Colony-Forming Unit/mL Research Articles

Revealing the Viable Microbial Community of Biofilm in a Sewage Treatment System Using Propidium Monoazide Combined with Real-Time PCR and Metagenomics.

Microbial community composition, function, and viability are important for biofilm-based sewage treatment technologies. Most studies of microbial communities mainly rely on the total deoxyribonucleic acid (DNA) extracted from the biofilm. However, nucleotide materials released from dead microorganisms may interfere with the analysis of viable microorganisms and their metabolic potential. In this study, we developed a protocol to assess viability as well as viable community composition and function in biofilm in a sewage treatment system using propidium monoazide (PMA) coupled with real-time quantitative polymerase chain reaction (qPCR) and metagenomic technology. The optimal removal of PMA from non-viable cells was achieved by a PMA concentration of 4 μM, incubation in darkness for 5 min, and exposure for 5 min. Simultaneously, the detection limit can reach a viable bacteria proportion of 1%, within the detection concentration range of 102-108 CFU/mL (colony forming unit/mL), showing its effectiveness in removing interference from dead cells. Under the optimal conditions, the result of PMA-metagenomic sequencing revealed that 6.72% to 8.18% of non-viable microorganisms were influenced and the composition and relative abundance of the dominant genera were changed. Overall, this study established a fast, sensitive, and highly specific biofilm viability detection method, which could provide technical support for accurately deciphering the structural composition and function of viable microbial communities in sewage treatment biofilms.

Capsicum Endophytic Bacterial Strain LY7 and Prochloraz Synergistically Control Chilli Anthracnose.

Chilli anthracnose is a major infectious disease of the genus Capsicum. Chemical control is the primary means of controlling this disease; however, the excessive use of chemical pesticides can adversely affect ecological security and human health. Here, our aim was to explore the synergistic effects of chemical and biological pesticides in the control of chilli anthracnose. The bacterial strain LY7, which is antagonistic to the anthracnose-causing fungus Colletotrichum scovillei, inhibited the growth of C. scovillei by 83.52%. Through morphological and genetic analyses, this strain was identified as Bacillus velezensis. Then, the compatibility of LY7 with three common chemical fungicides was determined. The in vitro protective and therapeutic efficacies of the 1 × 109 CFU/mL (colony-forming unit/mL) bacterial solution were 66.38% and 35.18%, respectively, but both were significantly lower than those of prochloraz, the most compatible fungicide. We then conducted field efficacy trials to elucidate the best combination of prochloraz and LY7; the highest control efficiency was achieved with a suspension of 1.0 × 108 CFU/mL of LY7 mixed with 0.75 g/L prochloraz (3:7 ratio). Electron microscopy revealed the inhibitory effects of LY7 and prochloraz on C. scovillei mycelial growth. These results suggest that an LY7-based biofungicide can partially replace prochloraz, serving as an integrated management strategy to control chilli anthracnose.

Antibacterial and Antibiofilm Efficacy of Copper-Doped Phosphate Glass on Pathogenic Bacteria.

the antibacterial activity was determined using microbroth dilution and time-kill assay. The antibiofilm activity was investigated using crystal violet and confocal laser scanning microscopy. Bacteria growing in absence of CDPG were used as controls. the MIC was ≥125 mg of CPDG/mL; the log10 CFU/mL reduction ranged from 2.66-3.14 to 6.23-9.65 after 4 and 24 h respectively. Generally, no growth was observed after 24 h of treatment with CDPG; the MBC was 250 mg/mL for L. casei and S. oralis while 500 mg/mL for the rest of the bacteria. The highest and lowest antibiofilm activity was observed against S. oralis and E. coli respectively. Three patterns of complete biofilm disruption were seen: (i) large areas with E. fecalis and S. oralis, (ii) medium-size pockets with S. aureus and P. aeruginosa, or (iii) small areas with E. coli and L. casei. CDPG can be potentially used as an antibacterial and an antibiofilm agent against oral biofilm-forming bacteria.

