Brown Spot Lesions Research Articles

First Report of Brown Leaf Spot Caused by Epicoccum tobaicum on Oat (Avena sativa) in Korea.

Oat (Avena sativa) is one of the Korean winter crops, and oat consumption has been increasing because it is widely perceived as a superfood. Recently, various fungal diseases have been reported likely due to climate changes during the winter season in Korea (Choi et al., 2018; Kim, 2020). During the 2020-2021 winter to spring, we surveyed new fungal diseases among cereals, including oats, in the southern region of Korea. In April 2021, brown leaf spots on oat leaves were observed in Gangjin, Jeollanam-do, Korea. These brown spots were irregular circles, ranging from 2-7 mm in diameter. Samples from three infected leaves were surface sterilized by treating them with 70% ethanol for 1 min and 1% NaOCl for 1 min. The samples were subsequently rinsed at least twice with distilled water and dried with a sterile paper towel before being placed on 1.5% water agar supplemented with 100 ppm streptomycin. Hyphal tips derived from infected tissues after incubation at 25C for 7 days were transferred to a fresh potato dextrose agar (PDA). Three isolates, labeled as KJO-AN2-S1, KJO-AN2-S2 and KJO-AN2-S3, were obtained via single hyphal tip purification. Colonies on PDA were pigmented vermilion and subsequently turned to saffron color with irregular margins after 7 days. Conidia produced on PDA were golden to dark brown, globose to subglobose, solitary, and measured 15.5-21.5 μm in diameter (n=50). Cultural and morphological characteristics suggested that these isolates belong to Epicoccum species (Chen et al. 2017). For identification by sequencing, the ITS (MW691866-68), tub2 (MW691872-74), and rpb2 (MW691869-71) regions of three isolates were amplified using the primer pairs ITS5/ITS4 (White et al., 1990), Btub2Fd/ Btub4Rd (Woudengerg et al., 2009), and RPB2-5F2 (Sung et al., 2007)/fRPB2-7cR (Liu et al., 1999), respectively. A maximum likelihood phylogenetic analysis based on the concatenated ITS, tub2, and rpb2 sequences placed the three isolates within a clade comprising E. tobaicum CBS 384.36. A mycelial plug (5 mm diameter) was inoculated onto wounded and unwounded leaves of healthy 12-day-old oat (cv. Choyang) seedlings. The control leaves were inoculated with a sterile PDA plug. All inoculated and control plants were placed in a plastic box and incubated at 20℃ in darkness with 100% humidity. After 1 day, the inoculated mycelial plug or sterile PDA plug from plants was removed; the plants in plastic boxes were then transferred to a growth chamber set at 20℃ with 12 h light and 60-70% humidity. While brown spot lesions were observed in both unwounded and wounded leaves 7 days post-inoculation, both wounded and unwounded control leaves remained asymptomatic. The pathogen was recovered from all symptomatic leaf tissues but could not be isolated from control leaves. The re-isolated pathogen was identified as E. tobaicum through morphological characterization and sequence-based identification, fulfilling Koch's postulates. This study is the first to report a causal relationship between E. tobaicum and brown leaf spot disease of oat in Korea. Identification of this newly emerging fungal disease on oats will help prepare for effectively managing this disease.

Dual effect of picolinic acid in the photosynthetic performance of rice plants infected by Bipolaris oryzae

