Publications by authors named "Steve Petrovski"

54 Publications

Novel Drexlerviridae bacteriophage KMI8 with specific lytic activity against Klebsiella michiganensis and its biofilms.

PLoS One 2021 7;16(9):e0257102. Epub 2021 Sep 7.

Department of Pharmacy and Biomedical Science, La Trobe Institute for Molecular Science, La Trobe University, Victoria, Australia.

The bacterial genus Klebsiella includes the closely related species K. michiganensis, K. oxytoca and K. pneumoniae, which are capable of causing severe disease in humans. In this report we describe the isolation, genomic and functional characterisation of the lytic bacteriophage KMI8 specific for K. michiganensis. KMI8 belongs to the family Drexlerviridae, and has a novel genome which shares very little homology (71.89% identity over a query cover of only 8%) with that of its closest related bacteriophages (Klebsiella bacteriophage LF20 (MW417503.1); Klebsiella bacteriophage 066039 (MW042802.1). KMI8, which possess a putative endosialidase (depolymerase) enzyme, was shown to be capable of degrading mono-biofilms of a strain of K. michiganensis that carried the polysaccharide capsule KL70 locus. This is the first report of a lytic bacteriophage for K. michiganensis, which is capable of breaking down a biofilm of this species.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0257102PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8423285PMC
September 2021

Draft Genome Sequence of Enterobacter asburiae NCR1, a Plant Growth-Promoting Rhizobacterium Isolated from a Cadmium-Contaminated Environment.

Microbiol Resour Announc 2021 Sep 2;10(35):e0047821. Epub 2021 Sep 2.

Department of Animal, Plant and Soil Sciences, La Trobe University, Bundoora, Victoria, Australia.

Enterobacter asburiae NCR1 is a plant growth-promoting rhizobacterium isolated from the rhizosphere of . We report the draft genome sequence of strain NCR1, which revealed many genes facilitating beneficial interactions with plant hosts.
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http://dx.doi.org/10.1128/MRA.00478-21DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8411915PMC
September 2021

Isolation and Characterisation of the Genus and Phylogenetic Investigation of the Bacteriophages.

Viruses 2021 Aug 6;13(8). Epub 2021 Aug 6.

Department of Physiology, Anatomy & Microbiology, La Trobe University, Melbourne, VIC 3086, Australia.

is a highly diverse genus containing over 200 species that can be problematic in both industrial and medical settings. This is mainly attributed to sp. being intrinsically resistant to an array of antimicrobial compounds, hence alternative treatment options are needed. In this study, two bacteriophages, PumA1 and PumA2 were isolated and characterized. Genome nucleotide analysis identified the two phages as novel at the DNA sequence level but contained proteins similar to phi29 and other related phages. Whole genome phylogenetic investigation of 34 phi29-like phages resulted in the formation of seven clusters that aligned with recent ICTV classifications. PumA1 and PumA2 share high genetic mosaicism and form a genus with another phage named WhyPhy, more recently isolated from the United States of America. The three phages within this cluster are the only candidates to infect . Sequence analysis of phage resistant mutants revealed that PumA1 and PumA2 require polymerized and peptidoglycan bound wall teichoic acid (WTA) for their infection. Bacteriophage classification is continuously evolving with the increasing phages' sequences in public databases. Understanding phage evolution by utilizing a combination of phylogenetic approaches provides invaluable information as phages become legitimate alternatives in both human health and industrial processes.
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http://dx.doi.org/10.3390/v13081557DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8402886PMC
August 2021

Draft Genome Sequence of Enterobacter mori Strain NSE2, Isolated from the Rhizosphere of a sp.

Microbiol Resour Announc 2021 Aug 26;10(34):e0048721. Epub 2021 Aug 26.

Department of Animal, Plant and Soil Sciences, La Trobe University, Bundoora, Victoria, Australia.

Enterobacter mori is an important plant pathogen. Here, we report the draft genome sequence of the plant-associated strain Enterobacter mori NSE2, which was found to harbor genes for promotive and pathogenic interactions with plants.
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http://dx.doi.org/10.1128/MRA.00487-21DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8388540PMC
August 2021

Genetic characterisation of antibiotic resistance transposons Tn6608 and Tn6609 isolated from clinical Pseudomonas strains in Cyprus.

J Glob Antimicrob Resist 2021 Sep 4;26:330-334. Epub 2021 Aug 4.

Department of Physiology, Anatomy & Microbiology, La Trobe University, Bundoora, Victoria 3086, Australia. Electronic address:

Objectives: Antibiotic therapy for Pseudomonas infections is becoming increasingly difficult. In this study, the transposons from two multidrug-resistant (MDR) clinical Pseudomonas strains containing related transposons responsible for giving rise to resistance determinants were characterised.

Methods: Two MDR clinical Pseudomonas isolates were obtained from a medical facility in Cyprus. The strains were identified as Pseudomonas putida C54 and Pseudomonas aeruginosa C69. DNA was extracted from both strains and was sequenced. Transposons were identified, annotated and compared with DNA sequences in GenBank.

