the absence of a dicarboximide spray programme. The incidence of benomyl resistance has not decreased since the last survey in 1984 in spite of a considerable reduction in the use of benzimidazole fungicides. There were no clear indications that the use of dichlofluanid influenced the incidence of benomyl or iprodione resistance. Disease control was poorer in crops with a higher incidence of iprodione resistance.     

Resistance to diethofencarb (NPC) in benomyl-resistant field isolates of Botrytis cinerea

Based on the 'negative cross-resistance' phenomenon between N -phenylcarbamates and benzimidazole fungicides, the fungicidal mixture of carbendazim + diethofencarb (MBC + NPC) was introduced, to control Botrytis cinerea phenotypes which are either sensitive to benzimidazoles and resistant to NPC (wild-type: Ben^S NPC^R). or resistant to benzimidazoles and sensitive to NPC(Ben^R NPC^S). At one out of four sites where the MBC + NPC mixture was used in commercial cucumber greenhouses, grey mould control failed and a new phenotype of B. cinerea was found. The new phenotype was resistant to benzimidazoles, as was 100% of the population screened in the four sites, but retained resistance to NPC. Accordingly, the new phenotype was designated Ben^R NPC^R. It was pathogenic on cucumber seedlings.     

Carbendazim resistance in the eyespot pathogen Pseudocercosporella herpotrichoides

Mycelial isolates (115) of Pseudocercosporella herpotrichoides were obtained from five field sites in England. Carbendazim-resistant isolates were detected by their mycelial growth on agar containing 1 μg/ml carbendazim. Resistant isolates were found at two of the five sites examined and one of these had never been treated with benzimidazole fungicides. Amongst the carbendazim- resistant isolates there was a predominance of isolates with pale mycelium, an irregular colony margin and a relatively slow growth rate; however, this association was not absolute. Large differences in the effects of carbendazim on mycelial growth of sensitive and resistant isolates were demonstrated; growth of sensitive isolates was completely inhibited at 0.5 μg /ml carbendazim whilst five of the six resistant isolates examined grew on agar containing 1000 μg/ml fungicide. The carbendazim-resistant isolates were cross-resistant to benomyl, thiophanate-methyl and to a Icsser degree thiabendazole, but not to prochloraz. Conidia of carbendazim-resistant isolates were as resistant. Carbendazim-resistant isolates were just as pathogenic to wheat as sensitive isolates. The implications of these results and other reports of benzimidazole resistance in P. herpotrichoides are discussed in relation to disease control.     

Biology and control of Polystigma ochraceum, the cause of almond red leaf blotch

Polystigma ochraceum is a major leaf pathogen of almond in Fars Province of Iran. Over a 4-year study period it was found that ascospore discharge began at flowering and continued for 4–5 weeks. The maximum discharge occurred at petal fall. The incubation period was estimated to be 4–5 weeks under experimental conditions. Although the mature ascospores could produce short germ tubes in distilled water or water agar, the fungus could not be cultured or grown on conventional media from either ascospores, pycnidiospores or stromatic tissues under laboratory conditions.
Of several systemic and non-systemic fungicides evaluated under field conditions, triforine at 100–400 μ/ml was most effective. Other fungicides which significantly reduced leaf infection were, in order of efficacy, copper oxychloride (2000 μg/ml), copper hydroxide (2000 mUg/ml), Bordeaux mixture (10 000 μg/ ml) and mancozeb (2000 μg/ml). Carbendazim and thiophanate methyl (500 μg/ml) increased the level of infection. One application of the fungicide at petal fall and then two at 14-day intervals were found to be effective in reducing the disease.     

