reference strain m abscessus atcc 19977 t  (ATCC)

Journal: Antimicrobial Agents and Chemotherapy

Article Title: Protective mechanism of action of the antifungal drug naftifine against Mycobacterium abscessus infection

doi: 10.1128/aac.01105-25

Figure Lengend Snippet: Naftifine inhibits intracellular M. abscessus growth in a concentration- and time-dependent manner. THP-1 macrophages were infected with ( A ) ATCC 19977 T smooth variant, ( B ) ATCC 19977 T rough variant, and clinical isolates ( C ) 23-S-10, ( D ) 23-S-12, and ( E ) 23-S-13. The ATCC 19977 T strains are sensitive to both clarithromycin and amikacin, while the clinical isolates are resistant to both antibiotics. Following infection, cells were treated with increasing concentrations of naftifine (3–90 μM) or 10 μM clarithromycin as a control. Intracellular bacterial burden was quantified by CFU enumeration at 24, 48, and 72 hours post-treatment. Data represent mean ± SD from three independent experiments. Statistical significance was determined by two-way ANOVA with Tukey’s multiple comparisons test for comparisons between each naftifine concentration and the DMSO control at corresponding time points. ** P < 0.01, *** P < 0.001, and **** P < 0.0001.

Article Snippet: The reference strain M. abscessus ATCC 19977 T was obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: Concentration Assay, Infection, Variant Assay, Control

Journal: Antimicrobial Agents and Chemotherapy

Article Title: Protective mechanism of action of the antifungal drug naftifine against Mycobacterium abscessus infection

doi: 10.1128/aac.01105-25

Figure Lengend Snippet: Dose-dependent cytotoxicity analysis in uninfected and M. abscessus -infected cells. ( A ) Representative flow cytometry dot plots show Annexin V and 7-AAD staining patterns in uninfected (UI) and infected (IN) cells treated with increasing concentrations (0, 10, 30, and 90 μM) of the test compound. THP-1 cells were infected with M. abscessus ATCC 19977 T smooth variants and treated with naftifine for 3 days. Gated regions indicate dead cells (Annexin V + , including early and late apoptotic and necrotic populations). ( B and C ) Quantification of cell death shows the percentage of Annexin V + population (both 7-AAD + and 7-AAD⁻) in uninfected ( B ) and infected ( C ) cells. Data represent mean ± SEM from at least three independent experiments. Statistical significance was determined by one-way ANOVA followed by Dunnett’s post hoc test for multiple comparisons against the control. ** P < 0.01 and **** P < 0.0001 compared to untreated control (0 μM).

Article Snippet: The reference strain M. abscessus ATCC 19977 T was obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: Infection, Flow Cytometry, Staining, Control

Journal: Antimicrobial Agents and Chemotherapy

Article Title: Protective mechanism of action of the antifungal drug naftifine against Mycobacterium abscessus infection

doi: 10.1128/aac.01105-25

Figure Lengend Snippet: Naftifine exhibits bacteriostatic activity against M. abscessus in axenic culture. Time-kill assays were performed to evaluate the antimicrobial activity of naftifine against different M. abscessus strains and morphological variants. ( A ) ATCC 19977 T smooth variant, ( B ) ATCC 19977 T rough variant, and clinical isolates ( C ) 23-S-10, ( D ) 23-S-12, and ( E ) 23-S-13. The ATCC 19977 T strains are sensitive to both clarithromycin and amikacin, while the clinical isolates are resistant to both antibiotics. Bacterial cultures in logarithmic growth phase were treated with naftifine at concentrations ranging from 2 to 64 µg/mL (equivalent to 6–198 μM) or DMSO vehicle control. Samples were collected at 24-hour intervals over a 96-hour incubation period, and bacterial viability was assessed by CFU enumeration following serial dilution plating. Data represent mean ± SEM from two independent experiments performed in triplicate. Statistical comparisons between naftifine-treated and DMSO control at each time point were performed using two-way ANOVA with Tukey’s multiple comparisons test. Statistical significance is indicated as * P < 0.05, ** P < 0.01, *** P < 0.001, and **** P < 0.0001.