Results: Two related nested transposons, here named Tn6608 (from P. putida C54) and Tn6609 (from P. aeruginosa C69), were characterised. The transposons are built on an ancestral Tn1403 base element (here named Tn1403A) that contains only the transposition module (tnpA and tnpR) and the associated cargo gene module (orfA, orfB, orfC and orfD) flanked by a 38-bp inverted repeat. The nested transposons identified in this study have evolved via acquisition of multiple transposons, adding multiple resistance genes to an ancestral transposon that originally lacked any resistance determinants.

Conclusion: Transposons related to Tn6608 and Tn6609 have evolved and are globally disseminated. Of particular interest is that most of these nested transposons are located within the same site in a genomic island, providing alternative avenues for dissemination.
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http://dx.doi.org/10.1016/j.jgar.2021.07.016DOI Listing
September 2021

Environmental hotspots for antibiotic resistance genes.

Microbiologyopen 2021 06;10(3):e1197

Department of Physiology, Anatomy and Microbiology, School of Life Sciences, College of Science, Health and Engineering, La Trobe University, Bundoora, Vic, Australia.

Bacterial resistance toward broad-spectrum antibiotics has become a major concern in recent years. The threat posed by the infectious bacteria and the pace with which resistance determinants are transmitted needs to be deciphered. Soil and water contain unique and diverse microbial communities as well as pools of naturally occurring antibiotics resistant genes. Overuse of antibiotics along with poor sanitary practices expose these indigenous microbial communities to antibiotic resistance genes from other bacteria and accelerate the process of acquisition and dissemination. Clinical settings, where most antibiotics are prescribed, are hypothesized to serve as a major hotspot. The predisposition of the surrounding environments to a pool of antibiotic-resistant bacteria facilitates rapid antibiotic resistance among the indigenous microbiota in the soil, water, and clinical environments via horizontal gene transfer. This provides favorable conditions for the development of more multidrug-resistant pathogens. Limitations in detecting gene transfer mechanisms have likely left us underestimating the role played by the surrounding environmental hotspots in the emergence of multidrug-resistant bacteria. This review aims to identify the major drivers responsible for the spread of antibiotic resistance and hotspots responsible for the acquisition of antibiotic resistance genes.
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http://dx.doi.org/10.1002/mbo3.1197DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8123917PMC
June 2021

Cocultivation of an ultrasmall environmental parasitic bacterium with lytic ability against bacteria associated with wastewater foams.

Nat Microbiol 2021 06 29;6(6):703-711. Epub 2021 Apr 29.

Department of Physiology, Anatomy, and Microbiology, La Trobe University, Melbourne, Victoria, Australia.

Many wastewater treatment plants around the world suffer from the operational problem of foaming. This is characterized by a persistent stable foam that forms on the aeration basin, which reduces effluent quality. The foam is often stabilized by a highly hydrophobic group of Actinobacteria known as the Mycolata. Gordonia amarae is one of the most frequently reported foaming members. With no currently reliable method for treating foams, phage biocontrol has been suggested as an attractive treatment strategy. Phages isolated from related foaming bacteria can destabilize foams at the laboratory scale; however, no phage has been isolated that lyses G. amarae. Here, we assemble the complete genomes of G. amarae and a previously undescribed species, Gordonia pseudoamarae, to examine mechanisms that encourage stable foam production. We show that both of these species are recalcitrant to phage infection via a number of antiviral mechanisms including restriction, CRISPR-Cas and bacteriophage exclusion. Instead, we isolate and cocultivate an environmental ultrasmall epiparasitic bacterium from the phylum Saccharibacteria that lyses G. amarae and G. pseudoamarae and several other Mycolata commonly associated with wastewater foams. The application of this parasitic bacterium, 'Candidatus Mycosynbacter amalyticus', may represent a promising strategy for the biocontrol of bacteria responsible for stabilizing wastewater foams.
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http://dx.doi.org/10.1038/s41564-021-00892-1DOI Listing
June 2021

Lytic Bacteriophage EFA1 Modulates HCT116 Colon Cancer Cell Growth and Upregulates ROS Production in an Co-culture System.

Front Microbiol 2021 31;12:650849. Epub 2021 Mar 31.

Department of Pharmacy and Biomedical Sciences, La Trobe Institute for Molecular Science, La Trobe University, Bendigo, VIC, Australia.

is an opportunistic pathogen in the gut microbiota that's associated with a range of difficult to treat nosocomial infections. It is also known to be associated with some colorectal cancers. Its resistance to a range of antibiotics and capacity to form biofilms increase its virulence. Unlike antibiotics, bacteriophages are capable of disrupting biofilms which are key in the pathogenesis of diseases such as UTIs and some cancers. In this study, bacteriophage EFA1, lytic against , was isolated and its genome fully sequenced and analyzed . Electron microscopy images revealed EFA1 to be a . The bacteriophage was functionally assessed and shown to disrupt biofilms as well as modulate the growth stimulatory effects of in a HCT116 colon cancer cell co-culture system, possibly via the effects of ROS. The potential exists for further testing of bacteriophage EFA1 in these systems as well as models.
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http://dx.doi.org/10.3389/fmicb.2021.650849DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8044584PMC
March 2021

Tn, a Carrier of Tn Family Transposons, Occurs in the Chromosome and in a Genomic Island of Clinical Strains.