Studies on metalaxyl-resistant Phytophthora infestans in potato crops in south-west Scotland

In 1981 metalaxyl-resistant Phytophthora infestans was found in 44% of the 63 crops examined in south-west Scotland. In 1982, when fungicides containing acylalanines (metalaxyl or ofurace) were not used in the area, it was detected in 15%. Metalaxyl-resistant strains were found in several crops which had been sprayed only with non-acylalanine fungicides or had received no fungicidal treatment. In neither year was resistance associated with a significant breakdown of disease control.
Metalaxyl-resistant isolates of the blight fungus were obtained from tubers from crops in which resistance was detected on the foliage.
In detached leaf experiments, resistant isolates retained their ability to grow on leaves treated with 100 μg/ml of metalaxyl after 52 weekly transfers on untreated leaves, and produced sporangia on leaves treated with up to 2000 μg/ml of metalaxyl or ofurace. In mixtures with sensitive sporangia the proportions remained stable after nine weekly transfers on untreated leaves. However, at concentrations as low as 50 μg/ml, metalaxyl and ofurace reduced the extent of colonization and the numbers of sporangia produced by resistant isolates by an average of 50%.     

The occurrence, characteristics and distribution in the United Kingdom of resistance to phenylamide fungicides in Bremia lactucae (lettuce downy mildew)

Failure to control Bremia lactucae (lettuce downy mildew) with metalaxyl in an intensive lettuce-producing region of Lancashire at the end of 1983 was shown to be due to the occurrence of a high level of resistance to this fungicide (isolates capable of growth at < 100 μg/ml metalaxyl). During most of 1984, metalaxyl-resistant isolates were obtained from numerous sites but all within a 20-km radius of the initial outbreak. Thereafter, at the end of 1984 and during 1985, metalaxyl-resistant isolates were recovered from most major lettuce-producing regions in the UK with protected crops more affected than field crops. AH metalaxyl-resistant isolates tested were identical in their response to fungicide, sexual compatibility type (B2) and virulence phenotype, probably representing a clone from a single origin. The resistant pathotype was virulent on resistance factors R 1-10 and 12-15 but lacked virulence for R 11 and 16-18. This was also the most common virulence phenotype among sensitive isolates collected at the same time. Cross-resistance to other phenylamide fungicides was demonstrated but isolates resistant and sensitive to phenylamide showed a similar response to the unrelated systemic fungicides propamocarb and fosetyl-Al. An F₁ sexual progeny isolate from a cross between a phenylamide-sensitive and a phenylamide-resistant isolate (presumed heterozygous at the locus or loci regulating response to phenylamide fungicides) exhibited an intermediate response to phenylamide fungicides. No isolates of this type were obtained from the field. At the high concentrations affecting spore germination, phenylamide fungicides exhibited lower activity against a resistant isolate compared with a sensitive isolate. The findings are discussed in relation to future control strategies, the population biology of the fungus and possible directions for lettuce breeding programmes.     

Phytotoxicity of destruxin B and its possible role in the pathogenesis of Alternaria brassicae

Destruxin B, the major phytotoxin produced by Allernaria brassicae , was not host-specific on 30 different plant species, causing necrotic and chlorotic symptoms both on host and non-host plants. Isolation of sufficient amounts of two minor destruxins, homodestruxin B and destruxin B₂, made it possible for the first time to show that they were both phytotoxic to leaves of Brassica napus. There were significant differences between taxonomic plant groups in their sensitivity to destruxin B. Brassica species were most sensitive to the toxin, and sensitivity decreased as relatedness of plant groups became more distant: thus it is suggested that destruxin B is host-selective in nature. The dilution-end-point of destruxin B was 0.2–3.8 μg/ml for the most sensitive host species, and 15 120 μg/ml for the least sensitive. The sensitivity of non-host species was between 15 and 750 μg/ml. The symptoms caused by destruxin B appear to be light-dependent. There were significant differences between taxonomic plant groups in their sensitivity to destruxin B. Brassica species were most sensitive to the toxin, and sensitivity decreased as relatedness of plant groups became more distant, thus it is suggested that destruxin B is host-selective in nature. Destruxin B appears to be a virulence factor, contributing to the aggressiveness of A. brassicae by conditioning the host tissue and thereby determining the susceptibility of the host.     

