Isolation of Arthrobotrys oligospora from soil of the Chinese Northern Tianshan Mountain slope pasture show predatory ability against Haemonchus contortus larvae

To understand the biocontrol ability of Arthrobotrys oligospora isolated from the Northern Tianshan Mountain slope pasture, Xinjiang region, China, on livestock gastrointestinal nematode diseases, we acquired 68 soil samples and isolated 26 nematode-trapping fungi using Haemonchus contortus L₃. Eight isolates were identified based on their morphological and molecular identification. The predacious activity against H. contortus was detected before and after passage through the sheep gastrointestinal tract. As a result, these eight isolates were identified as A. oligospora. They displayed predacious activities ranging from 90% to 97%. Six of the isolates could pass through the sheep gastrointestinal tract significantly reducing the number of H. contortus larvae by 76–79%. This study shows that A. oligospora isolated from the northern slope pasture of Tianshan Mountain has high predacious activity against H. contortus larvae and partly passing through the sheep gastrointestinal tract. This study also shows that conidial suspensions have no toxic side-effects on the sheep, indicating that they have the potential for the development of oral biological agents which prevent and control livestock gastrointestinal nematode diseases.     

Cloning and characterization of an extracellular serine protease from the nematode-trapping fungus <Emphasis Type="Italic">Arthrobotrys conoides</Emphasis>

An extracellular serine protease (Ac1) with a molecular mass of 35 kDa was purified from the nematode-trapping fungus Arthrobotrys conoides. The optimum activity of Ac1 is at pH 7.0 and 53.2°C (over 20 min). Ac1 can degrade a broad range of substrates including casein, gelatin, bovine serum albumin, collagen, and nematode cuticles. Moreover, the enzyme can immobilize the free-living nematode Panagrellus redivivus and the pine wood nematode Bursaphelenchus xylophilus, indicating Ac1 may be involved in infection against nematodes. The encoding gene of Ac1 contains one intron of 60-bp and two exons encoding a polypeptide of 411 amino acid residues. The deduced polypeptide sequence of Ac1 showed a high degree of similarity to two previously reported serine proteases PII and Mlx from other nematode-trapping fungi (81% aa sequence identity). However, three proteases Ac1, Aoz1 and Mlx showed optimum temperatures at 53.2, 45 and 65°C, respectively. Compared to PII, Ac1 appears to have a significantly higher activity against gelatin, bovine serum albumin, and non-denatured collagen. Moreover, our bioassay experiments showed that Ac1 is more effective at immobilizing P. redivivus than B. xylophilus.     

Predatory activity and passage of six nematophagous fungi species in gastrointestinal tract of trichostrongylide-infected sheep*

With the development of anthelmintic resistance of parasitic nematodes, screening the potential of predatory fungi candidates by in vitro and in vivo assay is necessary for biocontrol of parasitic nematodes in sheep. Fifteen native isolates of fungi species of six predators were evaluated through in vitro and in vivo experiment to evaluate the capacity of passing through the gastrointestinal tract of sheep. The results of the in vitro experiment showed that the reduction percentages of the third-stage larvae (L3) of trichostrongylides in faeces were 97.02–98.49% for five isolates of Arthrobotrys (Monacrosporium) sinense; 83.47–99.12% for three isolates of Arthrobotrys oviformis; 80.00–97.41% for four isolates of Arthrobotrys conoides; and 99.18%, 77.56%, and 75.72% for one isolate of Arthrobotrys microscaphoides, Arthrobotrys salina, and Dactylellina leptospora, respectively. In the in vivo assay, the results exhibited that the larval recoveries in faeces were significantly decreased (p?D. leptospora. After fungal treatment (within 24?h) of one isolate of A. salina, although the efficacy against L3 was 43.83%, tested fungus was detected in the faeces. The remaining isolates, except one isolate of D. leptospora and one isolate of A. conoides, were positive for either L3 reduction or fungal survival in faeces. The present study showed that nematophagous fungi could survive in the passage through the alimentary tract of sheep and should be potential candidates as a feed supplement.     

