Infectious diseases, in particular bacterial infections, are the leading cause of morbidity and mortality posing a global threat to human health. The emergence of antibiotic resistance has exacerbated the problem further. Hence, there is a need to search for novel sources of antibacterials. Herein, we explored gut bacteria of a variety of animals living in polluted environments for their antibacterial properties against multi-drug resistant pathogenic bacteria. A variety of species were procured including invertebrate species, Blaptica dubia (cockroach), Gromphadorhina portentosa (cockroach), Scylla serrata (crab), Grammostola rosea (tarantula), Scolopendra subspinipes (centipede) and vertebrate species including Varanus salvator (water monitor lizard), Malayopython reticulatus (python), Cuora amboinensis (tortoise), Oreochromis mossambicus (tilapia fish), Rattus rattus (rat), Gallus gallus domesticus (chicken) and Lithobates catesbeianus (frog). Gut bacteria of these animals were isolated and identified using microbiological, biochemical, analytical profiling index (API) and through molecluar identification using 16S rRNA sequencing. Bacterial conditioned media (CM) were prepared and tested against selected Gram-positive and Gram-negative pathogenic bacteria as well as human cells (HaCaT). The results revealed that CM exhibited significant broad-spectrum antibacterial activities. Upon heat inactivation, CM retained their antibacterial properties suggesting that this effect may be due to secondary metabolites or small peptides. CM showed minimal cytotoxicity against human cells. These findings suggest that gut bacteria of animals living in polluted environments produce broad-spectrum antibacterial molecule(s). The molecular identity of the active molecule(s) together with their mode of action is the subject of future studies which could lead to the rational development of novel antibacterial(s).
Pathogenic free-living amoebae are known to cause fatal central nervous system infections with extremely high mortality rates. High selectivity of the blood-brain barrier hampers delivery of drugs and untargeted delivery of drugs can cause severe side effects. Nanovehicles can be used for targeted drug delivery across the blood-brain barrier. Inorganic nanoparticles have been explored as carriers for various biomedical applications and can be modified with various ligands for efficient targeting and cell selectivity while lipid-based nanoparticles have been extensively used in the development of both precision and colloidal nanovehicles. Nanomicelles and polymeric nanoparticles can also serve as nanocarriers and may be modified so that responsiveness of the nanoparticles and release of the loads are linked to specific stimuli. These nanoparticles are discussed here in the context of the treatment of central nervous system infections due to pathogenic amoebae. It is anticipated that these novel strategies can be utilized in tandem with novel drug leads currently in the pipeline and yield in the development of much needed treatments against these devastating parasites.
Pollution, unhygienic conditions and organic waste are detrimental to human health. On the contrary, animals living in polluted environments, feeding on organic waste and exposed to noxious agents such as heavy metals must possess remarkable properties against contracting diseases. Species such as cockroaches and water monitor lizards thrive in unhygienic conditions and feed on decaying matter. Here, we investigated the antitumour properties of metabolites produced by gut bacteria isolated from Varanus salvator (Asian water monitor lizard). An adult water monitor lizard and a juvenile water monitor lizard were acquired, and dissected. Their aerobic gut bacteria were isolated and identificated through 16S rDNA sequencing. Next, bacterial conditioned media (CM) were prepared and utilised for subsequent assays. Growth inhibition, MTT 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide cell viability assay, cytotoxicity and cell survival assays were accomplished against a panel of cancer cells as well as a normal cell line. Furthermore, liquid chromatography-mass spectrometry (LC-MS) was employed to identify potential antitumour molecules. A plethora of bacteria were isolated from the gut of juvenile and adult V. salvator respectively. Moreover, CM prepared from selected bacteria exhibited antitumour activity. Of note, LC-MS results indicated the presence of several molecules with reported antitumour activity, namely, 3-butylidene-7-hydroxyphthalide, C75, enigmol, estrone 16-oxime, proglumide and S-allyl-L-cysteine. Furthermore, 356 potentially novel molecules from juvenile V. salvator and 184 from adult V. salvator were depicted. Thus, the gut microbiota of V. salvator might be considered as a great niche of antitumour molecules; however, further in vitro and in vivo studies are needed to assess the antitumour properties of these molecules.
