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7 March 2023

Annotation of the 12th Chromosome of the Forest Pathogen Fusarium circinatum

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1
Museum and Institute of Zoology, Polish Academy of Science, ul. Wilcza 64, 00-679 Warszawa, Poland
2
Institute for Sustainable Plant Protection (IPSP), National Research Council of Italy (CNR), Strada delle Cacce 73, 10135 Torino, Italy
3
Department of Entomology, Phytopathology and Molecular Diagnostics, Faculty of Agriculture and Forestry, University of Warmia and Mazury in Olsztyn, Pl. Łódzki 5, 10-727 Olsztyn, Poland
4
Faculty of Physics, Warsaw University of Technology, ul. Koszykowa 75, 00-662 Warszawa, Poland
Agronomy2023, 13(3), 773;https://doi.org/10.3390/agronomy13030773
This article belongs to the Special Issue Genetics and Molecular Biology of Pathogens in Agricultural Crops
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Abstract

The genus Fusarium comprises more than 300 species, and many of them are pathogens that cause severe diseases in agricultural, horticultural and forestry plants in both antropogenic and natural ecosystems. Because of their importance as plant pathogens, the genomes of several Fusarium spp. have been sequenced. Within this genus, Fusarium circinatum is one of the most harmful pathogens of pine trees attacking up to 60 Pinus species. Till now, the genomes of 13 strains of F. circinatum have been sequenced. The strain GL1327 we studied lacks a twelfth chromosome, which allows the study of virulence genes on this chromosome. Although the genome of several strains of F. circinatum has been sequenced, it is still almost completely unannotated, which severely limits the possibilities to further investigate the molecular mechanisms of virulence of Fusarium. Therefore, this study aimed to annotate the 12th chromosome of F. circinatum and integrate currently available resources. In silico annotation of the 12th chromosome of F. circinatum revealed the presence of 118 open reading frames (ORFs) encoding 141 proteins which were predicted using an ab initio gene prediction tool. The InterProScan and SMART analyses identified known domains in 30 proteins and eggNOG additionally in 12 of them. Among them, four groups can be distinguished: genes possibly related to heterokaryon incompatibility (4 genes), regulation of transcription (5 genes), plant cell wall degrading enzymes (7 genes) and trichothecene synthesis (3 genes). This study also integrated data of F.circinatum reference strain CMWF1803 assembled to chromosome level but not annotated with currently best annotated but assembled only to scaffold level strain NRRL 25331.

1. Introduction

Fusarium is a cosmopolitan genus of filamentous ascomycetes (Sordariomycetes: Hypocreales: Nectriaceae) that includes many toxin-producing plant pathogens of agricultural importance. The genus Fusarium includes over 300 phylogenetically distinct species [1]. Many of these species are plant pathogens that cause serious diseases on agricultural, horticultural and forestry plants in antropogenic and natural ecosystems [2].
Pine Pitch Canker Disease (PPC), a serious threat that attacks many pine species, is caused by the pathogenic fungus Fusarium circinatum Nirenberg & O’Donnell (teleomorph = Gibberella circinata). F. circinatum belongs to the EPPO A2 quarantine pathogen and causes one of the most devastating diseases in pine forests, afforestations and nurseries, not only in Europe but throughout the world [3]. The host range of F. circinatum is very broad and includes up to 60 Pinus species [4,5].
Although the whole genome sequence of about 44 Mb of F. circinatum has been determined [6], our knowledge of the fungal genes involved in its pathogenic behaviour is limited. Seven putative quantitative trait loci associated with mycelial growth and colony margins have been described [7]. Van Wyk et al. [8] discovered a locus that possibly determines the growth rate near the telomere of chromosome 3. The sequence of this locus is highly conserved in F. circinatum and its close relatives, except for a 12,000 bp insertion encoding five genes. An in silico analysis of the F. circinatum genome identified five candidate genes related to the growth (Fcfga1, Fcfgb1, Fcac, Fcrho1, and FcpacC) [9]. Functional studies of Fcrho1 deletion mutants, a Rho-type GTPase, showed significantly reduced growth in vitro than the corresponding ectopic and wild-type strains. The knockout mutant of Ras2, another gene encoding the GTPase, also produced significantly smaller lesions compared to the complementation mutants and wild-type strains. Growth studies showed also significantly smaller colonies and delayed germination of conidia in the knockout mutant strain [10].
Currently, the genomes of 13 strains of F. circinatum have been sequenced [11], providing a solid basis for comparative genomics. One of them (GL 1327) lacks a 12th chromosome [12], which provides favourable conditions for the identification and study of genes determining the virulence of F. circinatum.
The aim of this study was to identify ORFs localised on the 12th chromosome and subsequently characterise the encoded proteins.

2. Materials and Methods

Analysis In Silico

The sequence of the 12th chromosome (Assembly ASM2404739v1, Acc. No. CM043929.1) from the representative genome of F. circinatum Mexican strain CMWF1803 from Pinus patula [11] was retrieved from NCBI. Gene prediction was performed with the programme AUGUSTUS version 3.3.1 trained for F. graminearum with the ab initio gene prediction method [13,14].
The predicted protein sequences were analysed against the protein database NCBI-NR using BLASTp (default identity ≥ 40%, coverage ≥ 40%). Functional analysis of predicted protein sequences was performed using InterProScan 91.0 [15,16] against the integrated InterPro database consisting of PRINTS, SMART, Pfam, SUPERFAMILY, CATH -Gene3D, PANTHER and CDD databases [17], Simple Modular Architecture Research Tool (SMART v.9) [18] and against the unsupervised orthologous group database EggNOG v6.0 [19].

