Phylogeography and Evolution of Effector Genes in Phytophthora Species

Plant pathogens in the genus Phytophthora are known to cause disease on field crops, nursery plants, and forest trees. The best known example probably is Phytophthora infestans, which triggered the infamous Irish potato famine. Other important Phytophthora species include: P. ramorum (sudden oak death pathogen), P. sojae (soybean root rot pathogen) and P. cinnamomi (broad host range root rot pathogen). Knowledge and understanding of a pathogen’s host range, geographic distribution, genetic diversity, and evolutionary potential is a key to successful disease management. This thesis focused on using phylogeographic principles combined with genomics tools to answer questions regarding the expansion from the center of origin of P. infestans, identifying the putative center of origin for P. cinnamomi and evolution of RxLR (Arginine-any amino acid-Leucine-Arginine) effector genes in three Phytophthora species. A pathogen’s biology at its center of origin may be different compared to other geographical regions. This has been well demonstrated for the late blight pathogen P. infestans. Toluca Valley of Mexico is considered the center of origin of P. infestans, where it reproduces sexually and both mating types exist in a 1:1 ratio. Very few studies have looked into the population structure outside of Toluca Valley, but still in Mexico. We performed a detailed analyses using microsatellite markers and identified a gradient of genetic diversity in P. infestans populations outside of the Toluca Valley. We used evidence of equal mating type frequencies, high genetic diversity, sexual reproduction combined with phylogenetics to identify the putative center of origin of broad host range pathogen P. cinnamomi from a global population sample. Our result suggests Asia as the center of origin for the broad host range pathogen P. cinnamomi and subsequent migration out of Asia to Australia and Africa. Phytophthora species secrete numerous effectors during the process of infection which are thought to modulate the host defense system. One such class of effector is RxLR which help promote the infection and colonization of host cells. Some of these RxLR effectors can be targeted by plant resistance genes to initiate a defense response. The question of which of these genes have been conserved or lost during the evolution of Phytophthora species is largely unknown. Therefore, we used whole genome sequence data and a comparative genomics approach to test the hypothesis of variation in RxLR gene content in three Phytophthora clade 1C species. Our analysis demonstrated both conservation and proliferation of RxLR genes in these species along with high sequence diversity. We also observed emergence of a virulent allele for effector Avr3a in the modern isolates of P. infestans. Balancing selection has acted on the RxLR genes maintaining the high sequence diversity. The ascertainment bias due to mapping of short sequencing reads to the single reference genome was further quantified by assembling the unmapped reads and predicting genes shared by all isolates of P. infestans. Overall, this thesis broadens our phylogeographic and evolutionary understanding of pathogen Phytophthora species. A key to breeding for disease resistance is highly dependent on the effector repertoire of a pathogen and thus our work can help breeders to screen for resistance gene against the late blight pathogen

Letter to the Editor

Introduction: A medical humanities (MH) module, Sparshanam has been conducted for all first year undergraduate medical students at KIST Medical College, Nepal, since 2008. Knowledge, attitude and perceived skills in empathy, what it means to be sick in Nepal, the doctor, the patient, the family, doctor-patient relationship and professional values in medicine were studied at the beginning and conclusion of the module conducted from December 2011 to March 2012. Methods: A questionnaire was developed to study respondents’ perception regarding knowledge, attitude and perceived skill levels in the areas mentioned above. Total scores in different areas and overall score were calculated. All scores were normally distributed (one sample Kolmogorov-Smirnov test). Scores among different subgroups of respondents and before and after the module were compared using appropriate tests (

Notes for genera: basal clades of Fungi (including Aphelidiomycota, Basidiobolomycota, Blastocladiomycota, Calcarisporiellomycota, Caulochytriomycota, Chytridiomycota, Entomophthoromycota, Glomeromycota, Kickxellomycota, Monoblepharomycota, Mortierellomycota, Mucoromycota, Neocallimastigomycota, Olpidiomycota, Rozellomycota and Zoopagomycota)