AbstractBrown spot, caused by Bipolaris oryzae, is one of the most important diseases of rice. The non‐host toxin α‐picolinic acid (PA) has great potential to be used to enhance plant resistance against pathogen infection. The present study investigated the effect of spraying PA [0 (control), 1, 3, and 5 mg/mL] on the photosynthetic performance of rice plants (cultivar Metica‐1) infected or not with B. oryzae. Moreover, whether the PA treatment, especially at the highest concentration, could affect brown spot development was also evaluated. The chlorophyll a fluorescence parameters such as variable‐to‐maximum chlorophyll a fluorescence ratio (Fv/Fm), photochemical yield [Y(II)], yield for dissipation by down‐regulation [Y(NPQ)], the yield for non‐regulated dissipation [Y(NO)], and electron transport rate (ETR) as well as the concentration of photosynthetic pigments were determined. Based on the in vitro assay, PA inhibited mycelial growth of B. oryzae in a dose‐dependent manner and conidial germ tube length only decreased at 5 mg PA/mL. Conidia germination was not affected by the PA treatment. Necrotic lesions caused by PA were observed on leaves of non‐inoculated plants at 3 and 5 mg PA/mL. Symptoms of the brown spot were reduced on plants sprayed with 1 and 3 mg PA/mL compared to the control treatment. Brown spot lesions and those originating from PA toxicity overlapped for inoculated plants sprayed with 3 and 5 mg PA/mL. The photochemical performance of non‐inoculated plants was hampered by treatments with 3 and 5 mg PA/mL. Greater concentration of photosynthetic pigments and less impairment on the photosynthetic performance of inoculated plants sprayed with 1 mg PA/mL were noticed based on the values of Fv/Fm, Y(II), Y(NPQ), Y(NO), and ETR compared to inoculated plants non‐sprayed with PA. In conclusion, spraying rice plants with a low concentration of PA could decrease brown spot severity while preserving the photosynthetic capacity of the infected plants. The cellular damage generated by spraying the plants with the highest PA concentration did not favour the infection process of B. oryzae.

Morphological and Molecular Analysis of Fungal Species Associated with Blast and Brown Spot Diseases of Oryza sativa.

Fungal diseases blast and brown spot in rice cause severe yield losses worldwide. Blast is caused by Magnaporthe oryzae, and Bipolaris oryzae is reported as the main causal organism of brown spot. Both diseases cause leaf lesions that are difficult differentiate until the later stages. Early detection and differentiation of the lesions would help the adoption of disease management strategies specific to the pathogens and prevent reductions in the quality and quantity of rice yields. This study was conducted in the Northern Province of Sri Lanka over five consecutive rice cultivating seasons to characterize the causal fungi of rice blast and brown spot diseases by morphological and molecular means and to develop a visual guide to differentiate the two diseases. Disease incidence was recorded in 114 fields from 2017 to 2019, and fungal isolates associated with the lesions of both diseases were cultured and subjected to morphological and molecular characterization. Competitive growth interactions between M. oryzae and the more common individual fungal isolates of the brown spot lesions were evaluated. Fungal metagenomic analysis was conducted for the fungal species isolated from brown spot lesions. A suppression of blast accompanied by an increased incidence of brown spot disease was observed during the study period. M. oryzae was confirmed to be the causal organism of the blast, and >20 species of fungi were identified to be associated with brown spot lesions through morphological and molecular studies and metagenomic analyses. Fungal internal transcribed spacer region sequencing revealed genetic variation in the highly conserved region of DNA sequences of blast and brown spot fungal isolates. B. oryzae, Curvularia, and Microdochium species were commonly isolated from brown spot lesions. In vitro competitive growth interactions between the fungal isolates revealed growth suppression of M. oryzae by the fungal isolates associated with brown spot lesions. Similarly, it can be speculated that the abundance and severity of blast in the field may have an influence on brown spot-associated fungi. A simple visual guide was developed to differentiate blast and brown spot lesions. The findings would be highly useful in the timely management of these major fungal diseases affecting rice.

Incidence of Alternaria Species Associated with Watermelon Leaf Blight in Korea.

Alternaria leaf blight is one of the most common diseases in watermelon worldwide. In Korea, however, the Alternaria species causing the watermelon leaf blight have not been investigated thoroughly. A total of 16 Alternaria isolates was recovered from diseased watermelon leaves with leaf blight symptoms, which were collected from 14 fields in Korea. Analysis of internal transcribed spacer (ITS) region, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and RNA polymerase II second largest subunit (RPB2) were not competent to differentiate the Alternaria isolates. On the contrary, analysis of amplicon size of the histone H3 (HIS3) gene successfully differentiated the isolates into three Alternaria subgroups, and further sequence analysis of them identified three Alternaria spp. Alternaria tenuissima, A. gaisen, and A. alternata. Representative Alternaria isolates from three species induced dark brown leaf spot lesions on detached watermelon leaves, indicating that A. tenuissima, A. gaisen, and A. alternata are all causal agents of Alternaria leaf blight. Our results indicate that the Alternaria species associated watermelon leaf blight in Korea is more complex than reported previously. This is the first report regarding the population structure of Alternaria species causing watermelon leaf blight in Korea.