Linoleic acid — The nematicidal principle of several nematophagous fungi and its production in trap-forming submerged cultures

Linoleic acid was shown to be the only detectable nematicidal agent in the mycelial extracts of several predacious fungi of the genus Arthrobotrys. Although the compound is present in saprophytic cultures, induction of trap formation by nematodes or phenylalnyl-valine caused a significant increase in its production. In submerged cultures, the number of traps formed by Arthrobotrys conoides and Arthrobotrys oligospora was directly correlated to the increase of the concentration of linoleic acid. In A. conoides, the ratio of ergosterol to linoleic acid decreased from 2.6 in saprophytic cultures to 1.1 in trap-forming cultures induced with nematodes. Linoleic acid exhibited nematicidal activities towards the free-living nematode Caenorhabditis elegans with an LD₅₀ value of 5 g/ml.     

Establishment of Arthrobotrys flagrans as biocontrol agent against the root pathogenic nematode Xiphinema index

Plant-parasitic nematodes cause devastating agricultural damage worldwide. Only a few synthetic nematicides can be used and their application is limited in fields. Therefore, there is a need for sustainable and environment-friendly alternatives. Nematode-trapping fungi (NTF) are natural predators of nematodes. They capture and digest them with their hyphae and are starting to being used as bio-control agents. In this study, we applied the NTF Arthrobotrys flagrans (Duddingtonia flagrans) against the wine pathogenic nematode Xiphinema index. A. flagrans reduced the number of X. index juveniles in pot cultures of Ficus carica, an alternative host plant for X. index, significantly. Sodium-alginate pellets with A. flagrans spores were produced for vineyard soil inoculation under laboratory conditions. The NTF A. conoides, A. musiformis and A. superba were enriched from several soil samples, showing their natural presence. Trap formation is an energy-consuming process and depends upon various biotic and abiotic stimuli. Here, we show that bacteria of the genus Delftia, Bacillus, Pseudomonas, Enterobacter and Serratia induced trap formation in NTF like A. conoides and A. oligospora but not in A. flagrans in the absence of nematodes. The application of NTF along with such bacteria could be a combinatorial way of efficient biocontrol in nematode-infested soil.     

In vitro and in vivo activity of three sesquiterpenes against L(3) larvae of Anisakis type I

In order to investigate the possible use of terpenic derivatives to treat anisakiasis caused by L₃ larvae of Anisakis, we studied the in vitro and in vivo larvicidal activity of three sesquiterpenes (nerolidol, farnesol and elemol). In vitro experiments included the histological study of larval damage and in vivo studies the measurement of myeloperoxidase activity in rat gastrointestinal tract after administration of the sesquiterpenes. In the in vitro assays, the most active compound against the L₃ larvae was nerolidol, followed by farnesol; both caused the death of all nematodes, which showed cuticle changes and intestinal wall rupture. In the in vivo assays, only 20% of infected rats treated with nerolidol or farnesol showed gastric wall lesions in comparison to 86.6% of control animals. According to these results, nerolidol and farnesol are good candidates for further research as biocidal agents against L₃ larvae of Anisakis type I.     

Isolation,identification, and in vitro predatory activity of nematophagous fungus Arthrobotrys musiformis and its interaction with nematodes using scanning electron microscopy