Compared to the higher fungi (Dikarya), taxonomic and evolutionary studies on the basal clades of fungi are fewer in number. Thus, the generic boundaries and higher ranks in the basal clades of fungi are poorly known. Recent DNA based taxonomic studies have provided reliable and accurate information. It is therefore necessary to compile all available information since basal clades genera lack updated checklists or outlines. Recently, Tedersoo et al. (MycoKeys 13:1--20, 2016) accepted Aphelidiomycota and Rozellomycota in Fungal clade. Thus, we regard both these phyla as members in Kingdom Fungi. We accept 16 phyla in basal clades viz. Aphelidiomycota, Basidiobolomycota, Blastocladiomycota, Calcarisporiellomycota, Caulochytriomycota, Chytridiomycota, Entomophthoromycota, Glomeromycota, Kickxellomycota, Monoblepharomycota, Mortierellomycota, Mucoromycota, Neocallimastigomycota, Olpidiomycota, Rozellomycota and Zoopagomycota. Thus, 611 genera in 153 families, 43 orders and 18 classes are provided with details of classification, synonyms, life modes, distribution, recent literature and genomic data. Moreover, Catenariaceae Couch is proposed to be conserved, Cladochytriales Mozl.-Standr. is emended and the family Nephridiophagaceae is introduced

Genome-Wide Increased Copy Number is Associated with Emergence of Dominant Clones of the Irish Potato Famine Pathogen Phytophthora infestans

The plant pathogen implicated in the Irish potato famine, Phytophthora infestans, continues to reemerge globally. Understanding changes in the genome during emergence can provide insights useful for managing this pathogen. Previous work has relied on studying individuals from the United States, South America, Europe, and China reporting that these can occur as diploids, triploids, or tetraploids and are clonal. We studied variation in sexual populations at the pathogen’s center of origin, in Mexico, where it has been reported to reproduce sexually as well as within clonally reproducing, dominant clones from the United States and Europe. Our results newly show that sexual populations at the center of origin are diploid, whereas populations elsewhere are more variable and show genome-wide variation in gene copy number. We propose a model of evolution whereby new pathogen clones emerge predominantly by increasing the gene copy number genome-wide.The plant pathogen that caused the Irish potato famine, Phytophthora infestans, continues to reemerge globally. These modern epidemics are caused by clonally reproducing lineages. In contrast, a sexual mode of reproduction is observed at its center of origin in Mexico. We conducted a comparative genomic analysis of 47 high-coverage genomes to infer changes in genic copy number. We included samples from sexual populations at the center of origin as well as several dominant clonal lineages sampled worldwide. We conclude that sexual populations at the center of origin are diploid, as was the lineage that caused the famine, while modern clonal lineages showed increased copy number (3×). Copy number variation (CNV) was found genome-wide and did not to adhere to the two-speed genome hypothesis. Although previously reported, tetraploidy was not found in any of the genomes evaluated. We propose a model of dominant clone emergence supported by the epidemiological record (e.g., EU_13_A2, US-11, US-23) whereby a higher copy number provides fitness, leading to replacement of prior clonal lineages

Fatty Acid Analysis, Chemical Constituents, Biological Activity and Pesticide Residues Screening in Jordanian Propolis

Propolis is a resinous natural product collected by honeybees (Apis mellifera and others) from tree exudates that has been widely used in folk medicine. The present study was carried out to investigate the fatty acid composition, chemical constituents, antioxidant, and xanthine oxidase (XO) inhibitory activity of Jordanian propolis, collected from Al-Ghour, Jordan. The hexane extract of Jordanian propolis contained different fatty acids, which are reported for the first time by using GC-FID. The HPLC was carried out to identify important chemical constituents such as fatty acids, polyphenols and α-tocopherol. The antioxidant and xanthine oxidase inhibitory activities were also monitored. The major fatty acid identified were palmitic acid (44.6%), oleic acid (18:1∆9cis, 24.6%), arachidic acid (7.4%), stearic acid (5.4%), linoleic acid (18:2∆9–12cis, 3.1%), caprylic acid (2.9%), lignoceric acid (2.6%), cis-11,14-eicosaldienoic acid (20:2∆11–14cis, 2.4%), palmitoleic acid (1.5%), cis-11-eicosenoic acid (1.2%), α–linolenic acid (18:3∆9–12–15cis, 1.1%), cis-13,16-docosadienoic acid (22:2∆13–16cis, 1.0%), along with other fatty acids. The major chemical constituents identified using gradient HPLC-PDA analysis were pinocembrin (2.82%), chrysin (1.83%), luteolin-7-O-glucoside (1.23%), caffeic acid (1.12%), caffeic acid phenethyl ester (CAPE, 0.79%), apigenin (0.54%), galangin (0.46%), and luteolin (0.30%); while the minor constituents were hesperidin, quercetin, rutin, and vanillic acid. The percentage of α-tocopherol was 2.01 µg/g of the lipid fraction of propolis. Antioxidant properties of the extracts were determined via DPPH radical scavenging. The DPPH radical scavenging activities (IC50) of different extracts ranged from 6.13 to 60.5 µg/mL compared to ascorbic acid (1.21 µg/mL). The xanthine oxidase inhibition (IC50) ranged from 75.11 to 250.74 µg/mL compared to allopurinol (0.38 µg/mL). The results indicate that the various flavonoids, phenolic compounds, α-tocopherol, and other constituents which are present in propolis are responsible for the antioxidant and xanthine oxidation inhibition activity. To evaluate the safety studies of propolis, the pesticide residues were also monitored by LC-MS-MS 4500 Q-Trap. Trace amounts of pesticide residue (ng/mL) were detected in the samples, which are far below the permissible limit as per international guidelines

