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PubMed2006

Early days: genomics and human responses to infection.

DNA microarray-based gene transcript-profiling of the responses of primates to infection has begun to yield new insights into host-pathogen interactions; this approach, however, remains plagued by challenges and complexities that have yet to be adequately addressed. The rapidly changing nature over time of acute infectious diseases in a host, and the genetic diversity of microbial pathogens present unique problems for the design and interpretation of functional-genomic studies in this field. In addition, there are the more common problems related to heterogeneity within clinical samples, the complex, non-standardized confounding variables associated with human subjects and the complexities posed by the analysis and validation of highly parallel data. Whereas various approaches have been developed to address each of these issues, there are significant limitations that remain to be overcome. The resolution of these problems should lead to a better understanding of the dialogue between the host and pathogen.

Animals↗
PubMed2006

Microarray applications in microbial ecology research.

Microarray technology has the unparalleled potential to simultaneously determine the dynamics and/or activities of most, if not all, of the microbial populations in complex environments such as soils and sediments. Researchers have developed several types of arrays that characterize the microbial populations in these samples based on their phylogenetic relatedness or functional genomic content. Several recent studies have used these microarrays to investigate ecological issues; however, most have only analyzed a limited number of samples with relatively few experiments utilizing the full high-throughput potential of microarray analysis. This is due in part to the unique analytical challenges that these samples present with regard to sensitivity, specificity, quantitation, and data analysis. This review discusses specific applications of microarrays to microbial ecology research along with some of the latest studies addressing the difficulties encountered during analysis of complex microbial communities within environmental samples. With continued development, microarray technology may ultimately achieve its potential for comprehensive, high-throughput characterization of microbial populations in near real time.

Bacteria↗
PubMed2026

Genetic diversity and drug resistance profiles of Mycobacterium tuberculosis among Ethiopian children as determined by whole-genome sequencing.

UNLABELLED: Ethiopia ranks 30th among the tuberculosis (TB) burden countries, with children representing a significant yet understudied population group. This study aims to investigate the genetic diversity and drug-resistant profile among Ethiopian children. We included children under 15 years of age diagnosed with culture-confirmed pulmonary TB/drug-resistant TB between January 2017 and June 2023. Phenotypic drug susceptibility testing and whole-genome sequencing were conducted for 85 Mycobacterium tuberculosis (MTB) isolates. Demographic data were combined with genomic information. Lineage 4 was the most dominant (77.6%), while lineage 2 was less common (1%). Within lineage 4, several sub-lineages were identified, with lineage 4.2.2.2 being notably the most predominant (48%). Most of these cases were from Oromia (58%), including the hotspot areas for lineage 4 that were identified at a 99% confidence level. Among 17 MDR/pre-XDR-TB isolates, lineages 3 and 4.2.2.2 were the dominantly observed lineages/sub-lineages, with proportions of 29% and 65%, respectively. Of the 85 cases, 30.5% were drug-resistant TB to at least one of the five first-line anti-TB drugs tested by phenotypic drug susceptibility testing. Of these 26 drug-resistant TB cases, 23 were concordant with whole-genome sequencing characterization. The most frequent resistance mutations to rifampicin were found in the rpoB gene, specifically p.Ser450Leu (88%), followed by isoniazid in the katG gene, p.Ser315Thr (86%). Multidrug-resistant TB was strongly associated with MTB lineages (P = 0.007). This study identified high genetic diversity of M. tuberculosis and related drug-resistance mutations, with a strong concordance between whole-genome sequencing-based predictions and phenotypic drug susceptibility testing. IMPORTANCE: Our findings revealed a high genetic diversity of Mycobacterium tuberculosis among Ethiopian children, with the most common lineage being lineage 4, specifically lineage 4.2.2.2, in which a higher frequency of multidrug-resistant tuberculosis (TB) was observed. Additionally, we identified regional hotspots, suggesting ongoing community transmission. Moreover, whole-genome sequencing demonstrated high concordance with phenotypic drug susceptibility testing and identified mutation genes associated with first- and second-line anti-TB drugs, highlighting its usefulness in providing comprehensive results for resistance detection in children. Thus, it is essential for integrating genomic surveillance into childhood TB and drug resistance control.

Humans↗
PubMed1997

[Molecular epidemiological analysis of Pseudomonas aeruginosa by pulsed-field gel electrophoresis].