Evaluation of Antibacterial Efficacy of Randomly Selected Alcohol Based Hand Sanitizers Sourced from Grocery Shops within Lagos Metropolis on Some Local Bacterial Strains in COVID-19 Era

Background: Health experts promote the use of alcohol based hand rubs to contain the spread of microbes. The emergence of novel SARS-CoV-2 virus has brought a worsening public health challenge that re-enacted the importance of hand hygiene globally. Objective: This study evaluated the antimicrobial efficacy of locally made alcohol-based hand rubes sourced from grocery shops within Lagos metropolis, Nigeria. Methods: We conducted a laboratory based in vitro experiment, using 20 randomly sourced hand sanitizers against standard Escherichia coli (ATCC 25922) and three locally characterized Multi-drug-Resistant bacterial strains (Staphylococcus aureus (NIMR/NTCC/GP056), Klebsiella pneumonia (NIMR/NTCC/GN065) and Proteus stutzin - (NIMR/NTCC/ GN029). Reference standard, 60 % isopropanol was used as positive control. Test suspension method as per European standard PrEN12054 was employed. The Colony Forming Unit/mL (CFU/mL) at base line and after each contact time (15, 30 & 45 seconds) with samples was recorded. The Logarithmic reduction factor (RF) and percent reduction were computed and expressed using descriptive statistics. Results: Out of the 20 solutions tested (10 sprays and 10 gels), 11 (55%) had standard efficacy of 5-Log10 reduction factor (6.7- 6.8) recommended at 15 seconds exposure time on the 4 bacteria. Out of such 7 (64%) were spray solution (L1, L3, L5, L10, L11, L12 & L16), while 4 (36%) were gel solutions (L9, L15, L18 & L20). Another 2 (10%) had relative time based efficacy at between 30 to 45 seconds exposure (L2 spray and L14 gel). Seven (35%) (L7 & L17 spray; L4, L6, L8, L13 & L19 gel solutions) failed the test. Escherichia coli and Proteus stutzin were more susceptible to the samples tested and produced higher RF. Conclusion: About 45 % of the hand sanitizers had poor efficacy and this is quite high, especially in this era of pandemic. This report underscores the need for production policy review by the regulatory body. It is imperative to enforce quality management regime, particularly, internal and external production quality control. Periodic batch efficacy validation is necessary to ensure precision. Poor quality products must be actionable. We recommend this experiment be scaled up to national level and to cover major microbial pathogens.

MoS2 Functionalized Multicore Fiber Probes for Selective Detection of Shigella Bacteria Based on Localized Plasmon

Present study demonstrates the fiber-optic localized surface plasmon resonance (LSPR) based sensitive biosensor for detection of Shigella bacterial species. The proposed sensor is comprised of multi-core fiber (MCF) having seven cores arranged in a hexagonal shape and spliced with single-mode fiber (SMF) for efficient detection. An increase in evanescent waves (EWs) and coupling of modes between MCF cores was achieved by etching process in a controlled manner. The etching process also increases the refractive index sensitivity (RIS) of the proposed sensor. Further, coating with nanomaterials like gold nanoparticles (AuNPs) and molybdenum disulfide (MoS <sub xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">2</sub> ) helps in the excitation of localized plasmons. Here, Shigella specific oligonucleotide probes are used as a recognition element. The results demonstrate that the proposed sensor can successfully and efficiently detect the Shigella bacterial species with high sensitivity. Shigella in the range of 10 - 100 CFU/mL (colony-forming unit/mL) can cause serious intestinal infection and therefore, its detection in this range is critical. The proposed sensor demonstrates a linearity range from 1 to 10 <sup xmlns:mml="http://www.w3.org/1998/Math/MathML" xmlns:xlink="http://www.w3.org/1999/xlink">9</sup> CFU/mL with a detection time of 5 min and a limit of detection (LoD) of 1.56 CFU/mL. The proposed sensing methodology can be a potential alternative to the commercially existing ones in the near future.

Potassium Ferrate (VI) as the Multifunctional Agent in the Treatment of Landfill Leachate.

