First Report of Downy Mildew caused by Pseudoperonospora cannabina on Cannabis sativa in New York.

Hemp (Cannabis sativa <0.3% tetrahydrocannabinol) is an emerging crop used for grain, fiber, and cannabinoid production (Fike et al. 2020). In New York, hemp is grown both in controlled environment facilities, including greenhouses, and as a field crop. In August 2020, downy mildew-like symptoms were observed on leaves and inflorescence of hemp plants in a field research trial in Ithaca, NY. Several cultivars, including 'Auto CBD', were affected. Disease was severe with some plants reaching 75% disease severity at the individual plant level. In the most severely affected plots, there was no marketable yield. The disease was characterized by chlorotic and necrotic lesions producing sporangiophores under high humidity. Pigmented sporangia were produced on branched sporangiophores. On artificially inoculated leaves incubated at 18°C, 80% humidity, 12h light for 5d, sporangiophores produced 8-19 pigmented, lemon-shaped sporangia with mean ± SD dimensions of 25.2 ± 3.0 (18.9 to 30.4) x 18.2 ± 2.1 (14.6 to 23.2) µm (n=50). Each sporangium produced 2-5 zoospores after less than 45 min in water at room temperature (22°C). Sporangia were collected from sporulating lesions and DNA was extracted as outlined in Crowell et al. (2020). Fragments of the ribosomal internal transcribed spacer (ITS) region (White et al. 1990), the beta-tubulin ras-associated ypt1 gene (Moorman et al. 2002), and the mitochondrial cytochrome B oxidase subunit 2 (cox2) gene (Hudspeth et al. 2000) were amplified by PCR and sequenced bidirectionally. Sequences were deposited in GenBank under accession numbers OK086084, OM867581, and OM867580, respectively. BLAST searches using the amplified ITS and cox2 sequences resulted in 100% identity to Pseudoperonospora cannabina (HM636051.1, HM636003.1) with ypt1 aligning at 97.95% identity (382/390 bp) with P. cannabina (KJ651402.1). The molecular characterization identified the causal agent as P. cannabina. A representative isolate was deposited in the Cornell Plant Pathology Herbarium as CUP-070922. Sporangia were rinsed from detached leaves and used to confirm pathogenicity on whole plants. Ten 4-week-old 'Anka' plants were spray-inoculated until run off with a suspension of 1x104 sporangia mL-1. Ten control plants were sprayed with water. After inoculation, plants were placed in a 19˚C growth chamber with a 12-h photoperiod and misted for 30 min twice daily to maintain humidity above 80%. Sporangia and previously described symptoms were observed 7 days post-inoculation, while control plants were asymptomatic. The pathogen was reisolated onto detached leaves of 'Anka' from inoculated leaves where both sporangia and oospores were observed. The reisolated pathogen was confirmed morphologically and molecularly, through PCR amplification and bidirectional sequencing of the ITS, cox2, and ypt1 genes, as P. cannabina. To our knowledge, this is the first report of P. cannabina causing hemp downy mildew in New York. Depending on the severity and timing of infections, this disease could pose a significant threat to hemp production in the state. Other members of the genus, P. cubensis and P. humuli cause downy mildew on cucurbits and hops, respectively. As these can cause devastating diseases on their hosts, P. cannabina must be monitored with vigilance as an emerging pathogen (Purayannur et al. 2021; Savory et al. 2011). Literature Cited: Crowell, C. R., et al.2020. Plant Dis. 104:2949. DOI 10.1094/PDIS-04-20-0718-RE Fike, J. H., et al. 2020. Page 89 In: Sustainable Agriculture Reviews, vol 42. Springer, Cham, Switzerland. DOI 10.1007/978-3-030-41384-2_3 Hudspeth, D. S. S., et al. 2000. Mycologia 92:674. DOI 10.2307/3761425 Moorman, G. W., et al. 2002. Plant Dis. 86:1227. DOI 10.1094/PDIS.2002.86.11.1227 Purayannur, S., et al. 2021. Mol. Plant Pathol. 22:755. DOI 10.1111/mpp.13063 Savory, E. A., et al. 2011. Mol. Plant Pathol. 12:217. DOI 10.1111/j.1364-3703.2010.00670.x White, T. J., et al. 1990. Page 315 In: PCR Protocols. A Guide to Methods and Applications. Academic Press, San Diego, CA. DOI 10.1016/B978-0-12-372180-8.50042-1.