Phylogeography of the wide‐host range panglobal plant pathogen Phytophthora cinnamomi

Various hypotheses have been proposed regarding the origin of the plant pathogen Phytophthora cinnamomi. P. cinnamomi is a devastating, highly invasive soilborne pathogen associated with epidemics of agricultural, horticultural and forest plantations and native ecosystems worldwide. We conducted a phylogeographic analysis of populations of this pathogen sampled in Asia, Australia, Europe, southern and northern Africa, South America, and North America. Based on genotyping-by-sequencing, we observed the highest genotypic diversity in Taiwan and Vietnam, followed by Australia and South Africa. Mating type ratios were in equal proportions in Asia as expected for a sexual population. Simulations based on the index of association suggest a partially sexual, semi-clonal mode of reproduction for the Taiwanese and Vietnamese populations while populations outside of Asia are clonal. Ancestral area reconstruction provides new evidence supporting Taiwan as the ancestral area, given our sample, indicating that this region might be near or at the centre of origin for this pathogen as speculated previously. The Australian and South African populations appear to be a secondary centre of diversity following migration from Taiwan or Vietnam. Our work also identified two panglobal, clonal lineages PcG1-A2 and PcG2-A2 of A2 mating type found on all continents. Further surveys of natural forests across Southeast Asia are needed to definitively locate the actual centre of origin of this important plant pathogen.Agricultural Research Service, Grant/Award Number: 2072-22000-041-00-D; National Institute of Food and Agriculture, Grant/Award Number: 2011-68004-30154 and 2018-67013-27823info:eu-repo/semantics/publishedVersio

SARS-CoV-2 vaccination modelling for safe surgery to save lives: data from an international prospective cohort study

Background Preoperative SARS-CoV-2 vaccination could support safer elective surgery. Vaccine numbers are limited so this study aimed to inform their prioritization by modelling. Methods The primary outcome was the number needed to vaccinate (NNV) to prevent one COVID-19-related death in 1 year. NNVs were based on postoperative SARS-CoV-2 rates and mortality in an international cohort study (surgical patients), and community SARS-CoV-2 incidence and case fatality data (general population). NNV estimates were stratified by age (18-49, 50-69, 70 or more years) and type of surgery. Best- and worst-case scenarios were used to describe uncertainty. Results NNVs were more favourable in surgical patients than the general population. The most favourable NNVs were in patients aged 70 years or more needing cancer surgery (351; best case 196, worst case 816) or non-cancer surgery (733; best case 407, worst case 1664). Both exceeded the NNV in the general population (1840; best case 1196, worst case 3066). NNVs for surgical patients remained favourable at a range of SARS-CoV-2 incidence rates in sensitivity analysis modelling. Globally, prioritizing preoperative vaccination of patients needing elective surgery ahead of the general population could prevent an additional 58 687 (best case 115 007, worst case 20 177) COVID-19-related deaths in 1 year. Conclusion As global roll out of SARS-CoV-2 vaccination proceeds, patients needing elective surgery should be prioritized ahead of the general population.The aim of this study was to inform vaccination prioritization by modelling the impact of vaccination on elective inpatient surgery. The study found that patients aged at least 70 years needing elective surgery should be prioritized alongside other high-risk groups during early vaccination programmes. Once vaccines are rolled out to younger populations, prioritizing surgical patients is advantageous