3. Results

In Silico Characterisation of Putative F. circinatum Genes

The sequence of the 12th chromosome was processed for ab initio gene prediction using AUGUSTUS. A total of 118 putative genes were predicted, of which 56 are located on the plus strand and 62 on the minus strand (Table A1 in Appendix A). These genes can be transcribed into 141 transcripts. Thirty-six transcripts were intronless, while 41, 28, 10, 8, 7, 3, 2 and 1 transcripts have one, two, three, four, five, seven, either six or eight, and nine introns, respectively.
Six genes (g1, g19, g20, g31, g56 and g76) had two alternative transcription start sites, and the transcripts of 21 genes had no alternative splicing. The putative genes correspond to five scaffolds of F. circinatum strain NRRL 25331 (PRJNA565749): JAAQPE010000042.1-142267 bp; JAAQPE010000057.1-51020 bp; JAAQPE010000172.1-29236 bp; and JAAQPE010000262.1-90105 bp (Figure 1). The identity of the four scaffolds to the CM043929 sequence ranged from 99.09% to 99.81%, with only scaffold JAAQPE010000057.1 having 93.44%.
Figure 1. Localization of the scaffolds of F.circinatum strain NRRL 25331 on the 12th chromosome of the representative genome of Fusarium circinatum strain CMWF1803. Black bar—reference sequence of F.circinatum 12th chromosome CMWF1803 strain with coordinates of the sequence. Blue letters—names of F.circinatum strain NRRL 25331 scaffolds. Grey/red boxes—scaffolds of F.circinatum strain NRRL 25331. Insertions are marked with blue two hourglass-like triangles.
The predicted protein sequences were compared with sequences deposited in GenBank using BLASTp. Out of 141 queries, 130 sequences were highly identical (>90%) to F. circinatum strain NRRL 25331, seven proteins had lower identity (g99t2, 76.34%; g115t1, 81.17%; g16t1, 81.33%; G99t1, 84.22%; G92t2, 86.52%; G87t2, 87.94%; g78t1, 87.97%) and four (g48t1; g59t1; g74t1; G98t1) were not identical to F. circinatum but identical to other Fusarium species.
With the help of InterProScan and SMART, domains and protein architectures could be identified for 30 proteins (Table A3 in Appendix A). Among them, four groups can be distinguished: Genes possibly related to heterokaryon incompatibility, regulation of transcription, plant cell wall degrading enzymes and trichothecene synthesis.
Among the genes predicted by Augustus, four genes were found to be related to heterokaryon incompatibility (Figure 2). The protein g57t1 has two domains: Heterokaryon incompatibility (HET) and protein kinases (S-TKc). This protein sequence is identical to KAF5666823.1 (100% search coverage, 100% identity) of F. circinatum strain NRRL 25331, but is only referred to as serine-threonine kinase in GenBank. Four proteins (g26t1, g52t1 and g58t1 and g58t2) contain a NACHT nucleoside triphosphatase domain (named after the NAIP, CIITA, HET-E and TP-1 proteins) flanked by a varying number of ankyrin repeat domains. The alternative splicing of the g58 transcript has no effect on the protein architecture. In the g26t1 and g52t1 proteins, the nucleoside phosphorylase domain (PNP-UDP-1) is located proximal to NACHT; in addition, g52 contains the domain oxoglutarate/iron-dependent dioxygenase (2OG-FeII-Oxy).
Figure 2. Architecture of proteins potentially involved in heterokaryon incompatibility. Black rectangles—protein domains related to heterokaryon incompatibility. Green rectangles—ankyrin domain repeats. Blue polygon—protein kinases domain.
The incompatibility reaction is associated with massive transcriptional reprogramming. Four genes (g20, g55, g82 and g83) encoding putative transcription factors were found on the 12th chromosome. The protein g20t1 contains a Jumonji domain (JmjC). G82 contains a transcription factor domain specific for fungi (Fungal-trans). This protein sequence is identical to KAF5673552.1 (100% search coverage, 100% identity) of F. circinatum strain NRRL 25331, where it is designated cutinase transcription factor 1 alpha. The G55 transcript is subject to alternative splicing, but this does not affect the protein architecture. Both proteins (g55t1 and g55t2) have two domains: Fungal-trans and GAL4. The G83 protein (Table A4 in Appendix A) has a fungal binuclear Zn(2)-Cys(6) domain. In addition to the putative transcription factors, the g103t1 protein contains a SET domain typical of proteins involved in epigenetic regulation of gene expression.
During infection, Fusarium secretes various virulence factors, including effector proteins and plant cell wall degrading enzymes (CDWEs). The proteins encoded by g7 and g102 contain a lipase GDSL-2 domain. This domain is typical of SGNH hydrolase-type esterases that act as esterases and lipases. The carboxylesterase domain (COesterase) contains the proteins g81t1 and g82t2. Another type of hydrolases-peptidases-encode the genes g18 and g89. The protein g18t1 contains peptidase C1A, g98t1 and g89t2 peptidase C14 or caspase domain. In addition to hydrolases, there are two genes (g66 and g105) on the 12th chromosome that encode proteins containing a domain of the Major Facilitator Superfamily (MSF1).
One of the features of Fusarium is toxin synthesis. Three putative genes for trichothecene synthesis have been detected. G6t1 and g6t2 contain tyrosinase, g95t1-p450 and g70t1-three acyl-CoA domains. In addition to groups of putative genes related to heterokaryon incompatibility, transcriptional regulation, plant cell wall degrading enzymes and trichothecene synthesis, 13 proteins were found whose products may be involved in many metabolic processes of Fusarium metabolic processes. Another 12 proteins were annotated with eggNOG (Table A4 in Appendix A).

4. Discussion

4.1. Distribution of F. circinatum, the Causative Agent of PPC Disease

As mentioned in the introduction, the fungal pathogen F. circinatum is the causative agent of PPC disease [20]. Does the presence of the 12th chromosome cause the pathogen to severely attack a variety of pine species in forests and nurseries worldwide? The fungus can damage seedlings in nurseries and mature trees in forests. Symptoms in seedlings include wilting and in mature trees bleeding, resinous cankers on trunks or thick branches and tree death [20]. As F. circinatum has already been detected in Europe, it is considered a serious, potentially invasive forest pathogen that spreads via infected seeds, seedlings, wood, soil, wind, insect vectors and human activities. In Europe, the fungus has affected pine trees in northern Spain and Portugal and has also been detected in France and Italy. Research on the fungus (including its chromosomes) can therefore contribute to the understanding of its pathogenesis and thus to the development of an appropriate protection strategy. This should apply to young seedlings as well as to adult trees. Despite the economic importance of PPC disease, the worldwide distribution of the pathogen F. circinatum is poorly documented and the pathogenicity of its strains is even less known. It is likely that the genetic diversity and population structure of the pathogen influence the spread of PPC, including in Europe (models for the likely spread of the disease), and the susceptibility of hosts. Chromosome number could be important for the virulence of F. circinatum, which also depends on host species, tree age and environmental characteristics. Knowledge of the above factors is crucial for disease management, containment and mitigation strategies. The in silico analyses carried out should help countries that are currently free of F. circinatum to put in place effective procedures and restrictions and prevent the invasion of the pathogen.

4.2. Development of New Diagnostic Methods to Ensure Reduction of PPC

Fusarium circinatum is on the list of species recommended for regulation as a quarantine pest in Europe. More than 60 species of Pinus are susceptible to this pathogen, and it also attacks Douglas-fir (Pseudotsuga menziesii (Mirb.) Franco) and species from genera such as Picea and Larix. The European Food Safety Authority (EFSA) estimates the probability of reintroduction into the EU as very high [21]. Thanks to the possibility of early detection, continuous surveillance and inspections by quarantine services, outbreaks of F. circinatum in Italy and France have been officially eradicated. However, the global spread of F. circinatum suggests that the pathogen will continue to be found in new areas in the future. Rapid identification of the most virulent strains of the pathogen (using knowledge of their chromosomes) will be important in Europe and elsewhere to limit the spread of the disease. Currently, morphological identification methods are being replaced by molecular methods, which include conventional PCR with a specific target region in the intergenic interval and various real-time PCR protocols with varying specificity and sensitivity [22]. Perhaps the search for chromosome 12 will also be useful for pest risk assessment.

4.3. Pathways of Transmission and Potential Host Risk of the Pathogen

As F. circinatum is the causal agent of one of the most devastating forest diseases worldwide, its spread over long distances should be controlled, especially by monitoring infected seeds. On the other hand, at the regional level, seedlings, substrates and containers play an important role in the spread of the fungus [21]. The pathogen enters nurseries via infected seeds and is further spread by planting infected plants, especially since infected plants (asymptomatic) may appear without disease symptoms. Once established, F. circinatum is spread by rain, wind and insects. Natural spread of the pathogen is limited due to the short spore dispersal distances and the relatively short flight distances of the spreading insects. To understand how best to intervene in the development of the disease in nurseries and forests, we conducted annotation of twelfth chromosome.

