Fungal habitats include soil, water, and extreme environments. With around 100,000 fungus species already described, it is estimated that 5.1 million fungus species exist on our planet, making fungi one of the largest and most diverse kingdoms of eukaryotes. Fungi show remarkable metabolic features due to a sophisticated genomic network and are important for the production of biotechnological compounds that greatly impact our society in many ways. In this review, we present the current state of knowledge on fungal biodiversity, with special emphasis on filamentous fungi and the most recent discoveries in the field of identification and production of biotechnological compounds. More than 250 fungus species have been studied to produce these biotechnological compounds. This review focuses on three of the branches generally accepted in biotechnological applications, which have been identified by a color code: red, green, and white for pharmaceutical, agricultural, and industrial biotechnology, respectively. We also discuss future prospects for the use of filamentous fungi in biotechnology application.

BackgroundHeavy metal Resistance-Nodulation-Division (HME-RND) efflux systems help Gram-negative bacteria to keep the intracellular homeostasis under high metal concentrations. These proteins constitute the cytoplasmic membrane channel of the tripartite RND transport systems. Caulobacter crescentus NA1000 possess two HME-RND proteins, and the aim of this work was to determine their involvement in the response to cadmium, zinc, cobalt and nickel, and to analyze the phylogenetic distribution and characteristic signatures of orthologs of these two proteins.ResultsExpression assays of the czrCBA operon showed significant induction in the presence of cadmium and zinc, and moderate induction by cobalt and nickel. The nczCBA operon is highly induced in the presence of nickel and cobalt, moderately induced by zinc and not induced by cadmium. Analysis of the resistance phenotype of mutant strains showed that the ΔczrA strain is highly sensitive to cadmium, zinc and cobalt, but resistant to nickel. The ΔnczA strain and the double mutant strain showed reduced growth in the presence of all metals tested. Phylogenetic analysis of the C. crescentus HME-RND proteins showed that CzrA-like proteins, in contrast to those similar to NczA, are almost exclusively found in the Alphaproteobacteria group, and the characteristic protein signatures of each group were highlighted.ConclusionsThe czrCBA efflux system is involved mainly in response to cadmium and zinc with a secondary role in response to cobalt. The nczCBA efflux system is involved mainly in response to nickel and cobalt, with a secondary role in response to cadmium and zinc. CzrA belongs to the HME2 subfamily, which is almost exclusively found in the Alphaproteobacteria group, as shown by phylogenetic analysis. NczA belongs to the HME1 subfamily which is more widespread among diverse Proteobacteria groups. Each of these subfamilies present distinctive amino acid signatures.

Background Fluoroquinolones such as ciprofloxacin induce the mutagenic SOS response and increase the levels of intracellular reactive oxygen species (ROS). Both the SOS response and ROS increase bacterial mutagenesis, fuelling the emergence of resistant mutants during antibiotic treatment. Recently, there has been growing interest in developing new drugs able to diminish the mutagenic effect of antibiotics by modulating ROS production and the SOS response. Objectives To test whether physiological concentrations of N-acetylcysteine, a clinically safe antioxidant drug currently used in human therapy, is able to reduce ROS production, SOS induction and mutagenesis in ciprofloxacin-treated bacteria without affecting antibiotic activity. Methods The Escherichia coli strain IBDS1 and its isogenic mutant deprived of SOS mutagenesis (TLS−) were treated with different concentrations of ciprofloxacin, N-acetylcysteine or both drugs in combination. Relevant parameters such as MICs, growth rates, ROS production, SOS induction, filamentation and antibiotic-induced mutation rates were evaluated. Results Treatment with N-acetylcysteine reduced intracellular ROS levels (by ∼40%), as well as SOS induction (by up to 75%) and bacterial filamentation caused by subinhibitory concentrations of ciprofloxacin, without affecting ciprofloxacin antibacterial activity. Remarkably, N-acetylcysteine completely abolished SOS-mediated mutagenesis. Conclusions Collectively, our data strongly support the notion that ROS are a key factor in antibiotic-induced SOS mutagenesis and open the possibility of using N-acetylcysteine in combination with antibiotic therapy to hinder the development of antibiotic resistance.

