Characterization of isolated UV-C-irradiated mutants of microalga Chlorella vulgaris for future biofuel application.

Microalgae-based biofuel is considered as one of the most promising sources of alternative energy because it is sustainable and does not pose threats to the environment and food security. However, attempts in improving microalgal strains to attain the ideal characteristics for biofuel application are yet to unravel. In this study, random UV-C mutagenesis was employed to generate starch-deficient mutants of indigenous Chlorella vulgaris to enhance its productivity. Out of 872 colonies, two isolated mutants (cvm5 and cvm6) were isolated and showed significant increase in cell concentrations by > 1.47-fold and > 1.04-fold, respectively. However, mutant cells exhibited smaller in size which might contributed to the significant decrease in their biomass. Moreover, gathered data revealed that the total lipid content of cvm5 was enhanced significantly (75%, > 1.3-fold increase). Additionally, triacylglycerol (TAG) content of the said mutant constitutes 48% of the dry cell weight (DCW) while cvm6 consist of 41% of the DCW. These promising and novel findings suggest that the two generated and isolated mutants are good candidates for future commercial biofuel production, especially in the Philippines. In addition, these findings may contribute on the prior knowledge of the usage of UV-C for microalgal strain development.

Phylogroup typing and carbapenem resistance of Escherichia coli from agricultural samples in Metro Manila, Philippines.

Primary production environment is considered as reservoir of Escherichia coli contamination of produce. E. coli is classified into eight phylogroups which differ in ecological niches, evolutionary history, and phenotypic properties. To understand the population genetic structure and composition of E. coli in primary production environments in Metro Manila, Philippines, a total of 80 E. coli recovered from irrigation water, soil, vegetables, and feces of cat, carabao, chicken, dog, and goat were allocated into distinct phylogroups based on the presence and absence of genetic markers. Results showed that the most prevalent phylogroup was B1 (71.3%), followed by A (18.6%), D (6.3%), B2 (1.3%), E (1.3%), and an unknown phylogroup (1.3%). The most prevalent genetic marker was arpA, followed by TspE4.C2, yjaA, and chuA. The carbapenem resistance of 24 E. coli isolates representing different phylogroups was also evaluated. Intriguingly, all isolates exhibited uniform susceptibility. This is the first report to provide insights into the phylogroup structure and composition, as well as carbapenem resistance of E. coli from primary production in the Philippines, which highlights possible source of and solution for gastrointestinal and enteric diseases.

Influences of carbohydrate, nitrogen, and phosphorus sources on the citric acid production by fungal endophyte Aspergillus fumigatus P3I6.

Fungi are ecologically ubiquitous organisms on earth and regarded as one of the prolific sources of natural products. Fungal endophytes may provide essential prerequisite molecules to plant biochemical pathways which allow the efficient synthesis of primary and secondary metabolites. This study characterized the influences of various combinations of process parameters namely, carbohydrate, nitrogen, and phosphorus sources on citric acid (CA) production by the isolated fungal endophyte Aspergillus fumigatus P3I6 from Citrus microcarpa. Aspergillus fumigatus P3I6 had higher CA concentration of 9.2 (± 0.9) g L-1 and 9.0 (± 5.0 × 10-15) g L-1 when supplemented with sucrose and white refined sugar, respectively, than A. niger NRRL 599. Response Surface Methodology (RSM) had shown that A. fumigatus P3I6 produced the highest CA (23.8 g L-1) in Combination 4 (18.0% sucrose, 0.3 g L-1 ammonium sulfate, and 5.0 g L-1 dipotassium phosphate (K2HPO4)). Analysis of variance showed that when K2HPO4 concentrations were increased, CA content in fermentation media was significantly elevated. Hence, K2HPO4 was the most critical variable in the quadratic model (p < 0.05); however, sucrose concentration still has its role in production. Aside from using A. niger in most fermentation processes, this discovered fungal strain can be potentially used in biotechnological applications.