4.4. Risk of Establishing the Pathogen in New Regions in Europe

Pine trees as potential host plants are important components of native forests and plantations in Europe, where they play an important role both economically and ecologically. Pine diseases are mainly caused by fungal pathogens and can significantly affect the survival, vigour and yield of both individual trees and entire stands or plantations. PPC caused by F. circinatum, one of the most devastating pine diseases in the world, is an example of a new invasive disease in Europe.
The susceptibility of Scots pines in Poland (Pinus sylvestris L.) to infection by F. circinatum was tested in a greenhouse trial by [23]. Sixteen Polish pine cultivars were artificially inoculated with the 12th chromosome of F. circinatum and six other Fusarium species known to infect pine seedlings in nurseries. All pines were found to be highly susceptible to PPC and showed varying degrees of susceptibility to the other Fusarium species tested. The results suggest that the risk of establishment of the invasive pathogen F. circinatum may be high as a result of its accidental introduction in Poland.
In the future, the fungus is more likely to spread in the pine forests of southern Europe, but there is also the possibility of spread in central and northern Europe. In Lithuania, no occurrence of F. circinatum has been reported so far. In 2018, the susceptibility of three different native Lithuanian Pinus sylvestris provenances to this pathogen [24] was tested. For each origin, 38 pines were used and the soil was inoculated with a suspension of F. circinatum, and DNA was extracted from several plants that appeared unhealthy four weeks after soil inoculation. Using the real-time PCR method, F. circinatum could not be detected in these samples. However, the reason could be that the fungal biomass was too low in relation to the host biomass or that the strain was less pathogenic than others.

4.5. Possible Interactions between F. circinatum and Other Fungal Species

The impact of microbiome interactions on plant health and the possible role of the plant microbiome in disease expression have been the subject of several recent studies [24]. In Lithuania, the interaction between 12th chromosome strain of F. circinatum and several pine-inhabiting fungi such as Dothistroma septosporum, F. oxysporum and Lecanosticta acicola was also verified [24]. It was found that F. oxysporum grows slightly faster than F. circinatum and inhibits the growth rate of F. circinatum. D. septosporum produced dothistromin, which also appeared to slow the growth of the F. circinatum culture. In the meantime, L. acicola was displaced by F. circinatum.
Co-infection of trees with indigenous pathogenic fungi or alien oomycetes and F. circinatum is possible. Biotic interactions could play an important role in the establishment of the PPC pathogen in European nurseries and forests [25]. Available information on pine pathogens that may co-occur with F. circinatum in Europe will have an impact on pine survival and growth. Early and accurate identification of F. circinatum, a recently introduced pathogen currently being regulated in Europe, is crucial to prevent its introduction and spread in forests. Chromosome studies could provide valuable information in this regard if it is confirmed that the high pathogenicity of some strains of the fungus depends on them and others do not.