Possible use of potassium ferrate (VI) (K2FeO4) for the treatment of landfill leachate (pH = 8.9, Chemical Oxygen Demand (COD) 770 mg O2/L, Total Organic Carbon (TOC) 230 mg/L, Total Nitrogen (Total N) 120 mg/L, Total Phosphorus (Total P) 12 mg/L, Total Coli Count (TCC) 6.8 log CFU/mL (Colony-Forming Unit/mL), Most Probable Number (MPN) of fecal enterococci 4.0 log/100 mL, Total Proteolytic Count (TPC) 4.4 log CFU/mL) to remove COD was investigated. Central Composite Design (CCD) and Response Surface Methodology (RSM) were applied for modelling and optimizing the purification process. Conformity of experimental and predicted data (R2 = 0.8477, Radj2 = 0.7462) were verified using Analysis of Variance (ANOVA). Application of K2FeO4 using CCD/RSM allowed to decrease COD, TOC, Total N, Total P, TCC, MPN of fecal enterococci and TPC by 76.2%, 82.6%, 68.3%, 91.6%, 99.0%, 95.8% and 99.3%, respectively, by using K2FeO4 0.390 g/L, at pH = 2.3 within 25 min. Application of equivalent amount of iron (as FeSO4 × 7H2O and FeCl3 × 6H2O) under the same conditions allowed to diminish COD, TOC, Total N, Total P, TCC, MPN of fecal enterococci and TPC only by 38.1%, 37.0%, 20.8%, 95.8%, 94.4%, 58.2%, 90.8% and 41.6%, 45.7%, 29.2%, 95.8%, 92.1%, 58.2%, 90.0%, respectively. Thus, K2FeO4 could be applied as an environmentally friendly reagent for landfill leachate treatment.

Bacteria incorporated with calcium lactate pentahydrate to improve the mortar properties and self-healing occurrence

Concrete can be harmful to the environment due to its high energy consumption and CO2 emission and also has a potential crack formation, which can promote a drop in its strength. Therefore, concrete is considered as a non-sustainable material. The mechanisms by which bacterial oxidation of organic carbon can precipitate calcite that may fill the voids and cracks on cement-based materials have been extensively investigated to prevent and heal the micro-cracks formation. Hence, this study focused on utilizing a new alkaliphilic bacterial strain indigenous to an Indonesian site, Lysinibacillus sphaericus strain SKC/VA-1, incorporated with calcium lactate pentahydrate, as a low-cost calcium source, with various bacterial inoculum concentrations. The bacterium was employed in this study due to its ability to adapt to basic pH, thus improving the physical properties and rejuvenating the micro-cracks. Experimentally, the addition of calcium lactate pentahydrate slightly affected the mortar properties. Likewise, bacteria-incorporated mortar exhibited an enhancement in the physical properties of mortar. The highest improvement of mechanical properties (an increase of 45% and 36% for compressive and indirect tensile strength, respectively) was achieved by the addition of calcium lactate pentahydrate incorporated with 10% v/v bacterial inoculum [about 7 × 107 CFU/ml (colony-forming unit/ml)]. The self-healing took place more rapidly on bacterial mortar supplemented with calcium lactate pentahydrate than on the control specimen. XRD analysis demonstrated that the mineralogical composition of self-healing precipitates was primarily dominated by calcite (CaCO3), indicating the capacity of L. sphaericus strain SKC/VA-1 to precipitate calcite through organic carbon oxidation for self-healing the artificial crack on the mortar. To our knowledge, this is the first report on the potential utilization of the bacterium L. sphaericus incorporated with calcium lactate pentahydrate to increase the mortar properties, including its self-healing ability. However, further study with the water-cement ratio variation is required to investigate the possibility of using L. sphaericus and calcium lactate pentahydrate as an alternative method rather than reducing the water-cement ratio to enhance the mortar properties.

Antifungal photodynamic activities of phthalocyanine derivatives on Candida albicans.