In vivo confocal microscopy for the detection of canine fungal keratitis and monitoring of therapeutic response.

OBJECTIVE: To describe in vivo corneal confocal microscopy of dogs during the clinical course of fungal keratitis and correlate findings with clinical evaluations and an ex vivo experimental canine fungal keratitis model. ANIMALS STUDIED: Seven dogs with naturally acquired fungal keratitis and ex vivo canine corneas experimentally infected with clinical fungal isolates. PROCEDURES: Dogs with naturally acquired fungal keratitis were examined by in vivo laser scanning confocal microscopy. Initial confocal microscopic examinations were performed to assist in establishing the diagnosis of fungal keratitis. Serial confocal microscopic examinations were performed to guide antifungal chemotherapy. Confocal microscopy images of canine corneal fungal isolates were obtained by examination of experimentally infected ex vivo canine corneas to corroborate in vivo findings. RESULTS: Fungi cultured and detected by PCR from canine corneal samples included Candida albicans, Fusarium incarnatum-equiseti, Malassezia pachydermatis, and a Rhodotorula sp. Linear, branching, interlocking, hyperreflective structures were detected by confocal microscopy in dogs with filamentous fungal keratitis and round to oval hyperreflective structures were detected in dogs with yeast fungal keratitis. Antifungal chemotherapy was associated with a progressive reduction in the distribution and density of corneal fungal elements, alterations to fungal morphology, decreased leukocyte numbers, restoration of epithelial layers, and an increased number of visible keratocyte nuclei. No dogs had a recurrence of fungal keratitis following medication discontinuation. Confocal microscopic fungal morphologies were similar between in vivo and ex vivo examinations. CONCLUSIONS: In vivo corneal confocal microscopy is a rapid method of diagnosing fungal keratitis in dogs and provides a noninvasive mechanism for monitoring therapeutic response.

Malassezia pachydermatis keratomycosis in a dog.

A 13-year-old female Lhasa Apso was presented for blepharospasm and conjunctival hyperemia of the right eye. Ophthalmic examination revealed an anterior stromal ulcer associated with a raised yellow corneal plaque. In vivo confocal microscopy and cytology of the cornea identified neutrophilic inflammation and yeast cells. Malassezia pachydermatis was isolated from a corneal scraping. Treatment with topical voriconazole ophthalmic solution resolved the keratitis.

Impact of N-acetylcysteine on endothelial function, B-type natriuretic peptide and renal function in patients with the cardiorenal syndrome: a pilot cross over randomised controlled trial.

BACKGROUND: Both heart and renal failure are characterised by increased systemic oxidative stress and endothelial dysfunction and occur in the cardiorenal syndrome (CRS). The aim of the present study was to assess the impact of N-acetylcysteine (NAC), a potent antioxidant, on endothelial function, B-type natriuretic peptide (BNP) and renal function in patients with CRS. METHODS: In a double blind, placebo controlled manner, we randomised nine stable outpatients with both heart failure (LVEF<40% and NYHA class II or III) and renal failure (Cockroft Gault clearance of 20-60ml/min) to placebo or NAC (500mg orally twice daily) for 28 days followed by a wash out period (>7 days) and crossover to the other treatment. RESULTS: Eight patients completed the study and all data (N=9) was used in the analysis. Mean forearm blood flow improved significantly with NAC with mean ratio of improvement of 1.99 (SEM: ±0.49) for NAC and 0.73 (SEM: ±0.23) for placebo with a p-value of 0.047. There was no significant difference in BNP (p=0.25), renal function (p=0.71) or NYHA class (p=0.5). No deaths occurred during the trial. CONCLUSION: In this pilot trial of patients with CRS, NAC therapy was associated with improved forearm blood flow. This may represent a general improvement in endothelial function and warrants further investigation of antioxidant therapy in these patients.