Microorganisms 2020 Dec 15;8(12). Epub 2020 Dec 15.

Department of Physiology, Anatomy and Microbiology, La Trobe University, Bundoora, VIC 3086, Australia.

Transposons of the accessory gene pool contribute to phenotype and to genome plasticity. We studied local strains to ascertain the encroachment of -type site hunter transposons into clinical settings and their associations with other functional modules. Five different Tn family transposons were detected, all chromosomal. Some were solitary elements; one was in of Tn, a relative of a reported carrier of -type site hunters (class 1 integrons), but most were in of Tn, a new Tn-related transposon of unknown phenotype. Most of the Tn::Tn elements, and some Tn and Tn::Tn elements found in GenBank sequences, were at identical sites in an hypothetical gene of genomic island PAGI-5v. The island in clonally differing strains was at either of two tRNA loci, suggesting lateral transfer to these sites. This observation is consistent with the membership of the prototype PAGI-5 island to the ICE family of mobile genetic elements. Additionally, the site hunters in the nested transposons occupied different positions in the Tn carrier. This suggested independent insertion events on five occasions at least. Tn family members that were -/-defective were found in Tn- and Tn-like carriers in GenBank sequences of non-clinical spp. The transposition events in these cases presumably utilized functions in trans, as can occur with class 1 integrons. We suggest that in the clinical context, strains that carry Tn alone or in PAGI-5v can serve to disseminate functional site hunters; these in turn can provide the requisite -acting functions to assist in the dissemination of class 1 integrons, and hence of their associated antibiotic resistance determinants.
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http://dx.doi.org/10.3390/microorganisms8121997DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7765201PMC
December 2020

Characterization of Novel Lytic Bacteriophages of Isolated from a Pneumonia Patient.

Viruses 2020 10 8;12(10). Epub 2020 Oct 8.

Department of Pharmacy and Biomedical Science, La Trobe Institute for Molecular Science, La Trobe University, Bendigo, Victoria 3552, Australia.

spp. are becoming increasingly associated with lung infections in patients suffering from cystic fibrosis (CF). , which is closely related to , has been isolated from the lungs of CF patients and other human infections. This article describes the isolation, morphology and characterization of two lytic bacteriophages specific for an strain isolated from a pneumonia patient. This host strain was the causal agent of hospital acquired pneumonia-the first clinical report of such an occurrence. Full genome sequencing revealed bacteriophage genomes ranging in size from 45901 to 46,328 bp. Transmission electron microscopy revealed that the two bacteriophages AMA1 and AMA2 belonged to the family. Host range analysis showed that their host range did not extend to . The possibility exists for future testing of such bacteriophages in the control of infections such as those seen in CF and other infections of the lungs. The incidence of antibiotic resistance in this genus highlights the importance of seeking adjuncts and alternatives in CF and other lung infections.
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http://dx.doi.org/10.3390/v12101138DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7600146PMC
October 2020

The community compositions of three nitrogen removal wastewater treatment plants of different configurations in Victoria, Australia, over a 12-month operational period.

Appl Microbiol Biotechnol 2020 Nov 25;104(22):9839-9852. Epub 2020 Sep 25.

Department of Physiology, Anatomy and Microbiology, School of Life Sciences, College of Science, Health and Engineering, La Trobe University, Bundoora, Victoria, 3083, Australia.

Amplicon sequence fingerprinting of communities in activated sludge systems have provided data revealing the true level of their microbial biodiversity and led to suggestions of which intrinsic and extrinsic parameters might affect the dynamics of community assemblage. Most studies have been performed in China and Denmark, and comparatively little information is available for plants in other countries. This study looked at how the communities of three plants in Victoria, Australia, treating domestic sewage changed with season. All were designed to remove nitrogen microbiologically. They were all located close together to minimise any influence that climate and demographics might have on their operation, and samples were taken at weekly intervals for 12 months. 16S rRNA amplicon sequencing revealed that each plant community was distinctively different to the others and changed over the 12-month sampling period. Many of the factors suggested in other similar studies to be important in determining community composition in activated sludge systems could not explain the changes noted here. The most likely influential factors were considered to be temperature and influent composition reflecting changes in dietary intake by the populations served by each plant, since in all three, the most noticeable changes corresponded to seasonal shifts. KEY POINTS: • Monitoring microbial communities in 3 wastewater treatment plants removing nitrogen • Temperature is the most influential factor in dynamic changes in community composition.
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http://dx.doi.org/10.1007/s00253-020-10901-8DOI Listing
November 2020

Whole-genome analyses reveal gene content differences between nontypeable isolates from chronic obstructive pulmonary disease compared to other clinical phenotypes.

