Toxic effects of detergents on the alga Plagioselmis prolonga (Cryptophyta)

The effect of sodium dodecyl sulfate (SDS) and the household synthetic detergents (HSDs) Kristall and Tix (0.1, 1, and 10 mg/l) on cell motility, cell number dynamics, and the growth rate of the alga Plagioselmis prolonga (Cryptophyta) is studied. Algal cell motility proved to be the most sensitive indicator of detergent toxicity. SDS was the least toxic: 1 mg SDS/l caused a short-term loss of motility in 10% of the algal cells. The HSD Tix was the most toxic: only 70% of the cells recovered motility after a 24 h exposure to 1 mg/l. The substances tested in a concentration of 10 mg/l caused mortality of the P. prolonga population. According to their toxic effect on P. prolonga, the investigated toxicants can be arranged as follows: SDS < Kristall < Tix.     

The effect of liposomes on the growth and sensitivity of <Emphasis Type="Italic">Mycobacterium smegmatis</Emphasis> to isoniazide

The effects of the liposome form of isoniazide (IN) and liposomes without IN on the growth of Mycobacterium smegmatis were studied. Fluorescent assay demonstrated that the fraction of liposomes that interacted with M. smegmatis amounted to 1–3%. It was shown that the IN efficiency in a liposomal form decreased depending on the liposome composition and concentration as compared with the IN in water solution. A preincubation of mycobacteria with liposomes led to a decrease in their sensitivity to IN. An analogous effect was observed when incubating M. smegmatis with oleic acid. It was postulated that the relative resistance of M. smegmatis to the antibiotic when using lipids as a carbon substrate appeared due to a change in the agent’s metabolism and should be taken into account when testing in vitro the liposomal forms of antibiotics.     

Investigation of the antibacterial activity of a short cationic peptide against multidrug-resistant Klebsiella pneumoniae and Salmonella typhimurium strains and its cytotoxicity on eukaryotic cells

With the growing microbial resistance to conventional antimicrobial agents, the development of novel and alternative therapeutic strategies are vital. During recent years novel peptide antibiotics with broad spectrum activity against many Gram-positive and Gram-negative bacteria have been developed. In this study, antibacterial activity of CM11 peptide (WKLFKKILKVL-NH2), a short cecropin–melittin hybrid peptide, is evaluated against antibiotic-resistant strains of Klebsiella pneumoniae and Salmonella typhimurium as two important pathogenic bacteria. To appraise the antibacterial activity, minimal inhibitory concentration (MIC), minimal bactericidal concentration (MBC) and bactericidal killing assay were utilized with different concentrations (2–128 mg/L) of peptide. To evaluate cytotoxic effect of peptide, viability of RAJI, Hela, SP2/0, CHO, LNCAP cell lines and primary murine macrophage cells were also investigated with MTT assay in different concentrations (3–24 and 0.5–16 mg/L, respectively). MICs of K. pneumoniae and S. typhimurium isolates were in range of 8–16 and 4–16 mg/L, respectively. In bactericidal killing assay no colonies were observed at 2X MIC for K. pneumoniae and S. typhimurium isolates after 80–90 min, respectively. Despite the fact that CM11 reveals no significant cytotoxicity on RAJI, Hela, SP2/0, and CHO cell lines beneath 6 mg/L at first 24 and 48 h, the viability of LNCAP cells are about 50 % at 3 mg/L, which indicates strong cytotoxicity of the peptide. In addition, macrophage toxicity by MTT assay showed that LD₅₀ of CM11 peptide is 12 μM (16 mg/L) after 48 h while in this concentration after 24 h macrophage viability was about 70 %.     