Molecular identification of veterinary yeast isolates by use of sequence-based analysis of the D1/D2 region of the large ribosomal subunit.

Conventional methods of yeast identification are often time-consuming and difficult; however, recent studies of sequence-based identification methods have shown promise. Additionally, little is known about the diversity of yeasts identified from various animal species in veterinary diagnostic laboratories. Therefore, in this study, we examined three methods of identification by using 109 yeast samples isolated during a 1-year period from veterinary clinical samples. Comparison of the three methods-traditional substrate assimilation, fatty acid profile analysis, and sequence-based analysis of the region spanning the D1 and D2 regions (D1/D2) of the large ribosomal subunit-showed that sequence analysis provided the highest percent identification among the three. Sequence analysis identified 87% of isolates to the species level, whereas substrate assimilation and fatty acid profile analysis identified only 54% and 47%, respectively. Less-stringent criteria for identification increased the percentage of isolates identified to 98% for sequence analysis, 62% for substrate assimilation, and 55% for fatty acid profile analysis. We also found that sequence analysis of the internal transcribed spacer 2 (ITS2) region provided further identification for 36% of yeast not identified to the species level by D1/D2 sequence analysis. Additionally, we identified a large variety of yeast from animal sources, with at least 30 different species among the isolates tested, and with the majority not belonging to the common Candida spp., such as C. albicans, C. glabrata, C. tropicalis, and the C. parapsilosis group. Thus, we determined that sequence analysis of the D1/D2 region was the best method for identification of the variety of yeasts found in a veterinary population.

An fMRI motor activation paradigm demonstrates abnormalities of putamen activation in females with panic disorder.

BACKGROUND: The neurobiology of panic disorder is incompletely understood. The aim of this study was to determine if functional abnormalities of the putamen occur in panic disorder. METHODS: Activation patterns of 12 female subjects with panic disorder were compared to 18 female healthy controls using functional MRI at 3 T. A motor activation paradigm was used to probe putamen function. RESULTS: A complex motor activation paradigm for the non-dominant hand revealed decreased activation of the bilateral putamen among subjects with panic disorder. LIMITATIONS: The sample size was a relatively small cohort of non-depressed females. Further, some panic disorder subjects were taking medications and/or had comorbid conditions. However, second-level regression analyses did not reveal any correlations between medication use or comorbidity and activation patterns demonstrated by the non-dominant hand complex task. Finally, we used a post-hoc approach to determine the magnitude of global fMRI signal as a surrogate index of the global cerebral blood flow as a means of controlling for possible confounds from reduction of BOLD signal secondary to cerebral vasoconstriction resulting from possible hyperventilation among panic subjects. A more compelling approach would have been to record the respiratory data from subjects during scanning. CONCLUSIONS: Our findings suggest that putamen dysfunction occurs in at least some cases of panic disorder. We also provide preliminary evidence that a complex motor task for the non-dominant hand is a useful probe of putamen function in this disorder.

Putamen coactivation during motor task execution.

Models of corticostriatal motor circuitry have focused on the role of the circuit in the hemisphere of the motor cortex providing primary control (contralateral to the movement). We used functional magnetic resonance imaging and functional connectivity analyses to study circuit function in both the controlling and noncontrolling hemispheres. During the completion of a unilateral motor task with either hand, each putamen nucleus demonstrated strong coactivation with structures in both hemispheres. The putamen in the noncontrolling hemisphere (ipsilateral to the movement) coactivated more strongly with the controlling motor cortex than with the noncontrolling cortex. These findings suggest that the two corticostriatal circuits are functionally integrated. New circuit models based on functional connectivity may need to be developed.