Article Snippet: The reference strain M. abscessus ATCC 19977 T was obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: Activity Assay, Variant Assay, Control, Incubation, Serial Dilution

Journal: Antimicrobial Agents and Chemotherapy

Article Title: Protective mechanism of action of the antifungal drug naftifine against Mycobacterium abscessus infection

doi: 10.1128/aac.01105-25

Figure Lengend Snippet: Naftifine reduces M. abscessus burden in the lungs and spleens of immunosuppressed mice. BALB/c mice were immunosuppressed with dexamethasone and subsequently infected intranasally with 1,000 CFU of M. abscessus ATCC 19977 T rough variant. Treatment began 3 days post-infection and continued once daily for 14 days. Mice received either naftifine HCl (50 mg/kg of body weight, intraperitoneal), cefoxitin (200 mg/kg of body weight, subcutaneous, positive control), or PBS vehicle control (intraperitoneal). After the treatment period, mice were euthanized, and bacterial loads in the ( A ) lungs and ( B ) spleens were quantified by CFU enumeration and expressed as log 10 CFU per organ. Data represent individual mice with mean ± SEM. ( C ) Representative hematoxylin and eosin-stained lung sections from infected mice treated with PBS vehicle control (top panels) or naftifine (bottom panels) are shown at 4×, 10×, and 40× magnifications to illustrate histopathological changes. Statistical comparisons between treatment groups were performed with the Kruskal–Wallis test followed by Dunn’s multiple comparisons test. * P < 0.05; ** P < 0.01; *** P < 0.001 and ns, not significant.

Article Snippet: The reference strain M. abscessus ATCC 19977 T was obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: Infection, Variant Assay, Positive Control, Control, Staining

Journal: Antimicrobial Agents and Chemotherapy

Article Title: Protective mechanism of action of the antifungal drug naftifine against Mycobacterium abscessus infection

doi: 10.1128/aac.01105-25

Figure Lengend Snippet: Autophagy contributes to the intracellular efficacy of naftifine against M. abscessus . Intracellular bacterial survival (log 10 CFU) of ( A ) naftifine-sensitive M. abscessus ATCC 19977 T S (Mab S, blue) strain and naftifine-resistant M. abscessus strain (Mab-Naft R1, red) in wild-type RAW 264.7 macrophages (filled circles) or ( B ) ATG16L1 knockdown macrophages (open circles) treated with naftifine at 0, 5, 10, and 20 µM. Bacterial loads were quantified on days 1 (D1) and 2 (D2) post-infection. ( C and D ) Representative immunoblots showing LC3-I/LC3-II expression and β-actin (loading control) in macrophage lysates at day 1 post-infection with ( C ) Mab-Naft S or ( D ) Mab-Naft R1, treated with indicated naftifine concentrations (μM). Data represent mean ± SEM. Statistical significance was determined by two-way ANOVA. ** P < 0.01, *** P < 0.001, and **** P < 0.0001.

Article Snippet: The reference strain M. abscessus ATCC 19977 T was obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: Knockdown, Infection, Western Blot, Expressing, Control

Journal: Antimicrobial Agents and Chemotherapy

Article Title: Protective mechanism of action of the antifungal drug naftifine against Mycobacterium abscessus infection

doi: 10.1128/aac.01105-25

Figure Lengend Snippet: Synergistic interactions between naftifine and β-lactam antibiotics against M. abscessus . Checkerboard assays were performed to evaluate drug interactions between naftifine ( y -axis) and various antibiotics ( x -axis) against M. abscessus ATCC 19977 T smooth variant. Heat maps display the percentage of bacterial growth, with darker blue indicating greater growth. Twofold serial dilutions of each drug were tested in combination, and growth inhibition was assessed after incubation. Naftifine showed indifferent interactions with clarithromycin ( A ) and tigecycline ( E ), additive/partial synergistic effects with amikacin ( B ), but demonstrated synergistic effects when combined with the β-lactam antibiotics cefoxitin ( C ) and imipenem ( D ), particularly at sub-inhibitory concentrations. Synergy was defined as FICI ≤ 0.5, where the combined effect significantly exceeded the sum of individual drug activities. Data were analyzed and plotted using GraphPad Prism version 10.

