BACKGROUND AND OBJECTIVE

Nontuberculous mycobacteria (NTM) lung disease appears like tuberculosis infection but is resistant to primary anti-tuberculosis drugs. Hence, patients whose sputum sample tests positive for acid-fast bacilli (AFB) and bacterial culture for several times should be assessed for colonization or infection with NTM in a damaged lung secondary to TB. In such cases, though drug-resistant TB may be adequately treated, treatment may need to be directed towards the NTM as well. In NTM therapy, the duration and choice of treatment agent is based upon the specific organism and disease extent. This study used one-step multiplex PCR (mPCR) assay for rapid differentiation of solid cultures in Ogawa medium as Mycobacterium tuberculosis (MTB) and/or NTM.

A total of 80 stocked isolates obtained from the Lung Center of the Philippines from January to December 2018 were screened for NTM in terms of growth in Ogawa medium, acid fastness, and MPT64 TB antigen test result. These were from sputum specimens of multidrug-resistant tuberculosis (MDR-TB) patients. DNA was extracted from cultures (n=55) grown in Ogawa medium and one-step mPCR was performed to identify NTM to the species level.

Out of 80 samples screened, a total of 55 isolates were identified as NTM. One-step mPCR identified 12.73% (7/55) as M. abscessus, 34.55% (19/55) as M. massiliense, 1.82% (1/55) as M. kansasii, and 50.91% (28/55) were identified only up to genus Mycobacteria spp. Neither M. avium complex nor M. intracellulare was identified among the samples tested.

One-step mPCR was able to identify isolates as MTB or NTM coinciding with the initial screening using MPT64 TB antigen test. Multiplex PCR has given a more specific identificati on to the species level. The use of mPCR in identifying MTB and clinically significant NTM’s is suitable for the adequate treatment of mycobacterial infection.

BACKGROUND AND OBJECTIVE

The human nasal passages host major human pathogens. Recent research suggests that the microbial communities inhabiting the epithelial surfaces of the nasal passages play a key factor in maintaining a healthy microenvironment by affecting both resistance to pathogens and immunological responses. Colonization of the nasal cavity by different pathogens such as Staphylococcus aureus and Klebsiella aerogenes, is associated with a higher postoperative infection morbidity. Povidone-iodine (PVP-I) as an antiseptic has been proven to display high antibacterial, antiviral, and antifungal properties even at low concentrations, and was shown to be effective in the control of infections to limit their impact and spread. It can be used as a topical antiseptic for skin decontamination and wound management, as a nasal spray, or as a gargle. There are different methods in testing the efficacy of potential antimicrobial suspensions. This study aimed to determine the concentration of PVP-I that is most effective in nasal decolonization using microsuspension test and colorimetric minimum inhibitory concentration (MIC) determination assays, resazurin microtiter assay (REMA), and Dey-Engley (D/E) neutralizer assay. The findings of this study will contribute to knowledge regarding the intended use of PVP-I in microbial control, particularly in bacterial infections.

Several dilutions (2.0%, 1.0%, 0.5%, 0.25%, 0.1% and 0.09%) of commercially bought 10% (10 mg per 100 ml) povidone-iodine were prepared and tested against a standardized inoculum (1x105) of Staphylococcus aureus and Klebsiella aerogenes at different contacttimes (5 seconds, 10 seconds, 30 seconds, 1 minute, and 5 minutes). Microdilution suspension test was performed to determine the log reduction per variable, while REMA and D/E neutralizer assay were used to determine the MIC. A value of greater than or equal to 5 log reduction was considered effective for microdilution suspension test. Estimates of agreement statistics were used to interpret the results of the assay in which the overall percent agreement (OPA), positive percent agreement (PPA), negative percent agreement (NPA), and Cohen’s kappa statistics were calculated.

Povidone-iodine concentration of 0.25% exhibited ?5 log reduction against K. aerogenes at the minimum contact time of 5 seconds. On the other hand,  a slightly higher PVP-I concentration was required to achieve ?5 log reduction for S. aureus at 0.5% concentration and a minimum contact time of 1 minute. There was an observed concordance of the results of REMA and D/E neutralizer as MIC colorimetric indicators, which yielded an overall test percent agreement of 90.30% (95% CI: 84.73–94.36), and a strong level of agreement (? = 0.8, pCONCLUSION

