Prevalence and Recovery of Microorganisms from Containers used for the Collection of Forensic Biological Samples

Background: Microbes play a significant role in the degradation of biological evidence collected for forensic analysis. The present study is aimed to isolate and identify the microbes present inside the empty container used for the biological evidence collection. Methods: Bacterial isolation from the selected containers was done by cotton swab over the inner surface of the containers. Streaking was done on the surface of the three different culture plates as a Blood agar plate, Nutrient plate and MacConkey plate. The plates were placed in an incubator shaker at 37ºC for 48 hours. The colonies grown on the surface of the media were counted on and used for further study. Various biochemical assays were performed to characterize isolated bacteria. Results: Staining results suggested that the presence of Gram-positive stain (Staphylococcus, Bacillus, Corynebacterium, Clostridium) and Gram negative stain (E. coli, Enterobacteriaceae, Pseudomonas, Salmonella, Shigella, Stenotrophomonas, Bdellovibrio, Acetic acid bacteria). The Catalase and Coagulase test suggested the presence of Staphylococcus aureus, S. epidermis and S. sapropyticus. Moreover, the indole test suggested the presence of Citrobacter koseri, Kebsiella oxytoca, Proteus vulgaris etc. Some of the bacteria were urea metabolizing, including Proteus spp, Helicobacter pylori, Cryptococcus spp, Corynebacterium spp. Conclusion: This study recommends that there should be proper maintenance of the chain of custody from the collection to analysis so that evidence properly prevents degradation or contamination in the biological evidence. Extra care is needed for the collection and packing of biological evidence from the crime scene. Moreover, the collection containers, if left wide open, lead to contamination and degradation of biological evidence.

The Growth and Morphological Characterization Of Tropical Thermophilic Bacterium Parageobacillus caldoxylosilyticus ER4B

Parageobacillus caldoxylosilyticus is a rod-shaped thermophilic bacterium that can grow optimally at high temperatures. The thermophilicity of the bacterium is expected to be largely accounted for by the production of thermostable enzymes which has valuable applications in many fields. However, the species is poorly studied, hence, the growth conditions at high temperatures remained unclear until today. Therefore, this study aimed to determine the growth characterization of P. caldoxylosilyticus, including growth media preferences, optimal growth temperature, as well as minimum and maximum growth temperature. P. caldoxylosilyticus strain ER4B isolated from oil palm empty fruit bunch compost was used in this study. The bacterial strain was first identified using 16S rRNA sequencing, and the subsequent BLAST result showed that it is closest to P. caldoxylosilyticus strain UTM6. It is found that ER4B grew best in LB as compared to R2A, TSB, and NB medium. Further temperature tests determined the optimum growth temperature of the strain to be at 64°C Besides, the bacterium forms mucoid circular punctiform colonies that are yellowish in color on an agar plate, and the colony is usually 2 mm to 4 mm in diameter. The microscopic analysis also revealed that strain ER4B is a Gram-positive rod-shaped bacterium that has a length ranging from 3 µm to 6 µm, with a diameter of around 0.5 µm.

Morphology, microscopic anatomy and bacterial inhibition of extracts of Ludwigia adscendens (L.) H. Hara plants growing in Lang Sen Wetland Reserve, Long An province, Vietnam

The study aimed to supplement data on a plant that was considered a folk medicine of Vietnam. Morphological and microscopic anatomical characteristics of this species which were adapted to the frequently flooded habitat of the reserve was studied and analyzed. Alcoholic extracts of each stem, leaf and root part were recovered using a rotary evaporator. The inhibitory ability on bacterial strains including Bacillus cereus, B. subtilis and Escherichia coli was tested through agar plate diffusion method. The result showed that the stem and leaf extracts were effective against all three bacterial strains, while the root extracts had no effect against E. coli.

Nanofibrous Membrane with Encapsulated Glucose Oxidase for Self-Sustained Antimicrobial Applications

Polyvinyl alcohol (PVA) nanofibrous membrane, consisting of separately encapsulated glucose oxidase (GOx) and glucose (Glu) nanofibers, was prepared via simultaneously electrospinning PVA/GOx and PVA/Glu dopes. The as-prepared pristine membrane could self-sustainably generate hydrogen peroxide (H2O2) only in contact with an aqueous solution. The H2O2 production level was well maintained even after storing the dry membrane at room temperature for 7 days. Cross-linking the membrane via reaction with glutaraldehyde (GA) vapor could not only prevent the nanofibrous membrane from dissolving in water but also prolonged the release of H2O2. The sustained release of H2O2 from the membrane achieved antimicrobial capability equivalent to that of 1% H2O2 against both Escherichia coli and Staphylococcus aureus. Gram(+) S. aureus cells were more susceptible to H2O2 than Gram(−) E. coli and >99% of S. aureus were killed after 1 h incubation with the membrane. Pristine and GA-crosslinked nanofibrous membrane with in situ production of H2O2 were self-sterilized in which no microorganism contamination on the membrane could be detected after 2 weeks incubation on an agar plate. The GOx/Glu membrane may find potential application as versatile antimicrobial materials in the field of biomedicine, in the food and health industries, and especially challenges related to wound healing in diabetic patients.

