Airway microecology in rifampicin-resistant and rifampicin-sensitive pulmonary tuberculosis patients.

BACKGROUND: Pulmonary tuberculosis is a chronic infectious disease of the respiratory system. It is still one of the leading causes of death from a single infectious disease, but it has been stuck in the study of a single pathogen. Recent studies have shown that many diseases are associated with disruption of the native microbiota. In this study we investigated the occurrence of tuberculosis and the correlation between drug resistance and respiratory flora. High-throughput 16 S rRNA gene sequencing was used to characterize the respiratory microbiota composition of 30 tuberculosis (TB) affected patients and compared with 30 healthy (H) controls. According to their Gene Xpert results, 30 pulmonary tuberculosis patients were divided into 12 persons in the drug-sensitive group (DS0) and 18 persons in the drug-resistant group (DR0). The microbial flora of the two were compared with the H group. RESULTS: The data generated by sequencing showed that Firmicutes, Proteus, Bacteroides, Actinomyces and Fusobacterium were the five main bacterial phyla detected, and they constituted more than 96% of the microbial community. The relative abundances of Fusobacterium, Haemophilus, Porphyromonas, Neisseria, TM7, Spirochetes, SR1, and Tenericutes in the TB group was lower than that of the H group, and Granulicatella was higher than the H group. The PcoA diagrams of the two groups had obvious clustering differences. The Alpha diversity of the TB group was lower than that of the H group, and the Beta diversity was higher than that of the H group (P < 0.05). The relative abundance of Streptococcus in the DS0 group was significantly higher than that in the DR0 group (P < 0.05). CONCLUSION: Pulmonary tuberculosis can cause disorders of the respiratory tract microbial flora, in which the relative abundance of Streptococcus was significantly different between rifampicin-sensitive and rifampicin-resistant patients.

Role of pncA and rpsA gene sequencing in detection of pyrazinamide resistance in Mycobacterium tuberculosis isolates from southern China.

We sequenced pncA and rpsA genes plus flanking regions of 161 Mycobacterium tuberculosis isolates and found 10 new pncA and 3 novel rpsA mutations in pyrazinamide-resistant strains determined by the Bactec MGIT 960 system. The 3' end of rpsA might be added as the target of molecular detection of pyrazinamide susceptibility.

The beginning of the rpoB gene in addition to the rifampin resistance determination region might be needed for identifying rifampin/rifabutin cross-resistance in multidrug-resistant Mycobacterium tuberculosis isolates from Southern China.

We aimed to study the distribution and contribution of mutations in the rpoB whole gene in rifampin-resistant/rifabutin-resistant (RIF(r)/Rfb(r)) (or RIF/Rfb cross-resistant) clinical Mycobacterium tuberculosis isolates. One standard M. tuberculosis strain (H37Rv) and 392 other clinical M. tuberculosis isolates mainly from Guangdong Province of China whose susceptibilities to rifampin (RIF), rifabutin (Rfb), streptomycin (SM), ethambutol (EMB), and isoniazid (INH) were previously determined were subjected to DNA sequencing of their rpoB whole genes. H37Rv and the 30 drug-susceptible clinical isolates had no mutations in rpoB whole genes. In 43 rifampin-resistant/rifabutin-susceptible (RIF(r)/Rfb(s)) isolates, the most frequent mutation codons were 516 (62.80%), 526 (14.0%), and 533 (6.98%), but codon 531 had no mutation. Twenty-one of the 43 isolates (48.84%) had single mutations of H526L, H526S, D516V, D516Y, and D516F. In 319 RIF(r)/Rfb(r) isolates, the most frequent mutation codons were 531 (73.7%) and 526 (18.8%); the mutation frequency for codon 516 was 2.5%, and that for codon 533 was only 0.31%. A total of 82.8% (264/319) of them had single mutations of S531L, S531W, H526D, H526Y, H526R, Q513K, Q513P, Q510H, V176F, P206(T)R, Y314(T)C, and H323(T)Y (the superscript T indicates M. tuberculosis numbering; the remaining codons use the E. coli numbering), among which V176F, P206(T)R, Y314(T)C, and H323(T)Y were located in the beginning of rpoB, and all of them were present in 1.9% (6/319) of RIF(r)/Rfb(r) isolates. The multiple mutations in RIF(r)/Rfb(r) isolates and in RIF(r)/Rfb(s) isolates were also different from each other either in mutation positions or in types of mutation combinations. In conclusion, the mutations of rpoB in RIF-R/Rfb(s) and in RIF-R/Rfb-R isolates differ significantly from each other not only in the most frequent mutation codons (516, 531, and 533) but also in the most frequent single mutations (S531L, H526L, D516V, D516Y, and D516F), and the beginning of rpoB may confer a RIF/Rfb cross-resistance phenotype in M. tuberculosis. Molecular assays for identifying RIF/Rfb cross-resistance in M. tuberculosis might be improved in terms of accuracy by including this region, in addition to the rifampin resistance determination region.