Low povidone-iodine concentrations (i.e., 0.5% against S. aureus and 0.25% against K. aerogenes) were observed to have bactericidal activity of at least 5 log reduction as rapid as the minimum contact time of 5 seconds. Furthermore, D/E and REMA, as colorimetric indicators, had comparable performance (OPA = 90.30%; ? = 0.8, p
Human ; Bacteria ; Povidone-iodine ; Microbial Sensitivity Tests ; Anti-infective Agents, Local ; Enterobacter Aerogenes ; Staphylococcus Aureus

BACKGROUND AND OBJECTIVE

Methicillin-resistant Staphylococcus aureus (MRSA) is one of the leading causes of hospital and community-acquired infections, showing antimicrobial resistance (AMR), which is an increasing public health concern. One of the commonly-used methods to evaluate resistance include the Kirby-Bauer disk diffusion method. However, this test is found to be time-consuming, lacking in terms of mechanization and automation, alongside its non-applicability to certain antibiotics such as vancomycin. Thus, the Clinical Laboratory Standards Institute (CLSI) recommends using the broth microdilution method in the evaluation of antibacterial activities against S. aureus. A rapid laboratory identification of MRSA is important in the treatment of patients. Therefore, this study aims to optimize and evaluate the effectiveness of a rapid microplate assay using resazurin dye as a colorimetric indicator in determining antibacterial activity against clinical isolates of MRSA and methicillin-susceptible S. aureus (MSSA).

Clinical isolates of MRSA and MSSA were obtained from the Philippine General Hospital (PGH) Microbiology Section, and American Type Culture Collection (ATCC) controls of both strains (ATCC 25923 and ATCC 43300) were acquired. These were then subjected to identification and confirmation procedures. A standardization of bacterial inoculum was performed by comparing its 24-hr growth in Mueller Hinton Broth to 0.5 McFarland Standard. The resazurin microplate assay (REMA) was set-up using two-fold serial dilution of control antibiotics such as oxacillin, vancomycin, and cefoxitin. Each plate was inoculated with standardized bacterial growth of controls and clinical isolates. To determine the time needed for the reduction of the resazurin dye, a qualitative assessment was conducted by comparing the reaction time between a 6.75 mg/mL dye with a 0.01 mg/mL dye. The plates were also subjected to different incubation times and dye concentrations, and the optical densities of the plates were compared using a microplate reader.

Results showed that there were no significant differences between the optical densities of the wells of those incubated for 5 hours and for 24 hours (p >0.05). Furthermore, there was a significant reduction in the reaction time of the dye (from 18 hours to 1 hour) when the dye concentration was reduced from 6.75 mg/mL to 0.01 mg/mL. The optimized REMA showed a significant difference between the minimum inhibitory concentrations (MICs) of the different antibiotics against the control and isolate strains of MRSA and MSSA, showing a W of -2.98 (pCONCLUSION

Based on the results presented, the researchers determined the optimal condition for the resazurin microtiter assay, which was 0.01 g/mL concentration of resazurin dye, at a 5-hour incubation period. This study has shown that an optimized REMA is an efficient and fast method to determine the antimicrobial activities of oxacillin, cefoxitin, and vancomycin against MRSA and MSSA.

INTRODUCTION

Neurolisteriosis is caused by Listeria monocytogenes, a gram-positive microorganism. It usually affects vulnerable population including pregnant women, neonates, immunocompromised individuals, and elderly persons. This report describes a case of neurolisteriosis in a 31-year-old immunocompetent man.

This case involves a 31-year-old Filipino male who presented with decrease sensorium. A lumbar puncture was performed, and polymerase chain reaction (PCR) testing of the cerebrospinal fluid confirmed the presence of Listeria monocytogenes. On the fifth day of hospitalization, the patient developed unilateral sixth cranial nerve palsy and facial nerve palsy. He was treated with intravenous ampicillin for 21 days, resulting in significant improvement in the cranial nerve deficits.

It is the first neurolisteriosis case in this institution. There is only one published neurolisteriosis case in the Philippines which presented with brain abscess. Neurolisteriosis, although uncommon, is one of the differential diagnoses in patients presenting with fever, headache, and nuchal rigidity. Isolation of Listeria monocytogenes in the cerebrospinal fluid and blood culture is diagnostic. Neurolisteriosis is an invasive disease which can result in neurologic sequalae such as cranial nerve palsies. Targeted treatment aids in good clinical outcomes.