Motor deactivation in the human cortex and basal ganglia.

We used a functional magnetic resonance imaging motor activation paradigm for both hands and functional connectivity analyses to investigate motor deactivation. These analyses revealed ipsilateral (to the task) postcentral gyrus connectivity with the ipsilateral primary motor cortex as well as contralateral cerebellum for both hands. Analyses using default-mode network nodes as seed regions revealed connectivity patterns similar to previous studies of the default network and therefore provide evidence that this network is demonstrable using a synchronized motor activation paradigm. We did not find evidence suggesting that motor deactivation represents modulation of the default mode network. Therefore, motor deactivation is likely a motor-specific process. Finally, we found no evidence of basal ganglia circuit deactivation, which suggests that the two-pathway hypothesis of frontal-subcortical circuit function may be incomplete.

A preliminary longitudinal fMRI study of frontal-subcortical circuits in bipolar disorder using a paced motor activation paradigm.

BACKGROUND: Compelling evidence suggests abnormal functioning of frontal-subcortical (FSC) circuits in bipolar disorder, but it is unknown whether these are state or trait abnormalities. Longitudinal functional neuroimaging studies may help clarify this issue. However, studies to date have not determined which activation paradigms may be most useful for this purpose. A paced motor task has the potential to be more reliable than cognitive or emotional activation paradigms. METHODS: To evaluate the utility of a paced motor activation task as a longitudinal probe of FSC function, we conducted fMRI scans of 10 subjects with bipolar I disorder when euthymic. We compared activation patterns to the same subjects who had been previously scanned during an episode of depression. RESULTS: The paced motor task resulted in activation in the bilateral striatum which was consistent across mood states as well as greater activation among the subjects when euthymic in the right anterior cingulate and medial frontal gyrus. LIMITATIONS: The study sample was small (10 subjects) which limits generalizability of findings. CONCLUSIONS: To our knowledge, this is the first longitudinal study of bipolar illness utilizing a paced motor task. These findings suggest that a paced motor task is useful as a longitudinal probe of both state and trait function in bipolar disorder. Further, this study provides preliminary evidence that striatal functional abnormalities may represent a trait characteristic.

Point-of-service nerve conduction studies: an example of industry-driven disruptive innovation in health care.

Nerve conduction studies (NCS) and needle electromyography are useful and established diagnostic procedures for evaluating patients with signs and symptoms of neuromuscular disease. Although technological advances have occurred since the introduction of commercial electromyography instrumentation in the 1950s, most improvements have been evolutionary and were designed to benefit traditional users--neurologists and physiatrists specializing in electromyography. In the past seven years, instruments have been introduced that automate NCS and thereby enable a broader group of physicians, including internists and orthopedic surgeons, to perform these studies and utilize electromyographic data in the care of their patients. Automated NCS devices are an example of what Clayton Christensen terms a "disruptive innovation." In this article, automated NCS is contrasted with traditional electromyography, and the challenges and opposition to its widespread adoption are explored.

Elevated HDL cholesterol is functionally ineffective in cardiac transplant recipients: evidence for impaired reverse cholesterol transport.

BACKGROUND: Cardiac transplant recipients frequently have high plasma HDL levels but it is unclear whether these promote a cardioprotective profile. METHODS: Parameters of reverse cholesterol transport and endothelial function were compared in 25 cardiac transplant recipients with low (<1.4 mmol/L; n=11) or high (>1.4 mmol/L; n=14) plasma levels of HDL and in a reference healthy group. RESULTS: Patients with high HDL had lower levels of triglyceride and prebeta1-HDL and a higher proportion of large HDL particles. When normalized to apoA-I content, non-ABCA1-dependent cholesterol efflux from RAW 264.7 macrophage cells to plasma from high HDL patients was 33% lower when compared to plasma from patients with low HDL, whereas ABCA1-dependent cholesterol efflux was not impaired. Forearm vascular responses to acetylcholine and sodium nitroprusside were not influenced by HDL levels in these patients. Compared to a reference healthy group (n=26), cardiac transplant recipients had higher levels of triglyceride, lower levels of prebeta1-HDL and LCAT, and lower activities of cholesteryl ester transfer protein and phospholipid transfer protein. CONCLUSIONS: Hyperalphalipoproteinaemia in cardiac transplant recipients is associated with the formation of partially dysfunctional HDL. We conclude that high levels of HDL may not confer cardioprotection in this group of patients.

