Global climate change on our planet, together with the impacts of biotic, abiotic, and anthropogenic factors on ecosystems, leads to both negative and positive changes in the ecological and biological characteristics of microorganisms. Identifying the causes and underlying patterns of these changes makes the study of the systematics, geographical distribution, and biological characteristics of fungi one of the most pressing research priorities.

Currently available mycological data do not fully synthesize information on the distribution patterns, systematics, geography, and ecology of Micromycetes occurring across all geographical regions of the Republic of Uzbekistan. This is primarily due to the fact that the Mycobiota of higher plants in certain regions of the country have not been systematically studied; the Zarafshan National Nature Park is among such insufficiently investigated areas.

A comprehensive investigation of the species composition, geographical distribution, and biological characteristics of Micromycetes occurring on higher plants in the Zarafshan National Nature Park is therefore required. In addition, studying the ecological dependence and seasonal development patterns of Micromycetes associated with higher plants in the park is of significant scientific and practical importance.

At present, there are nine national nature parks in Uzbekistan. Among them, the Zarafshan National Nature Park is distinguished by its geographical location and its unique riparian (tugai) vegetation and floristic composition. The park was established in 2018 on the basis of the Zarafshan Nature Reserve, which had been founded in 1975.

The Zarafshan National Nature Park is located in the middle part of the Turkestan and Zarafshan mountain ranges. It is supplied with water by the Zarafshan River and its tributaries. The Zarafshan River is one of the largest river systems in Uzbekistan and Central Asia, extending approximately 770 km from east to west. Owing to the diversity of soil and climatic conditions, the Zarafshan Valley possesses a distinctive natural environment. For this reason, the Zarafshan River basin is divided into three parts: the upper, middle, and lower Zarafshan. The Zarafshan National Nature Park (with an area of approximately 2426.4 ha) is situated in the middle reaches of the Zarafshan River [1][2] (Figure 1).

Figure 1. Zarafshan National Nature Park (39˚40'10"N, 67˚05'43"E).

Groundwater in this area occurs at shallow depths, and four soil types are distinguished: marsh-meadow, alluvial-meadow, meadow-marsh, and marsh soils. The vegetation of the Zarafshan National Nature Park belongs to the tugai (riparian) plant group, and at present the park’s flora comprises 61 families, 229 genera, and 416 species of higher plants [1][2]. These plants form various formation associations, including herbaceous, shrubby, and arboreal communities, which create complex assemblages within the river valley. Tree and shrub vegetation is mainly composed of the following forest-forming species: Populus alba,Populuspruinosa,Populusdiversifolia,Populus nigra,Salixolgae,Salixturanica,Salixwilhelmsiana, Salixsongorica,Elaeagnus angustifolia,Platanusorientalis,Gleditsiatriacanthos, Robiniapseudoacacia,Ailanthus altissima,Acerplatanoides, and Tamarixpentandra, among others (Figure 1).

Field studies were conducted during the spring, summer, and autumn seasons. Mycological herbarium specimens collected during route-based field surveys were analyzed in the Laboratory of Botanical and Herbarium Research at Samarkand State University named after Sharof Rashidov. Macro- and micromorphological examinations of micromycete herbarium materials were carried out using trinocular MED D30T LCD and N-300M microscopes. The study of fungal morphological and disease symptoms, as well as species identification, was performed based on relevant scientific literature [3]-[6]. In addition, identification of the recorded pathogenic micromycetes was verified using the databases (https://www.mycobank.org, https://www.indexfungorum.org), while host plant nomenclature followed by GBIF (https://www.gbif.org/).

Field surveys were conducted during the growing seasons of 2024 and 2025 across four fixed routes covering diverse habitats within Zarafshan National Nature Park. Host plants were selected using a combination of the “random walk” method and targeted inspections of species known to be susceptible to rust fungi. A substantial sampling effort was made, totaling 250 field hours, to ensure a comprehensive inventory of the park’s Mycobiota.

According to the conducted research, 23 species of rust fungi belonging to 1 class, 1 order, 4 families, and 5 genera were recorded on higher plants in the Zarafshan National Nature Park. These fungi were found infecting 24 species of higher plants belonging to 6 families and 22 genera.

Rust fungi are obligate parasites and do not persist in nature as saprotrophs. They belong to the order Pucciniales, which comprises 28 families, approximately 214 genera, and about 9636 species (https://www.gbif.org/). In terms of host plant range and impact, Pucciniales represents one of the most important orders within the division Basidiomycota. All members of this group are plant parasites and are generally highly specialized with respect to their host plant species. They penetrate host plant tissues either through stomata or by directly piercing epidermal cells. Within the tissues, both primary and secondary mycelia grow intercellularly and form characteristic haustoria. At present, the systematics of rust fungi is based on the structure of their spores (spermatia, teliospores, aeciospores, urediniospores, and basidiospores) as well as their degree of specialization toward host plants. Within this order, the families Pucciniaceae (7,000 species), Raveneliaceae (397 species), Phragmidiaceae (321 species), Phakopsoraceae (314 species), Cloesporiaceae (204 species), Melampsoraceae (192 species), Pucciniastraceae (188 species), Uropyxidaceae (177 species), Cronartiaceae (132 species), and Gymnosporangiaceae (121 species) are dominant in terms of species richness (https://www.mycobank.org, https://www.indexfungorum.org, https://www.gbif.org/).