Microb Genom 2020 08 24;6(8). Epub 2020 Jul 24.

Department of Pharmacy and Biomedical Sciences, School of Molecular Sciences, College of Science, Health and Engineering, La Trobe University, Victoria, Australia.

Nontypeable (NTHi) colonizes human upper respiratory airways and plays a key role in the course and pathogenesis of acute exacerbations of chronic obstructive pulmonary disease (COPD). Currently, it is not possible to distinguish COPD isolates of NTHi from other clinical isolates of NTHi using conventional genotyping methods. Here, we analysed the core and accessory genome of 568 NTHi isolates, including 40 newly sequenced isolates, to look for genetic distinctions between NTHi isolates from COPD with respect to other illnesses, including otitis media, meningitis and pneumonia. Phylogenies based on polymorphic sites in the core-genome did not show discrimination between NTHi strains collected from different clinical phenotypes. However, pan-genome-wide association studies identified 79 unique NTHi accessory genes that were significantly associated with COPD. Furthermore, many of the COPD-related NTHi genes have known or predicted roles in virulence, transmembrane transport of metal ions and nutrients, cellular respiration and maintenance of redox homeostasis. This indicates that specific genes may be required by NTHi for its survival or virulence in the COPD lung. These results advance our understanding of the pathogenesis of NTHi infection in COPD lungs.
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http://dx.doi.org/10.1099/mgen.0.000405DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7641420PMC
August 2020

Draft Genome Sequence of an Isolate of Nontypeable Haemophilus influenzae from an Acute Exacerbation of Chronic Obstructive Pulmonary Disease in Tasmania.

Microbiol Resour Announc 2020 May 7;9(19). Epub 2020 May 7.

Tasmanian School of Medicine, College of Health and Medicine, University of Tasmania, Hobart, Tasmania, Australia

Nontypeable (NTHi) is an important cause of human illness, including pneumonia and acute exacerbations of chronic obstructive pulmonary disease (COPD). We report here the draft genome of an isolate of NTHi collected from the sputum of a patient presenting with COPD in Tasmania, Australia.
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http://dx.doi.org/10.1128/MRA.00375-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7206498PMC
May 2020

Exploring the operating factors controlling Kouleothrix (type 1851), the dominant filamentous bacterial population, in a full-scale A2O plant.

Sci Rep 2020 04 22;10(1):6809. Epub 2020 Apr 22.

Department of Physiology, Anatomy, and Microbiology, La Trobe University, Bundoora, VIC3086, Australia.

This study reveals that the abundance of the filament Kouleothrix (Eikelboom type 1851) correlated positively with poor settleability of activated sludge biomass in a Japanese full-scale nutrient removal wastewater treatment plant sampled over a one-year period. 16S rRNA amplicon sequence data confirmed that Kouleothrix was the dominant filament in the plant, with a relative abundance of 3.06% positively correlated with sludge volume index (SVI) (R = 0.691). Moreover, Kouleothrix (type 1851) appeared to form interfloc bridges, typical of bulking sludge, regardless of season. Together with earlier studies that indicated the responsibility of Kouleothrix (type 1851) on bulking events, these data suggest that their high relative abundances alone may be responsible for sludge bulking. 16S rRNA qPCR data for this filament showed changes in its relative abundance correlated with changes in several operational parameters, including mixed liquor temperature, sludge retention time, and suspended solids concentration, and it may be that manipulating these may help control Kouleothrix bulking.
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http://dx.doi.org/10.1038/s41598-020-63534-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7176654PMC
April 2020

Complete Genome Sequence of Moraxella osloensis Strain YV1, Isolated from an Australian Wastewater Treatment Plant.

Microbiol Resour Announc 2020 Apr 9;9(15). Epub 2020 Apr 9.

Department of Physiology, Anatomy, and Microbiology, La Trobe University, Bundoora, Australia

We report the complete genome sequence of strain YV1, which was isolated from a wastewater treatment plant in Australia. The YV1 genome comprises a 2,615,801-bp chromosome and four plasmids. strain YV1 displays the distinctive morphology of Eikelboom morphotype 1863.
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http://dx.doi.org/10.1128/MRA.00030-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7380525PMC
April 2020

Adaptive Evolution of Geobacter sulfurreducens in Coculture with Pseudomonas aeruginosa.

mBio 2020 04 7;11(2). Epub 2020 Apr 7.