In order to see the nosocomial infection by Pseudomonas aeruginosa (P. aeruginosa) in the Akita University Hospital, we analyzed the serotype and genotype for 150 strains isolated from various clinical specimens from 1994 to 1996. One hundred strains chosen at random were divided into 11 serotypes by serotyping and into 50 genotypes by pulsed-field gel electrophoresis of Spe I-digested P. aeruginosa genomic DNAs. Serotype E strains, most common, gave 17 genome patterns. Fifty strains separated periodically in certain wards had further 7 genome patterns. The strains isolated from one patient with different times or with different serotypes showed the same stable genotype. Genome patterns were inconsistent with susceptibility to antimicrobial agents. Both drug-susceptible and -resistant strains were found in strains with the same genome pattern. The genome pattern was identical, even though the susceptibility was different due to isolation time or storage. This suggested that the multidrug-resistant strains of P. aeruginosa expanded in the hospital were not derived from one original strain.

Cross Infection↗
PubMed1998

Analyzing genomes with cumulative skew diagrams.

A novel method of cumulative diagrams shows that the nucleotide composition of a microbial chromosome changes at two points separated by about a half of its length. These points coincide with sites of replication origin and terminus for all bacteria where such sites are known. The leading strand is found to contain more guanine than cytosine residues. This fact is used to predict origin and terminus locations in other bacterial and archaeal genomes. Local changes, visible as diagram distortions, may represent recent genome rearrangements, as demonstrated for two strains of Escherichia coli . Analysis of the diagrams of viral and mitochondrial genomes suggests a link between the base composition bias and the time spent by DNA in a single stranded state during replication.

Bacteria↗
PubMed2001

Catalyzing bacterial speciation: correlating lateral transfer with genetic headroom.

Unlike crown eukaryotic species, microbial species are created by continual processes of gene loss and acquisition promoted by horizontal genetic transfer. The amounts of foreign DNA in bacterial genomes, and the rate at which this is acquired, are consistent with gene transfer as the primary catalyst for microbial differentiation. However, the rate of successful gene transfer varies among bacterial lineages. The heterogeneity in foreign DNA content is directly correlated with amount of genetic headroom intrinsic to a bacterial species. Genetic headroom reflects the amount of potentially dispensable information--reflected in codon usage bias and codon context bias--that can be transiently sacrificed to allow experimentation with functions introduced by gene transfer. In this way, genetic headroom offers a potential metric for assessing the propensity of a lineage to speciate.

Bacteria↗
PubMed2026

Chemo-selective proteomics in microbial systems.

SUMMARYOver the past two decades, the field of bioorthogonal chemistry has transitioned from emerging to an established cornerstone of scientific inquiry. In parallel, advances in microbial and host-microbe research have highlighted the need for functional approaches that extend beyond genomic and transcriptomic analyses to directly interrogate protein-level activity. Despite this need, proteomic strategies capable of resolving dynamic, heterogeneous, and low-abundance protein populations remain underdeveloped in microbial systems. This review highlights the convergence of chemo-selective proteomic technologies with microbial biology, focusing on bioorthogonal non-canonical amino acid tagging (BONCAT), activity- or affinity-based protein profiling, and bioorthogonal post-translational modifications, and comments on possibilities for novel applications for the use of click chemistry-based tools in the functional interrogation of microbial systems. Together, these strategies enable spatiotemporal resolution of protein synthesis, selective profiling of microbial subpopulations, and direct characterization of protein activity and regulation in complex biological contexts, including single-species cultures, host-associated environments, and polymicrobial communities. Continued development and utilization of these technologies will enable deeper mechanistic insight into how microbial systems function and respond to environmental and host-derived cues.

bioorthogonal chemistry↗
PubMed2025

Genomic insights into Shigella species isolated from small ruminants and manure in the North West Province, South Africa.