Study Regarding the Capacity of Self-assembling Peptides to Remineralize the Acute and Chronic Incipient Caries Lesions

The aim of the study was to evaluate the effect of biomimetic self assembling peptides (P11 -4) on remineralization of acute and chronic incipient enamel lesions by microhardness analysis. Ten extracted permanent human teeth having incipient white spot lesions and ten extracted teeth with brown spot lesions on proximal surfaces were included in this study. Half of the extern surface of the lesions was covered with flowable composite resin (Vertise flow, Kerr Corporation) and served as negative control site. On the other half Curodont� Protect gel (Credentis AG, Windisch, Switzerland) was applied 2 times a week, 8 weeks (study). The teeth were cut mesio-distally using diamond disks under cooling water, perpendicularly to the extern surface of the lesions, in order that both areas (negative control and study) to be evaluated. The opposite proximal sound enamel on each section served as positive control. The subsurface microhardness of the enamel samples was determined using digital microhardness tester (MicroVickers Hardness System CV- 400DMTM, CV Instruments Namicon). On horizontal tooth sections indentations were made at 25�m (A1), 75 �m (A2), 125 �m (A3), 175 �m (A4), 225 �m (A5), and 275 �m (A6) bellow the extern surface of the lesions. For white spot and brown spot caries lesions, in both negative control and study groups, the VHN values increased from A1 to A6 areas. In study group VHN values were higher than in negative control group irrespective of the area of examination. The lowest VHN value was registered in negative control A1 area and the highest in study group A6 area. In all six areas of investigation, VHN values in negative control and study groups were lower than the values in positive control group. The use of self-assembling peptides on white spot and brown spot caries lesions increase the hardness of these lesions in the deeper layers of the lesions.

In Vitro Mutagenesis followed by Polymorphism Detection Using Start Codon Targeted Markers to Engineer Brown Spot Resistance in Kalanchoe

Kalanchoe (Kalanchoe blossfeldiana) is a common potted flower that is popular throughout the world. Brown spot (caused by Stemphylium lycopersici) is one of the common foliage diseases in kalanchoe. This disease tends to infect leaves of kalanchoe plants in hot and humid environments, reducing their aesthetic value. The current investigation aimed to generate mutations resistant to brown spot in ‘Mary’ kalanchoe through chemical mutagenesis followed by molecular marker identification. Putative mutants were developed by treating embryogenic calluses with ethyl methanesulfonate (EMS) at median lethal doses (LD50)–either a 0.8% concentration for 2 hours or a 1.0% concentration for 0.5 hours. Brown spot crude toxin solution was used as the selection agent to identify disease-resistant calluses during tissue culture. The optimal crude concentration (60%) was determined by soaking calluses with different concentrations of crude pathogen: 0%, 20%, 40%, 60%, and 80% (v/v). A total of 32 anti-brown spot lines were regenerated and tested for disease resistance with detached leaves. Three regenerated EMS mutant lines showed no obvious brown spot lesions on their leaves after the disease resistance assay and were subjected to polymorphism identification by start codon targeted (SCoT) molecular markers. Three (SCoT40, SCoT71, and SCoT72) of 45 selected primers were chosen to identify the mutants. This work may lay the foundation for further development of new disease-resistant cultivars of kalanchoe.

Micro-CT analysis of naturally arrested brown spot enamel lesions

ObjectivesThe aim of this study was to characterize the mineral density parameters through natural enamel brown spot lesions (BSLs) and to visualize and map their mineral distribution pattern in comparison to enamel whitespot lesions (WSLs). MethodsStudy specimens included seventeen proximal WSLs (ICDAS 1, 2), seventeen proximal BSLs and seventeen sound proximal specimens (ICDAS 0) collected from The Oral Surgery Department at Sydney Dental Hospital, Sydney, Australia. Imaging was undertaken using a high resolution, desktop micro-computed tomography system. A calibration equation was used to transform the grey level values of the images into true mineral density values. The qualitative analysis and the quantification of the lesion parameters including the mineral density and the thickness of the enamel lesion surface layer were performed using mineral density profiles plotted in FIJI and the visualized mineral maps in MATLAB respectively. ResultsThe maps of brownspot lesions revealed irregular demineralization patterns with faint boundaries and outlines. The regular triangular shape of proximal lesions was recognizable only in some parts of individual BSLs or was completely unrecognizable within the entire lesion. Scattered free-form areas of high density enamel were observed within or close to the surface of BSLs. A layer of high density enamel with a mineral density close to that of sound enamel was observed in all of the BSLs. The mean mineral density of the body of BSLs, including the scattered areas of high mineral density, was significantly higher than the corresponding values in white-spot lesions. The mean thickness of the surface layer in BSLs (79±15μm) was also significantly higher than white-spot lesions (51±11μm) (p<0.05). ConclusionThis study demonstrated that the mineralization parameters such as density and the thickness of the surface layer as well as distribution patterns through natural enamel brown spot lesions (BSLs) are different from enamel white-spot lesions (WSLs). The higher mineral density of the body of the lesion and the increased thickness of the surface layer in brown spot enamel lesions may suggest possible subsurface remineralization in the majority of naturally arrested BSLs.