4.6. In Silico Approach to the Identification and Characterisation of Genes

In this study, we used a genome-based in silico approach to identify and characterise genes located on the 12th chromosome of F. circinatum. Chromosome 12 has been shown to be the smallest of the chromosomes found in species of the F. fujikuroi complex. The size of these chromosomes varies considerably intra- and interspecifically and shows polymorphism in chromosome length compared to the other chromosomes [26].
Fungal cells can interact with each other either vegetatively or sexually. In ascomycete fungi, sexual interactions are controlled by the alleles at the mating type locus (MAT) and asexual interactions by the alleles at the loci vic (vegetative incompatibility) or het (heterokaryon incompatibility) [27]. In members of the F. fujikuroi species complex, 8 to 10 vic loci have been identified [28]. Vegetative incompatibility leads to programmed cell death. For programmed cell death associated with vegetative incompatibility, there are important proteins containing HET [29] and NACHT [30] domains. In F. circinatum we have found four putative proteins that contain a central NACHT domain. Two of them have an N-terminal PNP-UDP effector domain and all three have a C-terminal ANK repeat domain (Figure 2). This organisation is typical of Ascomycota, where 20% of proteins with NACH domains have N-terminal PNP-UDP and 42% have C-terminal ANK repeats [30]. Proteins containing the NACHT domain are involved in a process of non-self- recognition and programmed cell death of fungi called heterokaryon incompatibility [31,32].
The incompatibility response has been found to be associated with massive transcriptional reprogramming [33]. Transcription factors (TFs) play a key role in regulating gene expression by binding to DNA in a sequence-specific manner. TFs are usually classified according to their DNA-binding motif. Representatives of 80 TF families are typically found in fungal genomes. The largest of these is the zinc cluster (C6 zinc finger) family [34]. They play an important role in growth, development and pathogenicity [35,36,37]. Fusarium transcription factor 1 (FTF1) has been described as a potential regulator of effector expression in F. oxysporum f. sp. phaseoli and F. oxysporum f. sp. lycopersici [38]. Mahanty et al. [39] described that specialised C6-type TFs may act as major regulators of F. oxysporum f. sp. cepae pathogenicity during the development of Fusarium basal rot in onions.
Proteins with a zinc finger domain were found in the g24t1, g55t1 and g55t2 proteins. SMART identified a transcription factor specific to fungi in the g82t1 domain, while BLAST found sequence identity with KAF5673552.1, which was annotated as cutinase transcription factor 1 (CTF1). CTF1 belongs to the C6 zinc TFs. CTF regulates the expression of cutinases and fatty acid metabolism genes in F. solani f. sp. pisi [40] and Aspergillus nidulans [41]. Disruption of Ctf1 α eliminated the phytopathogenicity of F. solani [40]. F. oxysporum strains lacking a functional copy of the CTF1 gene are impaired in the induction of cutinase activity and in the expression of genes encoding cutinase and lipase [42].
Gene expression also depends on the methylation of histones. Acetylation of lysine (K) residues in histone 3 (H3) is associated with active transcription, while methylation of lysine or arginine (R) residues leads to a more complex outcome that depends on associated reader proteins [43]. H3K4 and H3K36 are considered to be hallmarks of euchromatin in yeast and higher eukaryotes [44]. In filamentous fungi, the picture appears to be more diverse, as data showed the ubiquitous presence of the H3K36 trimethylation mark (me3) in F. fujikuroi and F. graminearum [45,46]. Methylation of H3K4 has been shown to depend on the conserved SET domain-containing methyltransferase Set1 [47,48]. While Set1 is responsible for H3K4 methylation in the fungus, jumonji C is responsible for demethylation [49]. On Chr12 of F. circinatum we have discovered putative genes encoding both of these proteins: G103t1 contains the SET domain and g20t1 jumonji.
During the infection process, Fusarium uses a number of secretion systems and releases a variety of virulence factors such as mycotoxins, effector proteins and CWDEs to overcome the target host cells. CWDEs such as polygalacturonases, pectate lyases, xylanases, peptidases, peptide hydrolases, ribonucleases and cutinases may contribute to pathogenesis by degrading waxes, cuticles and cell walls to promote tissue invasion and pathogen spread [50,51]. Cutinases and lipases that catalyse the hydrolysis of ester bonds from fatty acid polymers, facilitating fungal invasion through the cuticle. Disruption of the lipase gene FGL1 in F. graminearum resulted in reduced extracellular lipolytic activity in culture and reduced virulence in both wheat and maize [52]. Disruption of another lipase gene, FgATG15, also greatly attenuated wheat head infection [53]. An active role of lipases in establishing full virulence has also recently been suggested for the plant pathogen F. oxysporum f. sp. lycopersici, where reduced lipolytic activity due to deletion of lipase regulatory genes resulted in reduced colonisation of tomato plants [54].
The secreted metalloprotease FoMep1 and the serine protease FoSep1 of F. oxysporum are involved in full virulence against tomato because they can reduce the antifungal activity of their host plant chitinases [55]. The FoAYP1 gene also encodes protease. Surprisingly, this protease is secreted by F. oxysporum but is localised in the nucleus in plant cells. The knockout strain of the FoAYP1 gene showed reduced virulence against tomato plants, but its mycelial growth and conidiation were unchanged [56]. The Major Facilitator Superfamily (MFS) is one of the largest known membrane transporter families. MFS transporters are currently the best characterised superfamily of secondary transmembrane transport proteins responsible for nutrient uptake, extrusion of metabolites and resistance to various toxic compounds, including not only secondary metabolites but also fungicides and antibiotics. On the other hand, MFS transporters play a role in the availability of nutrients for survival, including the transport of lipids, ions and small metabolites [57]. The transcript abundance of the MFS multidrug transporter was five times higher in pathogenic F. oxysporum than in non-pathogenic F. oxysporum. This transporter family regulates the movement of sugars, Krebs cycle metabolites, phosphorylated glycolytic intermediates, amino acids, peptides, osmoliths, iron siderophores, nucleosides, and organic and inorganic anions and cations [58]. In addition, MFS transporters have been linked to fungal pathogenicity by avoiding toxic compounds produced by the pathogen or protecting against plant defences [59]. On Chr12, not only the putative CTF gene but also putative genes encoding proteins containing lipase (g71, g102), COesterase (g81), peptidase (g18 and g89) and domains of MFS (g66 and g105) are localised.
Species of the genus Fusarium produce a wide variety of agriculturally important trichothecene toxins, which differ from each other in their pattern of oxygenation and esterification. Trichothecenes are a structurally diverse family of fungal sesquiterpene epoxides that cause mycotoxicosis in humans and animals and increase the virulence of some Fusarium species on crops. In F. sporotrichioides and F. graminearum, trichothecene biosynthetic genes are localised in a 40-kb gene cluster [60,61]. Genes in this cluster include trichodiene synthetase, P450 oxygenase, acetyltransferase, a toxin efflux pump and transcription factors containing a Cys2His2 zinc finger motif [62]. On Chr12, genes involved in trichothecene synthesis were found - a putative tyrosinase gene (g6), acyl-CoA dehydrogenase (g70) and p450 cytochrome oxidase (g95), but not organised in a cluster.
The gene g2 encodes a protein containing a GPI-anchored domain found at the N-terminus of a group of cell wall synthesis proteins involved in the synthesis of beta-1,6-glucan in the cell wall [63]. The cell wall shapes and protects the fungal cell. The 1,3-beta-glucan synthase is responsible for the synthesis of one of the main components of the fungal wall. This enzyme has been described in F. solani and many other Fusarium species [64]. Many attempts to delete the gene encoding this enzyme have been unsuccessful, suggesting that it may be a gene essential for cell life [65].
A comparison of the expression of serine/threonine protein kinase genes (ste12) in pathogenic and non-pathogenic strains of F. oxysporum f. sp. cubense showed a significant increase in the expression of ste12 in pathogenic strains [66]. Deletion of FgPTC1, a serine/threonine phosphatase, also attenuated the virulence of F. graminearum on wheat [67]. The mutant of F. verticillioides in which the fpk1 gene encoding the cAMP-dependent protein kinase was disrupted showed reduced vegetative growth, fewer and shorter aerial mycelia, severely impaired conidiation and reduced spore germination rate. After germination, the fresh hyphae were stout and unbranched. When inoculated into susceptible maize varieties, infection of the delta fpk1 mutant was delayed and infection efficiency was reduced compared to the wild-type strain [68]. Family of serine/threonine protein kinases and plays an important role in yeasts and other filamentous fungi. Deletion of FoIme2, which belongs to this family, in F. oxysporum reduced mycelial growth and conidia production. The mutants were hypersensitive to the osmotic stressor NaCl but less sensitive to the membrane stressor SDS. Deletion of FoIme2 also reduced pathogenicity [69]. The gene encoding the protein kinase (g56) is located on Chr 12.
In filamentous fungi, gene silencing by RNA interference (RNAi) affects many biological processes, including pathogenicity. Deletion of qde3, which encodes helicase, impaired conidiation and ascosporogenesis in F. graminaceum and contributes to sexual reproduction [70]. Chr. 12 contains the gene g74, which encodes a protein containing a helicase domain.

5. Conclusions

  • Overall, the knowledge gained in this study about the annotations of genes, ORFs and domains in the 12th chromosome of F. circinatum could make an important contribution to the management of PPC disease and to strategies for containment and mitigation strategies.
  • Our study can serve to clarify the phylogeny of the species and furthermore to develop new molecular detection tools.
  • The genomic organisation of virulence genes can be used to clarify the relationship between F. ciricantum and hosts.
  • We concluded that at least 14 genes are associated with pathogenesis/virulence.

Author Contributions

Conceptualization, T.M., S.M., A.O.; methodology, T.M., S.M., A.O.; software, T.M., P.B.; validation, T.M., P.B.; formal analysis, T.M., P.B.; investigation, T.M.; resources, A.O., T.M.; data curation, P.B.; writing—original draft preparation, T.M., T.O.; writing—review and editing, T.O., P.B., S.M.; visualization, P.B., T.M.; supervision, T.M., T.O.; project administration, T.M., T.O.; funding acquisition, A.O. All authors have read and agreed to the published version of the manuscript.

Funding

The publication was written as a part of result of the author’s (AO) internship in Slovak University of Agriculture in Nitra, co-financed by the European Union under the European Social Fund (Operational Program Knowledge Education Development), carried out in the project Development Program at the University of Warmia and Mazury in Olsztyn (POWR.03.05. 00-00-Z310/17).

Institutional Review Board Statement

Not applicable.

Data Availability Statement

Not applicable.

Conflicts of Interest

The authors declare no conflict of interest.