Isolation of an aboriginal bacterial community capable of utilizing cyanide,thiocyanate, and ammonia from metallurgical plant wastewater

An aboriginal bacterial community capable of degrading cyanide (10 mg/l) and thiocyanate (2 g/l) and eliminating ammonia (120 mg/l) had been isolated from recycled water samples after blast-furnace gas purification of a metallurgical plant wastewater. It was shown that the optimal conditions for this bacterial community were as follows: temperature, 34°C; pH, 8.8–9.0; available organic matter concentration (glucose equivalent), 5 g/l; and dissolved O₂ concentration, 8–10 mg/l. This aboriginal community was formed by the bacteria belonging to the genus Pseudomonas.     

Design, expression, and characterization of a novel targeted plectasin against methicillin-resistant Staphylococcus aureus

A novel specifically targeted antimicrobial peptide (STAMP) that was especially effective against methicillin-resistant Staphylococcus aureus (MRSA) was designed by fusing the AgrD1 pheromone to the N-terminal end of plectasin. This STAMP was named Agplectasin, and its gene was synthesized and expressed in Pichia pastoris X-33 via pPICZαA. The highest amount of total secreted protein reached 1,285.5 mg/l at 108 h during the 120-h induction. The recombinant Agplectasin (rAgP) was purified by cation exchange chromatography and hydrophobic exchange chromatography; its yield reached 150 mg/l with 94 % purity. The rAgP exhibited strong bactericidal activity against S. aureus but not Staphylococcus epidermidis or other types of tested bacteria. A bactericidal kinetics assay showed that the rAgP killed over 99.9 % of tested S. aureus (ATCC 25923 and ATCC 43300) in both Mueller–Hinton medium and human blood within 10 h when treated with 4× minimal inhibitory concentration. The rAgP caused only approximately 1 % hemolysis of human blood cells, even when the concentration reached 512 μg/ml, making it potentially feasible as a clinical injection agent. In addition, it maintained a high activity over a wide range of pH values (2.0–10.0) and demonstrated a high thermal stability at 100 °C for 1 h. These results suggested that this STAMP has the potential to eliminate MRSA strains without disrupting the normal flora.     

Comparison of antimicrobial efficacy of a fixed dose combination of ceftazidime + sulbactam with ceftazidime and sulbactam alone against five bacteria

The efficacy of the agents was evaluated on the basis of minimum inhibitory concentration (MIC) and time-kill curve analysis in Pseudomonas aeruginosa, Staphylococcus aureus, Acinetobacter baumannii, Mycobacterium smegmatis, and Citrobacter braakii; MIC was found to be 0.5, 4.0, 0.015625, 0.0078125, and 0.0625 mg/L in FDC of ceftazidime + sulbactam, respectively, which is lower than ceftazidime and sulbactam individually. Time-kill curve analysis demonstrated maximum killing of bacteria after 4 h. A fixed dose combination of ceftazidime + sulbactam was found to have stronger antibacterial properties than ceftazidime and sulbactam alone at in vitro analysis.     

Seasonal Changes in the Structure of the Anoxygenic Phototrophic Bacterial Community in Lake Mogilnoe,a Relict Lake on Kil'din Island in the Barents Sea

An anaerobic phototrophic bacterial community in Lake Mogilnoe, a relict lake on Kil'din Island in the Barents Sea, was studied in June 1999 and September 2001. Irrespective of the season, the upper layer of the anaerobic zone of this lake had a specific species composition of sulfur phototrophic bacteria, which were dominated by the brown-colored green sulfur bacterium Chlorobium phaeovibrioides. The maximum number of sulfur phototrophic bacteria was observed in June 1999 at a depth of 9 m, which corresponded to a concentration of bacteriochlorophyll (Bchl) e equal to 4.6 mg/l. In September 2001, the maximum concentration of this pigment (3.4 mg/l) was found at a depth of 10 m. In both seasons, the concentration of Bchl a did not exceed 3 μg/l. Purple sulfur bacteria were low in number, which can be explained by their poor adaptation to the hydrochemical and optical conditions of the Lake Mogilnoe water. In June 1999, the water contained a considerable number of Pelodictyon phaeum microcolonies and Prosthecochloris phaeoasteroides cell chains, which was not the case in September 2001. A 16S rDNA-based phylogenetic analysis of pure cultures of phototrophic bacteria isolated from the lake water confirmed that the bacterial community is dominated by Chl. phaeovibrioides and showed the presence of three minor species, Thiocystis gelatinosa, Thiocapsa sp., and Thiorhodococcus sp., the last of which is specific to Lake Mogilnoe.     