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.

Thermal and Sedimentation Stress Are Unlikely Causes of Brown Spot Syndrome in the Coral Reef Sponge, Ianthella basta

BackgroundMarine diseases are being increasingly linked to anthropogenic factors including global and local stressors. On the Great Barrier Reef, up to 66% of the Ianthella basta population was recently found to be afflicted by a syndrome characterized by brown spot lesions and necrotic tissue.Methodology/Principal FindingsManipulative experiments were undertaken to ascertain the role of environmental stressors in this syndrome. Specifically, the effects of elevated temperature and sedimentation on sponge health and symbiont stability in I. basta were examined. Neither elevated temperature nor increased sedimentation were responsible for the brown spot lesions, but sponges exposed to 32°C developed substantial discoloration and deterioration of their tissues, resulting in death after eight days and a higher microbial diversity in those samples. No shifts in the microbial community of I. basta were observed across a latitudinal gradient or with increased sedimentation, with three previously described symbionts dominating the community of all sponges (Alphaproteobacteria, Gammaproteobacteria and Thaumarchaea).Conclusions/SignificanceResults from this study highlight the stable microbial community of I. basta and indicate that thermal and sedimentation stress are not responsible for the brown spot lesions currently affecting this abundant and ecologically important sponge species.

Exploring the Role of Microorganisms in the Disease-Like Syndrome Affecting the Sponge Ianthella basta

A disease-like syndrome is currently affecting a large percentage of the Ianthella basta populations from the Great Barrier Reef and central Torres Strait. Symptoms of the syndrome include discolored, necrotic spots leading to tissue degradation, exposure of the skeletal fibers, and disruption of the choanocyte chambers. To ascertain the role of microbes in the disease process, a comprehensive comparison of bacteria, viruses, fungi, and other eukaryotes was performed in healthy and diseased sponges using multiple techniques. A low diversity of microbes was observed in both healthy and diseased sponge communities, with all sponges dominated by an Alphaproteobacteria, a Gammaproteobacteria, and a group I crenarchaeota. Bacterial cultivation, community analysis by denaturing gradient gel electrophoresis (Bacteria and Eukarya), sequencing of 16S rRNA clone libraries (Bacteria and Archaea), and direct visual assessment by electron microscopy failed to reveal any putative pathogens. In addition, infection assays could not establish the syndrome in healthy sponges even after direct physical contact with affected tissue. These results suggest that microbes are not responsible for the formation of brown spot lesions and necrosis in I. basta.

Prevalence of tissue necrosis and brown spot lesions in a common marine sponge

Sponges form a highly diverse and ecologically significant component of benthic communities. Despite their importance, disease dynamics in sponges remain relatively unexplored. There are reports of severe disease epidemics in sponges from the Caribbean and the Mediterranean; however, extensive sponge mortalities have not yet been reported from the Great Barrier Reef (GBR) and Torres Strait, north-eastern Australia. Marine sponge surveys were conducted in the Palm Islands on the central GBR and Masig Island, Torres Strait, to determine the health of the Demosponge Ianthella basta. Using tissue necrosis and the presence of brown lesions as a proxy of health, sponges were assigned to predetermined disease categories. Sponges with lesions were present at all sites with 43 and 66% of I. basta exhibiting lesions and symptoms of necrosis in the Palm Islands and Torres Strait, respectively. Sponges from the Torres Strait also showed a greater incidence of significant and extensive necrosis in comparison to sponges from Palm Island (11.5 v. 6%). These results indicate the widespread distribution of a disease-like syndrome affecting the health of I. basta, and highlight the critical need for regular monitoring programs and future research to assess patterns in disease dynamics and ascertain the etiological agents of infection.

