First Report Of Resistance Research Articles

Spring Barley Resistance and Tolerance to the Cereal Cyst Nematode Heterodera avenae.

Heterodera avenae is a cereal cyst nematode that reduces wheat yields in the Pacific Northwest of the United States. Barley is also susceptible but there were no previous reports of resistance or tolerance to H. avenae in the United States. Spring barley cultivars were assayed in H. avenae-infested fields over 2 years. Cultivars were planted in plots treated or not treated with aldicarb. Forty-five cultivars were evaluated for the market classes of two- and six-row feed barley cultivars and two- and six-row malt barley cultivars. One two-row feed barley ('Lenetah') was ranked as resistant and four were tolerant or very tolerant. In total, 1 two-row malt barley ('Odyssey') was very resistant and 10 were tolerant or very tolerant. Two six-row feed and two six-row malt barley cultivars were tolerant or very tolerant but none were resistant. Seven feed barley cultivars were ranked as having a balance of at least moderate resistance plus moderate tolerance: 'Champion', Lenetah, 'Xena', 'Idagold II', 'Transit', 'Millennium', and 'Goldeneye'. This is the first report of resistance and tolerance of barley in H. avenae-infested fields in the Pacific Northwest. Barley productivity can be improved by planting resistant plus tolerant cultivars or by using highly resistant and highly tolerant cultivars as parents in barley improvement programs.

First Report of Resistance to QoI Fungicides in Alternaria alternata Isolates from Potato in South Africa.

Since the first report of brown spot of potatoes (Solanum tuberosum) caused by Alternaria alternata (Fr.) Keissl in South Africa (3), disease intensity has steadily increased. No fungicides are registered for control of brown spot of potatoes in South Africa but growers attempt to control the disease with products registered for early blight, which include various QoI fungicides. Failure to control brown spot with QoI fungicides led to an investigation on putative development of resistance among A. alternata populations. In the summer of 2012, diseased leaves were collected from five potato growing regions. Isolations were made from the margin of brown spot lesions by plating surface disinfested tissue on V8 agar medium (200 ml V8 juice, 3 g CaCO3, 20 g agar). Plates were incubated at 25°C in darkness for 7 days, purified, and single-spore cultures transferred to fresh potato dextrose agar (PDA) plates. Identity of isolates was confirmed using conidial morphology and PCR amplification using species-specific primers AAF2 and AAR3 (1). Eight A. alternata isolates (PPRI 13607, 13608, 13609, 13610, 13611, 13612, 13613, and 13614) were obtained and screened for sensitivity to azoxystrobin in vitro by evaluating relative conidial germination on media amended with 0, 1.0, 2.5, 5.0, 10, 25, 50, 75, 85, and 100 μg of azoxystrobin per ml of media. The dose effect of the fungicide on germination and the EC50 of each isolate were computed using the probit analysis. Isolates were subjected to DNA extraction and the partial cytochrome b (cyt b) was amplified using primer pair CBF1 and CBR2 (2). PCR products were transformed into DH5α competent cells using a pGEM-T Easy vector. Both strands of the cloned fragments were sequenced using primers T7 and SP6 (4). Isolates PPRI 13611 and 13614 had low EC50 values of 0.11 and 0.23 μg/ml, respectively, and a mean EC50 of 0.17 μg/ml, showing their relative sensitivity to azoxystrobin. The other six isolates had EC50 values ranging from 51.88 to 114.92 μg/ml, and a mean EC50 of 71.60 μg/ml, showing their resistance to azoxystrobin. Sequence analysis of the partial cyt b gene showed strong association of resistance in isolates PPRI 13607, 13608, 13609, 13610, 13612, and 13613 to a base substitution resulting in an amino acid substitution at position 143 (G143A). Isolates PPRI 13611 and 13614 did not exhibit this mutation. Although resistance has been reported on other crops where QoI fungicides, including azoxystrobin, have been used to control different pathogens, this is the first report of resistance to a QoI fungicide in field isolates of A. alternata from potatoes in South Africa. Identification of resistance will help to explain failure to control this disease using QoI fungicides. Further monitoring of resistance to azoxystrobin and other QoI fungicides is warranted.