Table 1.Distribution of rust fungi on host plants and the degree of infection.

Twenty-three species of rust fungi belonging to the division Basidiomycota were analyzed and found to belong to the order Pucciniales and the families Pucciniaceae, Phragmidaceae, Melampsoraceae, and Gymnosporangiaceae. Based on the collected herbarium specimens, species of the genera Puccinia (13 species), Phragmidium (2 species), Melampsora (3 species), Uromyces (4 species), and Gymnosporangium (1 species) were confirmed to cause rust disease on their host plants (Table 1).

The degree of infection was assessed visually using a 4-point scale (0: absent, 1: low/scattered lesions, 2: moderate, 3: severe/extensive sori coverage). This standardized approach allowed for a rapid yet consistent estimation of pathogenicity across different host species.

PucciniaceaeChevall,PucciniaPers.

Spermogonia of Puccinia species are located on both sides of the host plant leaves, predominantly on the upper surface, beneath the epidermis; they are globose, contain paraphyses, and are colorless. Aecia occur on both sides of the host leaves, mainly on the lower surface, as well as on leaf veins and fruits. The peridium is cup-shaped or cylindrical, with margins often turned outward. Aeciospores are produced in chains, with a colorless or pigmented wall, appearing smooth or spiny. Uredinia develop on both sides of the host leaves, appearing powdery, sometimes forming concentric rings. They may be paraphysate or non-paraphysate. Urediniospores are single, borne on pedicels, with a wall that is spiny or smooth, and have germ pores. Telia are found on the leaves and stems of the host plant, appearing dense or powdery, and dark in color. Teliospores are single, borne on pedicels, typically 1 - 2 (sometimes 3 - 5) celled (16.6 - 26.4 × 19.8 - 29.7 mkm, Pucciniacousiniae). The wall is smooth or ornamented, occasionally with apical crown-like projections. Each cell contains one germ pore [3]-[6][8]-[12].

Uromyces(Link) Unger.

Spermogonia of Uromyces species are found on both sides of the host plant leaves, mainly on the upper surface, either in groups or scattered among the aecia, beneath the epidermis (16.5 - 23.31 × 16.6 - 19.9 mkm, Uromycespolygoni-avicularis). They are globose or flask-shaped, sometimes with a blunt-conical apex, pigmented, and paraphysate. Aecia develop on both sides of the host leaves, primarily on the lower surface, leaf veins, and stems, beneath the epidermis. They later appear powdery, forming clustered groups, and are pigmented. The peridium (wall) is cup-shaped or cylindrical, composed of cells of various shapes; the outer wall is usually thick, smooth, or striated, while the inner wall is thin and rough (spiny), often interrupted by teeth or fragments. Aeciospores are produced in chains, with smooth or finely spiny walls. Uredinia occur on both sides of the host leaves, mostly on the lower surface, leaf veins, and stems, developing beneath the epidermis. They later become powdery, scattered, grouped, or circular. Urediniospores are single, borne on pedicels, mixed with paraphyses or without, pigmented, with walls that are spiny or smooth, and possessing one or more germ pores. Telia are located on both sides of the host leaves, mainly on the lower surface, leaf veins, and stems, beneath the epidermis. They often penetrate deeply into the mesophyll, appearing open or closed, dense or porous, and dark in color. Teliospores are unicellular, borne singly on pedicels, pigmented, with a smooth or spiny wall, a single germ pore, and pedicels that may be colorless or pigmented, rigid or fragile [3]-[6][8]-[12].

PhragmidiaceaeCorda,PhragmidiumLink,Mag.

Spermogonia of Phragmidium species are located on both sides of the host plant leaves, mainly on the upper surface, developing beneath the cuticle. Their growth is indeterminate, and they consist of hymenia and peridia, occurring singly or in groups, small in size, and yellow in color. Aecia develop on both sides of the host leaves, primarily on the lower surface beneath the epidermis. They later appear powdery, rounded or elongated in shape, small, often merging with one another, and pigmented. They are surrounded by colorless or slightly pigmented cylindrical, bent paraphyses. Aeciospores are produced in chains or singly on pedicels, pigmented, with smooth or occasionally spiny walls. Uredinia develop on the lower surface of host leaves beneath the epidermis, later becoming powdery. They are rounded or elongated, small, and surrounded by bent, cylindrical or club-shaped paraphyses. Urediniospores occur singly, borne on pedicels, and are pigmented. In the studied specimens, urediniospores are pyriform (pear-shaped), elliptical, or subglobose in shape, measuring 19.9 - 21.7 × 15.8 - 17.6 mkm (Phragmidiumtuberculatum), with colorless or slightly pigmented spiny walls. Telia develops on the lower surface of host leaves beneath the epidermis, later appearing powdery, dark in color, with or without paraphyses. Teliospores are cylindrical, rarely club-shaped, two- or multicellular, with one, two, or more germ pores. They are dark-colored, with or without pedicels, and germinate immediately or after a dormancy period. The spore wall and pedicel have a thin, nearly colorless outer layer and a thick, pigmented, smooth or spiny inner layer. Pedicels vary in shape and size, are colorless, and are often hygroscopic, swelling in response to water [3]-[6][8]-[12].

