A three-dimensional (3D) printing approach to fabricate an isolation chip for high throughput in situ cultivation of environmental microbes.

The full plethora of environmental bacteria is often poorly represented in vitro as the majority remain difficult, if not impossible, to culture under standard laboratory settings. These bacteria often require native conditions for the formation of cell masses that collectively have higher chances of survival. With that, a 3D-printed version of the isolation chip (iChip) was used to cultivate bacteria from a tropical peat swamp in situ prior to growth and maintenance in vitro. Briefly, plates made from either acrylonitrile butadiene styrene (ABS), polylactic acid (PLA), or epoxy resin were tested in terms of their usability and durability under acidic conditions similar to those of peat matter. The epoxy resin plates were then found to be most optimal for the sampling conditions. Peat soil samples were collected from the base of a Koompassia malaccensis tree and reconstituted in molten 10% (wt/vol) tryptone soy agar (TSA) prior to inoculation. The iChips were subsequently assembled and buried in the site of origin. As a comparison, bacteria from the same soil sample were cultivated directly on TSA and incubated at 28 °C for two weeks. Thereafter, agar plugs from the iChip were transferred to TSA plates to allow microcolonies within each plug to grow. Each pure isolate from both cultivation approaches that grew was then pooled and extracted for total DNA prior to 16S rRNA gene amplification and sequencing via Illumina MiSeq. Taxonomic abundance comparison revealed that the bacterial taxa at the level of order were significantly different between the two approaches, particularly in the orders, Burkholderiales, Xanthomonodales, Enterobacteriales, and Actinomycetales (differences of 12.0, 7.1, 8.0, and 4.2%, respectively). This indicated that the 3D-printed iChips present a possible low-cost tool for the isolation of bacterial genera that may not be able to grow on media directly in vitro.

Extremely low prevalence in soil-transmitted helminth infections among a multi-ethnic community in Segamat, Malaysia.

Soil-transmitted helminth infections (STHs) are recognized as a major health issue among socio-economically deprived communities. However, information is still lacking regarding the prevalence rates of STHs in the broader community across different countries in the tropics. This community study aimed to determine the prevalence and risk factors for STHs in semi-rural communities in Segamat of Johor, Malaysia. A cross-sectional study was conducted with information collected from the study population through questionnaire. A total of 224 stool samples were examined for intestinal parasites through formalin-ether concentration and Kato-Katz techniques. Overall, only 1.8% (n = 4/224) of participants were infected with soil-transmitted helminths, the extremely low prevalence may be explained by the proper housing conditions with basic amenities and the practices of hygienic habits in daily life, highlighting the importance of adopting good hygienic practices.

Human intestinal parasitic infection: a narrative review on global prevalence and epidemiological insights on preventive, therapeutic and diagnostic strategies for future perspectives.

INTRODUCTION: Intestinal parasitic infection (IPI) is a global health concern among socioeconomically deprived communities in many developing countries. Many preventative strategies have been deployed to control IPI, however, there is a lack in standards on the techniques used to diagnose and monitor the prevalence of IPI. AREAS COVERED: The present article will review the diseases associated with IPI and discuss the current IPI control strategies such as the water, sanitation, and hygiene (WASH) interventions, community-led total sanitation (CLTS) approach, and regular anthelminthic treatments. For the first time, this review will also evaluate all currently practised diagnostic techniques for the detection of intestinal parasites and provide insights on future IPI control strategies. EXPERT OPINION: Advanced and improved diagnostic methods such as qPCR coupled with a high-resolution melting curve, aptamers, biosensors, and detection of extracellular vesicles can be used for detection of IPI. Vaccination against intestinal parasites can be made available to increase antibodies to interfere with the blood-feeding process by the parasites, which subsequently reduces the reproductive rates of the parasites. These methods collectively can serve as future management strategies for intestinal parasitic infections.

First reported case of Gilbertella persicaria in human stool: outcome of a community study from Segamat, Johor, Malaysia.

Species of fungi belonging to the order Mucorales can be found everywhere in the environment. Gilbertella persicaria, which belongs to this order, have often been isolated from fruits and in water systems. However, there has been no report of isolation of this fungus from human samples. During a gut mycobiome study, from the Segamat community, Gilbertella persicaria was isolated from a human fecal sample and was characterized through a series of morphological assessment, biochemical tests, and molecular techniques. The isolate produced a white velvety surface that turned grayish after 24 h. Although no biofilm production was observed, the results indicated that the isolate could form calcium oxalate crystals, produced urease, and was resistant to low pH. The isolate was sensitive to amphotericin but resistant to voriconazole and itraconazole. The features of this fungus that could help in its survival in the human gut are also discussed.

Draft Genome Sequence of Dyella sp. Strain C9, Isolated from a Malaysian Tropical Peat Swamp Forest.