Effect of COD to sulphate ratio and temperature in expanded-granular-sludge-blanket reactors for sulphate reduction

In an ethanol-fed expanded-granular-sludge-blanket (EGSB) reactor at 33°C, 80–90% of the sulphate load was removed at a rate of 4 g S/l d, provided that at least 6 g chemical oxygen demand (COD) per g sulphate-sulphur was supplied. The reactor started up in a matter of days. Gradually decreasing the ethanol to sulphate ratio (R) to about stoichiometry, resulted in 60–70% sulphate removal at rates of 7 g S/l d. Similar tendencies were observed with ethylene glycol as sole carbon and energy source. Total COD removal never reached more than 70–75%. This was related to a rather high biomass washout. The sulphate removal efficiency decrease when R was set at levels below 6, apparently because sulphate reducing bacteria (SRB) could not compete with methane producing bacteria (MPB) for acetate produced from the substrate dosed. Thermophilic operation at 55°C, after a stepwise increase in the reactor temperature over a period of 23 days, did not favour acetotrophic sulphate reduction. Yet, operation at 48°C and subsequently returning the temperature to 33°C clearly enhanced acetate conversion by SRB. In the case of an electron donor price of 0.035–0.075 USD/kg COD, the cost for operation at R=6 was found to be competitive to that at stoichiometry, i.e. R=2, provided the biogas produced was effectively used.     

A novel screening method based on menadione mediated rapid reduction of tetrazolium salt for testing of anti-mycobacterial agents

A microplate-based rapid, inexpensive and robust technique is developed by using tetrazolium salt 2, 3-bis [2-methyloxy-4-nitro-5-sulfophenyl]-2H-tetrazolium-5-carboxanilide (XTT) and menadione to determine the viability of Mycobacterium tuberculosis, Mycobacterium bovis BCG and Mycobacterium smegmatis bacilli in microplate format. In general, XTT reduction is an extremely slow process which takes almost 24 h to produce a detectable signal. Menadione could drastically induce this reduction to an almost equal extent within a few minutes in a dose dependent manner. The reduction of XTT is directly proportional to the cell concentration in the presence of menadione. The standardized protocol used 200 μM of XTT and 60 μM of menadione in 250 μl of cell suspension grown either in aerobic or anaerobic conditions. The cell suspension of M. bovis BCG and M. tuberculosis were incubated for 40 min before reading the optical density at 470 nm whereas M. smegmatis was incubated for 20 min. Calculated Signal/Noise (S/N) ratios obtained by applying this protocol were 5.4, 6.4 and 9.4 using M. bovis BCG, M. tuberculosis and M. smegmatis respectively. The calculated Z′ factors were > 0.8 for all mycobacterium bacilli indicating the robustness of the XTT Reduction Menadione Assay (XRMA) for rapid screening of inhibitors. The assay protocol was validated by applying 10 standard anti-tubercular agents on M. tuberculosis, M. bovis BCG and M. smegmatis. The Minimum Inhibitory Concentration (MIC) values were found to be similar to reported values from Colony Forming Unit (CFU) and REMA (resazurin microplate assay) assays. Altogether, XRMA is providing a novel anti-tubercular screening protocol which could be useful in high throughput screening programs against different physiological stages of the bacilli.     

