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Methods for detecting Gemmata spp. bacteremia in the microbiology laboratory.

ABSTRACT: Gemmata bacteria are fastidious, Gram-negative and aerobic. The only representatives are Gemmata obscuriglobus and Gemmata massiliana. These Planctomycetes appear to be a part of human digestive tract microbiome, and G. massiliana has been isolated from water. Further specific detection in the blood of two patients with febrile neutropenia suggests that Gemmata bacteremia may remain under-documented. The objective of this study was to develop an effective protocol to document Gemmata spp. bacteremia in the laboratory. Using mock-infected and control blood specimens, three methods for detecting Gemmata bacteremia, namely, automated microbial detection, culture on solid medium, and quantitative polymerase chain reaction (PCR), have been developed and studied.Gemmata spp. were undetected by automated blood culture system but culturing mock-infected blood on Caulobacter agar detected ? 102 G. obscuriglobus bacteria/mL and ? 104 G. massiliana bacteria/mL. Specific real-time PCR detected 102 Gemmata bacteria/mL. These protocols may be used to investigate the epidemiology of Gemmata spp. bacteremia.

SUBMITTER: Christen JR

PROVIDER: S-EPMC5759251 | biostudies-literature | 2018 Jan

REPOSITORIES: biostudies-literature

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Methods for detecting Gemmata spp. bacteremia in the microbiology laboratory.

Christen Jacques-Robert JR Edmond Edwin E Drancourt Michel M

BMC research notes 20180108 1

<h4>Objective</h4>Gemmata bacteria are fastidious, Gram-negative and aerobic. The only representatives are Gemmata obscuriglobus and Gemmata massiliana. These Planctomycetes appear to be a part of human digestive tract microbiome, and G. massiliana has been isolated from water. Further specific detection in the blood of two patients with febrile neutropenia suggests that Gemmata bacteremia may remain under-documented. The objective of this study was to develop an effective protocol to document G ...[more]

PMID: 29310696

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Project description:Background:Planctomycetes bacteria are known to be difficult to isolate, we hypothesized this may be due to missing iron compounds known to be important for other bacteria. We tested the growth-enhancement effect of complementing two standard media with Escherichia coli culture filtrate on two cultured strains of Gemmata spp. Also, the acquisition of iron by Gemmata spp. was evaluated by measuring various molecules involved in iron metabolism. Materials and Methods:Gemmata obscuriglobus and Gemmata massiliana were cultured in Caulobacter and Staley's medium supplemented or not with E. coli culture filtrate, likely containing siderophores and extracellular ferrireductases. We performed iron metabolism studies with FeSO4, FeCl3 and deferoxamine in the cultures with the E. coli filtrate and the controls. Results and Discussion:The numbers of G. obscuriglobus and G. massiliana colonies on Caulobacter medium or Staley's medium supplemented with E. coli culture filtrate were significantly higher than those on the standard medium (p < 0.0001). Agar plate assays revealed that the Gemmata colonies near E. coli colonies were larger than the more distant colonies, suggesting the diffusion of unknown growth promoting molecules. The inclusion of 10-4 to 10-3 M FeSO4 resulted in rapid Gemmata spp. growth (4-5 days compared with 8-9 days for the controls), suggesting that both species can utilize FeSO4 to boost their growth. In contrast, deferoxamine slowed down and prevented Gemmata spp. growth. Further studies revealed that the complementation of Caulobacter medium with E. coli culture filtrate and 10-4 M FeSO4 exerted a significant growth-enhancement effect compared with that obtained with Caulobacter medium supplemented with E. coli culture filtrate alone (p < 0.0122). Moreover, the intracellular iron concentrations in G. obscuriglobus and G. massiliana cultures in iron-depleted broth supplemented with the E. coli filtrate were 0.63 ± 0.16 and 0.78 ± 0.12 ?mol/L, respectively, whereas concentrations of 1.72 ± 0.13 and 1.56± 0.11 ?mol/L were found in the G. obscuriglobus and G. massiliana cultures grown in broth supplemented with the E. coli filtrate and FeSO4. The data reported here indicated that both E. coli culture filtrate and FeSO4 act as growth factors for Gemmata spp. via a potentiation mechanism.

Project description:Next-generation sequencing (NGS) of bacterial genomes has recently become more accessible and is now available to the routine diagnostic microbiology laboratory. However, questions remain regarding its feasibility, particularly with respect to data analysis in nonspecialist centers. To test the applicability of NGS to outbreak investigations, Ion Torrent sequencing was used to investigate a putative multidrug-resistant Escherichia coli outbreak in the neonatal unit of the Mercy Hospital for Women, Melbourne, Australia. Four suspected outbreak strains and a comparator strain were sequenced. Genome-wide single nucleotide polymorphism (SNP) analysis demonstrated that the four neonatal intensive care unit (NICU) strains were identical and easily differentiated from the comparator strain. Genome sequence data also determined that the NICU strains belonged to multilocus sequence type 131 and carried the bla(CTX-M-15) extended-spectrum beta-lactamase. Comparison of the outbreak strains to all publicly available complete E. coli genome sequences showed that they clustered with neonatal meningitis and uropathogenic isolates. The turnaround time from a positive culture to the completion of sequencing (prior to data analysis) was 5 days, and the cost was approximately $300 per strain (for the reagents only). The main obstacles to a mainstream adoption of NGS technologies in diagnostic microbiology laboratories are currently cost (although this is decreasing), a paucity of user-friendly and clinically focused bioinformatics platforms, and a lack of genomics expertise outside the research environment. Despite these hurdles, NGS technologies provide unparalleled high-resolution genotyping in a short time frame and are likely to be widely implemented in the field of diagnostic microbiology in the next few years, particularly for epidemiological investigations (replacing current typing methods) and the characterization of resistance determinants. Clinical microbiologists need to familiarize themselves with these technologies and their applications.

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Methods for detecting Gemmata spp. bacteremia in the microbiology laboratory.

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Methods for detecting Gemmata spp. bacteremia in the microbiology laboratory.

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