Antonio Starcevic - Academia.edu (original) (raw)

Papers by Antonio Starcevic

Microbiology Spectrum

The genomes of Streptomyces species are difficult to assemble due to long repeats, extrachromosom... more The genomes of Streptomyces species are difficult to assemble due to long repeats, extrachromosomal elements (giant linear plasmids [GLPs]), rearrangements, and high GC content. To improve the quality of the S. rimosus ATCC 10970 genome, producer of oxytetracycline, we validated the assembly of GLPs by applying a new approach to combine pulsed-field gel electrophoresis separation and GLP isolation and sequenced the isolated GLP with Oxford Nanopore technology.

Clustering and phylogenetic analysis of Acropora digitifera toxins. HMM based hierarchical cluste... more Clustering and phylogenetic analysis of Acropora digitifera toxins. HMM based hierarchical clustering (HHCompare): HHCompare clustering was performed at HMM-HMM similarity e-value of 1.0e-20. Following the clustering, sequences within each group were aligned using MUSCLE. For three sequence groups, phylogentic trees were constructed using Minimal Evolution method, while larger groups were analyzed using Maximum Likelihood (ML) method. In case of ML, evolutionary model was inferred by MEGA 6.0 model selection tool, based on Sample-corrected Akaike information criterion (AICc). Figure S1. HMM-based hierarchical clustering of coral toxins. Each split indicates HMM-HMM similarity with e-value below 1.0e-20. A: Group 1, ML analysis using LG + G model with 4 discrete gamma categories. B: Group 6, Minimal Evolution analysis. C: Group 9, Minimal Evolution analysis. D: Group 10, Minimal Evolution analysis. E: Group 11, ML analysis using JTT model with 4 discrete gamma categories. Figure S2. ...

The genome sequences of a small number of individuals have already been determined. “ New generat... more The genome sequences of a small number of individuals have already been determined. “ New generation” DNA sequencing technology should lead to rapidly falling prices and the widespread determination of individual genome sequences. A major problem will be the provision of expert advice to interpret the results, both for the individual and their physician. An integrated set of computer programs (ClustScan) was developed initially to annotate modular biosynthetic clusters in bacteria that are important for the production of antibiotics and other pharmaceutically important products. The concept of ClustScan is to use a top-down approach incorporating knowledge in an ordered way (using XML) to produce results in a form useful for Biologists including a special graphical user interface. The versatility of this program package has been demonstrated by annotating biochemical pathways in microbial, invertebrate animal and metagenomic datasets. The open architecture of ClustScan allows easy i...

Personalized Medicine in Healthcare Systems, 2019

Microbes living in and on us have been recognized in the last decade as important part of ourselv... more Microbes living in and on us have been recognized in the last decade as important part of ourselves and have been assigned a status of an organ with a wide range of functions—from metabolizing food to training our immune system. The intraindividual and interindividual variation of normal microbiota is vast and its composition is influenced by various external factors—most notable being diet, antibiotics and medication. External factors can cause a disturbance in microbiota frequently termed as dysbiosis which is often associated with a range of diseases—from inflammatory bowel disease and diabetes to obesity and cancer. Pioneering works in manipulating the microbiota for preventing and treating disease shows huge promise but also open a number of questions—from standardization and safety to personalization and reproducibility.

Frontiers in Cellular and Infection Microbiology, 2021

A decade ago, when the Human Microbiome Project was starting, urinary tract (UT) was not included... more A decade ago, when the Human Microbiome Project was starting, urinary tract (UT) was not included because the bladder and urine were considered to be sterile. Today, we are presented with evidence that healthy UT possesses native microbiota and any major event disrupting its “equilibrium” can impact the host also. This dysbiosis often leads to cystitis symptoms, which is the most frequent lower UT complaint, especially among women. Cystitis is one of the most common causes of antimicrobial drugs prescriptions in primary and secondary care and an important contributor to the problem of antimicrobial resistance. Despite this fact, we still have trouble distinguishing whether the primary cause of majority of cystitis cases is a single pathogen overgrowth, or a systemic disorder affecting entire UT microbiota. There are relatively few studies monitoring changes and dynamics of UT microbiota in cystitis patients, making this field of research still an unknown. In this study variations to...