Negative cross resistance between benzimidazole and N-phenylcarbamate fungicides and control of Botrytis cinerea on grapes

Isolates of Botrytis cinerea resistant to benzimidazoles (Ben^R), dicarboximides (Dic^R), or both (Ben^R Dic^R) were found on table grapes in vineyards in Israel. In vineyards treated for one or two seasons with benomyl and dicarboximides, 41% of the isolates were benomyl-resistant, 18% were dicarboximide-resistant and 21 % were resistant to both fungicides. The frequency of resistant strains was very low in non-treated vineyards. Negatively correlated cross resistance (NCCR) was manifested by 46 Ben^R isolates which were sensitive to isopropyl N -(3,4-diethoxyphenyl) carbamate (NPC) and 73 benomyl-sensitive (Ben^s) isolates which were insensitive to NPC; NCCR was not influenced by sensitivity to dicarboximides. A mixture of Ben^s Dic^R plus Ben^R Dic^s isolates, inoculated on rose petals, was inhibited by mixtures of benzimidazoles plus NPC but not by benomyl, NPC or iprodione alone. Grey mould, incited on maturing grape berries by a vineyard population composed of Ben^s and Ben^R strains, was not controlled by benomyl, while the mixture of NPC plus carbendazim was effective.     

Sensitivity of Fusarium moniliforme Isolates to Ipconazole

To estimate the sensitivity of Fusarium moniliforme to ipconazole, a sterol biosynthesis inhibitor (SBI), minimum inhibitory concentrations (MIC) were determined for isolates which were collected before the launch of ipconazole as a rice seed disinfectant. Research institutes from various prefectures in Japan supplied 211 isolates (group I) from their collections, and 84 isolates (group II) were isolated from rice paddy fields in Iwaki, Fukushima Prefecture. In group I, the MIC ranged from 0.10 to 6.25 μg/ml with a peak at 0.39 μg/ml. In group II, MIC values had the same range as group I, but the main peak was at 0.20 μg/ml. Ipconazole sensitivity did not differ significantly among groups I and II. Though the ranges of MIC values for ipconazole, pefurazoate and triflumizole were different in 60 isolates randomly chosen from group I, positive correlations were observed in their sensitivities to SBIs, suggesting a common mechanism in F. moniliforme for lowering sensitivities to SBIs. Among the 14 isolates tested, isolates with MIC values lower than 0.78 μg/ml for ipconazole were pathogenic to rice seedlings, and all the isolates with MIC values higher than or equal to 1.56 μg/ml were not pathogenic in the nursery test. Good protection against isolates causing “Bakanae” disease was obtained by dipping seeds for 24 hr in ipconazole. The pathogenic isolates can be controlled by the seed treatment with the practical dosage of ipconazole because of the adequate margin between the highest MIC value for the pathogenic isolates and the treatment concentration. In addition, the low or lack of pathogenicity of the isolates less sensitive to ipconazole may also contribute to the stable efficacy of ipconazole. Received 8 November 1999/ Accepted in revised form 18 April 2000     

福建省番茄灰霉病菌对氟啶胺敏感基线建立及与不同杀菌剂的交互抗性

为明确中国福建省番茄灰霉病菌对氟啶胺的敏感性及其与不同杀菌剂的交互抗性,采用菌丝生长速率法测定了106株采自福建省主要番茄产区的番茄灰霉病菌对氟啶胺的敏感性。结果表明,氟啶胺对福建省番茄灰霉病菌菌丝生长的EC50值在0.0037～0.0452 μg/mL之间,平均值为（0.0221 ±0.0098）μg/mL,其敏感性频率分布呈连续单峰曲线,符合正态分布,因此可将该EC50平均值（0.0221±0.0098）μg/mL作为福建省番茄灰霉病菌对氟啶胺的敏感基线,用于其田间抗药性监测。从106株菌株中选择15株对氟啶胺敏感性不同的菌株,测定了其对嘧霉胺、异菌脲、腐霉利和啶氧菌酯的敏感性。结果表明,供试5种杀菌剂对15株番茄灰霉病菌菌丝生长的平均抑制活性依次为氟啶胺 >异菌脲 >腐霉利 >啶氧菌酯 >嘧霉胺,氟啶胺与异菌脲、腐霉利、啶氧菌酯和嘧霉胺之间均不存在交互抗性。     

Sensitivity of Sclerotinia homoeocarpa to demethylation-inhibiting fungicides in Ontario, Canada, after a decade of use

