Confirmation of Itersonilia perplexans infecting pyrethrum (Tanacetum cinerariifolium) in Australia.

Pyrethrum (Tanacetum cinerariifolium (Trevir.) Sch. Bip.) is grown to extract pyrethrins which are active ingredients for insecticides (Greenhill 2007). The Australian pyrethrum industry supplies over 50% of the world market. Surveys of Tasmanian crops in spring 2013, detected the presence of a fungus putatively identified as Itersonilia perplexans Derx. on foliage in 54 of 86 surveyed fields (Hay et al. 2015). This fungus was associated with necrotic leaf tips often spreading to encompass whole leaves. However, pathogenicity to pyrethrum was not confirmed. To isolate, tissue was excised from foliar lesions, surface sterilised using 0.4% NaClO, placed onto 2% water agar and incubated at 20°C for 5 days. Colonies were pure-cultured by hyphal-tip transfer onto potato-dextrose agar. Eleven isolates were cultured onto yeast mold agar (YMA) for 14 days at 15°C in the dark (Horita and Yasuoka 2002). Colonies were slow growing (1.9 to 2.3 mm/day) white to buff on both surfaces, with a darker center visible on lower surfaces. Mycelia were straight and hyaline with clamp connections at the septa. Squares transferred from the edge of YMA colonies onto microscope slides produced ballistoconidia that were aseptate, granular and lunate, kidney or lemon-shaped after 24 h. Ballistoconidia lengths and widths (n = 50/isolate) ranged from 14.6 to 20.4 µm and 10.0 to 13.6 µm. Chlamydospores were not observed. These observations were consistent with descriptions of I. perplexans (Koike and Tjosvold 2001; Liu et al. 2015). All 11 isolates were sequenced across the internal transcribed spacer (ITS) region of rDNA (ITS; primers V9G/ITS4; de Hoog and van den Ende 1998; White et al. 1990), and large (LSU; primers LROR/LR7; Rehner and Samuels 1995), and small (SSU; NS1/NS4; White et al. 1990) subunits of rDNA (Genbank accession nos. KU563626 to KU563658). The ITS (673 bp), SSU (1,047 bp) and LSU (1,318 bp) differed by 3, 1 and 0 bp, respectively, across isolates. Maximum parsimony and maximum likelihood analyses of a concatenated 3 loci alignment with Cystofilobasidiales representatives (Liu et al. 2015) placed all isolates and the I. perplexans ex-neotype strain CBS 363.85 within a single monophyletic clade with 100% bootstrap support. Two representative isolates are stored at the Plant Pathology Herbarium (accession nos. BRIP 57986 and 57987). Leaves of 46-day-old pyrethrum plants (n = 45), generated from surface sterilised seed, were inoculated with a 1.5 × 105 ballistoconidia/ml suspension (equal mix of eight isolates) and maintained between 10 and 22°C under a 12-h photoperiod for 14 days. Brown necrotic leaf tips, consistent with reported field symptoms were observed on 71% of plants and I. perplexans was recovered from 69% of symptomatic plants. For flower inoculations, pyrethrum plants were removed from fields as vegetative plants in spring and maintained in a greenhouse set at 20:14°C and 14:10 h day:night. Open flowers (10 per plant) were dipped into a 1.2 × 104 ballistoconidia/ml suspension mix of three isolates. Brown withered ray florets were observed on 10/12 plants 18 days post-inoculation, matching those described in petal blight of chrysanthemum (McRitchie et al. 1973). I. perplexans was re-isolated from 11/12 inoculated plants and 1 control plant (of 12) which exhibited the same symptoms. In both experiments, I. perplexans was identified based on its distinctive morphology. This confirms the pathogenicity of I. perplexans to both pyrethrum leaves and flowers.

Evidence for Sexual Recombination in Didymella tanaceti Populations, and Their Evolution Over Spring Production in Australian Pyrethrum Fields.