Caries detection with DIAGNOdent and ultrasound

The objective of this study was to evaluate the repeatability of 2 nondestructive caries detection methods, ultrasound (ULS) and DIAGNOdent, for the detection of approximal caries lesions, and to evaluate 2 tips provided for the DIAGNOdent. White/brown-spot lesions (n = 42) were captured by a digital camera, and measured by test methods and confocal laser scanning microscope (CLSM). Intra- and inter-observer reliability, accuracy, and Cohen's kappa and Spearman's rho statistics were used. At DIAGNOdent, intra-observer agreements were 78.5%, 66.7% for A tip and 59.5%, 47.6% for B for 2 examiners, respectively. Accuracy of both tips was 50% for the first examiner; 45.3% for A and 47.6% for the B tip for the second examiner. All ULS measurements were accurate, reliable, and positively and significantly correlated between examiners. The A tip of the DIAGNOdent was found to be better than the B tip for the detection of enamel caries. All DIAGNOdent and ULS measurements were positive and significantly correlated for both examiners. Both methods demonstrated high repeatability and accuracy.

Microbiological Assessment of Occlusal Brown-Spot Lesions in Primary Molars

Conventional cultural methods were used to compare the plaque flora and the level of infection of the dentine underlying 51 occlusal brown-spot lesions and 21 sound occlusal sites on the primary dentition. Freshly extracted primary molar teeth were used, and occlusal brown-spot lesions and sound occlusal sites were identified using laser fluorescence (LF) and clinical visual methods. A standardized plaque sample was taken from each site, and an LF score was recorded for one discrete site per tooth. The teeth were carefully opened at each predetermined site to determine the clinical status of the underlying dentine, and samples were collected using a sterile bur. The microbiota of the plaque and dentine samples were enumerated and identified. The mean LF scores for the sound sites and brown-spot lesions were 1.2 and 30.5 (p < 0.001), and all the sound sites exhibited hard sound dentine, but 6 out of 51 brown spots exhibited softened dentine. Overall there was no significant (p > 0.1) difference between the level of infection of the dentine of the sound and brown-spot sites, although some sites in the brown-spot lesions yielded high numbers of bacteria. However, the numbers of bacteria as log₁₀(CFU per sample + 1) ± SE recovered from the plaque above the brown-spot lesions were significantly greater than above the sound sites, i.e. 2.89 ± 0.24 and 0.89 ± 0.33, respectively. These data indicate that brown-spot lesions may be more plaque retentive than sound sites and that they are either arrested or arresting lesions, which may require preventive intervention.

Quorum Sensing Regulates Exopolysaccharide Production, Motility, and Virulence in Pseudomonas syringae

The N-acyl homoserine lactone (AHL)-mediated quorum-sensing system in the phytopathogen Pseudomonas syringae pv. syringae requires the AHL synthase AhlI and the regulator AhlR, and is additionally subject to regulation by AefR. The contribution of quorum sensing to the expression of a variety of traits expected to be involved in epiphytic fitness and virulence of P syringae were examined. Both an aefR- mutant and an ahlI- ahlR- double mutant, deficient in AHL production, were significantly impaired in alginate production and had an increased susceptibility to hydrogen peroxide compared with the wild-type strain. These mutants were hypermotile in culture, invaded leaves more rapidly, and caused an increased incidence of brown spot lesions on bean leaves after a 48-h moist incubation. Interestingly, an aefR- mutant was both the most motile and virulent. Like the wild-type strain, the AHL-deficient mutant strains incited water-soaked lesions on bean pods. However, lesions caused by an ahlI- ahlR- double mutant were larger, whereas those incited by an aefR- mutant were smaller. In contrast, tissue maceration of pods, which occurs at a later stage of infection, was completely abolished in the AHL-deficient mutants. Both the incidence of disease and in planta growth of P syringae pv. tabaci were greatly reduced in transgenic tobacco plants that produced AHL compared with wild-type plants. These results demonstrate that quorum sensing in E syringae regulates traits that contribute to epiphytic fitness as well as to distinct stages of disease development during plant infection.