Appendix A. Localization and Annotation of Putative F. circinatum Proteins

Table A1. Localization of putative Open Reading Frames and transcripts, protein sizes, and number of introns derived from the F. circinatum 12th chromosome annotation.
Table A1. Localization of putative Open Reading Frames and transcripts, protein sizes, and number of introns derived from the F. circinatum 12th chromosome annotation.
GeneStrandLocalizationNo.
Introns		TranscriptLocalizationPredicted
Protein Size
1+31217–319602131217–31219196
+ 2231229–31960192
2+33294–340891 33294–34089238
3−36737–369891 36737–3698968
4+38485–392640 38485–39264259
5+40947–415010 40947–41501184
6−42288–447937142288–44793687
7242288–44793687
7+47832–503433 47832–50343712
8+50528–517720 50528–51772414
9−53269–536160 53269–53616115
10−53692–545010 53692–54501269
11−58288–595171 58288–59517393
12−67276–676740 67276–67674132
13−69092–695500 69092–69550152
14−71147–714300 71147–7143075
15+71985–725211 71985–72521136
16−76693–776991 76693–77699300
17+81870–838760 81870–83876668
18−83977–858270 83977–85827618
19−87569–882592 87569–88259194
20+89834–922870 89834–92287817
21+95681–963921195681–96392219
+ 1295777–96392187
22−101909–10304941101973–103049148
52101909–103049195
23+104774–1055652 104774–105565223
24+108785–1103710 108785–110371528
25+115888–1165352 115888–116535146
26−117728–1220842 117728–1220841379
27−130442–1316144 130442–131614323
28+133748–13495331133982–134953271
62133748–134953297
29−138883–1392270 138883–139227114
30−143831–1456375 143831–145637441
31+145727–14885111 145727–148851475
32+156385–1571221 156385–157122228
33−157739–1582840 157739–158284181
34−165252–1665410 165252–166541429
35−167760–1691282 167760–169128311
36+171554–17227111171554–172271178
2171578–172271170
37+173661–1748522 173661–174852364
38+177631–1794033 177631–179403368
39−179756–18072721179756–180727285
22179756–180727288
40+180965–1819191 180965–181919209
41−181947–1826032 181947–182603185
42−184433–18640331184433–186403436
42184433–186403461
43−197699–1985201 197699–198520131
44+201599–2022130 201599–202213204
45−206352–2067921 206352–206792114
46−212670–2131171 212670–213117117
47−217694–2182961 217694–218296134
48−219886–2204121 219886–220412153
49+241829–2430644 241829–243064292
50+249462–2499121 249462–249912132
51−255019–2554301 255019–255430117
52−260511–2676874 260511–2676871871
53+267833–2695060 267833–269506557
54+269924–2706100 269924–270610228
55−270814–27250341270814–272503442
42270814–272503433
56−279820–2819242 279820–281924636
57+284917–2889005 284917–2889001103
58+290740–29451281290740–294512993
82290740–2907421008
59−296603–2984231 296603–298423465
60−299021–3003370 299021–300337436
61+301149–3015350 301149–301535128
62+306378–3097893 306378–309789987
63+312081–3131330 312081–313133350
64−314421–3147181 314421–31471882
65−316363–3173842 316363–317384304
66+319615–32135511319636–321355550
12319615–321355557
67−321897–3227630 321897–322763288
68+324749–3259703 324749–325970339
69+327367–3281190 327367–328119250
70−328511–3297010 328511–329701396
71−333730–3345270 333730–334527165
72−335535–3364441 335535–336444141
73+339435–34012611339435–340042186
22339435–340126196
74−340698–3411472 340698–341147116
75−344219–3474772 344219–3474771042
76+349423–35051921349423–350519331
12349423–350519347
77−352248–3528791 352248–352879118
78+353875–3547071 353875–354707235
79+358678–3624235 358678–3624231046
80−364509–3655991 364509–365599346
81−368775–37223251368775–372232660
52368775–372232634
82−373134–3747751 373134–374775527
83+375893–3772341 375893–377234428
84−377746–3798692 377746–379869638
85+381383–3820240 381383–382024213
86+384422–38749531386043–387495418
02384422–385309295
53384422–387495754
87+389607–39303161389607–393031550
72389607–393031531
88+393067–3934110 393067–393411114
89+396581–39737811396650–397378167
12396581–397378190
90−399885–4012072 399885–401207504
91−403346–4038070 403346–403807153
92−405413–40578721405413–40578792
12405413–40573891
93−412668–4139391 412668–413939407
94+418703–4194152 418703–419415201
95−419654–4202680 419654–420268204
96−423279–4240952 423279–424095240
97−432389–4331830 432389–433183264
98−437147–4382631 437147–438263327
99−441782–44297411441782–442974379
22441782–442974349
100−444928–4465362 444928–446536502
101+447456–4488023 447456–448802392
102+451345–4520970 451345–452097250
103+459258–4610382 459258–461038552
104+462490–4628971 462490–462897102
105−465130–4687299 465130–468729888
106+476676–4780901 476676–478090448
107−481803–4820060 481803–48200667
108−486871–4873491 486871–487349141
109−490202–4927724 490202–492772528
110+490202–4927721 490202–492772156
111+497012–4981510 497012–498151379
112+499716–5001980 499716–500198160
113−502582–5043601 502582–504360508
114+506239–50744431506239–507444174
22506239–507444161
115+508757–50967721508757–509677198
32508757–509677239
116+511677–5129030 511677–512903408
117−513379–5143112 513379–514311137
118+515695–5166071 515695–516607249
Table A2. GenBank best matches of putative F. circinatum proteins.
Table A2. GenBank best matches of putative F. circinatum proteins.
ProteinLength (AA)Best Match IDAccession No. (Best Match)E ValueIdentity (%)
g1t1196Hypothetical protein FCIRC_1200 F. circinatumKAF5689737.1 5 × 10 − 129 100.00
g1t2192Hypothetical protein FCIRC_1200 F. circinatumXP_049150203.1 4 × 10 − 129 100.00
g2t1238Cell wall beta-glucan synthesis, FCIRC_1201, F. circinatumKAF5689738.1 1 × 10 − 170 100.00