Resistance in the mealybug Phenacoccus solenopsis Tinsley (Homoptera: Pseudococcidae) in Pakistan to selected organophosphate and pyrethroid insecticides

The mealybug Phenacoccus solenopsis is a destructive pest of cotton with the potential to develop resistance to most chemical classes of insecticides. Six populations of P. solenopsis from cotton crops at six different locations in Pakistan were evaluated for resistance to selected organophosphate and pyrethroid insecticides. Resistance ratios (RRs) at LC50 were in the range of 2.7–13.3 fold for chlorpyrifos, 11.6–30.2 fold for profenofos and for the three pyrethroids tested were 10.6–46.4 for bifenthrin, 5.8–25.2 for deltamethrin and 4.1–25.0 for lambda-cyhalothrin. This is the first report of resistance to organophosphate and pyrethroid insecticides in Pakistani populations of P. solenopsis. Regular insecticide resistance monitoring programs are needed to prevent field control failures. Moreover, integrated approaches including the judicious use of insecticides and rotation of insecticides with different modes of action are needed to delay the development of insecticide resistance in P. solenopsis.

First Report on Resistance to Pyraclostrobin, Thiophanate-methyl, Fenhexamid and Boscalid in Botrytis cinerea from Eucalyptus Seedlings in Florida Greenhouses

Botryotinia fuckeliana de Bary (anamorph Botrytis cinerea Pers.) is an ubiquitous plant pathogen causing gray mold disease on more than 200 crops grown in the field or in greenhouses. Eucalyptus seedlings originating from three different greenhouses showing stem lesions were submitted to the Gulf Coast Research and Education Center Disease Clinic in June 2012. Ten single spore isolates of B. cinerea were obtained and tested for sensitivity using spore germination and germ tube elongation assays described previously (4). Fungicides tested were pyraclostrobin at 100 μg/ml (Cabrio, BASF, Research Triangle Park, NC), thiophanate-methyl at 100 μg/ml (Topsin-M, UPI, King of Prussia, PA), fenhexamid at 1 and 50 μg/ml (Elevate, Arysta Life Sciences, Cary, NC), fludioxonil at 0.1 and 10 μg/ml (Medallion, Syngenta Crop Protection, Research Triangle Park, NC), and iprodione at 5 and 50 μg/ml (Rovral, Bayer CropScience, Greensboro, NC) on 1% malt extract agar (MEA, 10 g malt extract and 15 g agar), and to cyprodinil at 1 and 25 μg/ml (Vanguard, Syngenta Crop Protection) on 0.5% sucrose agar (4). Sensitivity to the succinate dehydrogenase inhibitors (SDHIs) boscalid at 5 μg/ml (Endura, BASF), penthiopyrad at 1 and 3 μg/ml (Fontelis, DuPont Crop Protection, Willington, DE), and fluopyram at 3 μg/ml (Luna Privilege, Bayer CropScience) was evaluated on yeast bacto acetate agar (YBA) (3). The discriminatory dose for boscalid was adapted from (2) whereas those used for penthiopyrad and fluopyram were developed in this study. Isolates were grown on malt yeast extract agar for 7 to 10 days and spore suspensions were prepared in sterile distilled water and diluted to 106 conidia/ml. Respective media in 9-cm petri dishes were seeded with 7-μl droplets from each isolate allowing testing for all isolates on one plate. Two plates were used for each fungicide and sensitivity tests were repeated twice. Germination and germ tube growth were assessed microscopically after 16 to 24 h incubation at 22°C. The frequency of isolates resistant to two, three, and four fungicides was 90, 60, and 10%, respectively. Nine isolates (90%) were resistant to thiophanate-methyl and pyraclostrobin, simultaneously, whereas six (60%) and two isolates (20%) were resistant to boscalid and fenhexamid, respectively. All boscalid-resistant isolates were also resistant to pyraclostrobin and thiophanate-methyl, but one fenhexamid-resistant isolate was sensitive to the other three fungicides. Eight isolates that germinated at 5 μg/ml iprodione but not at 50 μg/ml were considered sensitive. All isolates were sensitive to the SDHIs penthiopyrad and fluopyram as well as to cyprodinil and fludioxonil. To our knowledge, this is the first report of resistance to pyraclostrobin, thiophanate-methyl, fenhexamid, and boscalid in B. cinerea from eucalyptus seedlings in Florida. The absence of resistance to fludioxonil and iprodione is likely because these fungicides are not registered in nurseries as well as fluopyram and penthiopyrad which were developed only recently. Management practices should be developed to limit the selection and spread of additional resistant populations in eucalyptus nurseries as has occurred in Florida strawberries where multi-fungicide resistance is widespread (1).