Evaluation of an ultra-rapid antibiotic susceptibility testing method on positive blood cultures with Escherichia coli

Blood stream infection (BSI) is related to high mortality and morbidity. Early antimicrobial therapy is crucial in treating patients with BSI. The most common Gram-negative bacteria causing BSI is Escherichia coli. Targeted effective treatment of patients with BSI is only possible if it is based on antibiotic susceptibility testing (AST) data after blood culture positivity. However, there are very few methods available for rapid phenotypic AST and the fastest method takes 4 h. Here we analyzed the performance of a 30 min ultra-rapid method for AST of E. coli directly from positive blood cultures (BC). In total, 51 positive BC with E. coli were studied, and we evaluated the ultra-rapid method directly on positive BC as well as on E. coli colonies cultured on agar plates. The results obtained by the new method were compared with disk diffusion. The method provided accurate AST result in 30 min to Ciprofloxacin and Gentamicin for 92% and 84% of the positive BC samples, respectively. For E. coli isolates retrieved from agar plates, 86% and 96% of the AST results were accurate for Ciprofloxacin and Gentamicin, respectively, after 30 min of assay time. When time to result was modulated in-silico from 30 to 60 minutes for the agar plate samples, accuracy of AST results went up to 92% for Ciprofloxacin and to 100% for Gentamicin. The present study shows that the method is reliable and delivers ultra-rapid AST data in 30 minutes directly from positive BC and as well as from agar plates.

Evaluation of The Antifungal Activity of Nasturtium officinale (watercress) Oil with Calcium Hydroxide against Candida Albicans Isolated from Root Canal

Background: The interest in herbal extracts as antimicrobial agents has increased over the past few years in endodontic therapy. Nasturtium officinale (watercress) is a promising plant with great medicinal values. This study aimed to investigate the antifungal activity of watercress oil in combination with calcium hydroxide against Candida albicans as intracanal medicament. Materials and Methods: Candida albicans was isolated from patients with necrotic root canal or failed root canal treatment. The sensitivity of Candida albicans to different concentrations of watercress oil extract was determined by using the agar well diffusion method in comparison with calcium hydroxide paste. The agar plate method was used to determine the minimum fungicidal concentration (MFC) of the tested oil against the fungus. The combination of the oil extract of Nasturtium officinale with calcium hydroxide was evaluated and compared to calcium hydroxide paste with iodoform by using the agar well diffusion method. Results: The oil extract exhibited antifungal activity against Candida albicans, this activity was found to be increased as the concentration of extract increased. The tested combination of watercress oil extract with calcium hydroxide revealed larger inhibition zones than the ones formed by each tested agent individually. Conclusion: The oil extract of Nasturtium officinale is active against Candida albicans suggesting its potential to be used as an intracanal medicament alone or in combination with calcium hydroxide.

DILUCIÓN DE MIEL EN AGUA INDUCE HORMESIS EN LA ACTIVIDAD ANTIBACTERIANA SOBRE Staphylococcus aureus.

In pharmacology, hormesis is defined as the high efficacy of a drug at low doses and a decrease of efficacy at higher doses. In this study, the hormesis phenomenon on the antibacterial activity of honey was assessed. The activity of four honeys (A, D, C and D) were checked by agar plate diffusion method using S. aureus (ATCC 29213) as the test organism. Each honey was tested as follows; on the surface of an agar plate, five stainless steel cylinders with a diameter of 8,14 mm were radially disposed. Onto each cylinder 0,1 mL of the sample honey diluted in water at 50 – 25 - 12,5 -6,25 and 3,125 % v/v were placed. Plates were incubated at 35°C during 24 h. The surface of the cylinder diameter was set as reference antibacterial activity (Ra). The antibacterial activity of each honey was quantitated by the relationship between the surface of the inhibitory zone (Iz) and the surface of the reference activity (Iz/Ra). An increase in the antibacterial activity was observed at a concentration of water greater than 50% in honeys B and C. This hormetic behavior could be considered as indicator of the quality of the antibacterial activity of these honeys.