Clinical Proteomics, 2020

Background Reliable high-throughput microbial pathogen identification in human urine samples is c... more Background Reliable high-throughput microbial pathogen identification in human urine samples is crucial for patients with cystitis symptoms. Currently employed methods are time-consuming and could lead to unnecessary or inadequate antibiotic treatment. Purpose of this study was to assess the potential of mass spectrometry for uropathogen identification from a native urine sample. Methods In total, 16 urine samples having more than 105 CFU/mL were collected from clinical outpatients. These samples were analysed using standard urine culture methods, followed by 16S rRNA gene sequencing serving as control and here described culture-independent MALDI-TOF/TOF MS method being tested. Results Here we present advantages and disadvantages of bottom-up proteomics, using MALDI-TOF/TOF tandem mass spectrometry, for culture-independent identification of uropathogens (e.g. directly from urine samples). The direct approach provided reliable identification of bacteria at the genus level in monobact...

Current Opinion in Food Science, 2020

Probiotics are modern-day panacea with claims to alleviate or cure everything from diarrhea to ob... more Probiotics are modern-day panacea with claims to alleviate or cure everything from diarrhea to obesity and Alzheimer’s disease. They seem effective for numerous conditions (e.g. antibiotic-associated diarrhea), although these views have been challenged recently. Greater issue is the lack of confirmed efficacy for majority of claims provided by producers. While offering products without adequate clinical research, industry has more simplistic approach for probiotic treatments, that is, ‘one-formula-fits-all.’ In contrast, recent studies showed different efficacies of probiotic mixtures with multiple strains without fully understanding factors behind it. Furthermore, important parameter for prescribing probiotic treatment is the individual native microbiota. Interactions among probiotic strains and microbiota need more research, so there is a long way paved with ‘big data’ and chemometric analysis before it is possible to make full use of probiotics.

Food technology and biotechnology, 2017

The MEGGASENSE platform constructs relational databases of DNA or protein sequences. The default ... more The MEGGASENSE platform constructs relational databases of DNA or protein sequences. The default functional analysis uses 14 106 hidden Markov model (HMM) profiles based on sequences in the KEGG database. The Solr search engine allows sophisticated queries and a BLAST search function is also incorporated. These standard capabilities were used to generate the SCATT database from the predicted proteome of Streptomyces cattleya. The implementation of a specialised metagenome database (AMYLOMICS) for bioprospecting of carbohydrate-modifying enzymes is described. In addition to standard assembly of reads, a novel 'functional' assembly was developed, in which screening of reads with the HMM profiles occurs before the assembly. The AMYLOMICS database incorporates additional HMM profiles for carbohydrate-modifying enzymes and it is illustrated how the combination of HMM and BLAST analyses helps identify interesting genes. A variety of different proteome and metagenome databases have...

Food Technology and Biotechnology, Mar 29, 2013

The recent achievement of synthesising a functioning bacterial chromosome marks a coming of age f... more The recent achievement of synthesising a functioning bacterial chromosome marks a coming of age for engineering living organisms. In the future this should allow the construction of novel organisms to help solve the problems facing the human race, including health care, food, energy and environmental protection. In this minireview, the current state of the field is described and the role of synthetic biology in biotechnology in the short and medium term is discussed. It is particularly aimed at the needs of food technologists, nutritionists and other biotechnologists, who might not be aware of the potential significance of synthetic biology to the research and development in their fields. The potential of synthetic biology to produce interesting new polyketide compounds is discussed in detail.