First Report of Boscalid and Penthiopyrad-Resistant Isolates of Alternaria solani Causing Early Blight of Potato in Michigan.

Early blight of potato (Solanum tuberosum) is caused by Alternaria solani and occurs annually in Michigan. If left uncontrolled, it can result in yield losses exceeding 20% and impact stored potatoes. The disease is commonly managed using succinate dehydrogenase inhibitor (SDHI) fungicides (1). Unfortunately, recent studies have shown that SDHI resistance has increased dramatically over the past 2 years in A. solani populations (1,2). To investigate the occurrence of SDHI resistance in Michigan, potato leaves with early blight symptoms were collected from fields in Montcalm and Ionia counties, MI, in 2012. To obtain A. solani isolates from leaves, small pieces of leaf tissue (5 × 5 mm) were excised from the center of lesions and transferred on to water agar. Plates were incubated at 25°C overnight to allow conidia to germinate. Single germinated A. solani conidia were transferred to potato dextrose agar (PDA) and incubated at 25°C for 7 days. The identity of cultures was confirmed by colony and conidial morphology (3). Nineteen A. solani isolates were obtained and each was screened for sensitivity to the SDHI fungicides boscalid, penthiopyrad, and fluopyram, using a 50 ppm discriminatory dose based on EC50 values previously determined (2). Mycelial plugs (~5.5 mm) were transferred to amended and non-amended PDA plates that were incubated at 25°C for 7 days. An isolate was considered highly resistant if fungal growth relative to control plates exceeded 50%, moderately resistant if it was between 35 and 50%, and sensitive if it was less than 35% (2). A sensitive A. solani isolate (AS11) from Bonners Ferry, ID, was used as a control in these experiments. Of all isolates tested, 11% were highly resistant to both boscalid and penthiopyrad and 5% were moderately resistant to both fungicides, 21% were moderately resistant to penthiopyrad alone, and the remaining isolates (84 and 68% respectively) were sensitive to the two fungicides. None of the isolates tested were resistant to fluopyram. Recently, two major mutations, H227R in SdhB and H133R in SdhD, have been identified in highly resistant A. solani isolates in Idaho (2). Because the majority of the identified mutations occur near the 3' end of each subunit, this region was amplified and sequenced using the following primer sets: SdhB (5'-TACTGGTGGAACCAGGAGGAGTA-3' and 5'-CATACCACTCTAGGTGAAG-3'), SdhC (5'-CCAAATCACCTGGTACGCCTCG-3' and 5'-TCATCCGAGGAAGGTGTAGTAAAGGCTG-3'), and SdhD (5'-CCGACTCTATTCTCTGCGCCCT-3' and 5'-CTCGAAAGAGTAGAGGGCAAGACCCA-3'). In this study, all of the isolates that were highly resistant to both boscalid and penthiopyrad were found to contain the H133R mutation in SdhD, which correlated with the strongest resistance phenotype. To our knowledge, this is the first report of resistance to SDHI fungicides in populations of A. solani on potato in Michigan. These data are concerning as they indicate that the highly resistant isolates have already developed cross-resistance between boscalid and penthiopyrad, despite penthiopyrad not yet having regular use in Michigan. Although all of the isolates tested were sensitive to fluopyram, the discovery of isolates resistant to boscalid and penthiopyrad suggests that all SDHI fungicides should be considered at high risk of resistance development in Michigan.

Limited occurrence of resistant radish (Raphanus sativus) to AHAS-inhibiting herbicides in Argentina

Radish has developed feral and weedy biotypes, which is a concern for agriculture around the world. In Argentina, it is one of the most widespread and troublesome crop weeds. In Brazil, this species has developed herbicide-resistance to acetohydroxyacid synthase (AHAS) inhibiting herbicides. The objective of this study was to record the presence of herbicide-resistant weedy radish plants in Argentina. In spring 2008, we found a small population of radish at the end of the flowering stage in an imidazolinone-tolerant canola field treated with imazethapyr. Screening and dose-response tests were conducted to two successive generations. They proved the biotype resistant status, and showed extensive survival (between 50 and 80% of control) to the application of a double dose of four AHAS‑inhibiting herbicides from two different chemical families (imidazolinones and sulfonylureas). Dose-response assays exhibited very high resistance for imazethapyr (LD50 = 2452.5 g a.i. ha-1, GR50 = 2926.9 g a.i. ha-1) and intermediate for metsulfuron (LD50 = 3.0 g a.i. ha-1, GR50 = 43.2 g a.i. ha-1). The acquisition of cross-resistance to different herbicide families would confer an adaptive and invasive advantage in agricultural environments to this biotype. Due to the herbicide rotation conducted in the field, the dispersion of this biotype was restricted. This is the first report of resistance in weedy radish in Argentina.

