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BACKGROUND: In Malawi, the national pneumococcal conjugate vaccine (PCV13) demonstrated less herd immunity than the USA, likely due to higher natural pneumococcal carriage rates. We assessed PCV13 efficacy against experimental pneumococcal carriage in healthy Malawian adults. We explored how natural carriage (pneumococcal carriage of any other serotype apart from 6B) influenced experimental carriage rates and vaccine efficacy. METHODS: Healthy adults aged 18-40 were randomly assigned PCV13 (n=98) or saline (n=106), followed by intranasal SPN 6B inoculation at 20,000 (n=40), 80,000 (n=74), or 160,000 (n=90) CFU/100µl, 28 days post-vaccination. We evaluated natural and experimental pneumococcal carriage before and after vaccination on days 2, 7, and 14 post-inoculation using culture and multiplex qPCR targeting lytA/cpsA genes and compared carriage rates by vaccination status. RESULTS: Of 204 participants, 19.6% (40) exhibited experimental carriage, detected by culture and 25.5% (52) by qPCR. Vaccinated individuals had lower experimental carriage rates (10.2%, n=10/98) compared to the placebo group (28.3%, n=30/106). This difference in vaccine efficacy was more pronounced in participants without natural carriage (PCV13=8% n=6/75 vs. placebo=25.9%, n=21/81) compared to those with natural carriage (PCV13=14.8%, n=4/27 vs. placebo=26.5%, n=9/34). Using a log-binomial model, vaccine effectiveness (VE) was 62%, whether assessed by culture or qPCR. Natural carriers had a lower VE of 52% compared to participants with no natural carriage (VE=69%). CONCLUSION: We have shown that PCV13 VE estimate (62%) is robust whether carriage is assessed by culture or qPCR. PCV13 had lower VE in natural carriers compared to those without natural carriage at the inoculation visit.
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TB47, a new drug candidate targeting QcrB in the electron transport chain, has shown a unique synergistic activity with clofazimine and forms a highly sterilizing combination. Here, we investigated the sterilizing effects of several all-oral regimens containing TB47 plus clofazimine and linezolid as a block and the roles of fluoroquinolones and pyrazinamide in them. All these regimens cured tuberculosis within 4 to 6 months in a well-established mouse model, and adding pyrazinamide showed a significant difference in bactericidal effects.
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Mycobacterium tuberculosis , Tuberculose , Animais , Antituberculosos/uso terapêutico , Clofazimina/uso terapêutico , Modelos Animais de Doenças , Linezolida , Camundongos , Camundongos Endogâmicos BALB C , Tuberculose/tratamento farmacológicoRESUMO
BACKGROUND: Structural change in the gut microbiota is implicated in cancer. The beneficial modulation of the microbiota composition with probiotics and prebiotics prevents diseases. AIM: We investigated the effect of oligofructose-maltodextrin-enriched Lactobacillus acidophilus, Bifidobacteria bifidum, and Bifidobacteria infantum (LBB), on the gut microbiota composition and progression of colorectal cancer. METHODS: Sprague Dawley rats were acclimatized, given ampicillin (75 mg/kg), and treated as follows; GCO: normal control; GPR: LBB only; GPC: LBB+ 1,2-dimethylhydrazine dihydrochloride (DMH); and GCA: DMH only (cancer control). 16S V4 Pyrosequencing for gut microbiota analysis, tumor studies, and the expression of MUC2, ZO-1, occludin, TLR2, TLR4, caspase 3, COX-2, and ß-catenin were conducted at the end of experiment. RESULTS: Probiotic LBB treatment altered the gut microbiota. The relative abundance of genera Pseudomonas, Congregibacter, Clostridium, Candidactus spp., Phaeobacter, Escherichia, Helicobacter, and HTCC was decreased (P < 0.05), but the genus Lactobacillus increased (P < 0.05), in LBB treatment than in cancer control. The altered gut microbiota was associated with decreased tumor incidence (80 % in GPC vs. 100 % in GCA, P = 0.0001), tumor volume (GPC 84.23 (42.75-188.4) mm(3) vs. GCA 243 (175.5-344.5) mm(3), P < 0.0001) and tumor multiplicity/count (GPC 2.92 ± 0.26 vs. GCA 6.27 ± 0.41; P < 0.0001). The expression of MUC2, ZO-1, occludin, and TLR2 was increased, but expression of TLR4, caspase 3, Cox-2, and ß-catenin was decreased by LBB treatment than in cancer control GCA (P < 0.05). CONCLUSION: Administration of LBB modulates the gut microbiota and reduces colon cancer development by decreasing tumor incidence, multiplicity/count, and volume via enhanced TLR2-improved gut mucosa epithelial barrier integrity and suppression of apoptosis and inflammation.