g3t168Hypothetical protein FCIRC_1202, F. circinatumKAF5689739.1 3 × 10 − 40 100.00
g4t1258Hypothetical protein, FCIRC_1203 F. circinatumKAF5689740.10.0100.00
g5t1184Hypothetical protein FCIRC_1204 F. circinatumKAF5689741.1 9 × 10 − 135 100.00
g6t1687Tyrosinase precursor, FCIRC_1205, F. circinatumKAF5689742.10.0100.00
g6t2687Tyrosinase precursor, FCIRC_1205, F. circinatumKAF5689742.10.098.84
g7t1712Extracellular gdsl-like lipase, FCIRC_1206, F. circinatumKAF5689743.10.0100.00
g8t1414Hypothetical protein FCIRC_1207 F. circinatumKAF5689744.10.0100.00
g9t1115Hypothetical protein FCIRC_1208, F. circinatumKAF5689745.1 1 × 10 − 75 100.00
g10t1269Hypothetical protein FCIRC_1209, F. circinatumKAF5689746.10.0100.00
g11t1393Hypothetical protein FCIRC_1210, F. circinatumKAF5689747.10.0100.00
g12t1132Hypothetical protein FCIRC_1211, F. circinatumKAF5689748.1 2 × 10 − 90 100.00
g13t1152Hypothetical protein FCIRC_1212, F. circinatumKAF5689749.1 2 × 10 − 107 100.00
g14t175Hypothetical protein FCIRC_1213, F. circinatumKAF5689750.1 3 × 10 − 47 100.00
g15t1136Hypothetical protein FCIRC_1214, F. circinatumKAF5689751.1 1 × 10 − 94 100.00
g16t1300Serine threonine kinase, FCIRC_1215, F. circinatumKAF5689752.1 7 × 10 − 167 81.33
g17t1668Hypothetical protein CIRC_1216, F. circinatumKAF5689753.10.0100.00
g18t1616Hypothetical protein CIRC_1217, F. circinatumKAF5689754.10.0100.00
g19t1194Hypothetical protein CIRC_1218, F. circinatumKAF5689755.1 2 × 10 − 136 100.00
g20t1817Transcription factor jumonji, FCIRC_1219, F. circinatumKAF5689756.10.0100.00
g21t1219Hypothetical protein CIRC_1220, F. circinatumKAF5689757.1 6 × 10 − 154 92.80
g21t2187Hypothetical protein FCIRC_1220, F. circinatumKAF5689757.1 5 × 10 − 130 91.67
g22t1148Hypothetical protein FCIRC_1222, F. circinatumKAF5689759.1 2 × 10 − 88 99.23
g22t2195Hypothetical protein FCIRC_1222, F. circinatumKAF5228515.1 1 × 10 − 129 93.33
g23t1223Hypothetical protein CIRC_1223, F. circinatumKAF5689760.1 1 × 10 − 165 100.00
g24t1528Hypothetical protein CIRC_1224, F. circinatumKAF5689761.10.0100.00
g25t1146Hypothetical protein CIRC_1225, F. circinatumKAF5689762.1 5 × 10 − 102 100.00
g26t11379Ankyrin repeat protein, FCIRC_1226, F. circinatumKAF5689763.10.0100.00
g27t1323Hypothetical protein CIRC_1227, F. circinatumKAF5689764.10.0100.00
g28t1271Hypothetical protein FCIRC_1228, F. circinatumKAF5689765.10.0100.00
g28t2297Hypothetical protein FCIRC_1228, F. circinatumKAF5689765.10.0100.00
g29t1114Hypothetical protein FCIRC_1229, F. circinatumKAF5689766.1 3 × 10 − 77 100.00
g30t1441Hypothetical protein FCIRC_1230, F. circinatumKAF5689767.10.0100.00
g31t1475Translation initiation factor IF-2, FCIRC_1231, F. circinatumKAF5689768.1 4 × 10 − 179 96.51
g32t1228Hypothetical protein FCIRC_1232, F. circinatumKAF5689769.1 1 × 10 − 158 100.00
g33t1181Hypothetical protein FCIRC_1233, F. circinatumKAF5689770.1 2 × 10 − 131 100.00
g34t1429Hypothetical protein FCIRC_1234, F. circinatumKAF5689771.10.0100.00
g35t1311C2H2 transcription factor, FCIRC_1235, F. circinatumKAF5689772.10.0100.00
g36t1178Hypothetical protein FCIRC_1237, F. circinatumKAF5689774.1 1 × 10 − 126 100.00
g36t2170Hypothetical protein FCIRC_1237, F. circinatumKAF5689774.1 9 × 10 − 120 100.00
g37t1364Hypothetical protein FCIRC_1729, F. circinatumKAF5688719.10.0100.00
g38t1368Hypothetical protein FCIRC_1731, F. circinatumKAF5688721.10.0100.00
g39t1285Hypothetical protein FCIRC_1732, F. circinatumKAF5688722.10.0100.00
g39t2288Hypothetical protein FCIRC_1732, F. circinatumKAF5688722.10.098.61
g40t1209Hypothetical protein FCIRC_1733, F. circinatumKAF5688723.1 1 × 10 − 150 100.00
g41t1185Hypothetical protein FCIRC_1734, F. circinatumKAF5688724.1 2 × 10 − 135 100.00
g42t1436Hypothetical protein FCIRC_1735, F. circinatumKAF5688725.10.099.74
g42t2461Hypothetical protein FCIRC_1735, F. circinatumKAF5688725.10.099.74
g43t1131FK506-binding protein, FCIRC_1236, F. circinatumKAF5688726.1 8 × 10 − 93 100.00
g44t1204Hypothetical protein FCIRC_1737, F. circinatumKAF5688727.1 2 × 10 − 144 100.00
g45t1114Hypothetical protein FCIRC_1738 F. circinatumKAF5688728.1 5 × 10 − 78 100.00
g46t1117Hypothetical protein FCIRC_1739 F. circinatumKAF5688729.1 7 × 10 − 77 100.00
g47t1134Hypothetical protein FCIRC_1740, F. circinatumKAF5688730.1 1 × 10 − 92 100.00
g48t1153Arginine deiminase type-3, F. mexicanumKAF5555127.1 1 × 10 − 95 95.83
g49t1292Hypothetical protein FCIRC_10050, F. circinatumKAF5666814.10.0100.00
g50t1132Hypothetical protein FCIRC_10051, F. circinatumKAF5666815.1 1 × 10 − 92 100.00
g51t1117Sterol 3beta-glucosyltransferase, FCIRC_10052, F. circinatumKAF5666816.1 2 × 10 − 60 100.00
g52t11871NACHT ankyrin domain-containing protein, FCIRC_10053, F. circinatumKAF5666817.10.0100.00
g53t1557NCS1 nucleoside transporter, FCIRC_10054, F. circinatumKAF5666818.10.0100.00
g54t1228Asp glu hydantoin racemase, FCIRC_10055, F. circinatumKAF5666819.1 6 × 10 − 163 100.00
g55t1442C6 transcription factor, FCIRC_10056, F. circinatumKAF5666820.10.097.96
g55t2433C6 transcription factor, FCIRC_10056, F. circinatumKAF5666820.10.0100.00
g56t1638CMGC DYRK kinase, FCIRC_10058, F. circinatumKAF5666822.10.0100.00
g57t11103Serine threonine kinase, FCIRC_10059, F. circinatumKAF5666823.10.0100.00
g58t1993NACHT domain-containing protein, FCIRC_5226, F. circinatumKAF5682015.10.099.59
g58t21008NACHT domain-containing protein, FCIRC_5226, F. circinatumKAF5682015.10.092.73
g59t1465TPR domain-containing protein, F. denticulatumKAF5674688.10.055.26