Resistance to soybean aphid among wild soybean lines under controlled conditions

The soybean aphid, Aphis glycines Matsumura, is frequently a serious pest of soybean in the U.S. and Canada, and an occasional pest of soybean throughout its native range in Asia. Large infestations of this aphid cause economic loss to soybean by reductions in seed yield and oil concentration. Host-plant resistance is a potential alternative to insecticides for managing soybean aphid, and various sources may be evaluated, including adapted cultivars, landraces, and wild ancestors of the crop. To date, few studies have evaluated wild soybean for resistance to soybean aphid. In this study, initial screening assays indicated 20 wild soybean lines with resistance to soybean aphid, and three of the 20 lines had notable resistance in subsequent choice and no-choice assays. Significantly fewer soybean aphids settled on lines PI 468397 A and PI 479749 than on susceptible lines in choice assays, and aphid populations were low and moderately low on these two respective lines in no-choice assays. Populations of soybean aphid on a third line, PI 549046, were equal to or less than those on a resistant check over 3 wks in a no-choice assay. Based on results of the respective choice and no-choice assays, resistance to soybean aphid was manifested as both antixenosis and antibiosis in PI 468397 A and PI 479749, and as antibiosis in PI 549046. This is apparently the first report of resistance to soybean aphid in these three lines, and they provide soybean breeders and pest management practitioners new sources for developing aphid-resistant soybean lines.

Resistance to conventional insecticides in Pakistani populations of Musca domestica L. (Diptera: Muscidae): a potential ectoparasite of dairy animals

The house fly, Musca domestica L., is an important hygienic pest of humans and dairy animals with the potential to develop resistance to most chemical classes of insecticides. Six adult house fly strains from dairy farms in Punjab, Pakistan were evaluated for resistance to selected insecticides from organochlorine, organophosphate, carbamate and pyrethroid classes. For a chlorocyclodiene and two organophosphates tested, the resistance ratios (RR) at LC50 were in the range of 5.60-22.02 fold for endosulfan, 7.66-23.24 fold for profenofos and 2.47-7.44 fold for chlorpyrifos. For two pyrethroids and one carbamate, the RR values at LC50 were 30.22-70.02 for cypermethrin, 5.73-18.31 for deltamethrin, and 4.39-15.50 for methomyl. This is the first report of resistance to different classes of insecticides in Pakistani dairy populations of house flies. Regular insecticide resistance monitoring programs on dairy farms are needed to prevent field control failures. Moreover, integrated approaches including the judicious use of insecticides are needed to delay the development of insecticide resistance in house flies.

First Report of In-Vitro Boscalid-Resistant Isolates of Alternaria solani Causing Early Blight of Potato in Idaho.

Early blight of potato (Solanum tuberosum) is caused by Alternaria solani and occurs annually to some degree in Idaho. The timing of its appearance and rate of disease progress determine the impact on the potato crop. Though losses rarely exceed 20%, they can be higher and occur in stored potatoes if the disease is not controlled. Early blight is managed mainly by cultural practices such as plant nutrition, water management, and avoidance of plant stress, but also with the use of foliar fungicides. Currently, the main fungicides labeled for control of early blight are in the carboxamide and strobilurin fungicide groups. Development of resistance to some fungicide groups may contribute to the loss of control of early blight. Isolates of A. solani from Idaho potatoes were tested for resistance to boscalid in the carboxamide group. Diseased potato leaves with early blight symptoms were collected from fields near Parma, Rupert, and Aberdeen in southern Idaho in 2009 and 2010 and Bonners Ferry in northern Idaho in 2010. To obtain A. solani isolates from leaves, small pieces of leaf tissue (5 × 5 mm) were taken from the center of early blight lesions and streaked across the surface of a thin layer (3 mm) of water agar. Plates were incubated at 25°C overnight to allow spores to germinate. Single germinated A. solani spores were transferred to acidified potato dextrose agar and incubated in the dark at 25°C. Germinated spores were identified based on spore morphology. Spores of A. solani can be easily distinguished from other Alternaria spp. found on potato because they are ellipsoid to oblong and taper to a long beak that is usually as long as the spore body. The identity of cultures grown from single spores was confirmed by colony and spore morphology. Sensitivity of A. solani isolates to boscalid was determined by the spiral gradient endpoint method (2). For all isolates, the effective concentration for 50% reduction in growth was outside the range of the spiral plate dilution series (i.e., isolates were either completely insensitive or completely sensitive to boscalid). In total, 46 isolates (20 collected in 2009 and 26 collected in 2010) were tested against boscalid. Experiments were carried out twice with 2009 isolates using mycelial strips and conidial suspensions. Experiments with 2010 isolates were carried out three times using only conidial suspensions. Of the isolates from 2009, 15% were insensitive. There was no difference between the use of mycelial strips or conidial suspensions. In 2010, 62% of isolates were insensitive. By location, 72% of isolates from Parma, 73% from Rupert, 63% from Aberdeen, and 44% from Bonners Ferry were insensitive. Resistance to boscalid has been reported in A. alternata isolates from pistachio (1,3). However, to our knowledge, this is the first report of resistance to boscalid in isolates of A. solani on potato. These data suggest that resistance to boscalid is widespread in Idaho, even in areas like Bonners Ferry where potato cultivation is limited. Boscalid insensitivity in vitro may not translate directly to commercial production and currently there is no evidence to suggest that boscalid has failed to control early blight in Idaho. However, the discovery of insensitive isolates suggests that boscalid should be considered at high risk of resistance development.