This study investigated Shigella species' antibiotic resistance patterns and genomic characteristics from small ruminants and manure collected in Potchefstroom, North West, South Africa. Whole genome sequencing was used to determine resistome profiles of Shigella flexneri isolates from small ruminants' manure and Shigella boydii from sheep faeces. Comparative genomics was employed on the South African 261 S. flexneri strains available from GenBank, including the sequenced strains in this study, by investigating the serovars, antibiotic resistance genes (ARGs), and plasmid replicon types. The S. flexneri strains could not be assigned to known sequence types, suggesting novel or uncharacterized lineages. S. boydii R7-1A was assigned to sequence type 202 (ST202). Serovar 2A was the most common among South African S. flexneri strains, found in 96% of the 250 compared human-derived isolates. The shared mdf(A) was the most prevalent gene, identified in 99% of 261 S. flexneri genomes, including plasmid replicon types ColRNAI_1 (99%) and IncFII_1 (98%). Both species share a core set of resistance determinants mainly involving β-lactams (ampC1, ampC, ampH), macrolides (mphB), polymyxins (eptA, pmrF), multidrug efflux pumps (AcrAB-TolC, Mdt, Emr, Kpn families), and regulatory systems (marA, hns, crp, baeRS, evgAS, cpxA, gadX). However, S. boydii possesses additional resistance genes conferring resistance to tetracyclines (tet(A)), phenicols (floR), sulphonamides (sul2), and aminoglycosides (APH(3'')-Ib, APH(6)-Id), along with the acrEF efflux pump components (acrE, acrF). In contrast, S. flexneri harboured unique genes linked to polymyxin resistance (ugd) and regulatory functions (sdiA, gadW) that were absent in S. boydii. These findings highlight Shigella strains' genomic diversity and antimicrobial resistance potential in livestock-associated environments. Moreover, S. boydii highlights the potential risk of multidrug-resistant bacteria in farming and environmental routes. KEY POINTS: • First whole genome study of Shigella from manure and small ruminants in South Africa. • Shigella boydii strain carried multiple resistance genes to β-lactams and tetracycline. • Multidrug efflux pump gene mdf(A) was detected in 99% of South African Shigella flexneri strains.

Animals↗
PubMed2005

The metagenomics of soil.

Phylogenetic surveys of soil ecosystems have shown that the number of prokaryotic species found in a single sample exceeds that of known cultured prokaryotes. Soil metagenomics, which comprises isolation of soil DNA and the production and screening of clone libraries, can provide a cultivation-independent assessment of the largely untapped genetic reservoir of soil microbial communities. This approach has already led to the identification of novel biomolecules. However, owing to the complexity and heterogeneity of the biotic and abiotic components of soil ecosystems, the construction and screening of soil-based libraries is difficult and challenging. This review describes how to construct complex libraries from soil samples, and how to use these libraries to unravel functions of soil microbial communities.

DNA↗
PubMed2003

The biodiversity of microbial cytochromes P450.

The cytochrome P450 (CYP) superfamily of genes and proteins are well known for their involvement in pharmacology and toxicology, but also increasingly for their importance and diversity in microbes. The extent of diversity has only recently become apparent with the emergence of data from whole genome sequencing projects and the coming years will reveal even more information on the diversity in microbial eukaryotes. This review seeks to describe the historical development of these studies and to highlight the importance of the genes and proteins. CYPs are deeply involved in the development of strategies for deterrence and attraction as well as detoxification. As such, there is intense interest in pathways of secondary metabolism that include CYPs in oxidative tailoring of antibiotics, sometimes influencing potency as bioactive compounds. Further to this is interest in CYPs in metabolism of xenobiotics for use as carbon sources for microbial growth and as biotransformation agents or in bioremediation. CYPs are also current and potential drug targets; compounds inhibiting CYP are antifungal and anti-protozoan agents, and potentially similar compounds may be useful against some bacterial diseases such as tuberculosis. Of note is the diversity of CYP requirements within an organism, ranging from Escherichia coli that has no CYPs as in many bacteria, to Mycobacterium smegmatis that has 40 representing 1% of coding genes. The basidiomycete fungus Phanerochaete chrysosporium surprised all when it was found to contain a hundred or more CYPs. The functional genomic investigation of these orphan CYPs is a major challenge for the future.

Bacteria↗
PubMed2026

Sex-Dependent Microbial and Host Profiles Following Fecal Microbiota and Bifidobacterium longum Treatment in Stress-Induced Gut Dysbiosis.