Application of quantitative real-time PCR compared to filtration methods for the enumeration of Escherichia coli in surface waters within Vietnam.

Surface water samples in Vietnam were collected from the Saigon River, rural and suburban canals, and urban runoff canals in Ho Chi Minh City, Vietnam, and were processed to enumerate Escherichia coli. Quantification was done through membrane filtration and quantitative real-time polymerase chain reaction (PCR). Mean log colony-forming unit (CFU)/100 ml E. coli counts in the dry season for river/suburban canals and urban canals were log 2.8 and 3.7, respectively, using a membrane filtration method, while using Taqman quantitative real-time PCR they were log 2.4 and 2.8 for river/suburban canals and urban canals, respectively. For the wet season, data determined by the membrane filtration method in river/suburban canals and urban canals samples had mean counts of log 3.7 and 4.1, respectively. While mean log CFU/100 ml counts in the wet season using quantitative PCR were log 3 and 2, respectively. Additionally, the urban canal samples were significantly lower than those determined by conventional culture methods for the wet season. These results show that while quantitative real-time PCR can be used to determine levels of fecal indicator bacteria in surface waters, there are some limitations to its application and it may be impacted by sources of runoff based on surveyed samples.

Antimicrobial resistance in Escherichia coli and Salmonella spp. isolates from fresh produce and the impact to food safety.

Foodborne diseases associated with fresh produce consumption have escalated worldwide, causing microbial safety of produce of critical importance. Bacteria that have increasingly been detected in fresh produce are Escherichia coli and Salmonella spp., both of which have been shown to progressively display antimicrobial resistance. The study focused on the assessment of antimicrobial resistance of these enteric bacteria from different kinds of fresh produce from various open air markets and supermarkets in the Philippines. Using the disk diffusion assay on a total of 50 bacterial isolates obtained from 410 fresh produce surveyed, monoresistance to tetracycline was observed to be the most prevalent (38%), followed by multidrug resistance to tetracycline, chloramphenicol, ciprofloxacin, and nalidixic acid (4%), and lastly by dual resistance to tetracycline and chloramphenicol (2%). Using multiplex and simplex polymerase chain reaction (PCR) assays, tetA (75%) and tetB (9%) were found in tetracycline resistant isolates, whereas catI (67%) and catIII (33%) were detected in chloramphenicol resistant isolates. Sequence analysis of gyr and par genes from the ciprofloxacin and nalidixic acid resistant isolates revealed different mutations. Based on the results, fresh produce act as a reservoir of these antibiotic resistant bacteria which may pose health threat to consumers.

Detection of Class I and II integrons for the assessment of antibiotic and multidrug resistance among Escherichia coli isolates from agricultural irrigation waters in Bulacan, Philippines.

Contaminated irrigation water may greatly affect not only the quality of produce but also the people exposed to it. In this study, agricultural irrigation waters in Bulacan, Philippines were assessed and found to be contaminated with Escherichia coli (E. coli) ranging from 0.58 to 4.51 log10 CFU/mL. A total of 79 isolates of E. coli were confirmed through polymerase chain reaction (PCR) amplifying the uidA gene and were tested for phenotypic resistance using 10 antimicrobials through the Kirby-Bauer disc diffusion method. Forty-six isolates (58.22%) were noted to be multidrug resistant (MDR) with high resistance rate to cephalothin, tetracycline, streptomycin, ampicillin, trimethoprim, nalidixic acid, and chloramphenicol. Moreover, this study also examined the prevalence of Class I and II integrons accounting to 67.39% and 17.39%, respectively, of the MDR E. coli strains using multiplex PCR. The results imply that the agricultural water used in Bulacan is contaminated with the fecal material of man or other animals present in the area, and the presence of MDR bacteria, which pose a potential threat to individuals in these areas, is alarming. In addition, detection of integrons could be a good marker for the identification of MDR isolates. Lastly, this study could develop strategies for the proper management of farming sites leading to the detection of food-borne pathogens and prevention of infectious diseases.