First identification of a sweet potato begomovirus (sweepovirus) in Uganda: characterization, detection and distribution

Sweet potato begomoviruses diverge basally from all other begomoviruses and have been named sweepoviruses. In 2009, a sweepovirus was detected for the first time in sweet potato crops in Uganda by using the indicator plant Ipomoea setosa and generic primers in a polymerase chain reaction (PCR). An isolate was cloned and sequenced, the first fully sequenced genome of a sweepovirus from mainland Africa. At the nucleotide level, this isolate differed from other sweepoviruses by at least 13%, discriminating the Ugandan isolate as a new species which has been tentatively named Sweet potato leaf curl Uganda virus (SPLCUV). In infected sweet potato plants, SPLCUV showed an uneven distribution; it was detected more often in samples from the midrib and lamina of middle and lower leaves, and reversion to healthy tissue occurred, especially in shoots of cv. New Kawogo. This appears to be the first report of resistance to a sweepovirus in sweet potato. While it was only detected at relatively low efficiency by PCR, use of I. setosa plants as an indicator of sweepovirus infection in sweet potato plants was as efficient as using real‐time quantitative PCR (qPCR). Storage of dry leaves for 84 days and dried DNA extracts for 21 days did not affect the ability of PCR and qPCR to detect it. Sweepovirus(es) was detected frequently using generic primers in cultivars Ejumula, New Kawogo and 318L in eastern and central Uganda.

Resistance to Bt Maize in <I>Busseola fusca</I> (Lepidoptera: Noctuidae) From Vaalharts, South Africa

The first report of resistance of the maize stem borer [Busseola fusca (Fuller)] to Bt maize (MON810) was made in the Christiana area of South Africa during 2007. The objective of this study was to evaluate the status of resistance of other populations of B. fusca to Bt maize. One greenhouse and two laboratory studies were conducted. B. fusca populations were collected on Bt maize as well as the adjacent refugia (conventional maize and non-Bt maize) in the Vaalharts area, 50 km from the Christiana site. Control populations were collected from sites where Bt maize was not planted. In the greenhouse study 720 potted plants were each artificially infested with 10 neonate larvae of the F1-generation after the field collected populations were reared through to adults. Numbers of live larvae and larval mass per plant were determined at regular intervals over a 35-d period. Larvae of the Christiana conventional population (Bt-susceptible) on Bt maize (CHR08Con-Bt) and Bethal conventional population (Bt-susceptible) on Bt maize (BET08Con-Bt) did not survive on Bt maize for longer than 12 d. The populations collected from both Bt (VAA08Bt-Bt) maize and refuges (VAA08Ref-Bt) at Vaalharts were resistant and the subsequent generation of larvae completed their life cycle on Bt maize. Similar results were observed in the laboratory experiments. This study confirmed resistance of B. fusca to the Cry1Ab toxin (MON810). The geographical distribution of resistance was shown to include at least the Vaalharts area, in addition to the original report for the Christiana area. These observations that larvae collected from refugia at Vaalharts was resistant, show that the efficacy of the refuge strategy is compromised in this area because the contribution of refugia did not produce large enough numbers of susceptible individuals to mate with moths of which larvae survived inside Bt maize fields.