g60t1438TPR domain-containing protein, FCIRC_5228, F. circinatumKAF5682016.10.0100.00
g61t1128Hypothetical protein FCIRC_5229, F. circinatumKAF5682017.1 2 × 10 − 88 100.00
g62t1987Hypothetical protein FCIRC_5230, F. circinatumKAF5682018.10.092.40
g63t1350Hypothetical protein FCIRC_5231, F. circinatumKAF5682019.10.098.86
g64t182Hypothetical protein FCIRC_5232, F. circinatumKAF5682020.1 5 × 10 − 53 100.00
g65t1304Aspartate aminotransferase, FCIRC_5233, F. circinatumKAF5682021.1 1 × 10 − 136 100.00
g66t1550Multidrug resistance protein fnx1, FCIRC_8030, F. circinatumKAF5673567.10.0100.00
g66t2557Multidrug resistance protein fnx1, FCIRC_8030, F. circinatumKAF5673567.10.0100.00
g67t1288D-isomer specific 2-hydroxyacid dehydrogenase, FCIRC_8029, F. circinatumKAF5673566.10.0100.00
g68t1339Macrophomate synthase, FCIRC_8028, F. circinatumKAF5673565.10.0100.00
g69t1250Demethylmenaquinone methyltransferase family, FCIRC_8027, F. circinatumKAF5673564.10.0100.00
g70t1396Hypothetical protein FCIRC_8026 F. circinatumKAF5673563.10.0100.00
g71t1165Hypothetical protein FCIRC_8025, F. circinatumKAF5673562.1 1 × 10 − 119 100.00
g72t1141Hypothetical protein FCIRC_8024, F. circinatumKAF5673561.1 2 × 10 − 98 100.00
g73t1186Hypothetical protein FCIRC_8023, F. circinatumKAF5673560.1 3 × 10 − 133 100.00
g73t2196Hypothetical protein FCIRC_8023, F. circinatumKAF5673560.1 4 × 10 − 124 100.00
g74t1116SNF2 family domain containing protein, F. agapanthiKAF4497424.1 6 × 10 − 65 92.24
g75t11042Hypothetical protein FCIRC_8022, F. circinatumKAF5673559.10.0100.00
g76t1331Hypothetical protein FCIRC_8021, F. circinatumKAF5673558.10.0100.00
g76t2347Hypothetical protein FCIRC_8021, F. circinatumKAF5673558.10.095.39
g77t1118Hypothetical protein FCIRC_8020, F. circinatumKAF5673557.1 5 × 10 − 82 100.00
g78t1235Kinase-like domain-containing protein, FCIRC_8019, F. circinatumKAF5673556.1 9 × 10 − 167 87.97
g79t11046Hypothetical protein FCIRC_8018, F. circinatumKAF5673555.10.0100.00
g80t1346Hypothetical protein FCIRC_8017, F. circinatumKAF5673554.10.0100.00
g81t1660Para-nitrobenzyl esterase, FCIRC_8016, F. circinatumKAF5673553.10.0100.00
g81t2634Para-nitrobenzyl esterase, FCIRC_8016, F. circinatumKAF5673553.10.096.06
g82t1527Cutinase transcription factor 1 alpha, FCIRC_8015, F. circinatumKAF5673552.10.0100.00
g83t1428Hypothetical protein FCIRC_8014, F. circinatumKAF5673551.10.0100.00
g84t1638Hypothetical protein FCIRC_8013, F. circinatumKAF5673550.10.0100.00
g85t1213Hypothetical protein FCIRC_8012, F. circinatumKAF5673549.10.0100.00
g86t1418Hypothetical protein FCIRC_8010, F. circinatumKAF5673547.10.0100.00
g86t2295Hypothetical protein FCIRC_8011, F. circinatumKAF5673548.10.097.97
g86t3754SGL domain-containing protein, Fusarium sp. LHS14.1KAI8724150.1 2 × 10 − 95 52.38
g87t1550ATP synthase F1, FCIRC_8009, F. circinatumKAF5673546.10.091.65
g87t2531ATP synthase F1, FCIRC_8009, F. circinatumKAF5673546.10.087.94
g88t1114Hypothetical protein FCIRC_8008, F. circinatumKAF5673545.1 8 × 10 − 77 100.00
g89t1167Caspase, FCIRC_8007, F. circinatumKAF5673544.1 1 × 10 − 121 100.00
g89t2190Caspase, FCIRC_8007, F. circinatumKAF5673544.1 2 × 10 − 121 100.00
g90t1405Hypothetical protein FCIRC_8006, F. circinatumKAF5673543.10.0100.00
g91t1153Hypothetical protein FCIRC_8005, F. circinatumKAF5673542.1 1 × 10 − 105 100.00
g92t192Hypothetical protein FCIRC_8004, F. circinatumKAF5673541.1 8 × 10 − 60 100.00
g92t291Hypothetical protein FCIRC_8004, F. circinatumKAF5673541.1 3 × 10 − 46 86.52
g93t1407Transaldolase, FCIRC_7317, F. circinatumKAF5675701.10.0100.00
g94t1201Aromatic prenyltransferase, FCIRC_7316, F. circinatumKAF5675700.1 2 × 10 − 142 100.00
g95t1204Cytochrome P450 monooxygenase, FCIRC_7315, F. circinatumKAF5675699.1 2 × 10 − 150 100.00
g96t1240Nonribosomal peptide synthase, FCIRC_7314, F. circinatumKAF5675698.1 8 × 10 − 176 100.00
g97t1264Hypothetical protein FCIRC_7313, F. circinatumKAF5675697.10.0100.00
g98t1327Uncharacterized protein FSUBG_13770, F. subglutinansXP_036530762.1 7 × 10 − 158 64.23
G99t1379Hypothetical protein FCIRC_7311, F. circinatumKAF5675696.10.084.22
g99t2349Hypothetical protein FCIRC_7311, F. circinatumKAF5675696.10.076.34
g100t1502Rhs repeat-associated core domain-containing protein, FCIRC_7310, F. circinatumKAF5675695.10.0100.00
g101t1392Hypothetical protein FCIRC_7309, F. circinatumKAF5675694.10.0100.00
g102t1250Esterase SGNH hydrolase-type subgroup, FCIRC_7308,
F. circinatum		KAF5675693.10.0100.00
g103t1552SET domain-containing protein, FCIRC_7305, F. circinatumKAF5675692.10.0100.00
g104t1102Hypothetical protein FCIRC_7306, F. circinatumKAF5675691.1 3 × 10 − 67 100.00
g105t1888Major facilitator superfamily transporter, FCIRC_7305
F. circinatum		KAF5675690.10.0100.00
g106t1448Hypothetical protein FCIRC_7304, F. circinatumKAF5675689.10.095.12
g107t167Hypothetical protein FCIRC_7303, F. circinatumKAF5675688.1 4 × 10 − 39 100.00
g108t1141Hypothetical protein FCIRC_7302, F. circinatumKAF5675687.1 2 × 10 − 95 100.00
g109t1528Polyketide synthase FCIRC_7301, F. circinatumKAF5675686.10.0100.00
g110t1156Taurine dioxygenase family FCIRC_7300, F. circinatumKAF5675685.1 2 × 10 − 110 100.00
g111t1379Hypothetical protein FCIRC_7299, F. circinatumKAF5675684.10.0100.00
g112t1160Hypothetical protein FCIRC_7298, F. circinatumKAF5675683.1 6 × 10 − 116 100.00
g113t1508Hypothetical protein FCIRC_7297, F. circinatumKAF5675682.10.0100.00
