Medicinal plants from open-air markets in the State of Rio de Janeiro, Brazil as a potential source of new antimycobacterial agents.

ETHNOPHARMACOLOGICAL RELEVANCE: Several medicinal plants are traditionally traded in open-air markets in Rio de Janeiro State (Brazil) to treat tuberculosis (TB) and related symptoms. AIMS OF THE STUDY: Conduct a survey in the open-air markets of 20 cities of Rio de Janeiro State to find medicinal plants that are popularly used to treat tuberculosis and other related diseases and assess their in vitro antimycobacterial activity. MATERIALS AND METHODS: We used direct observation and semi-structured interviews and asked herbalists to list species (free listing) in order to gather data about the plant species most commonly used for lung problems. We calculated a Salience Index and acquired two species of "erva-de-passarinho" (mistletoe), Struthanthus marginatus and Struthanthus concinnus (Loranthaceae), commonly used to treat tuberculosis for a bioassay-guided isolation of the antimycobacterial active principles. Extracts, fractions and isolated compounds of both species were assayed in vitro against susceptible (H37Rv) and rifampicin-resistant (ATCC 35338) Mycobacterium tuberculosis strains. RESULTS: From the interviews, we generated a list of 36 plant species belonging to 12 families. The mistletoes Struthanthus marginatus and Struthanthus concinnus showed high Salience Index values among plants used to treat tuberculosis. Bioassay-guided fractionation of hexane extracts from both species led to the isolation and/or identification of steroids and terpenoids. The Minimum Inhibitory Concentration (MIC) of the extracts and isolated compounds ranged from 25 to 200 µg/mL. Some of the isolated compounds have been previously assayed against Mycobacterium tuberculosis, others are reported here for the first time (obtusifoliol: MIC H37Rv 50 µg/mL, MIC ATCC 35338 12.5 µg/mL; 3-O-n-acil-lup-20(29)-en-3ß,7ß,15α-triol: MIC H37Rv 200 µg/mL, MIC ATCC 35338 100 µg/mL). CONCLUSIONS: This study demonstrated the importance of ethnobotanical surveys in markets as a source for new drugs and also for scientific validation of folk medicine.

Biological cost in Mycobacterium tuberculosis with mutations in the rpsL, rrs, rpoB, and katG genes.

When bacteria develop drug-resistant mutations, there is often an associated biological cost; however, some strains can exhibit low- or no-cost mutations. In the present study, a quantitative resazurin reduction assay was used to measure the biological cost of Mycobacterium tuberculosis isolates that contained different mutations in the rpsL, rrs, rpoB, and katG genes, and showed different resistance profiles. Biological costs were determined by comparing the growth curves of drug-resistant isolates with drug-susceptible strains. Some strains, such as those with rpoB mutations other than S531L and strains with mutations in all of the studied genes, grew more slowly than did drug-susceptible strains. However, some strains grew more quickly than drug-susceptible strains, such as those that had only the rpsL K43R mutation. Strains with the mutation katG S315T presented heterogeneous biological costs. When analyzed individually, strains with the mutations rpsL43/katG315, rpoB531, and rpoB531/katG315 grew faster than drug-susceptible strains. The results suggest that some strains with the most common mutations correlated to a high resistance toward streptomycin, isoniazid and rifampicin can grow as well as or better than susceptible strains.

Streptomycin resistance and lineage-specific polymorphisms in Mycobacterium tuberculosis gidB gene.

Mutations related to streptomycin resistance in the rpsL and rrs genes are well known and can explain about 70% of this phenotypic resistance. Recently, the gidB gene was found to be associated with low-level streptomycin resistance in Mycobacterium tuberculosis. Mutations in gidB have been reported with high frequency, and this gene appears to be very polymorphic, with frameshift and point mutations occurring in streptomycin-susceptible and streptomycin-resistant strains. In this study, mutations in gidB appeared in 27% of streptomycin-resistant strains that contained no mutations in the rpsL or rrs genes, and they were associated with low-level streptomycin resistance. However, the association of certain mutations in gidB with streptomycin resistance needs to be further investigated, as we also found mutations in gidB in streptomycin-susceptible strains. This occurred only when the strain was resistant to rifampin and isoniazid. Two specific mutations appeared very frequently in this and other studies of streptomycin-susceptible and -resistant strains; these mutations were not considered related to streptomycin resistance, but as a polymorphism. We stratified the strains according to the different phylogenetic lineages and showed that the gidB(16) polymorphism (16G allele) was exclusively present in the Latin American-Mediterranean (LAM) genotype, while the gidB(92) polymorphism (92C allele) was associated with the Beijing lineage in another population. In the sample studied, the two characterized single-nucleotide polymorphisms could distinguish LAM and Beijing lineages from the other lineages.