Antimicrobial resistance is one of the most important causes of morbidity and mortality in the treatment of infectious diseases worldwide. Candida albicans is one of the most virulent and common species of fungi to cause invasive fungal infections on humans. Alternative treatment strategies, including photodynamic therapy, are needed for controlling these infectious diseases. The aim of this study was to investigate the antifungal photodynamic activities of phthalocyanine derivatives on C. albicans. The minimum inhibitory concentration (MIC) values of compounds were determined by the broth microdilution method. Uptake of the compounds in C. albicans and dark toxicity of the compounds were also investigated. Photodynamic inhibition of growth experiments was performed by measuring the colony-forming unit/mL (CFU/mL) of the strain. Maximum uptake into the cells was observed in the presence of 64 μg/mL concentration for each compound except for ZnPc. Compounds did not show dark toxicity/inhibitory effects at sub-MIC concentrations on C. albicans when compared to the negative control groups. Zn(II)Pc, ZnPc, and ZnPc-TiO2 showed fungicidal effect after irradiation with the light dose of 90 J/cm2 in the presence of the compounds. In addition to the fungicidal effects, SubPc, SubPc-TiO2, Es-SiPc, and Es-SubPc compounds were also found to have inhibitory effects on the growth of yeast cells after irradiation.

Microbiological Safety of Street-Vended Beverages in Chow Kit, Kuala Lumpur.

Improper handling, poor hygienic practices, and lack of environmental control affect the safety of street-vended beverages. The objective of this study is to determine the bacterial contamination level of three types of beverages (cordial-based drinks, milk-based drinks, fruit juices) sold by street vendors at Chow Kit, Kuala Lumpur. A total of 31 samples of beverages were analyzed to determine total viable count (TVC), total coliform, Escherichia coli, and Staphylococcus aureus counts via the standard plate count method. The results showed that only 9.7% of the total samples were not contaminated with the tested microorganisms. All milk-based drink samples were positive for TVC and also had the highest average bacterial counts at 5.30 ± 1.11 log Colony Forming Unit/mL (CFU/mL). About 71% of the samples were contaminated with total coliform with the average readings ranging between 4.30 and 4.75 log CFU/mL, whereas 58.1% of the samples were positive with S. aureus, with fruit juices having the highest average reading (3.42 ± 1.15 log CFU/mL). Only one sample (milk-based drink) was E. coli positive. This study showed that the microbiological safety level of street-vended beverages in Chow Kit, Kuala Lumpur was average and needs to be improved. Provision of food safety education and adequate sanitary facilities at vending sites are suggested to increase the safety of food products.

THE IMPACT OF GLOBAL WARMING ON THE PUBLIC HEALTH INCREASING THE BACTERIAL CAUSING INFECTIOUS DISEASES PERFORMED BY EXPERIMENT: VECTOR-BORNE DISEASES INSECTS, TAIF, KSA

The search conducted on "The impact of global warming (GW) on the public health (PH) increasing the bacterial causing infectious diseases (IDs) performed by experiment: Vector-borne diseases (VBDs) insects, Taif, KSA", the experiment used ants (Taif Tapinoma sessile), prepared, arranged appropriate nests and adjusted the temperature at (20, 25, 30, 35, 40 and 45°C), for a week of each zone. It revealed the behaviour as (normal, semi-normal and ab-normal), the mean of mortality rates were between (0-53.3%). The bacterial contents measured by the turbidity indicated the presence of multiplication, were between (0.109-0.328). The bacterial growth degrees by sings were between (+ - +++++) and percent between (12-100%). Colony Forming Unit/ml (CFU/ml) confined between (1.8X102-15.0X102)/mL. Through this experiment it turned out the GW had a significant role on the PH, helped the proliferation of bacterial pathogens that caused IDS. The conclusion wiped from the experiment that the extent degrees of GW disadvantages on the PH. The PH workers must take the "Preventive Health Prophylaxis Measures" (PHPMs) to protect the individuals from IDs by eliminating the VBDs of various types, monitoring the immunological situation of individuals, provided the vaccinations of IDs and preparing for complete PHPMs against any changes in the PH.