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Colo/efeitos dos fármacos , Neoplasias do Colo/metabolismo , Microbioma Gastrointestinal/efeitos dos fármacos , Probióticos/farmacologia , Receptor 2 Toll-Like/efeitos dos fármacos , 1,2-Dimetilidrazina/toxicidade , Animais , Bifidobacterium bifidum , Bifidobacterium longum subspecies infantis , Carcinógenos/toxicidade , Colo/metabolismo , Colo/microbiologia , Colo/patologia , Neoplasias do Colo/induzido quimicamente , Neoplasias do Colo/patologia , Ciclo-Oxigenase 2/genética , Modelos Animais de Doenças , Progressão da Doença , Imuno-Histoquímica , Lactobacillus acidophilus , Masculino , Mucina-2/genética , Neoplasias Experimentais/induzido quimicamente , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase em Tempo Real , Transdução de Sinais , Receptor 2 Toll-Like/genética , Receptor 4 Toll-Like/genética , Carga Tumoral/efeitos dos fármacosRESUMO
The increasing clinical significance of Mycobacterium abscessus is owed to its innate high-level, broad-spectrum resistance to antibiotics and therefore rapidly evolves as an important human pathogen. This warrants the identification of novel targets for aiding the discovery of new drugs or drug combinations to treat M. abscessus infections. This study is inspired by the drug-hypersensitive profile of a mutant M. abscessus (U14) with transposon insertion in MAB_1915. We validated the role of MAB_1915 in intrinsic drug resistance in M. abscessus by constructing a selectable marker-free in-frame deletion in MAB_1915 and complementing the mutant with the same or extended version of the gene and then followed by drug susceptibility testing. Judging by the putative function of MAB_1915, cell envelope permeability was studied by ethidium bromide accumulation assay and susceptibility testing against dyes and detergents. In this study, we established genetic evidence of the role of MAB_1915 in intrinsic resistance to rifampicin, rifabutin, linezolid, clarithromycin, vancomycin, and bedaquiline. Disruption of MAB_1915 has also been observed to cause a significant increase in cell envelope permeability in M. abscessus. Restoration of resistance is observed to depend on at least 27 base pairs upstream of the coding DNA sequence of MAB_1915. MAB_1915 could therefore be associated with cell envelope permeability, and hence its role in intrinsic resistance to multiple drugs in M. abscessus, which presents it as a novel target for future development of effective antimicrobials to overcome intrinsic drug resistance in M. abscessus. IMPORTANCE: This study reports the role of a putative fadD (MAB_1915) in innate resistance to multiple drugs by M. abscessus, hence identifying MAB_1915 as a valuable target and providing a baseline for further mechanistic studies and development of effective antimicrobials to check the high level of intrinsic resistance in this pathogen.