In late 2003, nine populations of Sclerotinia homoeocarpa in Ontario Canada (seven of which had been previously sampled in early 1994, prior to the registration of sterol demethylation-inhibiting (DMI) fungicides for turf disease control in Canada) were sampled and tested for sensitivity to propiconazole. Four of the nine populations had not been treated with DMI fungicides during the intervening years, and isolates from these locations were sensitive to propiconazole (geometric mean EC₅₀ values of 0·005–0·012 µ g mL⁻¹, compared with 0·005–0·008 µ g mL⁻¹ for the original 1994 populations). Among the five populations from 2003 that had been exposed to DMI fungicides, mean EC₅₀ values were significantly greater, ranging from 0·020 to 0·048 µ g mL⁻¹. A significant correlation of determination was found between estimated number of fungicide applications and log EC₅₀ ( R ² = 0·832, P  = 0·0001), and the equation predicted that 42·3 applications of propiconazole would be needed to bring a sensitive population (EC₅₀ < 0·01  µ g mL⁻¹) to a resistant level (EC₅₀ > 0·10  µ g mL⁻¹). Fungicide sensitivity vs. duration of fungicide efficacy was also tested, and it was found that isolates with decreased sensitivity were able to more quickly overcome the inhibitory effects of fungicide application, reducing the duration of control from 3 weeks to 2 weeks.     

河北省设施番茄灰霉病菌对啶酰菌胺和咯菌腈的敏感性

 为明确河北省设施番茄灰霉病菌对啶酰菌胺和咯菌腈的敏感性,2013-2016年,采用菌丝生长速率法测定了采自河北省八个地区的2 425株灰霉病菌对啶酰菌胺和咯菌腈的敏感性。2013-2016年监测数据显示,河北省设施番茄灰霉病菌对啶酰菌胺和咯菌腈的敏感性呈下降趋势。2016年从河北省不同地区采集的番茄灰霉病菌对啶酰菌胺具有相似的敏感性,且对啶酰菌胺产生了低水平的抗性,仅在唐山地区检测到了啶酰菌胺的高抗菌株;而不同地区采集的灰霉病菌对咯菌腈的敏感性具有差异,但均为咯菌腈的敏感菌株。因此,啶酰菌胺和咯菌腈仍可用于番茄灰霉病的防治,但应严格控制其使用频率,监测灰霉病菌对其敏感性变化动态。     

Occurrence of Corynespora cassiicola isolates resistant to boscalid on cucumber in Ibaraki Prefecture, Japan

A total of 651 isolates of cucumber corynespora leaf spot fungus ( Corynespora cassiicola ) collected from cucumber in Japan, either with (438 isolates) or without (213 isolates) a prior history of boscalid use, were tested for their sensitivity to boscalid by using a mycelial growth inhibition method on YBA agar medium. Additionally, seven isolates of C. cassiicola obtained from tomato, soybean, eggplant (aubergine) and cowpea in different locations in Japan were tested before boscalid registration. Minimum inhibitory concentration (MIC) and 50% effective concentration (EC₅₀) values for 220 isolates from crops without a prior history of boscalid use ranged from 0·5 to 7·5 μg mL⁻¹ and from 0·04 to 0·59 μg mL⁻¹, respectively. Two hundred and fourteen out of 438 isolates collected from ten cucumber greenhouses in Ibaraki Prefecture, Japan, which received boscalid spray applications showed boscalid resistance, with MIC values higher than 30 μg mL⁻¹. Moreover, resistant isolates were divided into two groups: a moderately resistant (MR) group consisting of 189 isolates with EC₅₀ values ranging from 1·1 to 6·3 μg mL⁻¹, and a very highly resistant (VHR) group consisting of 25 isolates with EC₅₀ values higher than 24·8 μg mL⁻¹. MR isolates were detected from all ten greenhouses, but VHR isolates were detected from only three. As a result of fungus inoculation tests which used potted cucumber plants, control failures of boscalid were observed against resistant isolates. Efficacy of boscalid was remarkably low against VHR isolates in particular. This is the first known report on boscalid resistance in Japan.     