MelampsoraceaeDietel,MelampsoraCastagne.

Spermogonia of Melampsora species are located on the upper or lower surfaces of host plant leaves, and sometimes on both sides, developing beneath the epidermis or cuticle. They are semi-globose or blunt-conical in shape, with a flat or slightly hemispherical hymenial layer. Spermatia are spherical in shape. Aecia develop on the lower surface of host leaves beneath the epidermis, are flat, cushion-shaped, and lack peridia and paraphyses. Their color ranges from dark orange-yellow to light yellow, fiery red, or yellow. Aeciospores are produced in chains, with a colorless wall, smooth or rod-like in structure. Uredinia develop on both sides of host leaves, mainly on the lower surface, beneath the epidermis, and later appear powdery. They may be without peridia or have a rudimentary peridium that is quickly lost, and are pigmented. Urediniospores occur singly, mixed with blunt or rod-shaped colorless paraphyses, and are pigmented. Their walls are colorless, smooth or spiny, sometimes with a smooth apex. In Melampsorapruinosae, urediniospores are typically spherical or elliptical in shape, measuring 23.1 - 36.3 × 19.8 - 26.4 mkm. Telia develops on both sides of host leaves, primarily on the lower surface, appearing as one- or multi-layered spots (black lesions). They occur beneath the epidermis or occasionally under the cuticle, dense, and brown to dark brown in color. Teliospores are 1(–2)-celled, germinating after a dormancy period; the wall is thin, pigmented, and smooth [3]-[6][8]-[12].

GymnosporangiaceaeChevall,GymnosporangiumR.Hedw. ex DC.

Spermogonia of Gymnosporangium species are located on the upper surface of host plant leaves, beneath the epidermis, and are globose or oval in shape. They are initially yellow, turning brown as they mature. Aecia develop on the lower surface of host leaves. Their peridia are slit almost to the base, with cells remaining connected laterally. Peridial cells are smooth or striated on the outside, and rough or finely spiny on the inside. Aeciospores are produced in chains, dark brown in color, with spiny walls. Uredinia occur on leaves or young shoots, developing beneath the epidermis and later appearing powdery. Urediniospores are single, with shape and size varying depending on internal structure. Telia develops on the annual shoots and stems of the host plant, forming fusiform (club-shaped) swellings. Teliospores are two- or multicellular, with pigmented, smooth walls, 2(–3) germ pores, and long, sturdy pedicels. In the examined specimens, teliospores are elliptical in shape, measuring 23.31 - 26.64 × 16.65 - 23.31 mkm (Gymnosporangiumconfusum) [3]-[6][8]-[12].

Figure 2. Some rust fungi distributed in the Zarafshan National Nature Park. 1: Puccinia graminis f. tritici Erikss., 2: Pucciniacousiniae P. Syd & Syd., 3: Puccinia graminis subsp.lolii W. L. Waterh., 4: Pucciniamalvacearum Bertro ex Mont., 5: Uromycespolygoni-avicularis Saccardo., 6: Pucciniataraxaci Plowr., 7: Pucciniabromina Erikss., 8: Pucciniaacroptili P. Syd & Syd.

Compared to earlier studies in the Samarkand region [3], our findings include 11 species that are newly recorded for Zarafshan National Nature Park. This update is crucial for understanding the current phytopathological status of the region’s vascular plants (Figure 2).

It was established that the 23 collected species of rust fungi occur on 24 plant species and exhibit pathogenicity. Analysis showed that 23 of these rust fungi belong to 1 division, 1 order, 4 families, and 5 genera: Melampsora (3 species), Gymnosporangium (1), Puccinia (13), Uromyces (4), and Phragmidium (2). These 22 rust fungi were found to parasitize 24 host plant species belonging to 6 families (Asteraceae, Malvaceae, Poaceae, Polygonaceae, Salicaceae, Rosaceae) and 22 genera (Rumex (1 species), Polygonum (1), Poa (1), Setaria (1), Lolium (1), Tripidium (1), Botriochloa (1), Triticum (1), Bromus (1), Calamagrostis (1), Arctium (1), Taraxacum (1), Rhaponticum (1), Populus (2), Carthamus (1), Cirsium (1), Carduus (2), Malva (1), Rosa (1), Rubus (1), Crataegus (1), Salix (1)). It was confirmed that these rust fungi are parasitic on their host plants and are capable of causing rust disease.