Pyruvate carboxylase from Mycobacterium smegmatis: stabilization,rapid purification,molecular and biochemical characterization and regulation of the cellular level

This is the first report on the purification and characterization of an anaplerotic enzyme from a Mycobacterium. The anaplerotic reactions play important roles in the biochemical differentiation of mycobacteria into non-replicating stages. We have purified and characterized a pyruvate carboxylase (PYC) from Mycobacterium smegmatis and cloned and sequenced its gene. We have developed a very rapid and efficient purification protocol that provided PYC with very high specific activities (up to 150 U/mg) that remained essentially unchanged over a month. The enzyme was found to be a homomultimer of 121 kDa subunits, mildly thermophilic, absolutely dependent on acyl-CoAs for activity and inhibited by ADP, by excess Mg²⁺, Co²⁺, and Mn²⁺, by aspartate, but not by glutamate and α-ketoglutarate. Supplementation of minimal growth medium with aspartate did not lower the cellular PYC level, rather doubled it; with glutamate the level remained unchanged. These observations would not fit the idea that the M. smegmatis enzyme fulfills a straightforward anaplerotic function; in a closely related organism, Corynebacterium glutamicum, PYC is the major anaplerotic enzyme. Growth on glucose provided 2-fold higher cellular PYC level than that observed with glycerol. The PYCs of M. smegmatis and Mycobacterium tuberculosis were highly homologous to each other. In M. smegmatis, M. tuberculosis and M. lepra, pyc was flanked by a putative methylase and a putative integral membrane protein genes in an identical operon-like arrangement. Thus, M. smegmatis could serve as a model for studying PYC-related physiological aspects of mycobacteria. Also, the ease of purification and the extraordinary stability could make the M. smegmatis enzyme a model for studying the structure–function relationships of PYCs in general. It should be noted that no crystal structure is available for this enzyme of paramount importance in all three domains of life, archaea, bacteria, and eukarya.     

Mycobacterium tuberculosis acyl carrier protein inhibits macrophage apoptotic death by modulating the reactive oxygen species/c-Jun N-terminal kinase pathway

Mycobacterial acyl carrier protein (AcpM; Rv2244) is a meromycolate extension acyl carrier protein of Mycobacterium tuberculosis (Mtb), which participates in multistep mycolic acid biosynthesis. However, the function of AcpM in host–mycobacterium interactions during infection remains largely uncharacterized. Here we show that AcpM inhibits host cell apoptosis during mycobacterial infection. To examine the function of AcpM during infection, we generated a recombinant Mycobacterium smegmatis (M. smegmatis) strain overexpressing AcpM (Ms_AcpM) and a strain transformed with an empty vector (Ms_Vec). Ms_AcpM promoted intracellular survival of M. smegmatis and led to a significant decrease in the death rate of primary bone marrow-derived macrophages (BMDMs). Importantly, Ms_AcpM showed significantly decreased reactive oxygen species (ROS) generation and activation of c-Jun N-terminal kinase (JNK) signaling compared with Ms_Vec. In addition, treatment of BMDMs with recombinant AcpM significantly inhibited the apoptosis and ROS/JNK signaling induced by M. smegmatis. Moreover, recombinant AcpM enhanced intracellular survival of Mtb H37Rv. Taken together, these results indicate that AcpM plays a role as a virulence factor by modulating host cell apoptosis during mycobacterial infection.     

The potential of synthetic hexaploid wheats to improve zinc efficiency in modern bread wheat

Synthetic hexaploid wheats (Triticum aestivum L) derived from crosses between durum wheat [Triticum turgidum ssp. durum (Desf.) Husn.] and diploid wheat (Aegilops tauschii Coss.) have been developed as a means of transferring desirable characteristics of Aegilops tauschii Coss. such as disease resistance and abiotic stress tolerance into modern bread wheat genotypes. In a growth room experiment using soil culture, we studied a group of 30 synthetic hexaploid wheat accessions together with modern wheat genotypes in order to identify new sources of zinc efficiency for further improvement of zinc efficiency in modern wheat genotypes. There was considerable genetic variation in expression of zinc deficiency symptoms (slight to severe), zinc efficiency (70–100%), shoot Zn concentration (5.8–10.5 and 33–53 mg/kg DW under deficient and sufficient Zn, respectively), shoot Zn content (3.8–10.6 and 34.0–64.6 μg/plant, under deficient and sufficient Zn, respectively) and Zn utilization (0.096–0.172 and 0.019-0.033 g DW/μg Zn under deficient and sufficient Zn, respectively) within synthetic accessions. The presence of synthetic accessions with greater zinc efficiency (100%) than zinc efficient modern wheat genotypes (85%) indicates that the synthetic hexaploids can be used to improve current levels of zinc efficiency in modern wheat genotypes. Synthetic hexaploids may also be a good source of high grain Zn concentration (28–66 mg Zn/kg seed DW).     