Department of Physiology, Anatomy and Microbiology, La Trobe University, Melbourne, Victoria, Australia

Interactions between microorganisms in mixed communities are highly complex, being either syntrophic, neutral, predatory, or competitive. Evolutionary changes can occur in the interaction dynamics between community members as they adapt to coexistence. Here, we report that the syntrophic interaction between and coculture change in their dynamics over evolutionary time. Specifically, sp. dominance increases with adaptation within the cocultures, as determined through quantitative PCR and fluorescence hybridization. This suggests a transition from syntrophy to competition and demonstrates the rapid adaptive capacity of spp. to dominate in cocultures with Early in coculture establishment, two single-nucleotide variants in the and genes emerged that were strongly selected for throughout coculture evolution with phenazine wild-type and phenazine-deficient mutants. Sequential window acquisition of all theoretical spectra-mass spectrometry (SWATH-MS) proteomics revealed that the variant cooccurred with the upregulation of an adenylate cyclase transporter, CyaE, and a resistance-nodulation-division (RND) efflux pump notably known for antibiotic efflux. To determine whether antibiotic production was driving the increased expression of the multidrug efflux pump, we tested -derived phenazine-1-carboxylic acid (PHZ-1-CA) for its potential to inhibit growth and drive selection of the and genetic variants. Despite its inhibitory properties, PHZ-1-CA did not drive variant selection, indicating that other antibiotics may drive overexpression of the efflux pump and CyaE or that a novel role exists for these proteins in the context of this interaction. and spp. cohabit many of the same environments, where spp. often dominate. Both bacteria are capable of extracellular electron transfer (EET) and play important roles in biogeochemical cycling. Although they recently in 2017 were demonstrated to undergo direct interspecies electron transfer (DIET) with one another, the genetic evolution of this syntrophic interaction has not been examined. Here, we use whole-genome sequencing of the cocultures before and after adaptive evolution to determine whether genetic selection is occurring. We also probe their interaction on a temporal level and determine whether their interaction dynamics change over the course of adaptive evolution. This study brings to light the multifaceted nature of interactions between just two microorganisms within a controlled environment and will aid in improving metabolic models of microbial communities comprising these two bacteria.
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http://dx.doi.org/10.1128/mBio.02875-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7157779PMC
April 2020

Novel Bacteriophages Capable of Disrupting Biofilms From Clinical Strains of .

Front Microbiol 2020 14;11:194. Epub 2020 Feb 14.

Department of Pharmacy and Biomedical Science, La Trobe Institute for Molecular Science, La Trobe University, Melbourne, VIC, Australia.

The increase in global warming has favored growth of a range of opportunistic environmental bacteria and allowed some of these to become more pathogenic to humans. is one such organism. Surviving in moist conditions in temperate climates, these bacteria have been associated with a range of diseases in humans, and in systemic infections can cause mortality in up to 46% of cases. Their capacity to form biofilms, carry antibiotic resistance mechanisms, and survive disinfection, has meant that they are not easily treated with traditional methods. Bacteriophage offer a possible alternative approach for controlling their growth. This study is the first to report the isolation and characterization of bacteriophages lytic against clinical strains of which carry intrinsic antibiotic resistance genes. Functionally, these novel bacteriophages were shown to be capable of disrupting biofilms caused by clinical isolates of The potential exists for these to be tested in clinical and environmental settings.
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http://dx.doi.org/10.3389/fmicb.2020.00194DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7033617PMC
February 2020

Comparative Analysis of Structural Variations Due to Genome Shuffling of Bacillus Subtilis VS15 for Improved Cellulase Production.

Int J Mol Sci 2020 Feb 14;21(4). Epub 2020 Feb 14.

Department of Biotechnology, K L E F University, Guntur 522 502, India.

Cellulose is one of the most abundant and renewable biomass products used for the production of bioethanol. Cellulose can be efficiently hydrolyzed by Bacillus subtilis VS15, a strain isolate obtained from decomposing logs. A genome shuffling approach was implemented to improve the cellulase activity of Bacillus subtilis VS15. Mutant strains were created using ethyl methyl sulfonate (EMS), N-Methyl-N' nitro-N-nitrosoguanidine (NTG), and ultraviolet light (UV) followed by recursive protoplast fusion. After two rounds of shuffling, the mutants Gb2, Gc8, and Gd7 were produced that had an increase in cellulase activity of 128%, 148%, and 167%, respectively, in comparison to the wild type VS15. The genetic diversity of the shuffled strain Gd7 and wild type VS15 was compared at whole genome level. Genomic-level comparisons identified a set of eight genes, consisting of cellulase and regulatory genes, of interest for further analyses. Various genes were identified with insertions and deletions that may be involved in improved celluase production in Gd7.. Strain Gd7 maintained the capability of hydrolyzing wheatbran to glucose and converting glucose to ethanol by fermentation with Saccharomyces cerevisiae of the wild type VS17. This ability was further confirmed by the acidified potassium dichromate (KCrO) method.
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http://dx.doi.org/10.3390/ijms21041299DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7072954PMC
February 2020

Molecular characterisation of a novel pathogenic avipoxvirus from the Australian magpie (Gymnorhina tibicen).

Virology 2020 01 7;540:1-16. Epub 2019 Nov 7.

School of Animal and Veterinary Sciences, Charles Sturt University, Wagga Wagga, NSW, 2678, Australia.