We isolated and identified nematophagous fungus Arthrobotrys musiformis from materials derived from cattle and sheep. We also molecularly characterised the native fungal isolates and evaluated the nematophagous activity of the isolates. A total of 19 isolates of A. musiformis were isolated from 1532 samples, and the detection rate of A. musiformis in all samples was 1.24%. These isolates were identified using a light microscope and their 5.8S rDNA, 18S rDNA, 28S rDNA, and the internal transcribed spacers 1 and 2 region. Interaction of the isolate (NPS045) with the nematode targets of the infective larvae (L₃) of Haemonchus contortus and the free-living nematode Caenorhabditis elegans was observed by scanning electron microscopy (SEM). SEM result showed that the two species of nematodes were initially captured at 5?h after being added to the isolates. L₃ was penetrated at 22?h after capture and completely destroyed by the fungus at 68?h. C. elegans was penetrated at 14?h post-capture and was completely digested by the fungus at 24?h. In vitro experimental assay of samples in 24-well plates showed that for the three fungal isolates, the L₃s of Trichostrongylus colubriforms were reduced by 94.80%, 90.17%, and 89.02%.     

广陈皮外源真菌的组成及产毒真菌分析

[背景]广陈皮为药食同源中药材,在高温、高湿且贮存不当的条件下容易发霉,从而产生毒素,严重威胁陈皮的质量安全.[目的]分析广陈皮表面外源真菌的组成及其产生毒素的真菌.[方法]采用平板稀释法分离广陈皮表面外源真菌,利用分生孢子形态特征及DNA序列分析进行真菌鉴定,采用高效液相色谱-三重串联四极杆质谱联用技术对青霉属和曲霉...     

多粘类芽胞杆菌KM2501-1杀南方根结线虫活性产物研究

【目的】南方根结线虫(Meloidogyne incognita)是一种危害严重的土传性植物病原线虫,给农业生产造成了巨大的经济损失,前期研究发现多粘类芽胞杆菌(Panebacillus polymyxa) KM2501-1具有很好的温室防治南方根结线虫效果,且可产生多种挥发性杀线虫活性物质,但对其非挥发性产物是否有杀线虫活性没有研究。本研究拟进一步分离鉴定其产生的杀线虫活性代谢产物,发掘新的杀线虫药物。【方法】对菌株KM2501-1进行液体发酵并离心收集发酵上清液,通过硅胶柱层析、高效液相色谱分离等方法得到高纯度的杀线虫活性物质,并通过液相色谱质谱联用分析、核磁共振等技术鉴定杀线虫活性物质的结构。【结果】生物活性检测显示,多粘类芽胞杆菌KM2501-1发酵上清液具有较强的南方根结线虫触杀活性,并能有效抑制南方根结线虫卵孵化,体外杀线虫效率高达87.66%,抑制卵孵化效率达92.26%。结构鉴定结果显示多粘类芽胞杆菌产生的杀线虫活性物质为环二肽类物质cyclo (Pro-Phe),800 mg/L的cyclo(Pro-Phe)杀线虫效率达84.75%。进一步的显微观测结果表明,与对照组相比,活性物质cyclo(Pro-Phe)处理后的根结线虫肠道组织紊乱、结构发生破坏。【结论】多粘类芽胞杆菌KM2501-1产生的cyclo (Pro-Phe)是一个具有杀线虫新功能的活性物质,其可能通过破坏线虫肠道杀死线虫。     

Callus induction and thallus regeneration from callus of phycocolloid yielding seaweeds from the Indian coast