BACKGROUND/AIMS: Irritable bowel syndrome (IBS) is a chronic functional gastrointestinal disorder influenced by stress, microbial dysbiosis, and immune activation. Microbiota-directed therapies, including fecal microbiota transplantation and probiotics, show promise, but their sex-specific effects remain unclear. We compared the therapeutic effects of lyophilized fecal microbiota (LFM) with Bifidobacterium longum BBH016 in male and female Wistar rats subjected to repeated water avoidance stress. METHODS: Fecal pellet output (FPO), colonic mast cell infiltration, and fecal short-chain fatty acids were measured. Gut microbial composition and function were analyzed by 16S rRNA sequencing and Kyoto Encyclopedia of Genes and Genomes pathway prediction. RESULTS: Both interventions significantly reduced FPO and mast cell infiltration in males but had less pronounced effects in females. Microbiota analyses revealed sex-dependent responses, with distinct microbial trajectories in each treatment group. Using linear discriminant analysis effect size, we identified seven key taxa with treatment- or sex-specific enrichment. Alistipes onderdonkii and Bacteroides uniformis consistently increased in both LFM- and B. longum-treated groups, regardless of sex. Bacteroides finegoldii and Barnesiella intestinihominis were specifically enriched in the LFM group. In males, Blautia faecis and Fusicatenibacter saccharivorans were enriched following the interventions, whereas Parabacteroides goldsteinii appeared exclusively in stressed males. Functional predictions revealed the enrichment of estrogen signaling and bile acid pathways in males and the attenuation of proinflammatory pathways in females following LFM. Correlations between microbial taxa and host outcomes were predominantly observed in male rats. CONCLUSIONS: These findings highlight sex-specific microbial and host responses to microbiota-targeted therapies in a stress-induced IBS model, emphasizing sex as a biological variable in designing personalized microbiome-based treatments.

Animals↗
PubMed2026

Oral cavity as a reservoir of respiratory pathogens: microbial dynamics in patients receiving mechanical ventilation.

OBJECTIVE: To investigate the prevalence, co-occurrence, and temporal dynamics of Staphylococcus aureus, Klebsiella pneumoniae, Streptococcus pneumoniae, and Pseudomonas aeruginosa in oral and tracheal samples from critically ill patients receiving mechanical ventilation. METHODS: This observational analytical study included 21 adult patients receiving mechanical ventilation who were admitted to an intensive care unit. Oral and tracheal samples were collected within the first 24 h after intubation (Day 0) and again after 48 h (Day 2). Bacterial genomic deoxyribonucleic acid was extracted and analyzed using quantitative realtime polymerase chain reaction. Microbial detection was evaluated qualitatively based on the presence or absence of each microorganism and quantitatively using relative bacterial load analysis. Statistical analyses assessed correlations between oral and tracheal colonization patterns and temporal changes in bacterial load. RESULTS: More than 43% of oral and tracheal samples tested positive for at least one investigated pathogen at both collection time points, indicating persistent microbial colonization during mechanical ventilation. Significant positive correlations for Staphylococcus aureus detection were observed between oral and tracheal samples on Day 0 (p=0.0028) and Day 2 (p=0.0021), suggesting a possible association between oral colonization and lower airway microbial presence. Streptococcus pneumoniae detected in initial oral samples showed a significant correlation with tracheal detection after 48 h (p=0.03). Quantitative analyses demonstrated no significant changes in bacterial load over time for most microorganisms. However, Klebsiella pneumoniae showed a significant increase in tracheal bacterial load between Day 0 and Day 2 (p=0.04), with higher tracheal loads than oral samples on Day 2 (p=0.01). CONCLUSION: The oral cavity may serve as an important reservoir of respiratory pathogens in patients receiving mechanical ventilation. The observed correlations between oral and tracheal colonization and pathogen-specific colonization patterns reinforce the relevance of oral health management in critical care settings. In particular, the increase in tracheal Klebsiella pneumoniae load suggests that lower airway proliferation may occur during mechanical ventilation independently of simultaneous increases in oral bacterial load.

Humans↗
PubMed2025

Molecular insights into the persistence and co-occurrence of two different carbapenem-resistant Pseudomonas aeruginosa lineages within a hospital setting.

UNLABELLED: Carbapenem-resistant Pseudomonas aeruginosa (CRPA) represents a critical-priority pathogen capable of causing life-threatening, multidrug-resistant infections. We performed susceptibility testing, whole-genome sequencing, and bioinformatic analyses on 137 CRPA isolates from a Guangdong hospital. We found that the major specimen types were respiratory specimens (57/137, 41.6%) and bronchoalveolar lavage (42/137, 30.7%). All isolates were carbapenem-resistant but had low resistance to polymyxin B (0.7%, 1/137). IncP-6-positive isolates exhibited &#x2265;2- to 32-fold higher resistance to 9/12 antibiotics (P < 0.05), with no difference to imipenem and meropenem. Fifty-four sequence types and 11 O-serogroups were identified, with ST1971 (6.6%) and O11 (29.9%) being predominant. Temporal and spatial patterns suggest persistent co-occurrence of clade 1 and clade 2 isolates, indicating potential nosocomial outbreak and clonal transmission. IMPORTANCE: The prevalence of carbapenem-resistant Pseudomonas aeruginosa (CRPA) has increased rapidly in recent years, yet few genetic and epidemiological studies on CRPA isolates have been performed. We performed susceptibility testing, whole-genome sequencing, and bioinformatic analyses on hospital isolates to investigate their resistance profiles and molecular epidemiology. These findings may offer new insights for developing effective global strategies to control CRPA and reduce untreatable infections in clinical settings.