PROPUESTA DE UNA METODOLOGÍA PARA CUANTIFICAR LA ACTIVIDAD ANTIBACTERIANA DE LA MIEL

In this study a standardized method for objectively comparing the antibacterial activity of honey was developed. The assay was performed using a strain of S. aureus and four honeys that have been shown antibacterial activity against this strain. The antibacterial activity of honeys were checked by using an agar plate diffusion method using S. aureus (ATCC 29213) as the test organism. On the surface of each agar plate six stainless steel cylinders with a diameter of 8,14 mm were radially disposed. Onto each cylinder 0,1 mL of sample honey at 10% v/v was placed and all Petri dishes were incubated at 35°C during 24 h. Each sample were analyzed by triplicate. The surface of the cylinder diameter was set as reference antibacterial activity equivalent to minimal inhibitory concentration (MIC). The antibacterial activity of each honey was quantitated by the relationship between the surface of the inhibitory zone and the surface of the reference activity expressed as multiples of MIC (x MIC). The method allowed establishing differences between the bacterial activities of the different honeys tested and could be used for identifying honeys that could be employed with medicinal purpose for the treatments of wounds.

Actinomadura violacea sp. nov., a madurastatin A1-producing strain isolated from lichen in Thailand

An actinomycete strain, LCR2-06T, isolated from a lichen sample on rock collected from Chiang Rai Province (Pong Phra Bat Waterfall), Thailand, was characterized using a polyphasic approach. The strain grew at 25–45 °C, pH 6–11 and on International Streptomyces Project 2 agar plate with 5 % (w/v) NaCl. It contained meso-diaminopimelic acid as the diamino acid in whole-cell hydrolysates. Rhamnose, ribose, xylose, madurose, glucose and galactose were detected as whole-cell sugar hydrolysates. Mycolic acids were absent. The N-acyl type of muramic acid was acetyl. The strain contained C16 : 0, TBSA 10-methyl C18 : 0 and 2-hydroxy C16 : 0 as the predominant fatty acids and MK-9(H6), MK-9(H4) and MK-9(H8) as the major menaquinones. The major polar lipids were diphosphatidylglycerol, phosphatidylinositol and unidentified phospholipid. The draft genome of strain LCR2-06T was closely related to Actinomadura barringtoniae TBRC 7225T (99.2 %), Actinomadura nitritigenes NBRC 15918T (98.8 %), Actinomadura montaniterrae TISTR 2400T (98.5 %) and Actinomadura physcomitrii JCM 33455T (97.9 %). The draft genome of LCR2-06T was 11.1 Mb with 10 588 coding sequences with an average G+C content of 72.7 mol%. Results of genomic analysis revealed that the ANIb and ANIm values between strain LCR2-06T and A. montaniterrae TISTR 2400T were 90.0 and 92.0 %, respectively. The digital DNA–DNA hybridization value was 43.9 % in comparison with the draft genome of A. montaniterrae TISTR 2400T. The strain produced an antibacterial compound active against Bacillus subtilis ATCC 6633 and Kocuria rhizophila ATCC 9341. The results of taxonomic analysis suggested that strain LCR2-06T represented a novel species of the genus Actinomadura for which the name Actinomadura violacea sp. nov. is proposed. The type strain is LCR2-06T (=JCM 33065T=KCTC 49547T=NBRC 114810T=LMG 32136T=TISTR 2935T).

Andrographis paniculata absorbed ZnO nanofibers as a potential antimicrobial agent for biomedical applications

The present study focuses on developing antimicrobial nanofibers with superhydrophobicity and insect repellent properties for wound dressing and biomedical applications. For the first time, Andrographis paniculata leaf extract was prepared and incorporated into ZnO nanofibers (AP-ZnO). The antifungal and antibacterial activities of AP-ZnO were tested using the agar plate method. Amid the bacteria and fungus strain tested, the prepared sample exhibited a greater efficiency against Staphylococcus aureus and Trichophyton rubrum respectively. AP-ZnO showed the highest larvicidal activity (100 ± 0.2) against Aedes aegypti, proving its insect repellent characteristics. The in vivo toxicity studies of AP-ZnO (100 μg ml−1) tested on Danio rerio ensure the biocompatibility of the prepared sample with maximum toxicity of 21.1% after 72 h, which is lower than commercial Prallethrin. Moreover, AP-ZnO-chitosan coated cotton fabric showed higher durability with contact angle (θ ∼ 151°) and was suggested to be used for self-cleaning applications in the biomedical sector. The examined results confirmed that AP-ZnO possesses different medicinal characteristics suitable for biomedical and pharmaceutical applications.