Avipoxviruses are significant pathogens infecting a wide range of wild and domestic bird species globally. Here, we describe a novel genome sequence of magpiepox virus (MPPV) isolated from an Australian magpie. In the present study, histopathologically confirmed cutaneous pox lesions were used for transmission electron microscopic analysis, which demonstrated brick-shaped virions with regular spaced thread-like ridges, indicative of likely infectious particles. Subsequent analysis of the recovered MPPV genome positioned phylogenetically to a distinct sub-clade with the recently isolated avipoxvirus genome sequences from shearwater and canary bird species, and demonstrates a high degree of sequence similarity with CNPV (96.14%) and SWPV-2 (95.87%). The novel MPPV complete genome is missing 19 genes with a further 41 genes being truncated/fragmented compared to SWPV-2 and contains nine predicted unique genes. This is the first avipoxvirus complete genome sequence that infects Australian magpie.
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http://dx.doi.org/10.1016/j.virol.2019.11.005DOI Listing
January 2020

The Phylogeny, Biodiversity, and Ecology of the in Activated Sludge.

Front Microbiol 2019 13;10:2015. Epub 2019 Sep 13.

Department of Physiology, Anatomy and Microbiology, La Trobe University, Bundoora, VIC, Australia.

It is now clear that several of the filamentous bacteria in activated sludge wastewater treatment plants globally, are members of the phylum They appear to be more commonly found in treatment plants designed to remove nitrogen (N) and phosphorus (P), most of which operate at long sludge ages and expose the biomass to anaerobic conditions. The seem to play an important beneficial role in providing the filamentous scaffolding around which flocs are formed, to feed on the debris from lysed bacterial cells, to ferment carbohydrates and to degrade other complex polymeric organic compounds to low molecular weight substrates to support their growth and that of other bacterial populations. A few commonly extend beyond the floc surface, while others can align in bundles, which may facilitate interfloc bridging and hence generate a bulking sludge. Although several recent papers have examined the phylogeny and physiology of in activated sludge plants in Denmark, this review takes a wider look at what we now know about these filaments, especially their global distribution in activated sludge plants, and what their functional roles there might be. It also attempts to outline why such information might provide us with clues as to how their population levels may be manipulated, and the main research questions that need addressing to achieve these outcomes.
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http://dx.doi.org/10.3389/fmicb.2019.02015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6753630PMC
September 2019

Bacteriophages in Natural and Artificial Environments.

Pathogens 2019 Jul 12;8(3). Epub 2019 Jul 12.

Department of Physiology, Anatomy & Microbiology, La Trobe University, Bundoora, VIC 3086, Australia.

Bacteriophages (phages) are biological entities that have attracted a great deal of attention in recent years. They have been reported as the most abundant biological entities on the planet and their ability to impact the composition of bacterial communities is of great interest. In this review, we aim to explore where phages exist in natural and artificial environments and how they impact communities. The natural environment in this review will focus on the human body, soils, and the marine environment. In these naturally occurring environments there is an abundance of phages suggesting a role in the maintenance of bacterial community homeostasis. The artificial environment focuses on wastewater treatment plants, industrial processes, followed by pharmaceutical formulations. As in natural environments, the existence of bacteria in manmade wastewater treatment plants and industrial processes inevitably attracts phages. The presence of phages in these environments can inhibit the bacteria required for efficient water treatment or food production. Alternatively, they can have a positive impact by eliminating recalcitrant organisms. Finally, we conclude by describing how phages can be manipulated or formulated into pharmaceutical products in the laboratory for use in natural or artificial environments.
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http://dx.doi.org/10.3390/pathogens8030100DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6789717PMC
July 2019

Genomic, morphological and functional characterisation of novel bacteriophage FNU1 capable of disrupting Fusobacterium nucleatum biofilms.

Sci Rep 2019 06 24;9(1):9107. Epub 2019 Jun 24.

Department of Pharmacy and Biomedical Sciences, La Trobe Institute for Molecular Science, La Trobe University, Victoria, Australia.

Fusobacterium nucleatum is an important oral bacterium that has been linked to the development of chronic diseases such as periodontitis and colorectal cancer. In periodontal disease, F. nucleatum forms the backbone of the polymicrobial biofilm and in colorectal cancer is implicated in aetiology, metastasis and chemotherapy resistance. The control of this bacteria may be important in assisting treatment of these diseases. With increased rates of antibiotic resistance globally, there is need for development of alternatives such as bacteriophages, which may complement existing therapies. Here we describe the morphology, genomics and functional characteristics of FNU1, a novel bacteriophage lytic against F. nucleatum. Transmission electron microscopy revealed FNU1 to be a large Siphoviridae virus with capsid diameter of 88 nm and tail of approximately 310 nm in length. Its genome was 130914 bp, with six tRNAs, and 8% of its ORFs encoding putative defence genes. FNU1 was able to kill cells within and significantly reduce F. nucleatum biofilm mass. The identification and characterisation of this bacteriophage will enable new possibilities for the treatment and prevention of F. nucleatum associated diseases to be explored.
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http://dx.doi.org/10.1038/s41598-019-45549-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6591296PMC
June 2019

Isolation and characterization of bacteriophage NTR1 infectious for Nocardia transvalensis and other Nocardia species.