Tan spot, caused by Didymella tanaceti, is one of the most important foliar diseases affecting pyrethrum in Tasmania, Australia. Population dynamics, including mating-type ratios and genetic diversity of D. tanaceti, was characterized within four geographically separated fields in both late winter and spring 2012. A set of 10 microsatellite markers was developed and used to genotype 774 D. tanaceti isolates. Isolates were genotypically diverse, with 123 multilocus genotypes (MLG) identified across the four fields. Fifty-eight MLG contained single isolates and Psex analysis estimated that, within many of the recurrent MLG, there were multiple clonal lineages derived from recombination. Isolates of both mating types were at a 1:1 ratio following clone correction in each field at each sampling period, which was suggestive of sexual recombination. No evidence of genetic divergence of isolates of each mating type was identified, indicating admixture within the population. Linkage equilibrium in two of the four field populations sampled in late winter could not be discounted following clone correction. Evaluation of temporal changes in gene and genotypic diversity identified that they were both similar for the two sampling periods despite an increased D. tanaceti isolation frequency in spring. Genetic differentiation was similar in populations sampled between the two sampling periods within fields or between fields. These results indicated that sexual reproduction may have contributed to tan spot epidemics within Australian pyrethrum fields and has contributed to a genetically diverse D. tanaceti population.

Imiquimod-induced hypopigmentation following treatment of periungual verruca vulgaris.

Imiquimod is a topical immunomodulating medication approved by the US Food and Drug Administration for the treatment of condyloma acuminata, actinic keratoses (AKs), and superficial basal cell carcinoma (BCC). Imiquimod commonly is used off label for its antiviral and antitumoral effects. We present a case of a 51-year-old man with vitiligolike depigmentation following treatment of periungual verruca vulgaris with imiquimod therapy.

Mycoflora Associated With Pyrethrum Seed and the Integration of Seed Steam Treatment Into Foliar Disease Management Strategies.

A complex of foliar diseases can affect pyrethrum in Australia, but those of greatest importance are ray blight, caused by Stagonosporopsis tanaceti, and tan spot, caused primarily by Didymella tanaceti. Isolation of fungi from pyrethrum seed lots produced over 15 years resulted in recovery of six known pathogens: S. tanaceti, D. tanaceti, Alternaria tenuissima, Colletotrichum tanaceti, Stemphylium botryosum, and Botrytis cinerea. The incidence of S. tanaceti and D. tanaceti isolated from seed varied between 0.9 and 19.5% (mean = 7.7%) and 0 and 24.1% (mean = 5.3%) among years, respectively. Commercial heat treatment of pyrethrum seed via steaming reduced the incidence of D. tanaceti from 10.9 to 0.06% and the incidence of S. tanaceti from 24.6% to nondetectable levels (<0.18%). In a second experiment, both species were reduced to nondetectable levels (<0.20%) from their initial incidences of 22.4 and 2.4%, respectively. In a field study in 2013, colonization of pyrethrum foliage by S. tanaceti was reduced from 21.1 to 14.3% in early winter when heat-treated seed was planted. However, isolation frequency of D. tanaceti was not affected significantly by seed treatment in this year. In a related experiment in 2015, the isolation frequency of D. tanaceti in plots planted from heat-treated seed depended on both prior application of an industry-standard fungicide program and proximity to another pyrethrum field in autumn. The fungus was recovered at a similar frequency in fungicide-treated and nontreated plots located near other pyrethrum fields (13.8 versus 16.3%, respectively), whereas recovery of the pathogen was reduced by fungicide applications in geographically remote pyrethrum fields (6.7 versus 1.4%, respectively). However, these differences in isolation frequency of D. tanaceti in autumn did not obviate the need for later fungicide applications to suppress foliar disease intensity in spring or flower yield in summer, independent of the proximity to other pyrethrum fields. This study suggests that steam treatment of seed can delay development of the foliar disease complex on pyrethrum, although an extremely low level of remaining infected seed or exogenous sources of inoculum necessitates the use of foliar fungicide applications in spring.

Pediatric nevoid basal cell carcinoma syndrome.