g114t1174Kinase-like (PK-like), FCIRC_7296 F. circinatumKAF5675681.1 8 × 10 − 88 100.00
g114t2161Kinase-like (PK-like), FCIRC_7296 F. circinatumKAF5675681.1 7 × 10 − 88 100.00
g115t1198Hypothetical protein FCIRC_7295, F. circinatumKAF5675680.1 4 × 10 − 132 81.17
g115t2239Hypothetical protein FCIRC_7295, F. circinatumKAF5675680.1 5 × 10 − 178 100.00
g116t1408Alpha beta-hydrolase, FCIRC_7294, F. circinatumKAF5675679.10.0100.00
g117t1137Hypothetical protein FCIRC_7293, F. circinatumKAF5675678.1 8 × 10 − 96 100.00
g118t1249Telomere-associated recQ-like helicase, FCIRC_7292, F. circinatumKAF5675677.10.0100.00
Table A3. Protein domains identified by InterPro scan and SMART in putative F. circinatum proteins.
Table A3. Protein domains identified by InterPro scan and SMART in putative F. circinatum proteins.
GenePfam Acc. No.InterPro Acc. No.Domain NameDomain Name
Abbreviation		Localization (AA)E Value
g2t1PF10342IPR018466Kre9/KNH-like N-terminal Ig-like domainGPI-anchored29–123 2.6 × 10 − 17
g6t1PF00264IPR002227Tyrosinase copper-bindingTyrosinase60–358 5.9 × 10 − 39
g6t2PF00264IPR002227Tyrosinase copper-bindingTyrosinase60–359 5.9 × 10 − 39
g7t1PF13472IPR013830SGNH hydrolase-type esteraseLipase_GDSL_2174–343 2.8 × 10 − 12
g18t1PF00112IPR000668Peptidase C1A, papain C-terminalPeptidase_C1434–607 4.9 × 10 − 7
g20t1-IPR003347JumonjiJmjC339–498 2.04 × 10 − 5
g26t1PF01048IPR000845Nucleoside phosphorylasePNP_UDP _142–358 7.5 × 10 − 12
g26t1PF05729IPR000845NACHT nucleoside triphosphataseNACHT407–589 7.4 × 10 − 7
g26t1-IPR002110Ankyrin repeatANK15 rpt. from 854 to 1374
g52t1PF03171IPR005123Oxoglutarate/iron-dependent dioxygenase2OG-FeII_Oxy49–158 2.8 × 10 − 17
g52t1PF01048IPR000845Nucleoside phosphorylasePNP_UDP _1175–472 3.3 × 10 − 7
g52t1PF05729IPR007111NACHT nucleoside triphosphataseNACHT575–742 1 × 10 − 6
g53t1PF02133IPR001248Purine-cytosine permeaseTransp_cyt_pur30–487 2.3 × 10 − 86
g54t1PF01177IPR015942Asp/Glu/hydantoin racemaseAsp_Glu_race42–219 4 × 10 − 10
g55t1PF04082IPR007219Transcription factor, fungiFungal_trans131–318 6 × 10 − 6
g55t1-IPR001138Zn(2)-C6 fungal-type DNA-bindingGAL46–54 4.19 × 10 − 4
g55t2PF04082IPR007219Transcription factor, fungiFungal_trans130–318 5.7 × 10 − 6
g55t2-IPR001138Zn(2)-C6 fungal-type DNA-bindingGAL46–54 4.19 × 10 − 4
g56t1-IPR000719Protein kinaseS_TKc29–359 9.33 × 10 − 9
g57t1-PR000719Protein kinaseS_TKc167–4660.142
g57t1PF06985IPR010730Heterokaryon incompatibilityHET693–821 5.4 × 10 − 7
g58t1PF05729IPR007111NACHT nucleoside triphosphataseNACHT291–469 9.7 × 10 − 8
g58t1-IPR002110Ankyrin repeatsANK6 rpt. from 711 to 963
g58t2PF05729IPR007111NACHT nucleoside triphosphataseNACHT291–469 9.9 × 10 − 8
g58t2-IPR002110Ankyrin repeatsANK6 rpt. from 726 to 978
p65t1PF00155IPR004839Aminotransferase, class I/class IIAminotran_1_227–208 1.8 × 10 − 25
p66t1PF07690IPR011701Major facilitator superfamilyMSF161–459 2 × 10 − 45
p66t2PF07690IPR011701Major facilitator superfamilyMSF186–466 2.1 × 10 − 45
p67t1PF02826IPR006140D-isomer specific 2-hydroxyacid dehydrogenase2-Hacid_dh_C60–256 1.1 × 10 − 46
p68t1PF03328IPR005000HpcH/HpaI aldolase/citrate lyaseHpcH_HpaI37–257 5.9 × 10 − 25
g70t1PF02771IPR013786Acyl-CoA dehydrogenase/oxidase, N-terminalAcyl-CoA_dh_N5–117 9.2 × 10 − 18
g70t1PF02770IPR006091Acyl-CoA dehydrogenase/oxidase, middleAcyl-CoA_dh_M121–222 6.9 × 10 − 18
g70t1PF00441IPR009075Acyl-CoA dehydrogenase/oxidase, C-terminalAcyl-CoA234–390 2.7 × 10 − 32
g74t1PF00271IPR001650Helicase, C-terminalhelicase_C14–116 1.8 × 10 − 8
g79t1-IPR002110Ankyrin repeatsANK9 rpt. from 275 to 275
g81t1PF00135IPR00201Carboxylesterase, type BCOesterase33–488 1.1 × 10 − 93
g81t2PF00135IPR00201Carboxylesterase, type BCOesterase33–364 4.9 × 10 − 83
g82t1-IPR007219Transcription factor, fungiFungal_trans242–314 1.16 × 10 − 6
g89t1PF00656IPR011600Peptidase C14, caspaseCaspase40–158 7 × 10 − 9
g89t2PF00656IPR011600Peptidase C14, caspaseCaspase63-181 1 × 10 − 8
g93t1PF03702IPR005338Anhydro-N-acetylmuramic acid kinaseAnmK2–382 1.9 × 10 − 60
g94t1PF11991IPR017795Aromatic prenyltransferase, DMATS-typeTrp_DMAT1–193 1.1 × 10 − 27
g95t1PF00067IPR001128Cytochrome P450p45024–182 6.1 × 10 − 15
g96t1PF00501IPR000873AMP-dependent synthetase/ligaseAMP-binding4–83 9.5 × 10 − 8
g100t1PF03534IPR003284Salmonella virulence plasmid proteinSpvB47–240 1.1 × 10 − 51
g102t1PF13472IPR013830SGNH hydrolase-type esteraseLipase_GDSL_222–239 1.5 × 10 − 11
g103t1-IPR001214SET domainSET7 –155 8.47 × 10 − 12
g105t1PF07690IPR011701Major facilitator superfamilyMSF138–416 3.5 × 10 − 38
g109t1PF00109IPR014030Beta-ketoacyl synthase, N-terminalKetoacyl-synt1–76 3.2 × 10 − 10
g109t1PF00109IPR014030Beta-ketoacyl synthase, N-terminalKetoacyl-synt72–166 4.2 × 10 − 23
g109t1PF02801IPR014031Beta-ketoacyl synthase, C-terminalKetoacyl-synt_C174–274 5.3 × 10 − 22
Table A4. Putative proteins of F. circinatum annotated by eggNOG.
Table A4. Putative proteins of F. circinatum annotated by eggNOG.
GeneeggNOGDescription
g9t1arCOG00379trimeric autotransporter adhesin
g10t17KF05fibrous sheath CABYR-binding protein
g24t1BKZCKZnF_C2H2
g27t15K2KNsjoegren syndrome nuclear autoantigen 1
g47t15J4GBanthrone oxygenases
g48t1KOG1724S-phase kinase-associated protein 1
g59t1KOG4626protein O-GlcNAc transferase
g60t1KOG4626protein O-GlcNAc transferase
g75t1KOG1546nicotinamide-nucleotide amidase
g81t17NBP8abhydrolase_1 alpha/beta hydrolase
g81t27NBP8abhydrolase_1 alpha/beta hydrolase
g101t17K74Hcupin domain

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