The tissue culture of phycocolloid yielding seaweeds included preparation of axenic explants, callus induction, subculture of excised callus and regeneration of plantlets from pigmented callus in the laboratory. Treatment of algal material with 0.1–0.5% detergent for 10 min and 1–2% betadine for 1–5 min and 3–5% antibiotic treatment for 48–72 h successively enabled viable axenic explants to be obtained as high as 60% for Gracilaria corticata, Sargassum tenerrimum and Turbinaria conoides and 10% for Hypnea musciformis. Callus induction was more conspicuous in T. conoides than in the other three species investigated. Of the irradiances investigated, 30 μmol photons m⁻² s⁻¹ produced calluses in as many as 40% explants in G. corticata and T. conoides and 10% in H. musciformis and S. tenerrimum. The explants cultured at 5 and 70 μmol photons m⁻² s⁻¹ did not produce any callus in all the species studied except for H. musciformis in which 10% explants developed callus at 5 μmol photons m⁻² s⁻¹. Most of the species investigated showed uniseriate filamentous Type of growths and buds from cut ends and from all over the surface of explants. Nevertheless, T. conoides had three Types of callus developments, namely (1) uniseriate filamentous Type of outgrowths from the centre of the cut end of explant, (2) bubbly Type of callus and (3) club-shaped callus clumps. The subculture of T. conoides callus embedded in 0.4% agar produced two Types of filamentous growth, namely filiform (with elongated cells) and moniliform filaments (with round cells) in the 2 months period after inoculation. Further, friable callus with loose cells was also found associated with excised callus. The moniliform filaments showed prolific growth of micro-colonies resembling to somatic embryo-like growth which, in liquid cultures, differentiated and developed into propagules with deformed shoots and distinct rhizoids. The shoots of these propagules remained stunted with abnormal leaf stalks without forming triangular shaped leaves as the parental plant and rhizoids had prolific growth in the laboratory cultures. The excised callus of G. corticata continued to grow when transferred to liquid cultures and showed differentiation of new shoots within 10 days. The shoots grew to a maximum length of 5–6 cm in the 2 months period in aerated cultures in the laboratory. Dedicated to the memory of Late Dr. Rangarajan.     

Bioactivity of intra and extracellular substances from cyanobacteria and lactic acid bacteria on “wood blue stain” fungi

Cyanobacteria (photoautotrophic prokariota) have potential for the control of pathogenic bacteria and fungi. The effect of intra and extracellular products from cyanobacterial strains on the growth of fungi isolated from “wood blue stain,” was tested. Extracellular products were obtained by concentration and sterilization of the culture medium where cyanobacteria were grown. Cyanobacterial substances promoted or inhibited fungal growth according to the fungal and cyanobacterial strains tested. Extracellular products from Nostoc muscorum 79a and the methanolic extract from Microchaete tenera 84b biomass inhibited growth of Sphaeropsis sapinea 2157 (64.7 and 775.6%, respectively). Extracellular products of Nostoc piscinale 59 and biomass methanolic extract from N. muscorum 79a produced the highest growth promotion of Trichoderma boningii 452 (105.0%) and T. viride 993 (136.7%). Extracellular products of the heterotrophic lactic acid bacterium Streptococcus termophilus were also tested and strongly inhibited (64–92%) all the fungal strains. The tested fungi have different sensitivity to the bioactive substances present in the biomass and/or the culture medium of the studied cyanobacteria and lactic acid bacterium. N. muscorum 79a, M. tenera 84b, and S. termophilus have potential to control the wood blue stain fungi by a friendly environmental alternative.     

In vitro evaluation of combination of Trichoderma harzianum and chitosan for the control of sapstain fungi

In vitro assays were undertaken to evaluate the control of two sapstain fungi, Leptographium procerum and Sphaeropsis sapinea by a combination of chitosan or chitosan oligomer and an albino strain of Trichoderma harzianum. Spore germination and hyphal growth of the test fungi were assessed on media amended with chitosan or chitosan oligomer with and without T. harzianum using either simultaneous inoculation with test fungus or inoculation 1, 2, or 3 days after pre-infection with test fungus.There was no mycelial growth of the test fungi regardless of chitosan concentrations used when either L. procerum or S. sapinea was simultaneously inoculated with T. harzianum. However, the dose–response of chitosan or chitosan oligomer on the test fungi was apparent when T. harzianum was not simultaneously inoculated with test fungus but introduced later. There was a greater growth reduction at higher concentrations (0.075–0.1% v/v) of chitosan, and overall chitosan oligomer was more effective than chitosan aqueous solution.Chitosan alone was able to restrict or delay the germination of spores but the combination of chitosan and T. harzianum inhibited spore germination and hence colony formation of test fungi regardless of time delay.     