BMC Genomics, 2015

Background: Gene duplication followed by adaptive selection is a well-accepted process leading to... more Background: Gene duplication followed by adaptive selection is a well-accepted process leading to toxin diversification in venoms. However, emergent genomic, transcriptomic and proteomic evidence now challenges this role to be at best equivocal to other processess. Cnidaria are arguably the most ancient phylum of the extant metazoa that are venomous and such provide a definitive ancestral anchor to examine the evolution of this trait. Methods: Here we compare predicted toxins from the translated genome of the coral Acropora digitifera to putative toxins revealed by proteomic analysis of soluble proteins discharged from nematocysts, to determine the extent to which gene duplications contribute to venom innovation in this reef-building coral species. A new bioinformatics tool called HHCompare was developed to detect potential gene duplications in the genomic data, which is made freely available (https://github.com/rgacesa/HHCompare). Results: A total of 55 potential toxin encoding genes could be predicted from the A. digitifera genome, of which 36 (65 %) had likely arisen by gene duplication as evinced using the HHCompare tool and verified using two standard phylogeny methods. Surprisingly, only 22 % (12/55) of the potential toxin repertoire could be detected following rigorous proteomic analysis, for which only half (6/12) of the toxin proteome could be accounted for as peptides encoded by the gene duplicates. Biological activities of these toxins are dominatedby putative phospholipases and toxic peptidases. Conclusions: Gene expansions in A. digitifera venom are the most extensive yet described in any venomous animal, and gene duplication plays a significant role leading to toxin diversification in this coral species. Since such low numbers of toxins were detected in the proteome, it is unlikely that the venom is evolving rapidly by preydriven positive natural selection. Rather we contend that the venom has a defensive role deterring predation or harm from interspecific competition and overgrowth by fouling organisms. Factors influencing translation of toxin encoding genes perhaps warrants more profound experimental consideration.

Microbiome R&D and Business Collaboration Forum

Peptide mass fingerprinting is a term which describes technique which utilizes ESI or MALDI MS fo... more Peptide mass fingerprinting is a term which describes technique which utilizes ESI or MALDI MS followed by tandem mass spectrometry sequencing. This technique has become a cornerstone for protein identification. Today, applications using peptide mass fingerprinting in biomedical analyses are a major driving force behind its rapid development. However, efficient and accurate analyses of generally big protein tandem mass spectrometry data sets require robust software. In terms of final goal, which is data interpretation, the role of software and underlying algortihms is at least equally important as the technique itself, a fact which is often neglected. High-throughput mass spectrometry instruments can readily generate hundreds of thousands of spectra. This fact combined with the ever growing size of genomic databases imposes tremendous demands for potential successful softvare solutions. In fact, it is the process of comparing large-scale mass spectrometry data with large databases t...

Systematic and Applied Microbiology, 2015

Samples were collected from sea sediments at seven sites in the northern Adriatic Sea that includ... more Samples were collected from sea sediments at seven sites in the northern Adriatic Sea that included six sites next to industrial complexes and one from a tourist site (recreational beach). The samples were assayed for alkanes and polycyclic aromatic hydrocarbons. The composition of the hydrocarbon samples suggested that industrial pollution was present in most cases. A sample from one site was also grown aerobically under crude oil enrichment in order to evaluate the response of indigenous bacterial populations to crude oil exposure. Analysis of 16S rRNA gene sequences showed varying microbial biodiversity depending on the level of pollution - ranging from low (200 detected genera) to high (1000+ genera) biodiversity, with lowest biodiversity observed in polluted samples. This indicated that there was considerable biodiversity in all sediment samples but it was severely restricted after exposure to crude oil selection pressure. Phylogenetic analysis of putative alkB genes showed high evolutionary diversity of the enzymes in the samples and suggested great potential for bioremediation and bioprospecting. The first systematic analysis of bacterial communities from sediments of the northern Adriatic Sea is presented, and it will provide a baseline assessment that may serve as a reference point for ecosystem changes and hydrocarbon degrading potential - a potential that could soon gain importance due to plans for oil exploitation in the area.

Genome announcements, Jan 8, 2013

Streptomyces rapamycinicus strain NRRL 5491 produces the important drug rapamycin. It has a large... more Streptomyces rapamycinicus strain NRRL 5491 produces the important drug rapamycin. It has a large genome of 12.7 Mb, of which over 3 Mb consists of 48 secondary metabolite biosynthesis clusters.

Genome announcements, Jan 17, 2014

The genome sequence of Streptomyces rimosus R6-500, an industrially improved strain which produce... more The genome sequence of Streptomyces rimosus R6-500, an industrially improved strain which produces high titers of the important antibiotic oxytetracycline, is reported, as well as the genome sequences of two derivatives arising due to the genetic instability of the strain.

