A Global Overview of Diversity and Phylogeny of the Rust Genus Uromyces

Uromyces is the second-largest plant pathogenic rust genus, is responsible for numerous diseases, and has major effects on both agricultural and non-agricultural plants. The genus is generally characterized by its unicellular teliospores that help to characterize it and distinguish it from another important rust genus, Puccinia. In this study, a global overview of the diversity and distribution of Uromyces is presented based on both online and offline resources. The information obtained was analyzed for numerical and graphical summaries to provide the diversity and distribution of the genus by country and continent. Besides this, broad taxonomical aspects, a brief life cycle, and other comparative aspects on diversity and distribution were also provided. In addition, a phylogenetic analysis based on the ITS and nLSU DNA sequence data available in GenBank and published literature was performed to examine the intergeneric relationships of Uromyces. The results obtained revealed that the rust genus is found distributed over 150 countries, territories, and occupancies of the world on around 647 plant genera belonging to 95 plant families. Phylogenetic studies based on LSU and ITS sequence data revealed that Uromyces species are polyphyletic and require more DNA-based analyses for a better understanding of their taxonomic placement.


Introduction
Uromyces (Link) Unger, a genus of rust fungi, was proposed by Unger (1833). The genus contains several important plant pathogens, parasitizes in both monocots and dicots throughout the world, and affects a range of crops, causing a varying amount of yield loss annually, with losses being very severe in many cases. This genus shares synonymy with numerous genera of fungi such as Alveomyces Bubák., Capitularia Rabenh., Coeomurus Gray., Dichlamys Syd. & P. Syd., Groveola Syd., Haplopyxis Syd. & P. Syd., Haplotelium Syd., Hypodermium subgen. Uromyces Link., Klebahnia Arthur., Nielsenia Syd., Poliotelium Syd., After the aecium is fully developed, the mature aeciospores infect the host surface by germinating and producing infection structures. Eventually, this infection leads to the production of uredia with urediospores. With the repeated infection of the host plant, these urediospores are produced in large quantities during the summer. Surprisingly, urediospores can disseminate thousands of kilometers with the help of the wind. When suitable hosts are found, these spores differentiate into telia, which ultimately produce unicellular diploid teliospores in the winter [17,18]. A general life cycle of the rust genus

Life Cycle
The species of rust genus Uromyces are generally macrocyclic in nature, i.e., exhibit all five spore forms known in Pucciniales. In addition, these fungi exhibit an autoecious life cycle, meaning that all spore forms are produced on a single host. However, the endocyclic or microcyclic nature with a heteroecious mode of the life cycle was observed in some species of Uromyces [2,5]. The general life cycle process of Uromyces spp. involves the germination of diploid teliospores in the spring with a metabasidium after overwintering on plant debris. The metabasidium produces four haploid basidiospores after meiosis. These basidiospores with two different mating types germinate and start infection by producing different infection structures on the surface of the host plant. Once an infection is established, the production of pycnia containing pycniospores of two mating types and receptive hyphae takes place. After, the spermatization of pycniospores of pycnia of different mating types and subsequent dikaryotization takes place in aecial primordia, along with a subsequent exchange.
After the aecium is fully developed, the mature aeciospores infect the host surface by germinating and producing infection structures. Eventually, this infection leads to the production of uredia with urediospores. With the repeated infection of the host plant, these urediospores are produced in large quantities during the summer. Surprisingly, urediospores can disseminate thousands of kilometers with the help of the wind. When suitable hosts are found, these spores differentiate into telia, which ultimately produce unicellular diploid teliospores in the winter [17,18]. A general life cycle of the rust genus Uromyces is presented in Figure 2 below. The biology and effectiveness of Uromyces heliotropii were studied by Hasan and Aracil [19], where they found the rust to effectively control an annual weed heliotrope (Heliotropium europaeum L.). They evaluated the biocontrol potential of the rust in a greenhouse and field inoculation experiments and observed that the rust rapidly killed infected plants and reduced or prevented seed production. Similarly, Anderson et al. [20] conducted a study to evaluate the potential of three rusts naturally infecting Chilean needle grass (Nassella neesiana) in Argentina, Uromyces pencanus, Puccinia graminella, and P. nassellae, as biocontrol agents. They found U. pencanus to be most effective due to the damage it inflicts on its host in the field. A list of species of Uromyces studied as biocontrol agents is given in Table 1.