Radiation-sensitive Gene A (RadA) Targets DisA,DNA Integrity Scanning Protein A,to Negatively Affect Cyclic Di-AMP Synthesis Activity in Mycobacterium smegmatis

Cyclic di-AMP has been recognized as a ubiquitous second messenger involved in the regulation of bacterial signal transduction. However, little is known about the control of its synthesis and its physiological role in bacteria. In this study, we report a novel mechanism of control of c-di-AMP synthesis and its effects on bacterial growth in Mycobacterium smegmatis. We identified a DisA homolog in M. smegmatis, MsDisA, as an enzyme involved in c-di-AMP synthesis. Furthermore, MsRadA, a RadA homolog in M. smegmatis was found to act as an antagonist of the MsDisA protein. MsRadA can physically interact with MsDisA and inhibit the c-di-AMP synthesis activity of MsDisA. Overexpression of MsdisA in M. smegmatis led to cell expansion and bacterial aggregation as well as loss of motility. However, co-expression of MsradA and MsdisA rescued these abnormal phenotypes. Furthermore, we show that the interaction between RadA and DisA and its role in inhibiting c-di-AMP synthesis may be conserved in bacteria. Our findings enhance our understanding of the control of c-di-AMP synthesis and its physiological roles in bacteria.     

Modulation of isoniazid susceptibility by flavonoids in Mycobacterium

In the course of a project to identify plant natural products which modulate the susceptibility of different strains of fast-growing mycobacteria to the first-line antituberculotic isoniazid (INH), several flavonoids without significant antimycobacterial activities at the tested concentrations were screened for their ability to decrease the minimum inhibitory concentrations (MICs) of INH. Flavonoids with different substitution patterns, namely epicatechin, isorhamnetin, kaempferol, luteolin, myricetin, quercetin, rutin and taxifolin were tested to examine structure–activity relationships (SARs) of these compounds. Different mycobacterial strains, i.e. Mycobacterium smegmatis (ATCC 14468), M. smegmatis mc²155 (ATCC 700084), M. smegmatis mc²2700, M. phlei (ATCC 11758) and M. fortuitum (ATCC 6841) were used. The strongest synergistic effects were observed in M. smegmatis mc²155 followed by M. phlei, whereas the tendency of INH potentiation by certain flavonoids remained the same within each strain. Myricetin was the most efficient intensifier of INH susceptibility in all tested strains causing a decrease of the MIC of INH up to 64-fold at 16 μg/ml, followed by quercetin. Structure–activity relationships of flavonoids as intensifiers of INH susceptibility in mycobacteria indicate that they overlap with SARs for their radical-scavenging properties, however the potentiation of INH activity cannot only be explained by their radical-scavenging activity alone.     

Interconnection of the mycobacterial heparin-binding hemagglutinin with cholesterol degradation and heme/iron pathways identified by proximity-dependent biotin identification in Mycobacterium smegmatis