Fecal contamination in irrigation water and microbial quality of vegetable primary production in urban farms of Metro Manila, Philippines.

Microbial contamination of fresh produce can present a severe risk to public health. By conducting a rigorous survey of irrigation waters, the impacts of fecal contamination on the quality of produce could be assessed. In this study, surface waters were observed to be contaminated with Escherichia coli, Salmonella spp., and somatic coliphages. Culture methods show that out of 373 irrigation water, soil, and vegetable samples collected for a 1-year period, 232 (62.20%) were found positive for E. coli, 213 (57.26%) for somatic coliphages, and 2 (0.53%) for Salmonella spp. Out of 190 water samples, 167 (87.9%) were found to have E.coli, 174 (91.6%) have somatic coliphages, and 1 (0.5%) with Salmonella spp. In soil samples, 36 of 91 (39.6%) have E. coli, 31 (34.0%) have somatic coliphages, and none with Salmonella spp. Lastly, out of 92 vegetable samples, 29 (31.5%), 8 (8.7%), and 1 (1.1%) were found to have E. coli, somatic coliphages, and Salmonella spp., respectively. Molecular analysis confirmed the presence of bacterial contaminants. Seasonal weather conditions were noted to have an effect on the presence and number of these fecal indicator organisms. The observed data suggest that contaminated irrigation water may greatly affect the quality of fresh produce from these agricultural operations.

Microbiological quality of fresh produce from open air markets and supermarkets in the Philippines.

This study is the first in the Philippines to conduct a comprehensive assessment of the prevalence of bacterial pathogens and somatic phages in retailed fresh produce used in salad preparation, namely, bell pepper, cabbage, carrot, lettuce, and tomato, using culture and molecular methods. Out of 300 samples from open air and supermarkets, 16.7% tested positive for thermotolerant Escherichia coli, 24.7% for Salmonella spp., and 47% for somatic phages. Results show that counts range from 0.30 to 4.03 log10 CFU/g for E. coli, 0.66 to ≥ 2.34 log10 MPN/g for Salmonella spp., and 1.30 to ≥ 3.00 log 10 PFU/g for somatic phages. Statistical analyses show that there was no significant difference in the microbial counts between open air and supermarkets (α = 0.05). TaqMan and AccuPower Plus DualStar real-time polymerase chain reaction (RT-PCR) was used to confirm the presence of these organisms. The relatively high prevalence of microorganisms observed in produce surveyed signifies reduction in shelf-life and a potential hazard to food safety. This information may benefit farmers, consumers, merchants, and policy makers for foodborne disease detection and prevention.

Prevalence of Escherichia coli in surface waters of Southeast Asian cities.

Surface water samples were collected from rivers which fed into large urban areas within Vietnam, Indonesia, Cambodia, and Thailand and were processed to enumerate Escherichia coli. Selected isolates were further characterized using PCR to detect the presence of specific virulence genes. Analyzing the four countries together, the approximate mean cfu/100 ml for E. coli counts in the dry season were log 4.3, while counts in the wet season were log 2.8. Of the 564 E. coli isolates screened for the presence of pathogenic genes, 3.9 % possessed at least one virulence gene. The most common pathogenic types found were Shiga toxin-producing E. coli isolates. These results reinforce the importance of monitoring urban surface waters for fecal contamination, that E. coli in these water environments may serve as opportunistic pathogens, and may help in determining the impact water usage from these rivers have on the public health of urban populations in Southeast Asia.

Do Your Kids Get What You Paid for? Evaluation of Commercially Available Probiotic Products Intended for Children in the Republic of the Philippines and the Republic of Korea.