Resistance to antibiotics is a global health problem. Activation of the SOS response, and the subsequent elevation in mutagenesis, contributes to the appearance of resistance mutations. Among currently used drugs, quinolones are the most potent inducers of the SOS response. In the present study, we show that amikacin inhibits ciprofloxacin-mediated SOS induction and mutagenesis in Pseudomonas aeruginosa.KEYWORDS Pseudomonas aeruginosa, SOS response, ciprofloxacin, recA A ntibiotics may cause genetic changes involving different pathways, and one of them is the induction of error-prone polymerases mediated by SOS response (1, 2). Ciprofloxacin (CIP), one of the antimicrobials of choice for the treatment of Pseudomonas aeruginosa, induces the SOS response (3-5) by interfering with gyrase or topoisomerase activity (6, 7). The SOS regulon controls 15 genes, including imuABC and dinB (4, 5), which encode error-prone polymerases (8,9). By inducing the SOS response, ciprofloxacin increases mutagenesis (e.g., see references 10 and 11), facilitating the appearance of drug resistance.We investigated the effect of antibiotic combinations on the SOS response and mutagenesis induced by ciprofloxacin in P. aeruginosa PAO1. recA is among the SOS-regulated genes in this organism (4,5). To analyze the induction of the SOS response, we constructed a chromosomal PrecA-lux reporter. The regulatory region upstream from recA (Ϫ501 bp relative to the start codon) was cloned into the pUC18T-mini-Tn7T-lux-Gm plasmid (12) and transferred to P. aeruginosa for integration at attTn7, resulting in strain attTn7::PrecA-lux. The effect of antibiotics on the expression of the reporter was evaluated in solid medium using disk-based qualitative assays. The PrecA-lux strain was diluted to an optical density at 260 nm (OD 600 ) of 0.1. Fifty microliters of this dilution was seeded on Mueller-Hinton (MH) agar, and test antibiotic disks and CIP (5 g) disks (Sensifar-Cefar, Brazil) were placed close to one another to observe the effect of the antibiotic interaction on recA expression. Luciferase activity was detected in the ChemiDoc MP system (Bio-Rad, USA).Amikacin (AMI), imipenem, meropenem, polymyxin B, ceftazidime, cefepime, and aztreonam were tested, representing different drug classes. We found that amikacin is not an inducer of the SOS response but is in fact a strong inhibitor of recA induction by sub-MICs of ciprofloxacin (Fig. 1A). Amikacin is an aminoglycoside derived from kanamycin (13). Aminoglycosides bind, with high affinity, to the A-site on the 16S rRNA of the 30S ribosome (14) and can cause mRNA decoding errors, block mRNA and tRNA translocation, and inhibit ribosome recycling (15).

The modulating effect of N-acetylcysteine (NAC) on the activity of different antibiotics has been studied in Pseudomonas aeruginosa. Our results demonstrate that, in contrast to previous reports, only the activity of imipenem is clearly affected by NAC. MIC and checkerboard determinations indicate that the NAC-based modulation of imipenem activity is dependent mainly on OprD. SDS-PAGE of outer membrane proteins (OMPs) after NAC treatments demonstrates that NAC does not modify the expression of OprD, suggesting that NAC competitively inhibits the uptake of imipenem through OprD. Similar effects on imipenem activity were obtained with P. aeruginosa clinical isolates. Our results indicate that imipenem-susceptible P. aeruginosa strains become resistant upon simultaneous treatment with NAC and imipenem. Moreover, the generality of the observed effects of NAC on antibiotic activity was assessed with two additional bacterial species, Escherichia coli and Acinetobacter baumannii. Caution should be taken during treatments, as the activity of imipenem may be modified by physiologically attainable concentrations of NAC, particularly during intravenous and nebulized regimes.A lthough its benefit has been questioned (1), N-acetylcysteine (NAC) is being used for the treatment of numerous disorders, including paracetamol intoxication, doxorubicin cardiotoxicity, ischemia-reperfusion cardiac injury, acute respiratory distress syndrome, bronchitis, chemotherapy-induced toxicity, HIV/ AIDS, heavy metal toxicity, and psychiatric disorders (2). This compound is also sold as a dietary supplement, with claims of antioxidant and liver-protecting effects. The antioxidant activity of NAC has been attributed to its reactivity with ·OH, ·NO2, CO 3 · Ϫ , and thiyl radicals, to its capacity for repair of oxidized key cellular molecules, and to its activity as a precursor for glutathione biosynthesis (2). Because of this mucolytic capacity, it has also been used to facilitate the processing of sputum specimens in bacteriology laboratories (3). The ability of NAC to reduce biofilms, alone or in combination with antimicrobials, has been addressed in several bacterial species (4-6). Furthermore, NAC has been proposed as a treatment in Helicobacter pylori infections (7). NAC utility in reducing sputum viscosity in patients with cystic fibrosis (CF) and chronic obstructive pulmonary disease (COPD) (8), likely due to its ability to break disulfide bonds, has also been claimed, although its mechanism of action is not well understood.As a result of an imbalance between the production of reactive oxygen species (ROS) caused by inflammation and their inactivation by the impaired antioxidant systems, CF patients with chronic Pseudomonas aeruginosa lung infection have increased oxidative stress. Therefore, antioxidant interventions, including the use of NAC, have been proposed to reduce the extent of oxidative lesions and the rate of lung deterioration (for a review, see reference 9). However, although controversial results on the improvement of lung f...