Citrus Black Rot is Caused by Phylogenetically Distinct Lineages of Alternaria alternata

ABSTRACT Phylogenetic analysis revealed that isolates of Alternaria alternata causing black rot of citrus were associated with six well-supported evolutionary lineages. Isolates recovered from brown spot lesions on Minneola tangelo, leaf spot lesions on rough lemon, and healthy citrus tissue and noncitrus hosts were related closely to isolates from black-rotted fruit. Phylogenies estimated independently from DNA sequence data from an endopolygalacturonase gene (endoPG) and two anonymous regions of the genome (OPA1-3 and OPA2-1) had similar topologies, and phylogenetic analysis was performed on the combined data set. In the combined phylogeny, isolates from diverse ecological niches on citrus and noncitrus hosts were distributed in eight clades. Isolates from all lineages, regardless of ecological or host association, caused black rot in fruit inoculation assays, demonstrating that small-spored Alternaria isolates associated with different ecological niches on citrus and other plant hosts are potential black rot pathogens. These data also indicated that the fungi associated with black-rotted fruit do not form a natural evolutionary group distinct from other Alternaria pathogens and saprophytes associated with citrus. The use of the name A. citri to describe fungi associated with citrus black rot is not justified and it is proposed that citrus black rot fungi be referred to as A. alternata.

Expansão de lesão por manchas foliares em cevada e sua interação com a aplicação foliar de fungicidas

A mancha-marrom (MM) e a mancha-reticular (MR), causadas pelos fungos Bipolaris sorokiniana e Pyrenophora teres, respectivamente, são as principais doenças foliares da cevada (Hordeum vulgare). A taxa de expansão da lesão pode ser um componente importante do progresso destas doenças. Em casa de vegetação e em campo, avaliou-se a expansão de lesão em cultivares de cevada, diante da aplicação preventiva ou curativa de um fungicida triazol (ciproconazole + propiconazole) e uma estrobilurina (azoxistrobim). O tamanho médio das lesões (MM) aumentou de 2,2 a 4,9 mm², com maior incremento entre 14 e 30 dias após a inoculação. As cultivares MN 698 e EMB 128 apresentaram taxas de expansão de lesão maiores que os demais. O tratamento preventivo com fungicida protegeu as plantas de infecção por B. sorokiniana por até 21 dias. Aplicações curativas, realizadas um, dois, quatro, seis, oito, dez ou 12 dias após a inoculação, determinaram lesões iniciais maiores (até 0,68 mm²) e mais numerosas (até 3,9 por folha), com ação apenas sobre a esporulação do fungo. Em campo, somente as primeiras aplicações, iniciadas entre 46 e 60 dias após a emergência das plantas, reduziram o tamanho das lesões de MR e a área abaixo da curva de progresso da doença (AACPD), com aumento no rendimento de grãos. Como os fungicidas apresentaram pouco ou nenhum efeito curativo sobre a expansão da lesão, sugere-se reavaliar as recomendações de controle baseadas em severidade de doença.

Comparative Fluorescence Spectroscopy Shows Differences in Noncavitated Enamel Lesions

The aim of this study was to compare emission spectra from noncavitated enamel caries with different degrees of discoloration under a wide range of excitation wavelengths. Freshly extracted human molars with white spot, light discolored and dark discolored brown spot enamel caries were selected (n = 4 each). Rectangular blocks (3.5 × 3.5 mm) were cut from the carious area and a corresponding sound area of the same tooth. Emission spectra were recorded from carious and the corresponding sound areas using a fluorescence spectrophotometer at excitation wavelengths from 360 nm up to 580 nm in steps of 20 nm. The specimens were submerged in water during measurement. The spectra were corrected and normalized to peak intensity for comparisons between spectra from sound and carious areas of each tooth. Excitation spectra were recorded for selected emission wavelengths that showed maximum intensity. Emission spectra of all types of carious lesions were shifted towards longer wavelengths (red shift) when compared to the spectra of the corresponding sound enamel. The red shift was highest for dark brown spot lesions and lowest for white spot lesions (p ≤ 0.001). Distinct fluorescence bands within 600–700 nm typical for porphyrin compounds were strongest for excitation wavelengths from 400 to 420 nm and present in most of the lesions investigated.