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Antituberculosos/uso terapêutico , Pneumonia/tratamento farmacológico , Tuberculose Resistente a Múltiplos Medicamentos/tratamento farmacológico , Adulto , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Pneumonia/etiologia , Estudos Retrospectivos , Resultado do Tratamento , Tuberculose Resistente a Múltiplos Medicamentos/complicaçõesRESUMO
BACKGROUND: The effect of childhood pneumococcal conjugate vaccine implementation in Malawi is threatened by absence of herd effect. There is persistent vaccine-type pneumococcal carriage in both vaccinated children and the wider community. We aimed to use a human infection study to measure 13-valent pneumococcal conjugate vaccine (PCV13) efficacy against pneumococcal carriage. METHODS: We did a double-blind, parallel-arm, randomised controlled trial investigating the efficacy of PCV13 or placebo against experimental pneumococcal carriage of Streptococcus pneumoniae serotype 6B (strain BHN418) among healthy adults (aged 18-40 years) from Blantyre, Malawi. We randomly assigned participants (1:1) to receive PCV13 or placebo. PCV13 and placebo doses were prepared by an unmasked pharmacist to maintain research team and participant masking with identification only by a randomisation identification number and barcode. 4 weeks after receiving either PCV13 or placebo, participants were challenged with 20â000 colony forming units (CFUs) per naris, 80â000 CFUs per naris, or 160â000 CFUs per naris by intranasal inoculation. The primary endpoint was experimental pneumococcal carriage, established by culture of nasal wash at 2, 7, and 14 days. Vaccine efficacy was estimated per protocol by means of a log-binomial model adjusting for inoculation dose. The trial is registered with the Pan African Clinical Trials Registry, PACTR202008503507113, and is now closed. FINDINGS: Recruitment commenced on April 27, 2021 and the final visit was completed on Sept 12, 2022. 204 participants completed the study protocol (98 PCV13, 106 placebo). There were lower carriage rates in the vaccine group at all three inoculation doses (0 of 21 vs two [11%] of 19 at 20â000 CFUs per naris; six [18%] of 33 vs 12 [29%] of 41 at 80â000 CFUs per naris, and four [9%] of 44 vs 16 [35%] of 46 at 160â000 CFUs per naris). The overall carriage rate was lower in the vaccine group compared with the placebo group (ten [10%] of 98 vs 30 [28%] of 106; Fisher's p value=0·0013) and the vaccine efficacy against carriage was estimated at 62·4% (95% CI 27·7-80·4). There were no severe adverse events related to vaccination or inoculation of pneumococci. INTERPRETATION: This is, to our knowledge, the first human challenge study to test the efficacy of a pneumococcal vaccine against pneumococcal carriage in Africa, which can now be used to establish vaccine-induced correlates of protection and compare alternative strategies to prevent pneumococcal carriage. This powerful tool could lead to new means to enhance reduction in pneumococcal carriage after vaccination. FUNDING: Wellcome Trust.
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Vacinas Pneumocócicas , Streptococcus pneumoniae , Adulto , Criança , Humanos , Malaui/epidemiologia , Vacinas Conjugadas , Sorogrupo , Vacinas Pneumocócicas/uso terapêuticoRESUMO