微杆菌Sneb159杀线虫活性物质的分离与鉴定

【目的】有关Microbacterium maritypicum在植物线虫生物防治方面的研究较少,探究菌株M. maritypicum Sneb159的杀线虫活性,明确菌株发酵液中具有杀线虫活性的物质,为生物农药的开发提供理论依据。【方法】本研究检测了菌株Sneb159发酵液对大豆胞囊线虫二龄幼虫的触杀活性;并以触杀活性为追踪,采用有机溶剂萃取、硅胶柱层析及半制备高效液相色谱等技术对菌株Sneb159发酵滤液中的活性物质进行分离纯化;采用核磁共振波谱对纯化物进行结构鉴定。【结果】菌株Sneb159具有杀线虫活性,在24 h和48 h处理组中线虫的死亡率均显著高于对照组。从菌株Sneb159发酵滤液中分离得到杀线虫活性物质A6,经结构鉴定确定该物质为苯乙酰胺。【结论】首次发现M. maritypicum对大豆胞囊线虫的触杀作用,并且明确活性物质为苯乙酰胺。结果表明菌株M. maritypicum Sneb159和苯乙酰胺在大豆胞囊线虫的生物防治方面具有较好的应用潜力。     

Response of nematode-trapping fungi to organic substrates in a coastal grassland soil

To understand why Arthrobotrys oligospora and other nematode-trapping fungi are common and sometimes abundant in the coastal grassland soils of the Bodega Marine Reserve (BMR, Sonoma County, CA), we examined how resident trapping fungi responded to the addition of eight organic substrates (lupine leaves, grass leaves, dead isopods, dead moth larvae, isopod faeces, deer faeces, shrimp shells, and powdered chitin). We were especially interested in the effects of dead isopods because isopods are abundant at BMR and because previous studies had documented strong responses of A. oligospora to other arthropods (dead moth larvae). Soil from BMR was packed into vials (40 g dry mass equivalent per vial with water potential at −230 kPa and bulk density at 0.9 g cm⁻³), and one substrate or no substrate was added to the soil surface. After 30 d at 20 °C, trapping fungi were quantified by dilution plating and most probable number procedures. The response of A. oligospora was inversely related to substrate carbon:nitrogen (C:N) ratio: substrates with low C:N ratios (dead isopods, lupine leaves, dead moth larvae) usually caused large increases in A. oligospora whereas those with higher C:N ratios (isopod faeces, deer faeces, grass leaves) did not. An exception was chitin powder, which had a low C:N ratio, but which did not cause A. oligospora to proliferate. Responses of A. oligospora were directly related to the quantity of nitrogen added with each substrate, and those substrates that caused large increases in resident nematodes usually caused large increases in A. oligospora. Other trapping fungi did not respond as strongly as A. oligospora.     

Preventive and therapeutic administration of an indigenous <Emphasis Type="Italic">Lactobacillus</Emphasis> sp. strain against <Emphasis Type="Italic">Proteus mirabilis</Emphasis> ascending urinary tract infection in a mouse model

Probiotics are increasingly being considered as non-pharmaceutical and safe potential alternatives for the treatment and prevention of a variety of pathologies including urinary tract infections. These are the most common infections in medical practice and are frequently treated with antibiotics, which have generated an intense selective pressure over bacterial populations. Proteus mirabilis is a common cause of urinary tract infections in catheterised patients and people with abnormalities of the urinary tract. In this work we isolated, identified and characterised an indigenous Lactobacillus murinus strain (LbO2) from the vaginal tract of a female mouse. In vitro characterisation of LbO2 included acid and bile salts tolerance, growth in urine, adherence to uroepithelial cells and in vitro antimicrobial activity. The selected strain showed interesting properties, suitable for its use as a probiotic. The ability of LbO2 to prevent and even treat ascending P. mirabilis urinary tract infection was assessed using an experimental model in the mouse. Kidney and bladder P. mirabilis counts were significantly lower in mice preventively treated with the probiotic than in non-treated mice. When LbO2 was used for therapeutic treatment, bladder counts of treated mice were significantly lower although no significant differences were detected in P. mirabilis kidney colonisation of treated and non-treated animals. These results are encouraging and prompt further research related to probiotic strains and the basis of their effects for their use in human and animal health.     