Virus Genes 2019 Apr 17;55(2):257-265. Epub 2018 Dec 17.

Department of Physiology, Anatomy & Microbiology, La Trobe University, Bundoora, VIC, Australia.

We describe here the isolation and characterization of the bacteriophage, NTR1 from activated sludge. This phage is lytic for Nocardia transvalensis, Nocardia brasiliensis and Nocardia farcinica. NTR1 phage has a genome sequence of 65,275 bp in length, and its closest match is to the Skermania piniformis phage SPI1 sharing over 36% of its genome. The phage belongs to the Siphoviridae family, possessing a long non-contractile tail and icosahedral head. Annotation of the genome reveals 97 putative open reading frames arranged in the characteristic modular organization of Siphoviridae phages and contains a single tRNA-Met gene.
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http://dx.doi.org/10.1007/s11262-018-1625-5DOI Listing
April 2019

Semi-Solid and Solid Dosage Forms for the Delivery of Phage Therapy to Epithelia.

Pharmaceuticals (Basel) 2018 Feb 26;11(1). Epub 2018 Feb 26.

Department of Pharmacy and Applied Science, La Trobe Institute for Molecular Science, La Trobe University Bendigo Campus, PO Box 199, Bendigo 3550, Australia.

The delivery of phages to epithelial surfaces for therapeutic outcomes is a realistic proposal, and indeed one which is being currently tested in clinical trials. This paper reviews some of the known research on formulation of phages into semi-solid dosage forms such as creams, ointments and pastes, as well as solid dosage forms such as troches (or lozenges and pastilles) and suppositories/pessaries, for delivery to the epithelia. The efficacy and stability of these phage formulations is discussed, with a focus on selection of optimal semi-solid bases for phage delivery. Issues such as the need for standardisation of techniques for formulation as well as for assessment of efficacy are highlighted. These are important when trying to compare results from a range of experiments and across different delivery bases.
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http://dx.doi.org/10.3390/ph11010026DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5874722PMC
February 2018

Characterization and formulation into solid dosage forms of a novel bacteriophage lytic against Klebsiella oxytoca.

PLoS One 2017 17;12(8):e0183510. Epub 2017 Aug 17.

La Trobe Institute for Molecular Science, La Trobe University, Bundoora, VIC, Australia.

Aim: To isolate and characterize bacteriophage lytic for the opportunistic pathogen Klebsiella oxytoca and their formulation into a range of solid dosage forms for in-vitro testing.

Methods And Results: We report the isolation, genomic and functional characterization of a novel bacteriophage lytic for Klebsiella oxytoca, which does not infect the closely related Klebsiella pneumoniae. This bacteriophage was formulated into suppositories and troches and shown to be released and lyse underlying Klebsiella oxytoca bacteria in an in-vitro model. These bacteriophage formulations were stable for at least 49 days at 4°C.

Conclusions: The successful in-vitro assay of these formulations here suggests that they could potentially be tested in-vivo to determine whether such a therapeutic approach could modulate the gut microbiome, and control Klebsiella oxytoca overgrowth, during antibiotic therapy regimes.

Significance And Impact Of The Study: This study reports a novel bacteriophage specific for Klebsiella oxytoca which can be formulated into solid dosage forms appropriate for potential delivery in testing as a therapy to modulate gut microbiome during antibiotic therapies.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0183510PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5560551PMC
October 2017

Bacteriophage formulated into a range of semisolid and solid dosage forms maintain lytic capacity against isolated cutaneous and opportunistic oral bacteria.

J Pharm Pharmacol 2017 Mar 29;69(3):244-253. Epub 2016 Dec 29.

La Trobe Institute for Molecular Science, La Trobe University, Bundoora, Vic., Australia.

Background: Resistance of bacteria to antimicrobial agents is of grave concern. Further research into the development of bacteriophage as therapeutic agents against bacterial infections may help alleviate this problem.

Objectives: To formulate bacteriophage into a range of semisolid and solid dosage forms and investigate the capacity of these preparations to kill bacteria under laboratory conditions.

Methods: Bacteriophage suspensions were incorporated into dosage forms such as creams, ointments, pastes, pessaries and troches. These were applied to bacterial lawns in order to ascertain lytic capacity. Stability of these formulations containing phage was tested under various storage conditions.

Key Findings: A range of creams and ointments were able to support phage lytic activity against Propionibacterium acnes. Assessment of the stability of these formulations showed that storage at 4 °C in light-protected containers resulted in optimal phage viability after 90 days. Pessaries/suppositories and troches were able to support phage lytic activity against Rhodococcus equi.

Conclusions: We report here the in-vitro testing of semisolid and solid formulations of bacteriophage lytic against a range of bacteria known to contribute to infections of the epithelia. This study provides a basis for the future formulation of diverse phage against a range of bacteria that infect epithelial tissues.
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http://dx.doi.org/10.1111/jphp.12673DOI Listing
March 2017

Dynamic interactions between prophages induce lysis in Propionibacterium acnes.