Species of Uromyces
Target Organism )Plants( Reference

The Rusts as Classical Biocontrol Agents
Uromyces is the causal agent of rust disease on numerous agricultural, horticultural, and forest plantations. Rust fungi of this genus are considered a major economic threat due to possible yield losses from reduced production. Rust diseases on pea, beans, lentils, polyhouse flower crops, clover, and many more are some of the hosts infected by this genus. In addition to their identity as the causative agent of rust, the Uromyces species also shows potential for the biological control of various phytopathogenic fungi, weeds, etc.
The biology and effectiveness of Uromyces heliotropii were studied by Hasan and Aracil [19], where they found the rust to effectively control an annual weed heliotrope (Heliotropium europaeum L.).
They evaluated the biocontrol potential of the rust in a greenhouse and field inoculation experiments and observed that the rust rapidly killed infected plants and reduced or prevented seed production. Similarly, Anderson et al. [20] conducted a study to evaluate the potential of three rusts naturally infecting Chilean needle grass (Nassella neesiana) in Argentina, Uromyces pencanus, Puccinia graminella, and P. nassellae, as biocontrol agents. They found U. pencanus to be most effective due to the damage it inflicts on its host in the field. A list of species of Uromyces studied as biocontrol agents is given in Table 1.  [26] Rust fungi as biocontrol agents are mainly studied in their use against plant weeds. The biocontrol ability of some species of Puccinia was investigated, such as as P. abrupta var. partheniicola on different growth stages of Parthenium hysterophorus [27,28], Puccinia arechavaletae on Cardiospermum grandiflorum [29], Puccinia komarovii var. glanduliferae on Impatiens glandulifera [30], and Prospodium transformans on Tecoma stans var. stans [31]. A number of studies on Uromyces species have been carried out by a number of researchers (see Table 1), but the assessment of species from other rust genera has not yet been well established. Furthermore, the broad aspects of the rust fungi as biocontrol agents based on biochemical and molecular approaches still need to be explored.

Data Collection and Compilation
This paper was compiled based on the information obtained from an extensive search of peer-reviewed publications, field guides, monographs, books, conference proceedings, project reports, and other offline and online resources. This information was updated as recently as December 2020. The information obtained was compiled firstly as a table depicting names of species of Uromyces, their hosts along with the family, the locality of occurrence, and the reference of scientific publication. The scientific names of the hosts and fungi were then cross-verified for scientific entities. The host name given in the original citation was sometimes changed to be consistent with the current taxonomy based on The Plant List (http://www.theplantlist.org; accessed on 20 April 2022). The names of rust fungi genus/species as reported in the cited publications were replaced by currently accepted names according to the websites MycoBank (www.mycobank.org; accessed on 20 April 2022) and Species Fungorum (www.speciesfungorum.org; accessed on 20 April 2022). Fungal Databases, US National Fungus Collections, ARS, USDA, an important online source of plant pathogens and their hosts, was also noted during the compilation [32]. An attempt was made to summarize all available literature on diversity and distribution of Uromyces spp.; only the most appropriate references were included in this study.

Analyses of Collected Data
After inserting the collected data into the primary database as a table, they were analyzed for numerical and graphical summaries. First, the information was analyzed by providing a comparative representation of the diversity and distribution of rust fungi (Uromyces) by country and continent. Thereafter, distribution patterns based on substrate types (herb, shrub, and tree) were constructed to understand the host preference of these rust fungi. In addition, the data of the host family were also presented. The publication indices of Uromyces spp. in terms of year, decade, and era are analyzed and presented in this paper. In addition, the references in other languages are translated into English so that the scientific community can understand them easily.