Serologic responses of dogs given a commercial vaccine against Leptospira interrogans serovar pomona and Leptospira kirschneri serovar grippotyphosa.

OBJECTIVE: To evaluate serum titers obtained by use of the microscopic agglutination test (ie, MAT titers) to Leptospira interrogans serovar pomona and autumnalis and Leptospira kirschneri serovar grippotyphosa in dogs given a commercial vaccine against serovars pomona and grippotyphosa. ANIMALS: Forty 12-week-old puppies and 20 mature Beagles. PROCEDURE: Puppies received a commercial vaccine against serovars pomona and grippotyphosa at 12 weeks of age, then received a booster vaccine and 3 weeks later; mature dogs received the vaccine once. Serum MAT titers to serovars pomona, autumnalis, and grippotyphosa were measured before vaccination and at 2, 4, 6, 10, and 16 weeks after the first or only vaccination. RESULTS: Of the 40 puppies vaccinated, 40, 0, and 40 developed MAT titers of > 100 after vaccination to serovars pomona, grippotyphosa, and autumnalis, respectively. Microscopic agglutination test titers to serovar autumnalis were higher than MAT titers to serovars pomona and grippotyphosa and persisted in some dogs for 16 weeks (6 weeks longer than for titers to serovar pomona). Of the 20 mature dogs, 13, 5, and 20 developed MAT titers of > 100 at 2 weeks to serovars pomona, grippotyphosa, and autumnalis, respectively. Titers to serovar pomona were higher and persisted in some dogs beyond 16 weeks after vaccination, compared with titers to serovars pomona and grippotyphosa, which persisted for 10 and 6 weeks, respectively. CONCLUSIONS AND CLINICAL RELEVANCE: Subunit vaccines against serovars pomona and grippotyphosa induce MAT titers not only to homologous antigens but also to serovar autumnalis, which could lead to a misdiagnosis of leptospirosis caused by serovar autumnalis.

In vivo and in vitro evidence for ACh-stimulated L-arginine uptake.

Whereas L-arginine is clearly recognized as the precursor for nitric oxide synthesis, and its entry into endothelial cells via system y(+) transport is established, few data exist regarding the acute regulation of this transport process. We specifically investigated the effect of ACh and isoprenaline (Iso) on L-arginine uptake in the human forearm and in cultured bovine aortic endothelial cells (BAEC). Sixteen healthy males were studied. During a steady-state intra-arterial infusion of [(3)H]L-arginine (100 nCi/min), the effects of ACh (9.25 and 37 microg/min), Iso (25-50 and 200 microg/min), and sodium nitroprusside (SNP) (1-2 and 8 microg/min) on forearm plasma flow (FPF), L-[(3)H]arginine uptake, and L-[(3)H]citrulline release were determined. In parallel experiments, the effects of ACh, Iso, and SNP on L-[(3)H]arginine uptake were studied in BAEC. L-Arginine uptake was inversely related to FPF (r = -0.50; P < 0.005). At a similar FPF (ACh 56.82 +/- 9.25, Iso 58.49 +/- 5.56, SNP 57.92 +/- 4.96 ml/min; P = ns), intra-arterial ACh significantly increased forearm uptake of L-[(3)H]arginine (54,655 +/- 8,018 dpm/min), compared with that observed with either Iso (40,517.23 +/- 6,841 dpm/min; P = 0.01) or SNP (36,816 +/- 4,650 dpm/min; P = 0.011). This was associated with increased ACh-induced L-[(3)H]citrulline release compared with Iso and SNP (P = 0.046). Similarly, in BAEC, ACh significantly increased L-[(3)H]arginine uptake compared with control, Iso, or SNP (ACh 12.0 x 10(7) +/- 1.83 x 10(7) vs. control 6.67 x 10(7) +/- 1.16 x 10(7) vs. Iso 7.35 x 10(7) +/- 1.63 x 10(7) vs. SNP 6.01 x 10(7) +/- 1.11 x 10(7) fmol.min(-1).mg(-1) at 300 micromol/l L-arginine; P = 0.043). Taken together, these data indicate that ACh stimulates L-arginine uptake in cultured endothelial cells and in human forearm circulation, indicating the potential for acute modulation of endothelial L-arginine uptake.