The bacterium Dyella sp. strain C9 was isolated from North Selangor Peat Swamp Forest, Malaysia, and studied using whole-genome sequencing. The putative genes involved in biogeochemical processes were annotated, and the genome sequence is publicly available in the NCBI database.

Current anti-biofilm strategies and potential of antioxidants in biofilm control.

INTRODUCTION: Biofilm formation is a strategy for microorganisms to adapt and survive in hostile environments. Microorganisms that are able to produce biofilms are currently recognized as a threat to human health. Areas covered: Many strategies have been employed to eradicate biofilms, but several drawbacks from these methods had subsequently raised concerns on the need for alternative approaches to effectively prevent biofilm formation. One of the main mechanisms that drives a microorganism to transit from a planktonic to a biofilm-sessile state, is oxidative stress. Chemical agents that could target oxidative stress regulators, for instance antioxidants, could therefore be used to treat biofilm-associated infections. Expert commentary: The focus of this review is to summarize the function and limitation of the current anti-biofilm strategies and will propose the use of antioxidants as an alternative method to treat, prevent and eradicate biofilms. Studies have shown that water-soluble and lipid-soluble antioxidants can reduce and prevent biofilm formation, by influencing the expression of genes associated with oxidative stress. Further in vivo work should be conducted to ensure the efficacy of these antioxidants in a biological environment. Nevertheless, antioxidants are promising anti-biofilm agents, and thus is a potential solution for biofilm-associated infections in the future.

Draft Genome Sequence of Dyella sp. Strain C11, Isolated from a Malaysian Tropical Peat Swamp Forest.

We report here the draft genome sequences of a bacterial isolate, Dyella sp. strain C11, which was isolated from a Malaysian tropical peat swamp forest. The putative genes for the biogeochemical processes were annotated, and the genome was deposited in an online database.

Draft Genome Sequence of Klebsiella sp. Strain C31 Isolated from a Malaysian Tropical Peat Swamp Forest.

We report here the draft genome of Klebsiella sp. strain C31, a bacterial isolate from the North Selangor peat swamp forest in Malaysia. The putative genes for the biogeochemical processes of the genome were annotated and investigated.

Draft Genome Sequence of Paraburkholderia sp. Strain C35, Isolated from a Malaysian Tropical Peat Swamp Forest.

We report the draft genome sequence of a bacterial isolate, Paraburkholderia sp. strain C35, which was isolated from a Malaysian tropical peat swamp forest. The putative genes for the biogeochemical processes were annotated and are publicly available in the online databases.

The role of reactive oxygen species in the antimicrobial activity of pyochelin.

The increase in prevalence of antimicrobial-resistant bacteria (ARB) is currently a serious threat, thus there is a need for new antimicrobial compounds to combat infections caused by these ARB. An antimicrobial-producing bacterium, Burkholderia paludis was recently isolated and was able to produce a type of siderophore with antimicrobial properties, later identified as pyochelin. The chelating ability of pyochelin has been well-characterized but not for its antimicrobial characteristics. It was found that pyochelin had MIC values (MBC values) of 3.13 µg/mL (6.26 µg/mL) and 6.26 µg/mL (25.00 µg/mL) against three Enterococcus strains and four Staphylococcus strains. Pyochelin was able to inhibit E. faecalis ATCC 700802 (a vancomycin-resistant strain) in a time and dose dependent manner via killing kinetics assay. It was demonstrated that pyochelin enhanced the production of intracellular reactive oxygen species (ROS) over time, which subsequently caused a significant increase in malondialdehyde (MDA) production (a marker for lipid peroxidation) and ultimately led to cell death by disrupting the integrity of the bacterial membrane (validated via BacLight assay). This study has revealed the mechanism of action of pyochelin as an antimicrobial agent for the first time and has shown that pyochelin might be able to combat infections caused by E. faecalis in the future.

Newly Isolated Paenibacillus tyrfis sp. nov., from Malaysian Tropical Peat Swamp Soil with Broad Spectrum Antimicrobial Activity.