Synchrotron X-Ray Microtomographic Investigation of Mineral Concentrations at Micrometre Scale in Sound and Carious Enamel

Synchrotron X-ray microtomography (XMT) was used to measure the linear attenuation coefficient (LAC) for 1.9-µm sidelength voxels within approximal brown spot lesions and sound human enamel. XMT demonstrated three-dimensional features, notably sheets with ∼30 µm periodicity having low LAC, identified as regions of demineralization corresponding to Retzius lines. Quantitative three-dimensional measurements of mineral concentration, derived from LAC with assumption of a single model composition, were consistent with previous measurements of sound and carious enamel from microradiographic projections. The uncertainty in measurements of mineral concentration and mineral fraction volume was investigated by modelling enamel with a range of composition and component densities. This analysis showed that, although mineral concentration can be determined from LAC with an error of <0.2 g cm^–3, the variation in pore fraction volume within caries lesions cannot be reliably determined from X-ray attenuation measurements alone.

An in vitro Comparison of a Combined FOTI/Visual Examination of Occlusal Caries with Other Caries Diagnostic Methods and the Effect of Stain on Their Diagnostic Performance

The aims of this in vitro study were to compare the combination of FOTI and visual inspection to other diagnostic methods for the detection and evaluation of the depth of occlusal caries and assess the effect of stain on diagnostic performance. Five diagnostic methods (combined FOTI/Visual (CFV), Visual, FOTI, DIAGNOdent and Electrical Caries Monitor) were considered. Histological validation was performed in the 152 occlusal molar sites (111 teeth) using 350-µm sections and a stereomicroscope. The rank correlation coefficients with histology ranged from 0.42 (DIAGNOdent) to 0.66 (CFV). The area under the ROC curves (AUC) for enamel lesions ranged from 0.82 (ECM) to 0.88 (CFV) and from 0.81 (DIAGNOdent) to 0.91 (CFV) for dentine lesions. The AUC at the dentine level for CFV and FOTI were similar (p > 0.05), but the AUC for CFV was significantly greater than for visual (p < 0.001), DIAGNOdent (p = 0.005) and the ECM (p = 0.04). FOTI was found to be particularly useful for the detection of dentine lesions. The exclusion of stain and brown spot lesions improved performance for all methods. It is concluded that CFV is useful for the determination of occlusal lesion depth and that in the presence of stain and brown spot lesions different cut-off points may be required for the ECM and DIAGNOdent to identify dentine lesions.

Worldwide Phylogeography of the Citrus Brown Spot Pathogen, Alternaria alternata

ABSTRACT Sixty-five isolates of Alternaria alternata were sampled from brown spot lesions on tangerines and mandarins (Citrus reticulata) and tangerine x grapefruit (C. reticulata x C. paradisi) hybrids in the United States, Colombia, Australia, Turkey, South Africa, and Israel to investigate the worldwide phylogeography of the fungus. Genetic variation was scored at 15 putative random amplified polymorphic DNA (RAPD) loci and 465 bp of an endo-polygalacturonase (endo-PG) gene was sequenced for each isolate. Cluster analysis of RAPD genotypes revealed significant differentiation between United State and Colombia isolates and Turkey, South Africa, Israel, and Australia isolates. Sequencing of endo-PG revealed 21 variable sites when the outgroup A. gaisen (AK-toxin-producing pathogen of Japanese pear) was included and 13 variable sites among the sampled isolates. Nucleotide substitutions at 10 of 13 variable sites represented silent mutations when endo-PG was translated in frame. Eight distinct endo-PG haplotypes were found among the sampled isolates and estimation of a phylogeny with endo-PG sequence data revealed three clades, each with strong bootstrap support. The most basal clade (clade 1) was inferred based on its similarity to the outgroup A. gaisen and consisted exclusively of pathogenic isolates from the United States and Colombia. Clade 2 consisted of pathogenic and nonpathogenic isolates from the United States, Australia, South Africa, and Israel and clade 3 contained pathogenic and nonpathogenic isolates from Australia, South Africa, Israel, and Turkey. Quantitative estimates of virulence (disease incidence) were obtained for isolates from the United States, Colombia, South Africa, Israel, and Turkey by spray inoculating detached citrus leaves and counting the number of lesions 24 h after inoculation. Large differences in virulence were detected among isolates within each location and isolates from the United States were significantly more virulent than isolates from other locations. Several isolates from Colombia, South Africa, Israel, and Turkey had low virulence and 8% of all isolates were nonpathogenic. All but one of the nonpathogenic isolates were found in clade 2 of the endo-PG phylogeny, which also included the most highly virulent isolates sampled.