A wide range of probiotic products is available on the market and can be easily purchased over the counter and unlike pharmaceutical drugs, their commercial distribution is not strictly regulated. In this study, ten probiotic preparations commercially available for children's consumption in the Republic of the Philippines (PH) and the Republic of Korea (SK) have been investigated. The analyses included determination of viable counts and taxonomic identification of the bacterial species present in each formulation. The status of each product was assessed by comparing the results with information and claims provided on the label. In addition to their molecular identification, safety assessment of the isolated strains was conducted by testing for hemolysis, biogenic amine production and antibiotic resistance. One out of the ten products contained lower viable numbers of recovered microorganisms than claimed on the label. Enterococcus strains, although not mentioned on the label, were isolated from four products. Some of these isolates produced biogenic amines and were resistant to one or several antibiotics. Metagenomic analyses of two products revealed that one product did not contain most of the microorganisms declared in its specification. The study demonstrated that some commercial probiotic products for children did not match their label claims. Infants and young children belong to the most vulnerable members of society, and food supplements including probiotics destined for this consumer group require careful checking and strict regulation before commercial distribution.

Detection of pathogenic bioaerosols and occupational risk in a Philippine landfill site.

Payatas landfill site is one of the major dumpsites in the Philippines. In this study, the landfill was evaluated regarding the presence and concentration of pathogenic bacterial aerosols through culture and molecular techniques. Bioaerosols are biological airborne particles that may or may not cause diseases. Active sampling was done using MAS-100 Air Sampler from both indoor and outdoor locations. To test the accuracy of the culture method, those that were presumptively identified as Staphylococcus aureus and Klebsiella pneumoniae were subjected to molecular analysis using 16s rRNA sequencing, and as a result, they were identified as different species. The exposure assessment done by interview showed matches among prevalent diseases. The results of this study can be used as a basis for creating standards and regulations for workers' and residents' safety and health in the landfill site.

Assessment of airborne bacteria in selected occupational environments in Quezon City, Philippines.

Exposure to bioaerosols has been associated with health deterioration among workers in several occupational environments. This highlights the need to study the microbiological quality of air of workplaces as no such study has been conducted yet in the Philippines. To detect and characterize the culturable mesophilic airborne bacteria in selected occupational environments we used passive sedimentation technique. It was observed that the number of colony-forming units was highest in junk shop, followed by the light railway transit station and last the office. By contrast, the bacterial composition was similar in all sites: Gram-positive cocci > Gram-positive bacilli > Gram-negative bacteria. Staphylococcus aureus and Bacillus spp. were also detected in all sites. These findings suggest that the presence of airborne bacteria may be a potential health hazard in urban occupational environments in the Philippines.

Antimicrobial activity, cytotoxicity, and phytochemical screening of Voacanga globosa (Blanco) Merr. leaf extract (Apocynaceae).

OBJECTIVE: To determine the antibacterial, antifungal, antiprotozoal, cytotoxic, and phytochemical properties of ethanol extracts of leaves of Voacanga globosa (Blanco) Merr. (V. globosa). METHODS: The extracts were tested against bacteria and fungus through disc diffusion assay; against protozoa through growth curve determination, antiprotozoal and cytotoxicity assays. RESULTS: The extract revealed antibacterial activities, inhibiting the growth of Staphylococcus aureus, Bacillus cereus, Pseudomonas aeruginosa, Micrococcus luteus, and Salmonella typhimurium. Antifungal assay showed that it inhibited Candida albicans. The antiprotozoal assay against Trichomonas vaginalis and Entamoeba histolytica showed that V. globosa can inhibit the parasites, wherein the action can be comparable to metronidazole. With the in situ cell death detection kit, Trichomonas vaginalis and Entamoeba histolytica exposed to V. globosa leaf extract was observed to fluoresce simultaneously in red and yellow signals signifying apoptotic-like changes. Preliminary phytochemical screening revealed the chemical composition of plant extract containing alkaloids, saponins, 2-deoxysugars, and hydrolysable tannins. CONCLUSIONS: Thus, this study provides scientific evidence on the traditional use of V. globosa leaf extract in treating microbial diseases. Further, the leaf extract can possibly be used to produce alternative forms of antimicrobials.