Emergence of antimicrobial resistance coupled with the slowdown in discovery of new antimicrobial compounds points to serious consequences for human health. Therefore, scientists are looking for new antimicrobial compounds from unique and understudied ecosystems such as tropical peat swamp forests. Over the course of isolating antimicrobial producing bacteria from North Selangor tropical peat swamp forest, Malaysia, a Gram variable, rod shaped, endospore forming, facultative anaerobic novel strain MSt1(T) that exerts potent and broad spectrum antimicrobial activity was isolated. Phylogenetic analysis using 16S rRNA gene sequences showed that strain MSt1(T) belonged to the genus Paenibacillus with the highest similarity to Paenibacillus elgii SD17(T) (99.5%). Whole genome comparison between strain MSt1(T) with its closely related species using average nucleotide identity (ANI) revealed that similarity between strain MSt1(T) with P. elgii B69 (93.45%) and Paenibacillus ehimensis A2 (90.42%) was below the recommended threshold of 95%. Further analysis using in silico pairwise DDH also showed that similarity between strain MSt1(T) with P. elgii B69 (55.4%) and P. ehimensis A2 (43.7%) was below the recommended threshold of 70%. Strain MSt1(T) contained meso-diaminopilemic acid in the cell wall and MK-7 as the major menaquinone. The major fatty acids of strain MSt1(T) were anteiso-C15:0 (48.2%) and C16:0 (29.0%) whereas the polar lipid profile consisted of phosphatidylglycerol, phosphatidylethanolamine, diphosphatidylglycerol, one unknown lipid, two unknown glycolipids, and one unknown phospholipid. Total DNA G+C content of strain MSt1(T) was 51.5 mol%. The extract from strain MSt1(T) exerted strong antimicrobial activity against Escherichia coli ATCC 25922 (MIC = 1.5 µg/mL), MRSA ATCC 700699 (MIC = 25 µg/mL) and Candida albicans IMR (MIC = 12.5 µg/mL). Partially purified active fraction exerted a strong effect against E. coli ATCC 25922 resulting in cell rupture when viewed with SEM. Based on distinctive taxonomic differences between strain MSt1(T) when compared to its closely related type species, we propose that strain MSt1(T) represents a novel species within the genus of Paenibacillus, for which the name Paenibacillus tyrfis sp. nov. (= DSM 100708(T) = MCCC 1K01247(T)) is proposed.

Burkholderia paludis sp. nov., an Antibiotic-Siderophore Producing Novel Burkholderia cepacia Complex Species, Isolated from Malaysian Tropical Peat Swamp Soil.

A novel Gram negative rod-shaped bacterium, designated strain MSh1T, was isolated from Southeast Pahang tropical peat swamp forest soil in Malaysia and characterized using a polyphasic taxonomy approach. The predominant cellular fatty acids (>10.0%) were C16:0 (31.7%), C17:0 cyclo (26.6%), and C19:0 cyclo ω8c (16.1%). The polar lipids detected were phosphatidylglycerol, phosphatidylethanolamine, and diphosphatidylglycerol. The predominant ubiquinone was Q-8. This revealed that strain MSh1T belongs to the genus Burkholderia. The type strain MSh1T can be differentiated from other Burkholderia cepacia complex (Bcc) species by phylogenetic analysis of 16S rRNA gene sequence, multilocus sequence analysis (MLSA), average nucleotide identity (ANI) and biochemical tests. DNA-DNA relatedness values between strain MSh1T and closely related type strains were below the 70% threshold value. Based on this polyphasic study of MSh1T, it can be concluded that this strain represents a novel species within the Bcc, for which the name Burkholderia paludis sp. nov. is proposed. The type strain is MSh1T (= DSM 100703T = MCCC 1K01245T). The dichloromethane extract of MSh1T exhibited antimicrobial activity against four Gram positive bacteria (Enterococcus faecalis ATCC 29212, E. faecalis ATCC 700802, Staphylococcus aureus ATCC 29213, S. aureus ATCC 700699) and a Gram negative bacteria (Escherichia coli ATCC 25922). Further purification work has led to the isolation of Compound 1, pyochelin. Pyochelin demonstrated antimicrobial activity against four S. aureus strains and three E. faecalis strains with MIC-values of 3.13 µg/ml and 6.26 µg/ml, respectively. SEM analysis showed that the cellular morphology of E. faecalis ATCC 700802 was not affected by pyochelin; suggesting that it might target the intracellular components. Pyochelin, a siderophore with antimicrobial activity might be useful in treating bacterial infections caused by S. aureus and E. faecalis, however further work has to be done.

Draft Genome Sequence of Paenibacillus sp. Strain MSt1 with Broad Antimicrobial Activity, Isolated from Malaysian Tropical Peat Swamp Soil.

We report the draft genome sequence of Paenibacillus sp. strain MSt1, which has broad-range antimicrobial activity, isolated from tropical peat swamp soil. Genes involved in antimicrobial biosynthesis are found to be present in this genome.

Draft Genome Sequences of Two Antimicrobial-Producing Burkholderia sp. Strains, MSh1 and MSh2, Isolated from Malaysian Tropical Peat Swamp Forest Soil.

We report the draft genome sequences of two antimicrobial-producing isolates, Burkholderia sp. strains MSh1 and MSh2, which were isolated from tropical peat swamp forest soil. Putative genes related to different antimicrobial production have been annotated in both genome sequences.