Objective: Rifampicin (RIF)-resistance, a surrogate marker for multidrug-resistant tuberculosis (TB), is mediated by mutations in the rpoB gene. We aimed to investigate the prevalence of mutations pattern in the entire rpoB gene of Mycobacterium tuberculosis clinical isolates and their association with resistance level to RIF. Methods: Among 465 clinical isolates collected from the Guangzhou Chest Hospital, drug-susceptibility of 175 confirmed Mtb strains was performed via the proportion method and Bactec MGIT 960 system. GeneXpert MTB/RIF and sanger sequencing facilitated in genetic characterization, whereas the MICs of RIF were determined by Alamar blue assay. Results: We found 150/175 (85.71%) RIF-resistant strains (MIC: 4 to >64 µg/mL) of which 57 were MDR and 81 pre-XDR TB. Genetic analysis identified 17 types of mutations 146/150 (97.33%) within RRDR (codons 426-452) of rpoB, mainly at L430 (P), D435 (V, E, G, N), H445 (N, D, Y, R, L), S450 (L, F) and L452 (P). D435V 12/146 (8.2%), H445N 16/146 (10.9%), and S450L 70/146 (47.94%) were the most frequently encountered mutations. Mutations Q432K, M434V, and N437D are rarely identified in RRDR. Deletions at (1284-1289 CCAGCT), (1295-1303 AATTCATGG), and insertion at (1300-1302 TTC) were detected within RRDR of three RIFR strains for the first time. We detected 47 types of mutations and insertions/deletions (indels) outside the RRDR. Four RIFR strains were detected with only novel mutations/indels outside the RRDR. Two of the four had (K274Q + C897 del + I491M) and (A286V + L494P), respectively. The other two had (G1687del + P454L) and (TT1835-6 ins + I491L) individually. Compared with phenotypic characterization, diagnostic sensitivities of GeneXpert MTB/RIF and sequencing analysis were 95.33% (143/150), and 100% (150/150) respectively. Conclusion: Our findings underscore the key role of RRDR mutations and the contribution of non-RRDR mutations in rapid molecular diagnosis of RIFR clinical isolates. Such insights will support early detection of disease and recommend the appropriate anti-TB regimens in high-burden settings.
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Drug-resistant tuberculosis (DR-TB) poses a new threat to global health; to improve the treatment outcome, therapeutic vaccines are considered the best chemotherapy adjuvants. Unfortunately, there is no therapeutic vaccine approved against DR-TB. Our study assessed the therapeutic efficacy of a recombinant drug-resistant BCG (RdrBCG) vaccine in DR-TB. We constructed the RdrBCG overexpressing Ag85B and Rv2628 by selecting drug-resistant BCG strains and transformed them with plasmid pEBCG or pIBCG to create RdrBCG-E and RdrBCG-I respectively. Following successful stability testing, we tested the vaccine's safety in severe combined immune deficient (SCID) mice that lack both T and B lymphocytes plus immunoglobulins. Finally, we evaluated the RdrBCG's therapeutic efficacy in BALB/c mice infected with rifampin-resistant M. tuberculosis and treated with a second-line anti-TB regimen. We obtained M. bovis strains which were resistant to several second-line drugs and M. tuberculosis resistant to rifampin. Notably, the exogenously inserted genes were lost in RdrBCG-E but remained stable in the RdrBCG-I both in vitro and in vivo. When administered adjunct to a second-line anti-TB regimen in a murine model of DR-TB, the RdrBCG-I lowered lung M. tuberculosis burden by 1 log10. Furthermore, vaccination with RdrBCG-I adjunct to chemotherapy minimized lung tissue pathology in mice. Most importantly, the RdrBCG-I showed almost the same virulence as its parent BCG Tice strain in SCID mice. Our findings suggested that the RdrBCG-I was stable, safe and effective as a therapeutic vaccine. Hence, the "recombinant" plus "drug-resistant" BCG strategy could be a useful concept for developing therapeutic vaccines against DR-TB.