两种蚕豆赤斑病菌对常用杀菌剂的抗药性检测及比较

对采自浙江、湖北和安徽3省的蚕豆赤斑病样品进行了病原菌的分离和鉴定,采用菌丝生长速率法检测了引起赤斑病的2种病原菌——蚕豆葡萄孢Botrytis fabae和灰葡萄孢B.cinerea 的抗药性发生情况,并在离体条件下通过抗药性诱导试验比较了二者的抗药性风险。结果共分离得到153个菌株,其中蚕豆葡萄孢122株(占79.7%),灰葡萄孢31株(占20.3%)。共检测到37株多菌灵高水平抗药性菌株(其中蚕豆葡萄孢9株)和42株异菌脲低水平抗药性菌株(其中蚕豆葡萄孢17株);嘧霉胺对153个菌株的EC50值在0.01~5.13 μg/mL之间,平均EC50值为0.72±0.15 μg /mL;表明蚕豆赤斑病菌对常见杀菌剂已表现出一定的抗药性,且灰葡萄孢的抗药性问题比蚕豆葡萄孢要严重得多。抗药性诱导试验进一步证实,灰葡萄孢的抗药性风险明显高于蚕豆葡萄孢。     

Smoke pollution and the effects of root-knot nematodes on the growth of eggplants

Single and combined effects of smoke pollution and the root-knot nematode, Meloidogyne incognita race 1, were studied on eggplant in 1988 and 1989 at two sites (K1 and K2) 1 and 2 km respectively away from a coal-fired thermal power plant, and a control site at the Department of Botany, Aligarh Muslim University (AMU). Mean concentrations of SO₂, NO₂ and suspended particulate matter (SPM) were respectively 172, 95 and 626 μg/m³ in 1988 and 144, 97 and 556 μg/m³ in 1989 at K1; and 258, 150 and 344 μg/m³ in 1988 and 226, 113 and 293 μg/m³ in 1989 at K2. Peak levels of the gases and SPM were recorded in the early (12.00–15.00) and late (15.00–18.00) afternoon respectively. Concentrations of the pollutants at a control site were very low. Marginal browning appeared on the leaves of eggplants grown at site K2 whether or not they were infected by the nematodes. The galling caused by nematodes was severe at both polluted sites but egg mass production was inhibited at K2. Egg laying was significantly reduced at both polluted sites. M. incognita significantly reduced plant growth, yield and leaf pigment production of eggplant at the control and polluted sites. Most non-infected plants also showed significant reductions in these plant varieties i.e. from the effects of pollution at K2. Foliar concentrations of sulphur were significantly increased at both polluted sites, being greater in infected plants. The interaction of smoke pollution and M. incognita infection led to greater suppressions at the polluted sites, which were mostly significant at K1. At the polluted sites, leaves had more and longer trichomes; also the number and size of stomata were decreased but their openings were wider especially in infected plants at the two polluted sites.     

Baseline sensitivity of Monilia yunnanensis to the DMI fungicides tebuconazole and triadimefon

Monilia yunnanensis was recently identified as a new species causing brown rot of peach in China. Sterol 14α-demethylase inhibitors (DMIs) continue to be important in the management of brown rot of Monilinia spp. worldwide. Tebuconazole and triadimefon are two kinds of DMI fungicides that may be used for brown rot control in China. To establish the baseline sensitivity of M. yunnanensis to these two DMI fungicides, 203 M. yunnanensis single spore isolates were collected. Measurements of sensitivity to the two fungicides were based on inhibition of mycelial growth. For both fungicides, the sensitivity distribution was a unimodal curve, with an EC₅₀ range (the effective concentration to inhibit mycelial growth by 50 %) of 0.0001–0.0644 μg/ml for tebuconazole and 0.2311–1.7477 μg/ml for triadimefon. The M. yunnanensis isolates were obtained from orchards where DMI fungicides have not been used for peach brown rot control, thus the fungicide sensitivity distribution established in this study can be considered as the baseline for monitoring the resistance development in M. yunnanensis once the DMI fungicides are used to control peach brown rot.