Extended-spectrum β-lactamase (ESBL)-producing Klebsiella pneumoniae has been associated with a wide range of infections in humans and animals. The objective of this study was to determine the genomic characteristics of two multiple drug resistant, ESBLs-producing K. pneumoniae strains isolated from a swine in 2013 (KP2013Z28) and a hospitalized patient in 2014 (KP2014C46) in Malaysia. Genomic analyses of the two K. pneumoniae strains indicated the presence of various antimicrobial resistance genes associated with resistance to β-lactams, aminoglycosides, colistin, fluoroquinolones, phenicols, tetracycline, sulfonamides, and trimethoprim, corresponding to the antimicrobial susceptibility profiles of the strains. KP2013Z28 (ST25) and KP2014C46 (ST929) harbored 5 and 2 genomic plasmids, respectively. The phylogenomics of these two Malaysian K. pneumoniae, with other 19 strains around the world was determined based on SNPs analysis. Overall, the strains were resolved into five clusters that comprised of strains with different resistance determinants. This study provided a better understanding of the resistance mechanisms and phylogenetic relatedness of the Malaysian strains with 19 strains isolated worldwide. This study also highlighted the needs to monitor the usage of antibiotics in hospital settings, animal husbandry, and agricultural practices due to the increase of β-lactam, aminoglycosides, tetracycline, and colistin resistance among pathogenic bacteria for better infection control.
Chikungunya virus is a reemerging arbovirus transmitted mainly by Aedes mosquitoes. As there are no specific treatments available, Chikungunya virus infection is a significant public health problem. This study investigated 120 extracts from selected medicinal plants for anti-Chikungunya virus activity. The plant materials were subjected to sequential solvent extraction to obtain six different extracts for each plant. The cytotoxicity and antiviral activity of each extract were examined using African monkey kidney epithelial (Vero) cells. The ethanol, methanol and chloroform extracts of Tradescantia spathacea (Commelinaceae) leaves showed the strongest cytopathic effect inhibition on Vero cells, resulting in cell viabilities of 92.6% ± 1.0% (512 μg/ml), 91.5% ± 1.7% (512 μg/ml) and 88.8% ± 2.4% (80 μg/ml) respectively. However, quantitative RT-PCR analysis revealed that the chloroform extract of Rhapis excelsa (Arecaceae) leaves resulted in the highest percentage of reduction of viral load (98.1%), followed by the ethyl acetate extract of Vernonia amygdalina (Compositae) leaves (95.5%). The corresponding 50% effective concentrations (EC50) and selectivity indices for these two extracts were 29.9 ± 0.9 and 32.4 ± 1.3 μg/ml, and 5.4 and 5.1 respectively. Rhapis excelsa and Vernonia amygdalina could be sources of anti-Chikungunya virus agents. [Int Microbiol 19(3):175-182 (2016)].
A biofilm is a community of microorganisms attached to a surface and embedded in a matrix of extracellular polymeric substances. Biofilms confer resistance towards conventional antibiotic treatments; thus, there is an urgent need for newer and more effective antimicrobial agents that can act against these biofilms. Due to this situation, various studies have been done to investigate the anti-biofilm effects of natural products including bioactive compounds extracted from microorganisms such as Actinobacteria. This review provides an insight into the anti-biofilm potential of Actinobacteria against various pathogenic bacteria, which hopefully provides useful information, guidance, and improvements for future antimicrobial studies. Nevertheless, further research on the anti-biofilm mechanisms and compound modifications to produce more potent anti-biofilm effects are required.