Induction of secretory aspartyl proteinase of <Emphasis Type="Italic">Candida albicans </Emphasis>by HIV-1 but not HSV-2 or some other microorganisms associated with vaginal environment

The most common type of candidiasis involves mucosal sites such as the oral cavity, the gastrointestinal tract and the vagina. Among many of virulence factors, the production of secretory aspartyl proteinase (Sap) by Candida albicans (C. albicans) has gained much attention, and factors leading to Sap induction are thus under intense study. The aim of this study was to examine whether some microorganisms such as Lactobacillus, Gardnerella vaginalis (G. vaginalis), human immunodeficiency virus type-1 (HIV-1) and human herpes simplex virus type-2 (HSV-2) had any Sap inducing effect on C. albicans. Here we showed that among the microorganisms tested in vitro only HIV-1 induced Sap production from C. albicans.     

In vivo Testing of Functional Properties of Three Selected Probiotic Strains

Summary Lactobacillus acidophilus M92, Lactobacillus plantarum L4 and Enterococcus faecium L3 were previously selected as probiotic strains on the base of in vitro selection criteria. To investigate functional properties of these three probiotic strains in vivo, Swiss albino mice were used as animal model. Survival, competition, adhesion and colonization were monitored in the gastrointestinal tract, as well as the immunomodulating capability of L. acidophilus M92, L. plantarum L4 and E. faecium L3. During the feeding of mice with probiotic strains with daily dose of 2 × 10¹⁰ rifampicin-resistant cells, the number of lactic acid bacteria in the faeces increased and reduction of enterobacteria and sulphite-reducing clostridia was observed. Rifampicin-resistant colonies of probiotic strains could be reisolated from the faeces of mice fed with the rifampicin-resistant cells. The similar results were obtained in homogenates of small and large intestine of mice on the first and fourteenth days after feeding with L. acidophilus M92, L. plantarum L4 and E. faecium L3. The adherence of the probiotic strains obtained in vitro correlated with their capability to adhere to mouse ileal epithelial cells in vivo. After oral immunization of mice with viable cells of L. acidophilus M92, L. plantarum L4 and E. faecium L3 with a daily dose of 2 × 10¹⁰ cells, the concentrations of serum IgA, IgG and IgM antibodies from all groups of mice were significantly higher in comparison to the control.     

Antagonism between entomopathogenic fungi and bacterial symbionts of entomopathogenic nematodes

Mutual effects between the symbiotic bacteria of entomopathogenic nematodes, Photorhabdus luminescens and Xenorhabdus poinarii, and entomopathogenic fungi were investigated in vitro. A dual culture assay on nutrient agar supplemented with bromothymol blue and triphenyltetrazolium chloride (NBTA) medium revealed that P. luminescens is antagonistic to Metarhizium anisopliae, Beauveria bassiana, B. brongniartii and Paecilomyces fumosoroseus by inhibiting their growth and conidial production; the fungal growth was not inhibited by X. poinarii. In a second laboratory experiment, crude extract produced by M. anisopliae was tested for its activity against P. luminescens and X. poinarii. Crude extract from M. anisopliae was antibacterial to P. luminescens and X. poinarii at 1000 g/ml and inhibited their growth on NBTA, but had no effect at 100 or 10 g/ml. The influence of the crude extract of M. anisopliae on the dispersal of infective juveniles (IJs) of Heterorhabditis megidis and Steinernema glaseri was assayed on Sabouraud Dextrose Agar (SDA) plates. Results showed that the crude extract of M. anisopliae had no toxic effects even at highest concentration (1000 g/ml).     