Plasma C-reactive protein, but not protein S, VCAM-1, von Willebrand factor or P-selectin, is associated with endothelium dysfunction in coronary artery disease.

Abnormal endothelium-dependent vasodilatation is well documented in coronary artery disease (CAD), as are significant increases of acute phase inflammatory proteins and other soluble hepatic and endothelium derived proteins. In the current study we investigated whether there is a relationship between endothelium-dependent vasodilatation and the plasma levels of such proteins. Further we examined the association between these proteins, together with age, cholesterol and endothelium function, and participant status (i.e. healthy, smoker, coronary artery disease). Three groups of participants were recruited: healthy controls (n = 20), cigarette smokers (n = 20) and patients with coronary artery disease (n = 20). Forearm vascular dilatation responses to the endothelium-dependent agonist acetylcholine were obtained using venous occlusion plethysmography. Blood was sampled at the time of study for the measurement of the serological proteins described above. Responses to acetylcholine were significantly dampened in patients with CAD when compared with control and smoker groups (forearm blood flow at acetylcholine 37 microg/min): control, smokers, CAD, 12.14 +/- 2.14, 12.35 +/- 09.4, 5.12 +/- 0.81, ANOVA, P < 0.05). Responses to acetylcholine were not different between controls and smokers. C-reactive protein (C-RP) levels (controls, smokers, CAD, 1.40 +/- 0.19, 2.27 +/- 0.71, 4.73 +/- 1.09, P < 0.05) and protein S levels were significantly higher in the CAD group compared with the other two groups. There was a significant inverse correlation between forearm blood flow responses to acetylcholine and C-reactive proteins. Both responses to acetylcholine and C-RP levels demonstrated a significant association with participant status with the r value increasing from 0.405 for responses to acetylcholine to 0.491 for the combination of responses to acetylcholine and C-RP. We conclude that there is an inverse correlation between C-reactive protein, but not protein S, VCAM-1, P-selectin nor von Willebrand factor, levels and abnormal endothelium-dependent vasodilation and further that responses to acetylcholine together with C-RP protein have a strong association with cardiovascular risk and disease.

Low-dose atorvastatin therapy does not augment endothelial function in active hypercholesterolaemic males.

AIMS: As statin therapy has been demonstrated to augment endothelial function in sedentary hypercholesterolaemia (HC), we aimed to investigate the effects of atorvastatin therapy on endothelial function in physically active, HC men. METHOD AND RESULTS: Eleven physically active, HC males were recruited. Endothelial function [forearm blood flow response to brachial artery infusion of acetylcholine (Ach)] was assessed twice in each subject following atorvastatin or no therapy in a randomized crossover design. In addition, endothelial function was compared with an active, normolipidaemic control group (C). Atorvastatin therapy reduced total and LDL cholesterol, but had no effect on basal blood flow or endothelial function (peak ACh mean difference +/- standard error 0.75 +/- 1.75 ml min(-1) per 100 ml tissue) [95% confidence interval (CI) -3.1, 4.6]. In addition, there was no difference in endothelial function between the HC and C groups (-1.14 +/- 2.60 ml min(-1) per 100 ml tissue; CI -6.53, 4.25). CONCLUSION: Statin therapy in HC patients with normal endothelial function does not augment endothelial function.