Deciphering protein–protein interactions is a critical step in the identification and the understanding of biological mechanisms deployed by pathogenic bacteria. The development of in vivo technologies to characterize these interactions is still in its infancy, especially for bacteria whose subcellular organization is particularly complex, such as mycobacteria. In this work, we used the proximity-dependent biotin identification (BioID) to define the mycobacterial heparin-binding hemagglutinin (HbhA) interactome in the saprophytic bacterium Mycobacterium smegmatis. M. smegmatis is a commonly used model to study and characterize the physiology of pathogenic mycobacteria, such as Mycobacterium tuberculosis. Here, we adapted the BioID technology to in vivo protein–protein interactions studies in M. smegmatis, which presents several advantages, such as maintaining the complex organization of the mycomembrane, offering the possibility to study membrane or cell wall-associated proteins, including HbhA, in the presence of cofactors and post-translational modifications, such as the complex methylation pattern of HbhA. Using this technology, we found that HbhA is interconnected with cholesterol degradation and heme/iron pathways. These results are in line with previous studies showing the dual localization of HbhA, associated with the cell wall and intracytoplasmic lipid inclusions, and its induction under high iron growth conditions.     

High production of D-β-hydroxyisobutyric acid from methacrylic acid by Candida rugosa and its mutant

Production of D-β-hydroxyisobutyric acid (D-HIBA) from methacrylic acid (MA) was investigated using Candida rugosa IFO 0750 and its mutant. Cell growth decreased as the MA concentration increased and was inhibited at D-HIBA concentrations higher than 30 g/l. Optimal MA concentration for D-HIBA production was in the range of 10–20 g/l. It was also noted that cell growth and D-HIBA production were inhibited by higher concen-trations of Na⁺, K⁺, and NH₄ ⁺, which were required for pH control during cultivation. With a suitably designed feeding mode of MA, the parent strain produced 65 g/l of D-HIBA after 120?h of fed-batch cultivation, but molar conversion yield of D-HIBA was less than 40%. The mutant, unable to assimilate propionic acid, produced as high as 70 g/l of D-HIBA in the same culture period with a molar conversion yield of more than 70%.     

Types of responses of a model population of microalga <Emphasis Type="Italic">Scenedesmus quadricauda</Emphasis> (Turp.) Breb. under different intoxication conditions

The main ways of reaction of a green chlorococcal alga Scenedesmus quadricauda (Turp.) Breb. model population to the toxic action of different chemicals (the fungicide imazalil sulfate and heavy metal Cr⁶⁺ as a potassium bichromate) are analyzed. Regularities of the change in the size-age and functional population characteristics are considered, and four types of universal microalgal response to the toxic action are demonstrated. In the presence of low concentrations (0.001 mg/l), the slowdown of the culture growth was caused by the long stop of cell division in part of the algal population rather than by cell death or a decrease in their growth. The lack of effect (or even culture growth stimulation under medium concentrations (0.1 mg/l)) is related to the cell division proliferation after a temporary stop. After increasing the toxicant concentration up to 1.0 mg/l, we observed a long cell division inhibition; however, the cell death was under high toxicant concentrations of 5.0–10.0 mg/l). A significant inhibition of functional cell parameters (photosynthetic intensity) took place only under high toxicant concentrations (5.0 mg/l and higher). It is shown that the microalgal population state can be estimated on the basis of an analysis of its structural and functional parameters.     

Functional Analysis of a c-di-AMP-specific Phosphodiesterase MsPDE from Mycobacterium smegmatis