Transgenic Bt maize: farmers’ perceptions, refuge compliance and reports of stem borer resistance in South Africa

AbstractBased on surface area, South Africa is currently ranked the eighth in planting genetically modified (GM) crops in the world. Bt maize (MON810) has been grown to control lepidopterous stem borers in South Africa since its first release during 1998. The first report of resistance of the African maize stem borer, Busseola fusca (Lepidoptera: Noctuidae) and poor compliance to refuge requirements that could have contributed to resistance development prompted this study which was done in the main maize production area of South Africa. Objectives were to evaluate farmer’s perceptions of the regulatory aspects guiding the planting of Bt maize and refugia and how it was applied between 1998 and 2010. This study involved 105 commercial farmers covering 87 778 ha of maize throughout the main maize production region on the Highveld of South Africa. A questionnaire survey was conducted and addressed signing of technical agreements upon purchasing GM seed, refuge compliance, pest management practices, perceived benefits and risks relating to Bt maize. Compliance with refugia requirements was low especially during the initial 5–7 years after release. An alarmingly high number of farmers applied insecticides as preventative sprays on Bt maize and refugia irrespective of stem borer infestation levels. A large proportion of farmers reported significant borer infestation levels on Bt maize and between 5% and 93% farmers in all districts applied insecticides to Bt maize to limit borer damage, indicating that the occurrence of resistance is more wide‐spread in the country than previously thought. This study shows irresponsible management of GM crop technology by farmers, chemical and seed companies. Concerns and perceptions of farmers in South Africa seem to differ from those in Europe. South African farmers perceived little, if any, negative impact on non‐target organisms and remain positive about the technology in spite of resistance development. Other studies indicated that European farmers were concerned about these aspects. A shared concern, however, was consumer acceptance of GM maize and marketability on the export market.

First Report of in vitro Fludioxonil-Resistant Isolates of Fusarium spp. Causing Potato Dry Rot in Michigan.

Fusarium dry rot of potato (Solanum tuberosum) is a postharvest disease caused by several Fusarium spp. Dry rot is managed primarily by reducing tuber bruising and promoting rapid wound healing. Dry rot symptomatic tubers were collected from Michigan seed lots in 2009 and 2010. The isolates may not have been exposed to fludioxonil because currently applications are restricted to seed not intended for seed production (3). Small sections were cut from the margins of necrotic regions with a scalpel, surface sterile in 10% sodium hypochlorite for 10 s, rinsed twice in sterile distilled water, and blotted with sterile filter paper. The tissue pieces were plated on half-strength potato dextrose agar (PDA) amended with 0.5 g/liter of streptomycin sulfate. The dishes were incubated at 23°C for 5 to 7 days. Cultures resembling Fusarium spp. were transferred onto water agar and hyphal tips from the margin of actively growing isolates were removed with a sterile probe and plated either on carnation leaf agar (CLA) or on half-strength PDA to generate pure cultures. Fusarium isolates were obtained and used for further studies. Among them, 54 were identified as Fusarium oxysporum and 23 as F. sambucinum. Identification was based on colony and conidial morphology on PDA and CLA, respectively. The identity was confirmed through DNA extraction followed by amplification and sequencing of the translation elongation factor (EF-1α) gene region. The Fusarium-ID v. (2) and the NCBI database were used to obtain the closest match to previously sequenced materials. Pathogenicity testing was done on disease-free potato tubers, cv. FL 1879. Tubers were surface sterilized for 10 min in 10% sodium hypochlorite and rinsed twice in distilled water. Three tubers per isolate were injected with 20 μl of a conidial suspension (106 conidia/ml) made from cultures grown on PDA for 7 days. Control tubers were injected with 20 μl of sterile distilled water. All tubers inoculated with F. sambucinum and F. oxysporum developed typical potato dry rot symptoms consisting of dry brown decay lesions. F. sambucinum and F. oxysporum were reisolated from all symptomatic tubers. An effective concentration for 50% reduction in growth (EC50) was determined for each F. sambucinum and F. oxysporum isolate for thiabendazole (TBZ), fludioxonil, and difenoconazole using the spiral gradient endpoint method (1). Sensitive and resistant F. sambucinum and F. oxysporum isolates were reported. Fifteen isolates of F. sambucinum and thirty-four of F. oxysporum were resistant to fludioxonil with EC50 greater than 130 mg/liter. The remainder was sensitive to fludioxonil with EC50 ranging from 0.8 to 4.9 mg/liter. To our knowledge, this is the first report of resistance to fludioxonil in isolates of F. sambucinum and F. oxysporum in Michigan. Fusarium insensitivity in laboratory studies may not translate directly to commercial production. This disparity may result from interactions not experienced in mixed populations or within a living host. There has been no compelling evidence to suggest that fludioxonil has failed to perform because of insensitivity to Fusarium. The occurrence of such isolated strains necessitates the development and registration of partner chemistries that can preempt any future concerns on lack of performance of products in use.