Molecular Data Analyzing
DNA sequence data of Uromyces species from the LSU and ITS rDNA were downloaded from GenBank and through earlier published literature. A checklist of molecular studies on Uromyces sp. along with the name of isolate and references were also prepared and presented in Table 2. The relevant publications on molecular analyses were also consulted [5,12,33,34]. Individual nucleotide sequences of LSU and ITS were aligned distinctly using MAFFT 7 (http://mafft.cbrc.jp/alignment/server/; accessed on 1 April 2022) [35], followed by manual checking and editing where necessary in BioEdit v. 7.0.9 [36]. The sequences of taxa containing poorly aligned portions, incomplete data, missing sequence data, and gaps were trimmed. The ITS and LSU sequences alignment was converted to NEXUS format (.nxs) using ClustalX 2.1 (http://www.clustal.org/clustal2/; accessed on 1 April 2022) for Phylogenetic Analysis Using PAUP (PAUP) analysis. The aligned LSU and ITS single-gene datasets and a concatenated dataset of LSU and ITS genes were analyzed with PAUP 4.0b10 [37]. These datasets were run after completing the program output tree in the Bootstrap.tre file. Maximum Likelihood bootstrap values greater than 60% are considered good bootstrap supports and are given above each node. Phylogenetic trees are visualized using the FigTree v1.4.0 program [38] and reorganized in Microsoft power point.

Phylogenetic Analyses
In the phylogenetic results, Uromyces were separated into two complexes in both ITS and LSU sequence data. Both complexes of ITS and LSU share many similar sequences. The incomplete sequences were mostly found in the Uromyces sequence dataset, e.g., ITS1 and 5.8S or ITS1, 5.8S complete, and ITS partial or 28S partially. Approximately 50% of the sequences had up to 300 nucleotides, while the remaining sequences had up to 800 nucleotides. Incomplete sequences can result in two complexes in a single genus. Therefore, complete gene sequences from ITS and LSU are needed to analyze these complex clades (Figures 3 and 4).     Bootstrap values of MP equal to or greater than 60% are given above branches, and 58 sequences are included in the phylogenetic analyses. The best maximum parsimony (MP) dataset consists of 873 total characters, of which 600 were constant, 125 parsimony-informative, and 148 parsimonynon-informative. The parsimony analysis of the data matrix showed 1000 equally parsimonious trees with a length of 625 steps in the first tree (CI = 0.550, RI = 0.708, RC = 0.389, HI = 0.450).