Article Snippet: The reference strain M. abscessus ATCC 19977 T was obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: Variant Assay, Inhibition, Incubation

Journal: Antimicrobial Agents and Chemotherapy

Article Title: Protective mechanism of action of the antifungal drug naftifine against Mycobacterium abscessus infection

doi: 10.1128/aac.01105-25

Figure Lengend Snippet: Naftifine inhibits intracellular M. abscessus growth in a concentration- and time-dependent manner. THP-1 macrophages were infected with ( A ) ATCC 19977 T smooth variant, ( B ) ATCC 19977 T rough variant, and clinical isolates ( C ) 23-S-10, ( D ) 23-S-12, and ( E ) 23-S-13. The ATCC 19977 T strains are sensitive to both clarithromycin and amikacin, while the clinical isolates are resistant to both antibiotics. Following infection, cells were treated with increasing concentrations of naftifine (3–90 μM) or 10 μM clarithromycin as a control. Intracellular bacterial burden was quantified by CFU enumeration at 24, 48, and 72 hours post-treatment. Data represent mean ± SD from three independent experiments. Statistical significance was determined by two-way ANOVA with Tukey’s multiple comparisons test for comparisons between each naftifine concentration and the DMSO control at corresponding time points. ** P < 0.01, *** P < 0.001, and **** P < 0.0001.

Article Snippet: The tested strains included the reference ATCC 19977 T strain in both smooth and rough morphotypes and clinical isolates of M. abscessus subsp. abscessus harboring mutations that confer resistance to amikacin and clarithromycin ( ).

Techniques: Concentration Assay, Infection, Variant Assay, Control

Journal: Antimicrobial Agents and Chemotherapy

Article Title: Protective mechanism of action of the antifungal drug naftifine against Mycobacterium abscessus infection

doi: 10.1128/aac.01105-25

Figure Lengend Snippet: Dose-dependent cytotoxicity analysis in uninfected and M. abscessus -infected cells. ( A ) Representative flow cytometry dot plots show Annexin V and 7-AAD staining patterns in uninfected (UI) and infected (IN) cells treated with increasing concentrations (0, 10, 30, and 90 μM) of the test compound. THP-1 cells were infected with M. abscessus ATCC 19977 T smooth variants and treated with naftifine for 3 days. Gated regions indicate dead cells (Annexin V + , including early and late apoptotic and necrotic populations). ( B and C ) Quantification of cell death shows the percentage of Annexin V + population (both 7-AAD + and 7-AAD⁻) in uninfected ( B ) and infected ( C ) cells. Data represent mean ± SEM from at least three independent experiments. Statistical significance was determined by one-way ANOVA followed by Dunnett’s post hoc test for multiple comparisons against the control. ** P < 0.01 and **** P < 0.0001 compared to untreated control (0 μM).

Article Snippet: The tested strains included the reference ATCC 19977 T strain in both smooth and rough morphotypes and clinical isolates of M. abscessus subsp. abscessus harboring mutations that confer resistance to amikacin and clarithromycin ( ).

Techniques: Infection, Flow Cytometry, Staining, Control

Journal: Antimicrobial Agents and Chemotherapy

Article Title: Protective mechanism of action of the antifungal drug naftifine against Mycobacterium abscessus infection

doi: 10.1128/aac.01105-25

Figure Lengend Snippet: Naftifine exhibits bacteriostatic activity against M. abscessus in axenic culture. Time-kill assays were performed to evaluate the antimicrobial activity of naftifine against different M. abscessus strains and morphological variants. ( A ) ATCC 19977 T smooth variant, ( B ) ATCC 19977 T rough variant, and clinical isolates ( C ) 23-S-10, ( D ) 23-S-12, and ( E ) 23-S-13. The ATCC 19977 T strains are sensitive to both clarithromycin and amikacin, while the clinical isolates are resistant to both antibiotics. Bacterial cultures in logarithmic growth phase were treated with naftifine at concentrations ranging from 2 to 64 µg/mL (equivalent to 6–198 μM) or DMSO vehicle control. Samples were collected at 24-hour intervals over a 96-hour incubation period, and bacterial viability was assessed by CFU enumeration following serial dilution plating. Data represent mean ± SEM from two independent experiments performed in triplicate. Statistical comparisons between naftifine-treated and DMSO control at each time point were performed using two-way ANOVA with Tukey’s multiple comparisons test. Statistical significance is indicated as * P < 0.05, ** P < 0.01, *** P < 0.001, and **** P < 0.0001.

Article Snippet: The tested strains included the reference ATCC 19977 T strain in both smooth and rough morphotypes and clinical isolates of M. abscessus subsp. abscessus harboring mutations that confer resistance to amikacin and clarithromycin ( ).