Human leptospirosis is considered as one of the most widespread and potentially fatal zoonotic diseases that causes high mortality and morbidity in the endemic regions of tropical and subtropical countries. The infection can arise from direct or indirect exposure of human through contaminated environment that contains leptospires or animal reservoirs that carry leptospires. The clinical manifestations during human leptospirosis ranges from asymptomatic, mild infections to severe and life-threatening complications involving multi-organ failures with kidneys, lungs and liver severely affected. Despite much efforts have been put in to unravel the pathogenesis during human leptospirosis, it remains obscure to which extent the host factors or the pathogen itself contribute towards the pathogenesis. Host innate immunity, especially, polymorphonuclear neutrophils and complement system are involved in the first line of defense during human leptospirosis. However, pathogenic Leptospira has acquired diverse evasion strategies to evade from host immunity and establish infection in infected hosts. Hence, in this review, we focus on organs pathology during human leptospiral infection and host evasion strategies employed by Leptospira. A profound understanding on leptospiral immunity and how Leptospira subvert the immune system may provide new insights on the development of therapeutic regimens against this species in future.
Silver nanoparticles (SN) have been recently developed as a new class of antimicrobial agents against numerous pathogenic microorganisms. SN have also been used as efficient drug delivery systems and have been linked with increasing drug potency. Here, we demonstrated the enhanced antifungal efficacy of nystatin (NYT) and fluconazole (FLU) after conjugation with SN. The antifungal bioactivity of NYT- and FLU-coated SN was evaluated against Candida albicans ATCC 10231 and Aspergillus brasiliensis ATCC 16404 by the agar tube dilution method. The aim of this study was to determine and compare the antifungal efficacy of NYT and FLU with their SN and, finally, the combination of both nanoparticles as NYT-SN + FLU-SN against pathogenic fungi. The results indicated that all test samples showed a dose-dependent response against tested fungi. SN significantly enhanced the antifungal effects of NYT and FLU as compared to drugs alone. We observed a remarkable increase in the percent inhibition of both fungi (90-100%) when treated with a combination of both nanoparticles NYT-SN + FLU-SN at 200 μg/mL only. Furthermore, the morphological modifications occurred at the surface of fungal species were also analyzed by atomic force microscopy (AFM) and scanning electron microscopy (SEM). While tested against primary human cell line, all SN showed negligible cytotoxicity. Hence, these results suggest that the combination of SN with NYT and FLU may have clinical implications in the treatment of fungal infections. However, in vivo studies are needed before recommending the use of these nanoparticles safely in clinical situations.
Acanthamoeba keratitis is a sight-endangering eye infection, and causative organism Acanthamoeba presents a significant concern to public health, given escalation of contact lens wearers. Contemporary therapy is burdensome, necessitating prompt diagnosis and aggressive treatment. None of the contact lens disinfectants (local and international) can eradicate Acanthamoeba effectively. Using a range of compounds targeting cellulose, ion channels, and biochemical pathways, we employed bioassay-guided testing to determine their anti-amoebic effects. The results indicated that acarbose, indaziflam, terbuthylazine, glimepiride, inositol, vildagliptin and repaglinide showed anti-amoebic effects. Compounds showed minimal toxicity on human cells. Therefore, effects of the evaluated compounds after conjugation with nanoparticles should certainly be the subject of future studies and will likely lead to promising leads for potential applications.
Paddy is an important crop in Malaysia. There are various pathogens able to infect paddy causing a loss in yield's production. In this study, dual culture method, volatile organic compound (VOC) analysis, and non-volatile compound analysis were used to assess the ability of mushroom to control fungal rice pathogens including Curvularia lunata, Bipolaris panici-miliacei, and Nigrospora sp. Four mushroom isolates were further analysed for their antagonistic activity against rice pathogen. The highest percentage inhibition of radial growth (PIRG) was recorded between 45.55 and 73.68% observed in isolate 42b. The 4 isolates with the highest PIRG based on the dual culture analysis were then tested for their production of VOCs and non-volatile compound. Internal transcribed spacer (ITS) region analysis of the 4 mushroom isolates revealed their identity as Coprinellus disseminates (isolate 12b), Marasmiellus palmivorus (isolate 42b), Trametes maxima (isolate 56e), and Lentinus sajor-caju (isolate 60a). This study showed that mushroom isolates have the potential of antagonistic effect on various fungal rice pathogens tested by the production of secondary metabolites and mycoparasitic interaction.