Rapid colorimetric sensing platform for the detection of Listeria monocytogenes foodborne pathogen

Listeria monocytogenes is a serious cause of human foodborne infections worldwide, which needs spending billions of dollars for inspection of bacterial contamination in food every year. Therefore, there is an urgent need for rapid, in-field and cost effective detection techniques. In this study, rapid, low-cost and simple colorimetric assay was developed using magnetic nanoparticles for the detection of listeria bacteria. The protease from the listeria bacteria was detected using D-amino acid substrate. D-amino acid substrate was linked to the carboxylic acid on the magnetic nanoparticles using EDC/NHS chemistry. The cysteine residue at the C-terminal of the substrate was used for the self-assembled monolayer formation on the gold sensor surface, which in turn the black magnetic nanobeads will mask the golden color. The color will change from black to golden color upon the cleavage of the specific peptide sequence by the Listeria protease. The sensor was tested with serial dilutions of Listeria bacteria. It was found that the appearance of the gold surface area is proportional to the bacterial concentrations in CFU/ml. The lowest detection limit of the developed sensor for Listeria was found to be 2.17×102 colony forming unit/ml (CFU/ml). The specificity of the biosensor was tested against four different foodborne associated bacteria (Escherichia coli, Salmonella, Shigella flexnerii and Staphylococcus aureus). Finally, the sensor was tested with artificially spiked whole milk and ground meat spiked with listeria.

Study on the mechanism of antibacterial action of magnesium oxide nanoparticles against foodborne pathogens.

BackgroundMagnesium oxide nanoparticles (MgO nanoparticles, with average size of 20 nm) have considerable potential as antimicrobial agents in food safety applications due to their structure, surface properties, and stability. The aim of this work was to investigate the antibacterial effects and mechanism of action of MgO nanoparticles against several important foodborne pathogens.ResultsResazurin (a redox sensitive dye) microplate assay was used for measuring growth inhibition of bacteria treated with MgO nanoparticles. The minimal inhibitory concentrations of MgO nanoparticles to 104 colony-forming unit/ml (CFU/ml) of Campylobacter jejuni, Escherichia coli O157:H7, and Salmonella Enteritidis were determined to be 0.5, 1 and 1 mg/ml, respectively. To completely inactivate 108−9 CFU/ml bacterial cells in 4 h, a minimal concentration of 2 mg/ml MgO nanoparticles was required for C. jejuni whereas E. coli O157:H7 and Salmonella Enteritidis required at least 8 mg/ml nanoparticles. Scanning electron microscopy examination revealed clear morphological changes and membrane structural damage in the cells treated with MgO nanoparticles. A quantitative real-time PCR combined with ethidium monoazide pretreatment confirmed cell membrane permeability was increased after exposure to the nanoparticles. In a cell free assay, a low level (1.1 μM) of H2O2 was detected in the nanoparticle suspensions. Consistently, MgO nanoparticles greatly induced the gene expression of KatA, a sole catalase in C. jejuni for breaking down H2O2 to H2O and O2.ConclusionsMgO nanoparticles have strong antibacterial activity against three important foodborne pathogens. The interaction of nanoparticles with bacterial cells causes cell membrane leakage, induces oxidative stress, and ultimately leads to cell death.

Assessment of a semi-automated protocol for multiplex analysis of sepsis-causing bacteria with spiked whole blood samples.