Resistance to Citrus psorosis virus in transgenic sweet orange plants is triggered by coat protein–RNA silencing

The lack of naturally occurring resistance to Citrus psorosis virus (CPsV) has demanded exploitation of a transgenic approach for the development of CPsV-resistant sweet orange plants. Transgenic sweet orange plants producing intron-hairpin RNA transcripts (ihpRNA) corresponding to viral cp, 54K or 24K genes were generated and analyzed at the molecular and phenotypic levels. Two independent CPsV challenge assays demonstrated that expression of ihpRNA derived from the cp gene (ihpCP) provided a high level of virus resistance, while those derived from 54K and 24K genes (ihp54K and ihp24K) provided partial or no resistance. The presence of small interfering RNA molecules (siRNAs) in the ihpCP transgenic sweet orange plants prior to virus challenge, indicated that CPsV resistance was due to pre-activated RNA silencing, but siRNAs accumulation level was not directly correlated to the degree of the triggered virus resistance among the different lines. However, pre-activation of the RNA-silencing machinery and a certain minimum accumulation level of siRNA molecules targeting the viral genome are key factors for creating virus-resistant plants. This is the first report of resistance in citrus plants against a negative-strand RNA virus as CPsV.

Identification of Resistance to the Large Raspberry Aphid in Black Raspberry

The large raspberry aphid (Amphorophora agathonica Hottes) is an important vector of viruses in Rubus L. across North America. Although breeding for aphid resistance has long been recognized as an important tool for protecting red raspberries (Rubus idaeus L.) from viral infection, this is the first report of resistance to A. agathonica in black raspberry (Rubus occidentalis L.). Seedlings from 132 wild populations of black raspberries, representing the species' native range, were screened for resistance to A. agathonica. Strong resistance was found in three of these populations, one from Ontario (ORUS 3778), one from Maine (ORUS 3817), and one from Michigan (ORUS 4109). Resistance to the large raspberry aphid in ORUS 3778 and ORUS 3817 is dominant and appears to be conferred by different genes. We propose that the genes for resistance in ORUS 3778 and ORUS 3817 be designated Ag4 and Ag5, respectively. Resistance to A. agathonica in ORUS 4109 also appears to be controlled by a dominant allele at a single locus, but cannot be differentiated from Ag4 at this time.

Resistance phenomena to amitraz from populations of the ectoparasitic mite Varroa destructor of Argentina

In Argentina, Varroa destructor resistance to coumaphos has been previously reported. However, the status of mite susceptibility to other hard acaricides is still unknown. At present, high infestation levels of V. destructor are being detected in colonies of Apis mellifera after treatment with amitraz. The aim of the present study was to determine the LC(50) of amitraz in V. destructor from three apiaries with high mite density after treatment with the acaricide. The LC(50) values were 3.9, 3.5, and 3.7μg/Petri dish for mites from three different apiaries. Significant LC(50) differences were detected between resistant and susceptible mites. LC(50) increased 35-39-fold when compared to the corresponding baseline, suggesting the development of resistance. These results are the first report of resistance to amitraz in V. destructor in Argentina and extend the knowledge according to the status of acaricides resistance in the country.

Oseltamivir‐resistant pandemic (H1N1) 2009 influenza in a severely ill patient: the first Australian case

After a 10-day course of oral oseltamivir for pandemic (H1N1) 2009 influenza infection, a renal transplant recipient developed rapid-onset severe primary viral pneumonia due to oseltamivir-resistant virus. Respiratory failure progressed despite high-dose oral oseltamivir, nebulised zanamivir and cessation of immunosuppressive medications, but his condition improved with intravenous zanamivir. He subsequently died of non-respiratory complications. This is the first case of oseltamivir-resistant pandemic (H1N1) 2009 in Australia and the first report of resistance in a solid organ transplant recipient.