[Antimicrobial resistance of clinical isolates of Stenotrophomonas maltophilia].

OBJECTIVE: To investigate the antimicrobial resistance of clinical isolates of Stenotrophomonas matophilia (SMA) and the mechanisms of their drug resistance. METHODS: Disc diffusion method (NCCLS) was used to detect the resistant patterns of 88 initial SMA isolates resistant to 12 antibiotics isolated from a local hospital in the past 4 years. PCR was used to detect the 7 aminoglycosides modifying enzymes genes (AME) against amikacin and gentamicin. Metal-beta-lactamases (MBLs) were screened by synergic method, and extended-spectrum beta-lactamases (ESBLs) were detected by double-disk synergy test. RESULTS: The resistance rates of the SMA isolates were 0%-9.7% to minocycline, 12.5%-22.6% to ticarcillin-clavulanic acid, 12.5%-28.6% to levofloxacin, 18.8%-33.3% to doxycycline, 18.8%-40% to sulfamethoxazole compound, 50%-65.7% to ciprofloxacin, 50%-66.7% to cehazindme, 54.8%-66.7% to amikacin, 75%-100% to gentamicin, 81.3%-100% to piperacillin, 87.5%-100% to aztreonam and 93.5%-100% to imipenem. Aac(3)-I and ant(4')-II were not detected in these strains. The positive rates of the other 5 AME genes of aac(3)-II, ant(2'')-I, aac(6')-I, aac(3)-III, aac(3)-IV were 2.3%, 5.7%, 8%, 10%, and 10%, respectively. SMA strains producing ESBLs were found at the rate of 38.6%; 25% of the strains were MBL-producing, and 13.6% produced both ESBLs and MBLs. CONCLUSION: Most of the SMAs we isolated are multidrug-resistant through various mechanisms. The choice of antibiotics should be made according to the susceptibility results.

[Detection of pathogenicity island-associated genes in enterococcal isolates].

OBJECTIVE: To investigate the presence of pathogenicity island (PAI)-associated genes in the enterococcal isolates. METHODS: Using PCR and hybridization methods, PAI-associated genes were detected in 155 enteococcal strains isolated from clinical patients and healthy individuals. RESULTS: Among the 155 enterococcal isolates, 137 (88.39%) carried at least one of PAI-associated genes, namely hyd (positivity rate of 81.94%), psaA (78.06%), nuc (57.42%), esp (53.55%), cylB (52.90%), and gls24-like (38.06%) genes. Expect for esp gene, the other 5 genes showed higher positivity rates in the E. faecalis strains than in the E. faecium strains, and this difference was statistically significant for the genes nuc, cylB, and gls24-like. The positivity rates and the number of these genes in the E. faecalis from clinical isolates were both significantly higher than those in the strains isolated from healthy individuals. CONCLUSION: The data show a wide distribution of the PAI-associated genes among the enterococcal strains, and E. faecalis strains are more likely than E. faecium strains to be positive for the 6 genes, which are present at significant higher rates in the clinically isolated samples than in that from healthy individuals.