Sepsis is associated with high morbidity and mortality rates worldwide. Rapid and reliable diagnostic methods are needed for efficient and evidence-based treatment of septic patients. Recently, new molecular tools have emerged to complement the conventional culture-based diagnostic methods. In this study, we used spiked whole blood samples to evaluate together two ready-to-use molecular solutions for the detection of sepsis-causing bacteria. We spiked whole blood with bacterial species relevant in sepsis and extracted bacterial DNA with the NorDiag Arrow device, using the SelectNA Blood pathogen DNA isolation kit. DNA extracts were analyzed by the polymerase chain reaction (PCR)- and microarray-based Prove-it™ Bone and Joint assay, resulting in correctly identified bacterial species with detection limits of 11–600 colony-forming unit/mL (CFU/mL). To understand the recovery losses of bacterial DNA during the sample preparation step and the capability of the PCR- and microarray-based platform to respond to the sensitivity requirements, we also determined the analytical sensitivity of the PCR and microarray platform to be 1–21 genome equivalents for the tested bacterial species. In addition, the inclusivity of the Prove-it™ Bone and Joint assay was demonstrated with methicillin-resistant Staphylococcus aureus (MRSA) clones carrying SCCmec types I, II, IV, or V and a nontypable SCCmec type. The proof-of-concept for accurate multiplex pathogen and antibacterial resistance marker detection from spiked whole blood samples was demonstrated by the selective bacterial DNA extraction method combined with the high-throughput PCR- and microarray-based platform. Further investigations are needed to study the promising potential of the concept for sensitive, semi-automated identification of sepsis-causing pathogens directly from whole blood.

Prevalence of Asymptomatic Bacteriuria in HIV Infected Patients in a Tertiary Hospital in Lagos, Nigeria

Background: People living with Human Immunodeficiency Virus (HIV) are more predisposed to urinary tract infections due to suppression of their immunity by the virus. Asymptomatic bacteriuria is associated with an increased risk of symptomatic urinary tract infection and the latter being an important contributor to development of chronic renal failure, hypertension and toxaemia of pregnancy. The aim of this study was to determine the prevalence of asymptomatic bacteriuria in HIV-infected patients and proffer a recommendation on the need or otherwise to screen. Methods: This was a cross sectional study of treatment-naive HIV-infected patients attending the HIV clinics of Lagos State University Teaching Hospital (LASUTH), Ikeja. A single voided aseptically collected mid-stream urine (MSU) was obtained from each patient and all samples processed immediately, were sent for urinalysis and culture. Isolates were considered significant if there were ≥105 colony forming unit/mL (CFU/mL) with 2 or less isolates, doubtful significance if 5 CFU/mL. Significant isolates were selected for identification. Data were analyzed using SPSS version 16.0 (Statistical Package for Social Sciences, Inc., Chicago, Ill). Results: A total of 156 consenting participants were recruited into the study. The mean age was 36.45 ± 8.65 years. There were 60 of 156 (38.4%) males and 96 of 156 (61.56%) females. Only 33 of 156 (21.2%) had significant bacteriuria, out of the 33 participants, 19 (57.8%) were asymptomatic, while 14 of 33 (42.4%) had significant growth but were symptomatic or on antibiotics. E. coli was isolated in 9 of 19 (47.3%), followed by Staph aureus 4 of 19 (21.05%). Conclusion: More than half of participants who had significant growth had asymptomatic bacteriuria, while one-fifth of all patients had significant growth. Considering this statistics, screening for or treatment of asymptomatic bacteriuria may be recommended in all HIV-infected patients.

The Role of Trichoderma in the Rhizosphere of Tomato Plants

It is well established that microorganisms are closely associated with the roots of plants can directly influence plant growth and development. Species of Trichoderma, on the other hand are primarily studied for their ability to control plant disease. The ability of species of Trichoderma to directly promote or inhibit plant growth has been noted for many years.Tomatoes were treated with different Trichoderma strains by seed treatment and soil inoculation. The Trichoderma were grown on malt-agar medium and conidia were washed off by sterile water for making suspension which contained 107 CFU/ml (colony forming unit/ml). The suspension was used for seed treatment and for the soil inoculation by watering as well. The artificial soil inoculation was made by Trichodermas growing on grounded maize were mixed in mould and tomato seed were sown in it. Tomato seeds were also sown in bags made of close-meshed material which allowed the soil microorganisms to colonize the roots but it simultaneously protected the roots from soil contamination. Roots were put on Trichoderma selective medium to check the root colonization of the Trichoderma.The tomato plants were bedded out in a field in four repetition. After harvesting by hand the results supported by statistics shown that there was significant differences between the yield of the untreated and treated tomato plants by Trichoderma strains.