Species Distribution and Resistance Patterns to Growth-Promoting Antimicrobials of Enterococci Isolated from Pigs and Chickens in Korea

A total of 147 Enterococcus faecium and 165 Enterococcus faecalis isolates from fecal samples of chickens and pigs at slaughterhouses in Korea were tested for their resistance to 8 growth-promoting antimicrobials commonly used in animals and quinupristin and dalfopristin. Resistance to most antimicrobials was very common among both E. faecalis and E. faecium. In particular, E. faecalis showed almost no susceptibility to all the antimicrobials tested except penicillin and flavomycin, to which 1.4% and less than 24% showed resistance, respectively. Although the prevalence of resistance was lower than in E. faecalis, E. faecium showed relatively uniform resistance to all the agents tested. Among the antimicrobials tested, virginiamycin and penicillin were the most effective against E. faecium isolates: less than 31% and 41% showed resistance to those 2 antimicrobials, respectively. Penicillin was the only agent that showed relatively strong activity against both E. faecalis and E. faecium. Resistance observed in E. faecalis and E. faecium against most antimicrobials used for growth promotion was more prevalent in Korea than in European countries. The current study is the first report of resistance against feed additive antimicrobials in enterococcal isolates from livestock in Korea.

Expression of a Chitinase Gene from Metarhizium anisopliae in Tobacco Plants Confers Resistance against Rhizoctonia solani

The chit1 gene from the entomopathogenic fungus Metarhizium anisopliae, encoding the endochitinase CHIT42, was placed under the control of the CaMV 35S promoter, and the resulting construct was transferred to tobacco. Seventeen kanamycin-resistant transgenic lines were recovered, and the presence of the transgene was confirmed by polymerase chain reactions and Southern blot hybridization. The number of chit1 copies was determined to be varying from one to four. Copy number had observable effects neither on plant growth nor development. Substantial heterogeneity concerning production of the recombinant chitinase, and both general and specific chitinolytic activities were detected in leaf extracts from primary transformants. The highest chitinase activities were found in plants harboring two copies of chit1 inserts at different loci. Progeny derived from self-pollination of the primary transgenics revealed a stable inheritance pattern, with transgene segregation following a mendelian dihybrid ratio. Two selected plants expressing high levels of CHIT42 were consistently resistant to the soilborne pathogen Rhizoctonia solani, suggesting a direct relationship between enzyme activity and reduction of foliar area affected by fungal lesions. To date, this is the first report of resistance to fungal attack in plants mediated by a recombinant chitinase from an entomopathogenic and acaricide fungus.

Morphological and molecular characterization of melon accessions resistant to Fusarium wilts

Fusarium wilt incited by Fusarium oxysporum f. sp. melonis (F.o.m) is one of the most widespread and devastating melon diseases. While resistance to physiological races 0, 1, and 2 is relatively frequent in different botanical varieties, sources of resistance to race 1,2 are restricted to a few Far-Eastern accessions. In this work, the results of a screening for resistance to F.o.m. race 1,2 among 32 accessions are presented. Three Japanese accessions (‘Kogane Nashi Makuwa’, ‘C-211’, and ‘C-40’) showed the highest resistance levels, but useful levels of resistance were also detected in one Russian ‘C-160’ and two Spanish (‘C-300’ and ‘Mollerusa-7’) accessions. These resistant materials, together with other accessions previously described as resistant to F.o.m. races 0, 1, and/or 2 have been morphologically and molecularly characterized. Based on cluster analysis, these accessions have been grouped according to the botanical subspecies they belong to. Assessment of genetic diversity indicated that the resistant accessions to races 0, 1 and 2, are scattered along the established clusters. On the other hand, high levels of resistance to the race 1,2 could be found only among accessions belonging to Cucumis melo subsp. agrestis, nevertheless, a certain degree of resistance to this race could also be found within some accessions belonging to subsp. melo. As far as we know, this is the first report of resistance to F.o.m race 1,2 found out from the Far-Eastern melon material. Based on fruits characteristics, it appears that several inodurus and cantalupensis accessions could be exploited in breeding programs as resistance sources to F.o.m races 0, 1 and/or 2 for the improvement of these melon types. The accessions with the highest levels of resistance to the race 1,2 appeared to be very distant both molecularly and morphologically from the commercial types. Nevertheless ‘C-160’, ‘C-300’, and ‘Mollerusa-7’ classified as var. inodorus are morphologically very similar to the Spanish commercial types and might be used as resistant sources in breeding these melon types.