Early secreted antigenic target of 6 kDa protein (ESAT-6) is the major virulence factor of Mycobacterium tuberculosis (MTB), which can resist the clearance of MTB in bodies by inhibiting macrophage phagocytosis and autophagy reaction, thus impeding the immune defense function of the body against MTB infection. In addition, ESAT-6-induced apoptosis of macrophage and massive necrosis of innate immune cells can foster MTB proliferation and colonization, leading to systemic MTB infection. Moreover, ESAT-6 hampers the protective immune response of Th1 cells, reducing the secretion of pro-inflammatory cytokines and contributing to immune dysfunction, thus accelerating the course of MTB infection. During the process, the high immunogenicity of ESAT-6 can be leveraged as a dominant antigen in the development of new TB vaccines, making it a promising candidate with broad prospects for further development.
Humans ; Mycobacterium tuberculosis ; Vaccines ; Cytokines ; Apoptosis ; Autophagy ; Sepsis

Background and Objective: Methicillin-resistant Staphylococcus aureus (MRSA) is one of the leading causes of hospital and community-acquired infections, showing antimicrobial resistance (AMR), which is an increasing public health concern. One of the commonly-used methods to evaluate resistance include the Kirby-Bauer disk diffusion method. However, this test is found to be time-consuming, lacking in terms of mechanization and automation, alongside its non-applicability to certain antibiotics such as vancomycin. Thus, the Clinical Laboratory Standards Institute (CLSI) recommends using the broth microdilution method in the evaluation of antibacterial activities against S. aureus. A rapid laboratory identification of MRSA is important in the treatment of patients. Therefore, this study aims to optimize and evaluate the effectiveness of a rapid microplate assay using resazurin dye as a colorimetric indicator in determining antibacterial activity against clinical isolates of MRSA and methicillin-susceptible S. aureus (MSSA). Methods: Clinical isolates of MRSA and MSSA were obtained from the Philippine General Hospital (PGH) Microbiology Section, and American Type Culture Collection (ATCC) controls of both strains (ATCC 25923 and ATCC 43300) were acquired. These were then subjected to identification and confirmation procedures. A standardization of bacterial inoculum was performed by comparing its 24-hr growth in Mueller Hinton Broth to 0.5 McFarland Standard. The resazurin microplate assay (REMA) was set-up using two-fold serial dilution of control antibiotics such as oxacillin, vancomycin, and cefoxitin. Each plate was inoculated with standardized bacterial growth of controls and clinical isolates. To determine the time needed for the reduction of the resazurin dye, a qualitative assessment was conducted by comparing the reaction time between a 6.75 mg/mL dye with a 0.01 mg/mL dye. The plates were also subjected to different incubation times and dye concentrations, and the optical densities of the plates were compared using a microplate reader. Results: Results showed that there were no significant differences between the optical densities of the wells of those incubated for 5 hours and for 24 hours (p >0.05). Furthermore, there was a significant reduction in the reaction time of the dye (from 18 hours to 1 hour) when the dye concentration was reduced from 6.75 mg/mL to 0.01 mg/mL. The optimized REMA showed a significant difference between the minimum inhibitory concentrations (MICs) of the different antibiotics against the control and isolate strains of MRSA and MSSA, showing a W of -2.98 (p <0.05) using the Wilcoxon Rank-Sum non-parametric test. Furthermore, the REMA has shown better illustration of anti-MRSA and anti-MSSA activities as compared to the Kirby Bauer disk diffusion method. Conclusion Based on the results presented, the researchers determined the optimal condition for the resazurin microtiter assay, which was 0.01 g/mL concentration of resazurin dye, at a 5-hour incubation period. This study has shown that an optimized REMA is an efficient and fast method to determine the antimicrobial activities of oxacillin, cefoxitin, and vancomycin against MRSA and MSSA.
Methicillin Resistant Staphylococcus aureus

Objective: To determine the point prevalence of, and risk factors associated with MRSA carriage among resident physicians of surgical departments at the Ospital ng Maynila Medical Center. Methods: Design: Cross-sectional Study. Setting: Tertiary Government Training Hospital. Participants:51 resident physicians from different surgical departments (general surgery, obstetrics and gynecology, ophthalmology, otorhinolaryngology – head and neck surgery and dermatology) underwent nasal and pharyngeal swabs with microbial culture and sensitivity testing to identify MRSA carriers. Fisher Exact Test and logistic regression were utilized to determine associations between MRSA carriage and various risk factors including frequency of hand washing and departmental affiliation. Results: Overall prevalence rate of MRSA carriage was 9.8%. Otorhinolaryngology residents had the highest combined prevalence of MRSA of 42.9%, significantly higher compared to other departments and were used as a reference in logistic regression analyses. Notably, handwashing only once daily was associated with a 20-fold increase in the risk of MRSA carriage (OR 20.5, 95% CI: 1.82 to 230, p = .014). Other departments did not demonstrate statistically significant differences in MRSA carriage rates. Conclusions Otorhinolaryngology resident physicians had the highest combined prevalence of MRSA and nasal MRSA was found only in otorhinolaryngology residents. The surgical subspecialty and frequency of handwashing of the healthcare worker were identified as important risk factors to develop MRSA carriage. Targeted interventions (including enhanced infection control protocols and regular screening) are needed especially in high-risk departments.
Methicillin-Resistant Staphylococcus aureus ; Surgical Wound Infection