Cyclic di‑AMP (c-di-AMP) is a second signaling molecule involved in the regulation of bacterial physiological processes and interaction between pathogen and host. However, the regulatory network mediated by c-di-AMP in Mycobacterium remains obscure. In M. smegmatis, a diadenylate cyclase (DAC) was reported recently, but there is still no investigation on c-di-AMP phosphodiesterase (PDE). Here, we provide a systematic study on signaling mechanism of c-di-AMP PDE in M. smegmatis. Based on our enzymatic analysis, MsPDE (MSMEG_2630), which contained a DHH-DHHA1 domain, displayed a 200-fold higher hydrolytic efficiency (k_cat/K_m) to c-di-AMP than to c-di-GMP. MsPDE was capable of converting c-di-AMP to pApA and AMP, and hydrolyzing pApA to AMP. Site-directed mutations in DHH and DHHA1 revealed that DHH domain was critical for the phosphodiesterase activity. To explore the regulatory role of c-di-AMP in vivo, we constructed the mspde mutant (Δmspde) and found that deficiency of MsPDE significantly enhanced intracellular C₁₂-C₂₀ fatty acid accumulation. Deficiency of DAC in many bacteria results in cell death. However, we acquired the M. smegmatis strain with DAC gene disrupted (ΔmsdisA) by homologous recombination approach. Deletion of msdisA reduced bacterial C₁₂-C₂₀ fatty acids production but scarcely affected bacterial survival. We also provided evidences that superfluous c-di-AMP in M. smegmatis could lead to abnormal colonial morphology. Collectively, our results indicate that MsPDE is a functional c-di-AMP-specific phosphodiesterase both in vitro and in vivo. Our study also expands the regulatory network mediated by c-di-AMP in M. smegmatis.     

Porins Increase Copper Susceptibility of Mycobacterium tuberculosis

Copper resistance mechanisms are crucial for many pathogenic bacteria, including Mycobacterium tuberculosis, during infection because the innate immune system utilizes copper ions to kill bacterial intruders. Despite several studies detailing responses of mycobacteria to copper, the pathways by which copper ions cross the mycobacterial cell envelope are unknown. Deletion of porin genes in Mycobacterium smegmatis leads to a severe growth defect on trace copper medium but simultaneously increases tolerance for copper at elevated concentrations, indicating that porins mediate copper uptake across the outer membrane. Heterologous expression of the mycobacterial porin gene mspA reduced growth of M. tuberculosis in the presence of 2.5 μM copper by 40% and completely suppressed growth at 15 μM copper, while wild-type M. tuberculosis reached its normal cell density at that copper concentration. Moreover, the polyamine spermine, a known inhibitor of porin activity in Gram-negative bacteria, enhanced tolerance of M. tuberculosis for copper, suggesting that copper ions utilize endogenous outer membrane channel proteins of M. tuberculosis to gain access to interior cellular compartments. In summary, these findings highlight the outer membrane as the first barrier against copper ions and the role of porins in mediating copper uptake in M. smegmatis and M. tuberculosis.     

The ethnobotany,leaf anatomy,essential oil variation and biological activity of Pteronia incana (Asteraceae)

The only available ethnobotanical information on Pteronia incana has been recorded by the Montagu Museum in the Western Cape Province of South Africa. It was reported that the plant is used to treat influenza, fever, kidney ailments and backache. In common with other species of Pteronia, the plant contains an essential oil reminiscent of pine turpentine oil with β-pinene, limonene, 1,8-cineole, myrcene, spathulenol, p-cymene and methyleugenol as main compounds present in all or most of the samples, with smaller amounts of α-pinene, sabinene, γ-terpinene, terpinen-4-ol, biclogermacrene, globulol and α-bisabolol in some of the distillates. We investigated the oil composition of 11 individual plants collected at three geographically distant localities but found limited variation, both within and between populations. Leaf sections of P. incana showed that it is anatomically similar to P. divaricata in the presence of a secretory duct associated with the main vascular bundle (and often other bundles as well), in addition to glandular and non-glandular trichomes on both leaf surfaces. One yeast (Cryptococcus neoformans), two Gram-negative bacteria (Moraxella catarrhalis and Klebsiella pneumoniae) and one Gram-positive bacterium (Mycobacterium smegmatis) were selected for antimicrobial activity studies using the minimum inhibitory concentration (MIC) microtitre plate method. The results showed that methanol:dichloromethane (MeOH:CH₂Cl₂) extracts were active against M. smegmatis (lowest MIC values of 0.5–0.8 mg/ml) and C. neoformans (lowest MIC values of 0.5–2.0 mg/ml). The essential oil was most active against C. neoformans (lowest MIC value of 0.3 mg/ml). These results provide a scientific rationale for the use of P. incana in Cape herbal medicine.