Res Microbiol 2017 Feb - Mar;168(2):103-112. Epub 2016 Sep 14.

Department of Physiology, Anatomy and Microbiology, La Trobe University, Bundoora, VIC 3086, Australia. Electronic address:

Progress in next-generation sequencing technologies has facilitated investigations into microbial dynamics. An important bacterium in the dairy industry is Propionibacterium freudenreichii, which is exploited to manufacture Swiss cheeses. A healthy culture of these bacteria ensures a consistent cheese with formed 'eyes' and pleasant flavour profile, and the investigation of prophages and their interactions with these bacteria could assist in the maintenance of the standard of this food product. Two bacteriophages, termed PFR1 and PFR2, were chemically induced using mitomycin C from two different dairy strains of P. freudenreichii. Both phages have identical genomes; however, PFR2 was found to contain an insertion sequence, IS204. Host range characterisation showed that PFR1 was able to form plaques on a wild type Propionibacterium acnes strain, whereas PFR2 could not. The lytic plaques observed on P. acnes were a result of PFR1 inducing the lytic cycle of a pseudolysogenic phage in P. acnes. Further investigation revealed that both PFR1 and PFR2 could infect P. acnes but not replicate. This study demonstrates the dynamic interactions between phages, which may alter their lytic capacity under certain conditions. To our knowledge, this is the first report of two phages interacting to kill their host.
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http://dx.doi.org/10.1016/j.resmic.2016.09.004DOI Listing
March 2017

Locating and Activating Molecular 'Time Bombs': Induction of Mycolata Prophages.

PLoS One 2016 3;11(8):e0159957. Epub 2016 Aug 3.

Department of Physiology, Anatomy and Microbiology, La Trobe University Bundoora, VIC, Australia.

Little is known about the prevalence, functionality and ecological roles of temperate phages for members of the mycolic acid producing bacteria, the Mycolata. While many lytic phages infective for these organisms have been isolated, and assessed for their suitability for use as biological control agents of activated sludge foaming, no studies have investigated how temperate phages might be induced for this purpose. Bioinformatic analysis using the PHAge Search Tool (PHAST) on Mycolata whole genome sequence data in GenBank for members of the genera Gordonia, Mycobacterium, Nocardia, Rhodococcus, and Tsukamurella revealed 83% contained putative prophage DNA sequences. Subsequent prophage inductions using mitomycin C were conducted on 17 Mycolata strains. This led to the isolation and genome characterization of three novel Caudovirales temperate phages, namely GAL1, GMA1, and TPA4, induced from Gordonia alkanivorans, Gordonia malaquae, and Tsukamurella paurometabola, respectively. All possessed highly distinctive dsDNA genome sequences.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0159957PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4972346PMC
August 2017

Genome Sequences of Pseudomonas oryzihabitans Phage POR1 and Pseudomonas aeruginosa Phage PAE1.

Genome Announc 2016 Jun 16;4(3). Epub 2016 Jun 16.

Department of Physiology, Anatomy and Microbiology, La Trobe University, Bundoora, Victoria, Australia

We report the genome sequences of two double-stranded DNA siphoviruses, POR1 infective for Pseudomonas oryzihabitans and PAE1 infective for Pseudomonas aeruginosa The phage POR1 genome showed no nucleotide sequence homology to any other DNA phage sequence in the GenBank database, while phage PAE1 displayed synteny to P. aeruginosa phages M6, MP1412, and YuA.
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http://dx.doi.org/10.1128/genomeA.01515-15DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4911491PMC
June 2016

The Formulation of Bacteriophage in a Semi Solid Preparation for Control of Propionibacterium acnes Growth.

PLoS One 2016 10;11(3):e0151184. Epub 2016 Mar 10.

La Trobe Institute for Molecular Sciences, La Trobe University, Bundoora, VIC, Australia.

Aims: To isolate and characterise phage which could lyse P. acnes and to formulate the phage into a delivery form for potential application in topical treatment of acne infection.

Methods And Results: Using standard phage isolation techniques, ten phage capable of lysing P. acnes were isolated from human skin microflora. Their genomes showed high homology to previously reported P. acnes phage. These phage were formulated into cetomacrogol cream aqueous at a concentration of 2.5x108 PFU per gram, and shown to lyse underlying P. acnes cells grown as lawn cultures. These phage formulations remained active for at least 90 days when stored at four degrees Celsius in a light protected container.

Conclusions: P. acnes phage formulated into cetomacrogol cream aqueous will lyse surrounding and underlying P. acnes bacteria, and are effective for at least 90 days if stored appropriately.

Significance And Impact Of The Study: There are few reports of phage formulation into semi solid preparations for application as phage therapy. The formulation method described here could potentially be applied topically to treat human acne infections. The potential exists for this model to be extended to other phage applied to treat other bacterial skin infections.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0151184PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4786141PMC
November 2016
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