Morphological Diversity and Distribution
The results compiled on diversity and distribution revealed that the rust genus Uromyces comprised a total of 1500 species that occurred worldwide as obligate parasitic fungi on vascular plants [3,4]. After combing through the different online databases [1, 32,73], a total of 988 species were included in this paper. Similar to all fungi, its distribution shows great variations in different parts of the world. The tremendously changing climates across the world lead to diversified flora, resulting in a wide diversity and distribution of rust fungi. With regard to global diversity, it is pertinent to note here that Uromyces varies in diversity among countries and continents. These rust fungi generally show a macrocyclic nature and autoecious mode of the life cycle [2,5], which confirms their morphological diversity on specific hosts in particular regions. The broad host range in microcyclic and heteroecious life cycles was also found in Uromyces species.
The genus Uromyces predominantly showed its great diversity in North America in comparison to other continents. Almost 834 (30%) species are described here, which is the highest among all continents. The diversity of the genus is known in other continents of the world as follows: Asia, with 633 (23%) species described; Europe, with 622 (23%) species; North America, with 321 (12%) species; Africa, with 313 (11%) described species; and Australia, with 32 (1%) species described. The genus appears to be well-represented in North American, Asian, and European counties. After dispersal by various modes, such as wind, water, or insect vectors, the propagules of rust fungi germinate and infect plant tissues of specific hosts. Entry of these pathogens takes place either by natural openings such as lenticels and stomata or by wounds or injuries caused by various physical agents [74].
Human anthropogenic activities also play an important role in the global distribution of these organisms. So far, Uromyces species have been found on every land on earth except Antarctica. To understand the distribution of the Uromyces species, we analyzed their distributions across continents and terrestrial ecoregions. More than 150 countries and territories or occupancies showed the distribution of this rust genus. Although only 73 sequences of ITS and nLSU Uromyces species were identified based on molecular characteristics, respectively, the majority of the species are still identified morphologically. This may impact the number and distribution pattern of the Uromyces species as molecularbased research on the rust fungi progress. In comparison to all continents, the highest distribution of 834 species of Uromyces was recorded over 66 regions of different countries and dependencies of North America. The distribution pattern observed in other continents was observed as follows: 633 species in 26 countries and dependencies of Asia; 622 species in 33 countries and dependencies of Europe; 321 species in 17 countries and dependencies of South America; 313 species in 27 countries and dependencies of Africa; and 37 species in 3 countries, islands, or dependencies of Oceania. The array of this global distribution of Uromyces species reveals their vast diversity and justifies its position as the second-largest genus of rust fungi ( Figures 5 and 6). Human anthropogenic activities also play an important role in the global distribution of these organisms. So far, Uromyces species have been found on every land on earth except Antarctica. To understand the distribution of the Uromyces species, we analyzed their distributions across continents and terrestrial ecoregions. More than 150 countries and territories or occupancies showed the distribution of this rust genus. Although only 73 sequences of ITS and nLSU Uromyces species were identified based on molecular characteristics, respectively, the majority of the species are still identified morphologically. This may impact the number and distribution pattern of the Uromyces species as molecular-based research on the rust fungi progress. In comparison to all continents, the highest distribution of 834 species of Uromyces was recorded over 66 regions of different countries and dependencies of North America. The distribution pattern observed in other continents was observed as follows: 633 species in 26 countries and dependencies of Asia; 622 species in 33 countries and dependencies of Europe; 321 species in 17 countries and dependencies of South America; 313 species in 27 countries and dependencies of Africa; and 37 species in 3 countries, islands, or dependencies of Oceania. The array of this global distribution of Uromyces species reveals their vast diversity and justifies its position as the second-largest genus of rust fungi ( Figures 5 and  6).

Distribution Patterns of Rust Fungi (Uromyces) by Substrate Types
Uromyces, being the second-largest rust genus after Puccinia, contains a number of important plant pathogens. The species of Uromyces attacks nearly all categories of plants and causes great damage to both plant and their products. Uromyces is a genus of rust fungi that infects both monocots and dicots throughout the world. Analyses of the available literature on host diversity of Uromyces revealed that a total of 647 plant genera belonging to 95 plant families were found to be infected by these rust fungi.  [75] revealed that nearly 180 plant species belonging to 85 genera and 32 families were found to be infected with Uromyces spp. Among all families, Fabaceae and Poaceae were found to be the most infected with different species of Uromyces. A family-wise comparison of the genera of infected host plants is shown in Figure 7.   Figure 8, while detailed information on the diversity, host range, and distribution of Uromyces species is summarized in Table 3.  Figure 8, while detailed information on the diversity, host range, and distribution of Uromyces species is summarized in Table  3.                        [87,88,171,172,185,191,310,415]                      Uromyces seseli-graminis E. Fisch. Arrhenatherum elatius, Melica ciliate, and Poae sp. Europe [106] Uromyces seselis Sousa da Câmara Seseli tortuosum Portugal [722] Uromyces sesseae Lagerh. Sessea sp. Ecuador [325] Uromyces setariae-italicae Yoshino Brachiaria spp., Chaetochloa spp., Eriochloa spp., Lasiacis spp., Setaria spp., and Urochloa spp. Worldwide [294] Uromyces shahrudensis Petr.