Toxicon : official journal of the International Society on Toxinology, 2015

An important mechanism for the evolution of toxins in venomous animals is believed to be the acqu... more An important mechanism for the evolution of toxins in venomous animals is believed to be the acquisition of genes encoding proteins that switch from physiological to toxic roles following gene duplication. The 'reverse recruitment' hypothesis pertains that these genes can also revert back to physiological functions, although such events are thought to be rare. A non-supervised homology searching method was developed which allowed the peptide diversity of animal toxins to be described as combinations between limited numbers of amino-acid sequence blocks we called 'tox-bits'. Taking the phospholipase A2 (PLA2) protein family as an example, a Bernoulli Trial was used to test if 'tox-bits' were robust enough to distinguish between peptides with physiological or toxin functions. The analysis revealed that discrimination was indeed possible, and supports the very recent 'restriction' hypothesis whereby genes with the potential to encode toxic functions have...

Journal of Biotechnology, 2014

Molecular & cellular proteomics : MCP, Jan 5, 2015

Shipboard experiments were performed over a two day period to examine the proteomic response of t... more Shipboard experiments were performed over a two day period to examine the proteomic response of the symbiotic coral Acropora microphthalma exposed to acute conditions of high temperature / low light or high light / low temperature stress. During these treatments, corals had noticeably bleached. The photosynthetic performance of residual algal endosymbionts was severely impaired but showed signs of recovery in both treatments by the end of the second day. Changes in the coral proteome were determined daily and, using recently available annotated genome sequences, the individual contributions of the coral host and algal endosymbionts could be extracted from these data. Quantitative changes in proteins relevant to redox state and calcium metabolism are presented. Notably, expression of common antioxidant proteins was not detected from the coral host but present in the algal endosymbiont proteome. Possible roles for elevated carbonic anhydrase in the coral host are considered: to restor...

PLoS ONE, 2010

Background: The success of tropical reef-building corals depends on the metabolic cooperation bet... more Background: The success of tropical reef-building corals depends on the metabolic cooperation between the animal host and the photosynthetic performance of endosymbiotic algae residing within its cells. To examine the molecular response of the coral Acropora microphthalma to high levels of solar irradiance, a cDNA library was constructed by PCR-based suppression subtractive hybridisation (PCR-SSH) from mRNA obtained by transplantation of a colony from a depth of 12.7 m to near-surface solar irradiance, during which the coral became noticeably paler from loss of endosymbionts in sun-exposed tissues. Methodology/Principal Findings: A novel approach to sequence annotation of the cDNA library gave genetic evidence for a hypothetical biosynthetic pathway branching from the shikimic acid pathway that leads to the formation of 4deoxygadusol. This metabolite is a potent antioxidant and expected precursor of the UV-protective mycosporine-like amino acids (MAAs), which serve as sunscreens in coral phototrophic symbiosis. Empirical PCR based evidence further upholds the contention that the biosynthesis of these MAA sunscreens is a 'shared metabolic adaptation' between the symbiotic partners. Additionally, gene expression induced by enhanced solar irradiance reveals a cellular mechanism of light-induced coral bleaching that invokes a Ca 2+-binding synaptotagmin-like regulator of SNARE protein assembly of phagosomal exocytosis, whereby algal partners are lost from the symbiosis. Conclusions/Significance: Bioinformatics analyses of DNA sequences obtained by differential gene expression of a coral exposed to high solar irradiance has revealed the identification of putative genes encoding key steps of the MAA biosynthetic pathway. Revealed also by this treatment are genes that implicate exocytosis as a cellular process contributing to a breakdown in the metabolically essential partnership between the coral host and endosymbiotic algae, which manifests as coral bleaching.

Organic & Biomolecular Chemistry, 2006

Post-translationally modified ribosomal peptides are unusual natural products and many have poten... more Post-translationally modified ribosomal peptides are unusual natural products and many have potent biological activity. The biosynthetic processes involved in their formation have been delineated for some, but the patellamides represent a unique group of these metabolites with a combination of a macrocycle, small heterocycles and D-stereocentres. The genes encoding for the patellamides show very low homology to known biosynthetic genes and there appear to be no explicit genes for the macrocyclisation and epimerisation steps. Using a combination of literature data and large-scale molecular dynamics calculations with explicit solvent, we propose that the macrocyclisation and epimerisation steps are spontaneous and interdependent and a feature of the structure of the linear peptide. Our study suggests the steps in the biosynthetic route are heterocyclisation, macrocyclisation, followed by epimerisation and finally dehydrogenation. This study is presented as testable hypothesis based on literature and theoretical data to be verified by future detailed experimental investigations.