Background: Ardisia serrata (Aunasin) is an endemic Philippine plant of the family Primulaceae, with several studiesshowing the genus Ardisia as having potential antibacterial, antiangiogenic, cytotoxic, and antipyretic properties. Objective: This study aims to determine the antibacterial and antibiofilm-forming activity of Ardisia serrata ethanolic and aqueous extracts on Escherichia coli, Methicillin-Sensitive Staphylococcus aureus (MSSA), and Methicillin-Resistant Staphylococcus aureus (MRSA). Methods: This is an experimental study testing the activity against bacterial strains of E. coli, MSSA, and MRSA using ethanolic and aqueous extracts of A. serrata leaves. Microtiter susceptibility and biofilm inhibition assays were done with two-fold dilutions of the extract against the selected strains using spectrophotometry with optical density (OD) at 600 nm and 595 nm, respectively, to quantify bacterial growth and biofilm inhibition. The bacterial susceptibility and biofilm inhibition activity was reported as percent inhibition (PI). Minimum inhibitory concentration (MIC), and minimum biofilm inhibition concentration (MBIC) values were obtained using logarithmic regression of the PI values. Results: A. serrata ethanolic extracts showed weak growth inhibitory activity against MSSA and MRSA with minimum inhibitory concentration (MIC) values of 2.6192 and 3.2988 mg/mL, respectively, but no biofilm inhibition activity was noted, while the aqueous extracts exhibited negligible biofilm inhibition activity against MSSA and MRSA with minimum biofilm inhibition concentration (MBIC) values of 13.5972 and 8964.82 mg/mL, respectively, and with no growth inhibition activity. Both ethanolic and aqueous extracts showed no growth inhibition and biofilm inhibition activities against E. coli. Conclusion Staphylococcus aureus is susceptible to the bioactivity of the leaf extracts of A. serrata and has potential to be used as an antibacterial in the treatment of infectious diseases.
Methicillin-resistant Staphylococcus aureus ; Escherichia coli ; natural product ; biological products

OBJECTIVE

The aim of this study was to investigate the protective effects of Lactobacillus brevis BIOTECH 1766 against oxidative damage in the brain, liver, and kidneys induced by aluminum (Al) poisoning in ICR mice.

Twenty mice were divided into four groups (n = 5): (I) control, (II) Al, (III) citric acid (CA), and (IV) L. brevis BIOTECH 1766 group. A 14-day treatment period was implemented, wherein groups I and II received sterile water, while groups III and IV received 10 mg/kg bw of CA and 1 x 109 cfu/kg bw of L. brevis BIOTECH 1766, respectively. On day 15, all except the control group received a single oral dose of 1438 mg/kg bw of AlCl3. 6H2O. After 24 h, mice were euthanized to collect the brain, liver, and kidneys for the oxidative stress marker analyses and histopathological examination.

Acute intoxication of Al led to a significant increase in tissue malondialdehyde (MDA) and a significant decrease in the tissue's reduced glutathione (GSH), catalase (CAT), and superoxide dismutase (SOD). Mice pretreated with CA or L. brevis BIOTECH 1766 have markedly reduced CAT activity in the liver, and SOD in all three organs. Extensive organ injuries were also prevented by CA and L. brevis BIOTECH 1766 pretreatment, with the latter providing better protection against liver damage.

The findings showed that L. brevis BIOTECH 1766 provides a protective effect against acute Al poisoning in mice by ameliorating oxidative damage in the brain, liver, and kidneys.

Humans ; Retrospective Studies ; Bronchiectasis ; Mycobacterium avium Complex ; Syndrome ; Nocardia Infections/drug therapy* ; Nocardia ; Mycobacterium avium-intracellulare Infection