This study characterized and identified the antimicrobial compounds from an endophytic fungus (Fusarium incarnatum (C4)) isolated from the orchid, Cymbidium sp. Chromatographic techniques were employed to separate the bioactive compounds from the crude extracts of F. incarnatum (C4). Following bio-guided fractionation, two fractionated extracts (fractions 1 and 2) of F. incarnatum (C4) exhibited antibacterial and antifungal activities against Bacillus cereus (MIC: 0.156 mg/mL) and Ganoderma boninense (MIC: 0.3125 mg/mL), respectively. The active fractions were discovered to comprise of a variety of bioactive compounds with pharmacological importance (alkaloids, flavonoids, phenolic compounds, terpenoids, peptides and fatty acids). Liquid chromatography mass-spectrometry (LCMS) analysis detected the presence of antibacterial (kanzonol N, rifaximin, linoleic acid (d4), cannabisativine, docosanedioic acid, and stearamide) and antifungal components (3-methyl-quinolin-2-ol, prothiocarb, kanzonol N, peganine, 5Z-tridecene, and tetronasin) in fractions 1 and 2, respectively, which may have contributed to the antimicrobial effects. Findings from this study highlighted the important potential of fungal endophytes from medicinal hosts as producers of antimicrobials and antibiotics.
Methicillin-susceptible Staphylococcus aureus (MSSA) is an important nosocomial pathogen worldwide. This study aims to investigate the in vitro biofilm-forming ability of clinical MSSA isolated from various sources in the main public tertiary referral hospital in Terengganu, Malaysia and to detect the presence of biofilm-associated and regulatory genes among these isolates. A total of 104 MSSA isolates [pus (n = 75), blood (n = 24), respiratory secretions (n = 2), eye (n = 2), and urine (n = 1)] were investigated for slime production and biofilm formation using Congo red agar and crystal violet microtitre plate, respectively. Fifteen MSSA isolates with varying degrees of biofilm formation were selected for validation via a real-time cell analyser. All isolates were screened for microbial surface components recognising adhesive matrix molecules (MSCRAMM) and accessory gene regulator (agr) using polymerase chain reaction assay. A total of 76.0% (79/104) isolates produced slime layer, while all isolates developed biofilm as follows: 52.8% (55/104) strong biofilm producers, 40.4% (42/104) intermediate biofilm producers, and 6.7% (7/104) weak biofilm producers. A total of 98.1% (102/104) isolates carried at least one of the screened MSCRAMM gene(s) with the eno gene detected at the highest rate (87.5%, 91/104), while the sasG gene was significantly associated with strong biofilm production (p = 0.015). Three agr groups, 1, 2, and 3, were detected among the MSSA isolates with a predominance of agr-3 (32.7%, 34/104). In conclusion, biofilm formation varied greatly among clinical MSSA isolates, and the presence of sasG gene and agr-1 may play important role in initiating MSSA infections via biofilm formation.