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Antituberculosos/farmacologia , Vacina BCG/imunologia , Farmacorresistência Bacteriana/genética , Mycobacterium bovis/genética , Mycobacterium tuberculosis/efeitos dos fármacos , Tuberculose Pulmonar/prevenção & controle , Vacinas Sintéticas/imunologia , Amicacina/farmacologia , Amicacina/uso terapêutico , Animais , Antígenos de Bactérias/biossíntese , Antígenos de Bactérias/genética , Antígenos de Bactérias/imunologia , Antituberculosos/uso terapêutico , Vacina BCG/biossíntese , Vacina BCG/genética , Vacina BCG/uso terapêutico , Modelos Animais de Doenças , Levofloxacino/farmacologia , Levofloxacino/uso terapêutico , Camundongos Endogâmicos BALB C , Camundongos SCID , Mycobacterium bovis/química , Mycobacterium bovis/efeitos dos fármacos , Mycobacterium tuberculosis/patogenicidade , Plasmídeos , Protionamida/farmacologia , Protionamida/uso terapêutico , Pirazinamida/farmacologia , Pirazinamida/uso terapêutico , Tuberculose Pulmonar/tratamento farmacológico , Tuberculose Pulmonar/patologia , Vacinas Sintéticas/biossíntese , Vacinas Sintéticas/genética , Vacinas Sintéticas/uso terapêutico , VirulênciaRESUMO
Multidrug-resistant tuberculosis (MDR-TB) remains a serious public health threat worldwide. To date, the anti-TB activity of TB47 (T), an imidazopyridine amide class of antibiotics targeting QcrB in the electron transport chain, has not been systematically evaluated, especially in a new regimen against MDR-TB. This study employed both macrophage infection and a mouse model to test the activity of T alone or in combination with other antimicrobial agents. Different regimens containing amikacin (A), levofloxacin (L), ethambutol (E), and pyrazinamide (Z) + clofazimine (C)/T were evaluated in the mouse model. The bacterial burdens of mice from different groups were monitored at different time points while relapse was assessed 6 months after treatment cessation. Colonies obtained at relapse underwent drug susceptibility testing. We found that T exhibited highly synergistic bactericidal activity with C in all models. Adding T to ALEZC might shorten the MDR-TB treatment duration from ≥ 9 months to ≤ 5months, as five months of treatment with ALEZCT achieved zero relapse rates in 2 animal experiments. These findings indicate that T exhibits a highly synergistic sterilizing activity when combined with C. All isolates from relapsing mice remained sensitive to each drug, suggesting that the relapse was not due to drug resistance but rather associated with the type of regimen.
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Antituberculosos/farmacologia , Clofazimina/farmacologia , Mycobacterium tuberculosis/efeitos dos fármacos , Tuberculose Resistente a Múltiplos Medicamentos/tratamento farmacológico , Animais , Antituberculosos/administração & dosagem , Antituberculosos/química , Clofazimina/administração & dosagem , Modelos Animais de Doenças , Sinergismo Farmacológico , Feminino , Imidazóis/administração & dosagem , Imidazóis/farmacologia , Camundongos , Camundongos Endogâmicos BALB C , Testes de Sensibilidade Microbiana , Mycobacterium tuberculosis/isolamento & purificação , Piridinas/administração & dosagem , Piridinas/farmacologia , Recidiva , Tuberculose Resistente a Múltiplos Medicamentos/microbiologiaRESUMO
Buruli ulcer (BU) is an emerging infectious disease that causes disfiguring skin ulcers. The causative agent, Mycobacterium ulcerans, secretes toxin called mycolactone that triggers inflammation and immunopathology. Existing treatments are lengthy and consist of drugs developed for tuberculosis. Here, we report that a pyrazolo[1,5-a]pyridine-3-carboxamide, TB47, is highly bactericidal against M. ulcerans both in vitro and in vivo. In the validated mouse model of BU, TB47 alone reduces M. ulcerans burden in mouse footpads by more than 2.5 log10 CFU compared to the standard BU treatment regimen recommended by the WHO. We show that mutations of ubiquinol-cytochrome C reductase cytochrome subunit B confer resistance to TB47 and the dissimilarity of CydABs from different mycobacteria may account for their differences in susceptibility to TB47. TB47 is highly potent against M. ulcerans and possesses desirable pharmacological attributes and low toxicity that warrant further assessment of this agent for treatment of BU.
