Clonostachys rosea is a necrotrophic mycoparasitic fungus, used for biological control of plant pathogenic fungi. A better understanding of the underlying mechanisms resulting in successful biocontrol is important for knowledge‐based improvements of the application and use of biocontrol in agricultural production systems. Transcriptomic analyses revealed that C. rosea responded with both common and specific gene expression during interactions with the fungal prey species Botrytis cinerea and Fusarium graminearum. Genes predicted to encode proteins involved in membrane transport, biosynthesis of secondary metabolites and carbohydrate‐active enzymes were induced during the mycoparasitic attack. Predicted major facilitator superfamily (MFS) transporters constituted 54% of the induced genes, and detailed phylogenetic and evolutionary analyses showed that a majority of these genes belonged to MFS gene families evolving under selection for increased paralog numbers, with predicted functions in drug resistance and transport of carbohydrates and small organic compounds. Sequence analysis of MFS transporters from family 2.A.1.3.65 identified rapidly evolving loop regions forming the entry to the transport tunnel, indicating changes in substrate specificity as a target for selection. Deletion of the MFS transporter gene mfs464 resulted in mutants with increased growth inhibitory activity against F. graminearum, providing evidence for a function in interspecific fungal interactions. In summary, we show that the mycoparasite C. rosea can distinguish between fungal prey species and modulate its transcriptomic responses accordingly. Gene expression data emphasize the importance of secondary metabolites in mycoparasitic interactions.

Secondary metabolites produced by biological control agents may influence the outcome of their interactions with plant pathogenic microorganisms and plants. In the present study, we investigated the role of the nonribosomal peptide synthetase gene nps1 expressed by the biocontrol fungus Clonostachys rosea. A gene expression analysis showed that nps1 was induced during confrontations with the plant pathogenic fungus Botrytis cinerea. Gene deletion strains of nps1 displayed increased growth rates and conidiation. However, the nematicidal activity of culture filtrates from C. rosea Δnps1 strains was significantly weaker than that from wild-type filtrates (P ≤ 0.001); after 24 h of incubation with culture filtrates from nps1 deletion strains, only 13 to 33% of a mixed community of nematodes were dead compared with 42% of nematodes incubated with wild-type culture filtrates. The Δnps1 strains also showed reduced biocontrol efficacy during pot experiments, thus failing to protect wheat seedlings from foot rot disease caused by the plant pathogenic fungus Fusarium graminearum. Furthermore, C. rosea Δnps1 strains were not able to reduce populations of plant-parasitic nematodes in soil or in roots of wheat as efficiently as the wild-type strain. Both C. rosea wild-type and Δnps1 strains increased the dry shoot weight and shoot length of wheat by 20 and 13%, respectively. We showed that NPS1, a putative nonribosomal peptide synthetase encoded by nps1, is a biocontrol factor, presumably by producing a hitherto unknown nonribosomal peptide compound with antifungal and nematicidal properties that contributes to the biocontrol properties of C. rosea.

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Biological control is a promising approach to reduce plant diseases caused by nematodes. We tested the effect of the fungus Clonostachys rosea strain IK726 inoculation on nematode community composition in a naturally nematode infested soil in a pot experiment, and the effect of C. rosea on plant health. The numbers of plant-parasitic nematode genera extracted from soil and plant roots decreased by 40 to 73% when C. rosea was applied, while genera of nonparasitic nematodes were not affected. Soil inoculation of C. rosea increased fresh shoot weight and shoot length of wheat plants by 20 and 24%, respectively, while only shoot dry weight increased by 48% in carrots. Light microscopy of in vitro C. rosea-nematode interactions did not reveal evidence of direct parasitism. However, culture filtrates of C. rosea growing in potato dextrose broth, malt extract broth and synthetic nutrient broth exhibited toxicity toward nematodes and immobilized 57, 62, and 100% of the nematodes, respectively, within 48 h. This study demonstrates that C. rosea can control plant-parasitic nematodes and thereby improve plant growth. The most likely mechanism responsible for the antagonism is antibiosis through production of nematicidal compounds, rather than direct parasitism.

Mycobacterium bovis is a pathogen of cattle. The unpasteurized milk of affected cattle is a source of infection in humans. Despite the screening of cattle and the pasteurization of milk, M bovis has not been eradicated. A high index of clinical suspicion is needed in symptomatic patients with a history of possible exposure. At risk groups include animal workers, farmers, meat packers, vets and zoo keepers. Humans are usually infected by the aerosol route. We present two cases of human bovine tuberculosis. One was a presumptive case and the second was a confirmed case. Both responded well to antituberculous therapy. In the confirmed case, there was evidence of transmission to the partner living in the same house. Rifampicin prophylaxis was given to the exposed case. The M. bovis from the confirmed case was isoniazid resistant, in addition to having the well known resistance to pyrazinamide. Isoniazid resistance has been described before in those who are immunocompromised. We describe it in an immunocompetent patient. Case reports Case 1A 50-year-old male farm worker presented to our clinic in July 2008 with a history of tightness of the chest, dyspnoea and erythema nodosum in March of the same year. His antistreptolysin O titre was elevated at that time at 545 IU ml 21. There was no history of night sweats, weight loss, cough or constitutional symptoms. In the family history, his sister had erythema nodosum during her pregnancy. He was a welder by occupation. His past medical history included childhood asthma and allergic rhinitis. On examination, his temperature was 36.5uC, his blood pressure 125/73 mmHg, pulse 66 beats min 21 and oxygen saturation was 96 % on ambient air. His cardiovascular system was normal, a respiratory examination was unremarkable and his abdomen was soft and non-tender. There was no cervical and axillary lymph node enlargement. Leg examination revealed red healing macules on the anterior aspect of both shins associated with some dry skin and scaling. There was no pitting oedema.Laboratory examination showed a normal full blood count with an erythrocyte sedimentation rate of 15 mm h A chest X-ray showed a prominent right hilum. A computed tomography (CT) scan of the thorax showed enlargement of the mediastinal lymph nodes along with nodular and linear opacification in the right upper lobe (Fig. 1).The standard Mantoux test, consisting of 2 tuberculin units Statens Serum Institute tuberculin RT23 in 0.1 ml solution injected intradermally, was strongly positive at 4 cm. Bronchoscopy results were normal. Direct microscopy with Ziehl-Neelsen staining and tuberculosis (TB) culture on sputum and bronchial washings were negative on two occasions. In view of a history of exposure to bovine TB (bTB) in the past, a presumptive diagnosis of bTB was made. The patient was commenced on antituberculous therapy (ATT). He was treated with isoniazid, rifampicin and ethambutol for 2 months, followed by isoniazid and rifampicin for a further 7 months. After 9 months of treatment the patient was asymptomatic. T...