Soil salinity in rice cultivation areas is considered a severely limiting factor that adversely affects the quantity and quality of rice production in wetlands. Recently, the alternative use of salt-tolerant plant growth-promoting rhizobacteria (PGPR) inhabiting extreme saline conditions has gained remarkable attention and had positive effects on soil and crops. Therefore, a study has been initiated to develop a liquid biofertilizer formulation from locally isolated multi-strain salt-tolerant PGPR strains such as Bacillus tequilensis and Bacillus aryabhattai, using glycerol (5 mM), trehalose (10 mM), and polyvinylpyrrolidone (PVP) at 1% as additives to prolong the shelf-life of the bacteria. After 3 months of incubation, the bacterial population in the trehalose-supplemented mixed strain was highest at 9.73×107 CFU/mL, followed by UPMRE6 and UPMRB9 at 9.40×107 CFU/mL and 8.50×107 CFU/mL respectively. The results showed that the optimal trehalose concentration successfully prolonged the shelf-life of bacteria with minimal cell loss. Validation of quadratic optimization by response surface methodology revealed that the cell density of the mixed strain was 4.278×107 log CFU/mL after 24 h. The precision ratio was 99.7% higher than the predicted value in the minimized medium formulation: 0.267 g/mL trehalose, 1% glycerol, at 120 rpm agitation using the data analysis tools of Design Expert software. The population study confirmed the better and longer survival of salt-tolerant PGPR fortified with 10 mM trehalose, which was considered the best liquid biofertilizer formulation. Moreover, the optimized trehalose-glycerol liquid formulation can be used commercially as it is cost-effective.
Acanthamoeba are free living amoebae that are the causative agent of keratitis and granulomatous amoebic encephalitis. Alpha-Mangostin (AMS) is a significant xanthone; that demonstrates a wide range of biological activities. Here, the anti-amoebic activity of α-Mangostin and its silver nano conjugates (AMS-AgNPs) were evaluated against pathogenic A. castellanii trophozoites and cysts in vitro. Amoebicidal assays showed that both AMS and AMS-AgNPs inhibited the viability of A. castellanii dose-dependently, with an IC50 of 88.5 ± 2.04 and 20.2 ± 2.17 μM, respectively. Both formulations inhibited A. castellanii-mediated human keratinocyte cell cytopathogenicity. Functional assays showed that both samples caused apoptosis through the mitochondrial pathway and reduced mitochondrial membrane potential and ATP production, while increasing reactive oxygen species (ROS) and nicotinamide adenine dinucleotide phosphate (NADPH) cytochrome-c reductase in the cytosol. Whole transcriptome sequencing of A. castellanii showed the expression of 826 genes, with 447 genes being up-regulated and 379 genes being down-regulated post treatment. The Kyoto Encyclopedia of Genes and Genomes analysis showed that the majority of genes were linked to apoptosis, autophagy, RAP1, AGE-RAGE and oxytocin signalling pathways. Seven genes (PTEN, H3, ARIH1, SDR16C5, PFN, glnA GLUL, and SRX1) were identified as the most significant (Log2 (FC) value 4) for molecular mode of action in vitro. Future in vivo studies with AMS and nanoconjugates are needed to realize the clinical potential of this work.
Sulfitobacter is a bacterium recognized for its production of AMP-independent sulfite oxidase, which is instrumental in the creation of sulfite biosensors. This capability underscores its ecological and economic relevance. In this study, we present a newly discovered phage, Sulfitobacter phage vB_SupP_AX, which was isolated from Maidao of Qingdao, China. The vB_SupP_AX genome is linear and double-stranded and measures 75,445 bp with a GC content of 49%. It encompasses four transfer RNA (tRNA) sequences and 79 open reading frames (ORFs), one of which is an auxiliary metabolic gene encoding thioredoxin. Consistent with other N4-like phages, vB_SupP_AX possesses three distinct RNA polymerases and is characterized by the presence of four tRNA molecules. Comparative genomic and phylogenetic analyses position vB_SupP_AX and three other viral genomes from the Integrated Microbial Genomes/Virus v4 database within the Rhodovirinae virus subfamily. The identification of vB_SupP_AX enhances our understanding of virus-host interactions within marine ecosystems.