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Antibacterianos/uso terapêutico , Úlcera de Buruli/tratamento farmacológico , Úlcera de Buruli/microbiologia , Mycobacterium ulcerans/efeitos dos fármacos , Mycobacterium ulcerans/patogenicidade , Animais , Complexo III da Cadeia de Transporte de Elétrons/genética , Camundongos , Mutação , Mycobacterium ulcerans/genéticaRESUMO
Ethionamide (ETA) and prothionamide (PRO) are interchangeably used in tuberculosis (TB) chemotherapy regimens. Subtle discrepancies between biochemical and genetic information on the modes of sensitivity and resistance of isoniazid (INH) and ETA warrants further studies. We report a new mutation - EthAW21R - in Mycobacterium bovis Bacillus Calmette-Guérin that corresponds with co-resistance to both PRO and ETA, which to the best of our knowledge has not been reported before. Our findings suggest that mutation EthAW21R could be used as a marker site for testing PRO and ETA cross-resistance.
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Our research group previously isolated and identified a strain of pathogenic Escherichia coli from clinical samples called E. coli O124 K72. The present study was aimed at determining the potential effects of E. coli O124 K72 on intestinal barrier functions and structural proteins integrity in guinea pig. Guinea pigs were grouped into three groups; control (CG); E. coli O124 K72 (E. coli); and probiotics Lactobacillus rhamnosus (LGG). Initially, we create intestinal dysbiosis by giving all animals Levofloxacin for 10days, but the control group (CG) received the same volume of saline. Then, the animals received either E. coli O124 K72 (E. coli) or Lactobacillus rhamnosus (LGG) according to their assigned group. E. coli O124 K72 treatment significantly affected colon morphology and distorted intestinal barrier function by up-regulating Claudin2 and down-regulating Occludin. In addition, E. coli upregulated the mRNA expression of MUC1, MUC2, MUC13 and MUC15. Furthermore, suspected tumor was found in the E. coli treated animals. Our results suggested that E. coli O124 K72 strain has adverse effects on intestinal barrier functions and is capable of altering integrity of structural proteins in guinea pig model while at same time it may have a role in colon carcinogenesis.
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Escherichia coli/fisiologia , Intestinos/microbiologia , Intestinos/patologia , Proteínas de Junções Íntimas/metabolismo , Animais , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Claudina-2/metabolismo , Eletroforese em Gel de Gradiente Desnaturante , Modelos Animais de Doenças , Disbiose/tratamento farmacológico , Disbiose/metabolismo , Disbiose/microbiologia , Disbiose/patologia , Microbioma Gastrointestinal , Cobaias , Intestinos/efeitos dos fármacos , Mucina-2/metabolismo , Ocludina/metabolismoRESUMO
BACKGROUND/AIM: Probiotics have been suggested as prophylactic measure in colon carcinogenesis. This study aimed at determining the potential prophylactic activity of Lactobacillus rhamnosus GG CGMCC 1.2134 (LGG) strain on colorectal carcinogenesis via measuring its effect on Nuclear factor kappa B (NFκB) inflammatory pathway and apoptosis. MATERIALS AND METHODS: 64 Sprague Dawley rats were grouped into four as follows; Group 1 (Healthy control), Group 2 (LGG), Group 3 (cancer control Dimethyl hydrazine (DMH)) and Group 4 (LGG+DMH). LGG was administered orally to LGG and LGG+DMH groups. Colon carcinogenesis was chemically induced in LGG+DMH and DMH groups by weekly injection of 40mg/kg DMH. Animals were sacrificed after 25 weeks of experiment and tumor characteristics assessed. The change in expression of NFκB-p65, COX-2, TNFα, Bcl-2, Bax, iNOS, VEGFα, ß-catenin, Casp3 and p53 were evaluated by western blotting and qRT-PCR. RESULTS: LGG treatment significantly reduced tumor incidence, multiplicity and volume in LGG+DMH treatment group compared to DMH cancer control group. Also, LGG treatment reduced the expression of ß-catenin and the inflammatory proteins NFκB-p65, COX-2 and TNFα; the anti-apoptotic protein Bcl-2, but increased the expression of the pro-apoptotic proteins Bax, casp3 and p53 compared with DMH group. CONCLUSION: LGG have a potential protection effect against colon carcinogenesis; inducing apoptosis and ameliorating inflammation, and may hold a promise as bio-therapeutic dietary agent.