Synergies between biological and neonicotinoid insecticides for the curative control of the white grubs Amphimallon majale and Popillia japonica

Synergistic combinations of biological and chemical insecticides might yield promising alternatives for soil insect pest management. In turfgrass of the Northeast U.S., control of root-feeding scarab larvae is highly dependent on conventional insecticides. Studies on interactions between entomopathogenic nematodes and neonicotinoid insecticides, however, demonstrate the feasibility of synergies as an approach for reduced-risk curative control. To understand the breadth of potential synergies, we screened numerous combinations of biological control agents with sublethal doses of neonicotinoids against third instars. Interactions were characterized as synergistic, additive or antagonistic. The most promising combinations identified in laboratory bioassays were advanced to greenhouse pot studies and then to field trials featuring microplots with artificially infested populations. To reveal variation across scarab species, trials were conducted on Amphimallon majale and Popillia japonica. Synergies were consistent across trials and specific to white grub species. For A. majale, synergistic combinations of Heterorhabditis bacteriophora with imidacloprid and clothianidin were discernible in laboratory, greenhouse and field trials. For P. japonica, synergistic combinations of Beauveria bassiana and Metarhizium anisopliae with both neonicotinoids were discernible in the laboratory and greenhouse, but not in the field. For both species, antagonistic interactions were discernible between Bt-products and both neonicotinoids. While nematode-neonicotinoid synergies among scarab larvae have been examined before, fungi-neonicotinoid synergies are unreported. In the context of previous studies, however, no patterns emerge to explain variation across target species or control agent. Further study of non-additive interactions will guide how biological and chemical products could be combined to enhance soil insect pest management.     

New fungal genera, Tectonidula gen. nov. for Calosphaeria-like fungi with holoblastic-denticulate conidiogenesis and Natantiella gen. nov. for three species segregated from Ceratostomella

Two morphologically similar groups of ascomycetes with globose to subglobose perithecia, elongate necks, unitunicate asci floating freely at maturity, and hyaline ascospores currently placed in Calosphaeria s. lat. and Ceratostomella s. lat., respectively, are studied. The Calosphaeria-like fungi have groups of perithecia growing between cortex and wood, arranged in circular groups with converging necks and piercing the cortex in a common point; the asci with a shallow apical ring and U- to horseshoe-shaped hyaline ascospores are compared with Calosphaeria pulchella, the type species of the genus. Conidiogenesis of the investigated Calosphaeria-like fungi is holoblastic-denticulate; ramichloridium-like and sporothrix-like conidiophores and conidia were formed in vitro. Ascospore and ascus morphology, structure of the ascal apex, ascogenous system, mode of conidiogenesis and the large subunit rRNA sequences of this group differ considerably from C. pulchella and both groups are unrelated. Thus a new genus, Tectonidula, is described with two accepted species, T. hippocrepida and T. fagi; they are separated by ascospore and ascus morphology and holoblastic-denticulate conidiogenesis from the core species of Calosphaeria. The placement of Tectonidula among perithecial ascomycetes is discussed. The relationship of Tectonidula with Barbatosphaeria and two ramichloridium-like hyphomycetous genera Rhodoveronaea and Myrmecridium is investigated. Three species formerly attributed to Ceratostomella are studied. The revision of the herbarium type specimen and fresh material of Ceratostomella ligneola revealed that it is conspecific with Ceratostomella ampullasca and Ceratostomella similis. The LSU phylogeny clearly separated C. ligneola from Ceratostomella s. str. and morphologically similar Lentomitella. On the basis of molecular sequence data and detailed comparison of morphology of asci, ascospores and ascogenous system the genus Natantiella is described for C. ligneola with C. ampullasca and C. similis as its synonyms. Natantiella produced sterile mycelium in vitro.