Nevoid basal cell carcinoma syndrome (NBCCS) is a rare, autosomal-dominant, cancer-predisposing, multisystem disorder. The clinical manifestations of NBCCS include multiple basal cell carcinomas (BCCs), odontogenic keratocysts, palmar or plantar pits, and calcification of the falx cerebri. We present a case of an 11-year-old boy with Fitzpatrick skin type V who presented with multiple facial lesions and a history of maxillary keratocysts. Skin biopsy was consistent with pigmented BCC of the right nasolabial fold. Further clinical workup revealed multiple pigmented BCCs, palmoplantar pits, and calcification of the tentorium. Genetic testing revealed a heterozygous mutation in the patched 1 gene, PTCH1, consistent with NBCCS. This case highlights the treatment considerations in pediatric cases of NBCCS in Fitzpatrick skin type V patients.

The Tricky Tear Trough: A Review of Topical Cosmeceuticals for Periorbital Skin Rejuvenation.

There is a growing demand for noninvasive anti-aging products for which the periorbital region serves as a critical aspect of facial rejuvenation. This article reviews a multitude of cosmeceutical ingredients that have good scientific data, specifically for the periorbital region. Topical treatment options have exponentially grown from extensively studied retinoids, to recently developed technology, such as growth factors and peptides. With a focus on the periorbital anatomy, the authors review the mechanisms of action of topical cosmeceutical ingredients, effectiveness of ingredient penetration through the stratum corneum, and validity of clinical trials.

Changes in Distribution and Frequency of Fungi Associated With a Foliar Disease Complex of Pyrethrum in Australia.

In Australia, pyrethrum (Tanacetum cinerariifolium) is affected by a foliar disease complex that can substantially reduce green leaf area and yield. Historically, the most important foliar disease of pyrethrum in Australia has been ray blight, caused by Stagonosporopsis tanaceti, and other fungi generally of minor importance. Temporal fluctuations in the frequency of fungi associated with foliar disease were quantified in each of 83 fields in northern Tasmania, Australia, during 2012 and 2013. Sampling was conducted throughout winter (April to July), spring (August to September), and summer (November) representing different phenological stages. Microsphaeropsis tanaceti, the cause of tan spot, was the pathogen most prevalent and isolated at the highest frequency, irrespective of sampling period. The next most common species was S. tanaceti, whose isolation frequency was low in winter and increased in spring and summer. Known pathogens of pyrethrum, Alternaria tenuissima, Colletotrichum tanaceti, and Stemphylium botryosum were recovered sporadically and at low frequency. Two species of potential importance, Paraphoma chrysanthemicola and Itersonilia perplexans, were also found at low frequency. This finding suggests a substantial shift in the dominant pathogen associated with foliar disease, from S. tanaceti to M. tanaceti, and coincides with an increase in defoliation severity in winter, and control failures of the spring fungicide program. Factors associated with this finding were also investigated. Sensitivity of M. tanaceti and S. tanaceti populations to the fungicides boscalid and cyprodinil collected prior to and following disease control failures in the field were tested under in vitro conditions. A high proportion (60%) of the M. tanaceti isolates obtained from fields in which no response to the spring fungicide program was found were insensitive to 50 µg a.i./ml boscalid. This represented a 4.2-fold increase in the frequency of this phenotype within the M. tanaceti population over 2 years. No shifts in sensitivities to cyprodinil of M. tanaceti and S. tanaceti, or S. tanaceti to boscalid, were observed. Considering the increase in defoliation severity over winter, the benefits of applying fungicides in autumn, in addition to the commercial standard (spring only), were quantified in 14 individual field trials conducted in 2011 and 2012. Mixed-model analysis suggested fungicide application in autumn may improve pyrethrum growth during late winter and early spring, although effects on defoliation and yield were minimal. The increasing prevalence and isolation frequency of M. tanaceti and boscalid resistance within the population is of concern and highlights the urgent need for adoption of nonchemical methods for disease management in Australian pyrethrum fields.