Techniques: Activity Assay, Variant Assay, Control, Incubation, Serial Dilution

Journal: Antimicrobial Agents and Chemotherapy

Article Title: Protective mechanism of action of the antifungal drug naftifine against Mycobacterium abscessus infection

doi: 10.1128/aac.01105-25

Figure Lengend Snippet: Naftifine reduces M. abscessus burden in the lungs and spleens of immunosuppressed mice. BALB/c mice were immunosuppressed with dexamethasone and subsequently infected intranasally with 1,000 CFU of M. abscessus ATCC 19977 T rough variant. Treatment began 3 days post-infection and continued once daily for 14 days. Mice received either naftifine HCl (50 mg/kg of body weight, intraperitoneal), cefoxitin (200 mg/kg of body weight, subcutaneous, positive control), or PBS vehicle control (intraperitoneal). After the treatment period, mice were euthanized, and bacterial loads in the ( A ) lungs and ( B ) spleens were quantified by CFU enumeration and expressed as log 10 CFU per organ. Data represent individual mice with mean ± SEM. ( C ) Representative hematoxylin and eosin-stained lung sections from infected mice treated with PBS vehicle control (top panels) or naftifine (bottom panels) are shown at 4×, 10×, and 40× magnifications to illustrate histopathological changes. Statistical comparisons between treatment groups were performed with the Kruskal–Wallis test followed by Dunn’s multiple comparisons test. * P < 0.05; ** P < 0.01; *** P < 0.001 and ns, not significant.

Article Snippet: The tested strains included the reference ATCC 19977 T strain in both smooth and rough morphotypes and clinical isolates of M. abscessus subsp. abscessus harboring mutations that confer resistance to amikacin and clarithromycin ( ).

Techniques: Infection, Variant Assay, Positive Control, Control, Staining

Journal: Antimicrobial Agents and Chemotherapy

Article Title: Protective mechanism of action of the antifungal drug naftifine against Mycobacterium abscessus infection

doi: 10.1128/aac.01105-25

Figure Lengend Snippet: Autophagy contributes to the intracellular efficacy of naftifine against M. abscessus . Intracellular bacterial survival (log 10 CFU) of ( A ) naftifine-sensitive M. abscessus ATCC 19977 T S (Mab S, blue) strain and naftifine-resistant M. abscessus strain (Mab-Naft R1, red) in wild-type RAW 264.7 macrophages (filled circles) or ( B ) ATG16L1 knockdown macrophages (open circles) treated with naftifine at 0, 5, 10, and 20 µM. Bacterial loads were quantified on days 1 (D1) and 2 (D2) post-infection. ( C and D ) Representative immunoblots showing LC3-I/LC3-II expression and β-actin (loading control) in macrophage lysates at day 1 post-infection with ( C ) Mab-Naft S or ( D ) Mab-Naft R1, treated with indicated naftifine concentrations (μM). Data represent mean ± SEM. Statistical significance was determined by two-way ANOVA. ** P < 0.01, *** P < 0.001, and **** P < 0.0001.

Article Snippet: The tested strains included the reference ATCC 19977 T strain in both smooth and rough morphotypes and clinical isolates of M. abscessus subsp. abscessus harboring mutations that confer resistance to amikacin and clarithromycin ( ).

Techniques: Knockdown, Infection, Western Blot, Expressing, Control

Journal: Antimicrobial Agents and Chemotherapy

Article Title: Protective mechanism of action of the antifungal drug naftifine against Mycobacterium abscessus infection

doi: 10.1128/aac.01105-25

Figure Lengend Snippet: Synergistic interactions between naftifine and β-lactam antibiotics against M. abscessus . Checkerboard assays were performed to evaluate drug interactions between naftifine ( y -axis) and various antibiotics ( x -axis) against M. abscessus ATCC 19977 T smooth variant. Heat maps display the percentage of bacterial growth, with darker blue indicating greater growth. Twofold serial dilutions of each drug were tested in combination, and growth inhibition was assessed after incubation. Naftifine showed indifferent interactions with clarithromycin ( A ) and tigecycline ( E ), additive/partial synergistic effects with amikacin ( B ), but demonstrated synergistic effects when combined with the β-lactam antibiotics cefoxitin ( C ) and imipenem ( D ), particularly at sub-inhibitory concentrations. Synergy was defined as FICI ≤ 0.5, where the combined effect significantly exceeded the sum of individual drug activities. Data were analyzed and plotted using GraphPad Prism version 10.

Article Snippet: The tested strains included the reference ATCC 19977 T strain in both smooth and rough morphotypes and clinical isolates of M. abscessus subsp. abscessus harboring mutations that confer resistance to amikacin and clarithromycin ( ).

Techniques: Variant Assay, Inhibition, Incubation