Discussion
Uromyces is the second-largest rust genus, the species of which are phytopathogenic to any category of plants, causing severe damage and reducing growth and yields. The present study provides literature-based complete information on this rust in a single compilation. In addition to being distributed worldwide on vascular plants, Uromyces species cause several damaging diseases on major agricultural crops such as alfalfa (Medicago sativa), bean (Phaseolus vulgaris), carnation (Dianthus caryophyllus), chickpea (Cicer arietinum), clover (Trifolium sp.), and pea (Pisum sativum). This study contributes to a better understanding of the taxonomy of these rust fungi in terms of their taxonomic placement, biology, pathogenicity, life cycle, diversity, and distribution. The information presented in this study helps to better understand all possible aspects of Uromyces in a single document.
The genus Uromyces is distributed globally on around 647 plant genera belonging to 95 plant families. Poaceae and Fabaceae are the most affected families, with the occurrence of more than 100 species of Uromyces. However, these fungi infect about 95 species; their occurrence on Poaceae and Fabaceae reflects the specificity of these rusts to grasses and legumes. In addition to host diversity, the distribution of species of Uromyces exhibited a wide range across the globe. The distribution of this genus extends to over 150 countries, territories, and occupancies of the world. In its continental diversity distribution, North America is followed by Asia, Europe, South America, Africa, and Oceania, respectively. A large variation in the geographical distribution along with the vast diversity of hosts demonstrated the impact of significantly changing climatic zones on rust fungi. Besides this, more than 400 Uromyces species are endemic to more than 100 countries, provinces, and islands. This may be due to the climatic conditions and precise distribution of hosts. Available studies on global diversity and distribution are rare; however, regional descriptions are available. A checklist of rust fungi of New Zealand provided by Mckenzie [313] reported the occurrence of 31 species of Uromyces. Similarly, Bahcecioglu and Kabaktepe [80] reported 74 species from Turkey, while Afshan and Khalid [536] reported 15 species of the grass family Poaceae from Pakistan. In India, 97 species of Uromyces have been reported on various hosts [75].
A total of 61 Uromyces species were reported from Portugal, whereas about 91 were from Iran [430]. The occurrence of species of Uromyces reported from different countries also supports the broad diversity of these fungi on a wide range of hosts.
Although 988 species of Uromyces investigated in the present study are found all over the world, only 73 species are known to have DNA sequence data. As in the case of other rust fungi, the species of Uromyces are also difficult to culture, which may be one of the major factors behind the reduced availability of molecular data. In addition, the isolation of DNA directly from rust fungi present on a natural host and then its sequencing is not simple or easy, which also affects the molecular studies of these fungi. Phylogenetic studies based on LSU and ITS sequence data revealed that Uromyces species are polyphyletic taxa and required more DNA-based analyses for a better understanding of their taxonomic placement. The polyphyletic nature of Uromyces species was also confirmed by Aime and McTaggart [5] in their study to propose a higher ranking classification for rust fungi, with notes on genera. Similarly, this was also proposed by Gautam et al. [34] during their study on Indian Pucciniales with the description of the taxonomic outline, including important descriptive notes. Overall, the present study proposes the requirement of fresh collections of Uromyces species and their molecular characterization to generate molecular data so that their phylogenetic relationships can be explained more precisely. The development of a universal digital platform exclusively for global rust fungi should be developed for the benefit of researchers working on this specific group of fungi.

Conclusions
Being the second-largest plant pathogenic rust genus, Uromyces showed a great variation with respect to its diversity and distribution. After a complete analysis of information gathered in the present study, it was concluded that the species of rust genus Uromyces are distributed globally. Their distribution has been reported in over 150 countries and territories or occupancies of the world. However, the genus Uromyces predominantly showed its great diversity in North America in comparison to other continents. Approximately 647 plant genera belonging to 95 plant families are reported to be affected by these rust pathogens. Apart from this, the endemic nature of this genus is also revealed, which concluded that more than 400 species of Uormyces are found to be endemic in more than 100 countries. The biocontrol nature of some species of Uormyces is also elucidated in this study. Moreover, analyses of LSU and ITS sequence data revealed the polyphyletic nature of species of Uromyces. Further DNA-based analyses of rust disease caused by Uromyces are still required to develop a better understanding of their taxonomic placement.

Acknowledgments:
The authors wish to thank their respective organizations for providing the necessary laboratory facilities and valuable support during the study. The publication of this article was funded by Chiang Mai University, Thailand.

Conflicts of Interest:
The authors declare no conflict of interest.