Pseudomonas aeruginosa↗
PubMed1997

High frequency transposition of the Tn5 derivative Tn5493 in Streptomyces lividans.

The mini-Tn5 derivative transposon Tn5493 transposes at a frequency as high as 3% in the Gram-positive bacterium Streptomyces lividans. The use of a thiostrepton-resistance gene and a temperature-sensitive delivery system carrying the transposase gene allows an easy selection for stable transposition events. Insertions into the S. lividans genome seem to be fairly random, as shown by Southern blot and pulsed-field gel electrophoresis (PFGE). Transpositions were easily mapped by PFGE using the restriction sites of rare-cutting enzymes present in the transposon. Because of its characteristics, Tn5493 is a promising tool for genetic and molecular analysis and manipulation of the antibiotic-producing streptomycetes.

Base Sequence↗
PubMed2006

Hybridization of genomic DNA to microarrays: a challenge for the analysis of environmental samples.

The use of DNA microarrays for detection and identification of bacteria and genes of interest from various environments (e.g. soil, sediment, water column...) is a major challenge for microbiologists working on functional diversity. So far, most of the genomic methods that have been described rely on the use of taxonomic markers (such as 16S rRNA) that can be easily amplified by PCR prior to hybridization on microarrays. However, taxonomical markers are not always informative on the functions present in these bacteria. Moreover, genes for which sequence database is limited or that lack any conserved regions will be difficult to amplify and thus to detect in unknown samples. Furthermore, PCR amplification often introduces biases that lead to inaccurate analysis of microbial communities. An alternative solution to overcome these strong limitations is to use genomic DNA (gDNA) as target for hybridisation, without prior PCR amplification. Though hybridization of gDNA is already used for comparative genome hybridization or sequencing by hybridization, yet to the high cost of tiling strategies and important data filtering, its adaptation for use in environmental research poses great challenges in terms of specificity, sensitivity and reproducibility of hybridization. Considering the very faint number of publications that have described hybridization of gDNA to microarrays for environmental applications, we confront in this review the different approaches that have been developed so far, and propose alternative strategies that may contribute to improve the development of microarrays for studying the microbial genetic structure and composition of samples of high environmental and ecological value.

Bacteria↗
PubMed1997

Genetic relationship of penicillin resistant Streptococcus pneumoniae serotype 19B strains in Japan.

Pulsed field gel electrophoresis (PFGE) of the genomic DNA of penicillin resistant serotype 19B Streptococcus pneumoniae was carried out. Thirteen strains form the Nagasaki area and 12 strains from other areas in Japan were examined. Twenty-three strains were resistant to erythromycin, tetracycline and trimethoprim/sulfamethoxazole but susceptible to chloramphenicol. Eight strains were resistant to ceftriaxone. All strains were multiply resistant. Five strains isolated from Nagasaki were indistinguishable from each other by using restriction enzymes Apa I and Sma I. Two strains isolated from other areas were indistinguishable from the above five strains. We could classify 13 Nagasaki strains into 3 groups and the total of 25 Japanese strains into 6 groups. These results suggest that the increasing prevalence of multiply drug resistant S. pneumoniae serotyped 19B in Japan is not due to a single clone, but at least one clone has spread widely in Japan.

DNA, Bacterial↗
PubMed2004

Biocatalytic degradation of pollutants.

Microbial reactions play key roles in biocatalysis and biodegradation. The recent genome sequencing of environmentally relevant bacteria has revealed previously unsuspected metabolic potential that could be exploited for useful purposes. For example, oxygenases and other biodegradative enzymes are benign catalysts that can be used for the production of industrially useful compounds. In conjunction with their biodegradative capacities, bacterial chemotaxis towards pollutants might contribute to the ability of bacteria to compete with other organisms in the environment and to be efficient agents for bioremediation. In addition to the bacterial biomineralization of organic pollutants, certain bacteria are also capable of immobilizing toxic heavy metals in contaminated aquifers, further illustrating the potential of microorganisms for the removal of pollutants.

Bacteria↗