The influence of light regulation on fungal growth and enzyme production was tested on endophytic isolates of Fusarium proliferatum (CCH), Colletotrichum boninense (PL1, PL9, OL2), Colletotrichum gloeosporiodes (OL3) and Colletotrichum siamense (PL3). The isolates were treated with blue, red, green, and yellow light, while white fluorescent light (12 h light/12 h dark photoperiod) and 24 h dark conditions were applied as control. Results revealed that coloured light treatments induced formation of circadian rings, while exposure to white light and dark conditions showed less pronounced circadian rings. Growth and sporulation of endophytes were not significantly influenced by light. By contrast, enzyme production was affected by coloured light treatments, notably with red (amylase), blue (cellulase) and yellow (cellulase, xylanase, L-asparaginase) light, resulting in lower enzyme levels for certain isolates. Under control conditions, enzyme production was relatively higher for amylase, cellulase, xylanase (for cultures incubated in the dark), and for L-asparaginase (for cultures incubated in white fluorescent light). Among the endophytic isolates, F. proliferatum (CCH) showed better response to coloured light treatment as higher sporulation and enzyme production was detected, although growth was significantly suppressed. On the contrary, C. gloeosporiodes (OL3) showed better growth but significantly lower enzyme production and sporulation when treated with the various coloured light. This study revealed that coloured light may have the potential to manipulate growth, sporulation and enzyme production in certain fungal species as strategies for fungal control or for harnessing of valuable enzymes.
Soil salinity has been one of the significant barriers to improving rice production and quality. According to reports, Bacillus spp. can be utilized to boost plant development in saline soil, although the molecular mechanisms behind the interaction of microbes towards salt stress are not fully known. Variations in rice plant protein expression in response to salt stress and plant growth-promoting rhizobacteria (PGPR) inoculations were investigated using a proteomic method and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Findings revealed that 54 salt-responsive proteins were identified by mass spectrometry analysis (LC-MS/MS) with the Bacillus spp. interaction, and the proteins were functionally classified as gene ontology. The initial study showed that all proteins were labeled by mass spectrometry analysis (LC-MS/MS) with Bacillus spp. interaction; the proteins were functionally classified into six groups. Approximately 18 identified proteins (up-regulated, 13; down-regulated, 5) were involved in the photosynthetic process. An increase in the expression of eight up-regulated and two down-regulated proteins in protein synthesis known as chaperones, such as the 60 kDa chaperonin, the 70 kDa heat shock protein BIP, and calreticulin, was involved in rice plant stress tolerance. Several proteins involved in protein metabolism and signaling pathways also experienced significant changes in their expression. The results revealed that phytohormones regulated the manifestation of various chaperones and protein abundance and that protein synthesis played a significant role in regulating salt stress. This study also described how chaperones regulate rice salt stress, their different subcellular localizations, and the activity of chaperones.
Enzymes play a crucial role in various industrial sectors. These biocatalysts not only ensure sustainability and safety but also enhance process efficiency through their unique specificity. Lipases possess versatility as biocatalysts and find utilization in diverse bioconversion reactions. Presently, microbial lipases are gaining significant focus owing to the rapid progress in enzyme technology and their widespread implementation in multiple industrial procedures. This updated review presents new knowledge about various origins of microbial lipases, such as fungi, bacteria, and yeast. It highlights both the traditional and modern purification methods, including precipitation and chromatographic separation, the immunopurification technique, the reversed micellar system, the aqueous two-phase system (ATPS), and aqueous two-phase flotation (ATPF), moreover, delves into the diverse applications of microbial lipases across several industries, such as food, vitamin esters, textile, detergent, biodiesel, and bioremediation. Furthermore, the present research unveils the obstacles encountered in employing lipase, the patterns observed in lipase engineering, and the application of CRISPR/Cas genome editing technology for altering the genes responsible for lipase production. Additionally, the immobilization of microorganisms' lipases onto various carriers also contributes to enhancing the effectiveness and efficiencies of lipases in terms of their catalytic activities. This is achieved by boosting their resilience to heat and ionic conditions (such as inorganic solvents, high-level pH, and temperature). The process also facilitates the ease of recycling them and enables a more concentrated deposition of the enzyme onto the supporting material. Consequently, these characteristics have demonstrated their suitability for application as biocatalysts in diverse industries.