Multiplex PCR for rapid differentiation of three species in the “Clostridium clostridioformegroup”

Abstract

Clostridium clostridioforme is a relatively antimicrobial resistant, phenotypically heterogeneous anaerobe that has been involved in a variety of infections. 16S rDNA sequencing analysis revealed three principal species in what has been called Clostridium clostridioforme - Clostridium bolteae, C. clostridioforme, and Clostridium hathewayi. Based on the 16S rDNA sequence information we obtained, we developed a cost-effective, timesaving one-step multiplex PCR assay for rapid and accurate differentiation of these three species. The established multiplex PCR identification scheme was applied to the identification of 88 clinical isolates that had previously been identified phenotypically as C. clostridioforme. The identification obtained from multiplex PCR assays showed 100% agreement with 16S rDNA sequencing identification. This scheme will permit more accurate assessment of the role of these three Clostridium species in infection and of the degree of antimicrobial resistance in each of the species.

Selective Growth Inhibitory Effect of Biochanin A Against Intestinal Tract Colonizing Bacteria

“…The results summarized in Table 1 show that the most susceptible strains with MIC 64 μg/mL were the type culture as well as the clinical isolates of Cl. clostridioforme , a relatively antimicrobial resistant anaerobe that has been involved in a variety of infections, including bacteraemia [ 43 ]. The inhibition activity at MIC level 256 μg/mL was observed for type cultures of Cl.…”

“…Although clostridia are generally considered to be Gram-positive, this genus is not uniform in its Gram-staining. According to Yuli et al [ 43 ] Cl. clostridioforme may stand as Gram-negative; Cl.…”

Quantitative and dynamic profiling of human gut core microbiota by real-time PCR

“…Species-specific genetic markers and primers were selected in two steps. First, we conducted a comprehensive literature survey to search and verify previously reported specific primers that were used to detect these microbes (Ahmed et al 2019 ; Chen et al 2019 ; Gui et al 2020 ; Laue et al 2006 ; Li et al 2020 ; Park et al 2013 ; Schriefer et al 2018 ; Takahashi et al 2016 ; Tong et al 2011 ; Yu et al 2017 ; Yuli et al 2005 ). Second, we performed the following in silico analysis for some bacteria with no available or qualified specific primers.…”

Temporal Regulation of the Bacterial Metabolite Deoxycholate during Colonic Repair Is Critical for Crypt Regeneration

“…Stool was collected from the treated groups of mice at day 6 post-injury and DNA was isolated using the QIAamp DNA Stool Mini Kit (Qiagen). The following primers were used for PCR-based amplification (Nava et al, 2011; Steed et al, 2017; Wells et al, 2003; Yuli et al, 2005). baiCD forward (5′-GGWTTCAGCCCRCAGATGTTCTTTG-3′), baiCD reverse (5′-GAATTCCGGGTTCATGAACATTCTKCKAAG-3′ Clostridium clostridioforme forward (5’-GCCGCATGGCAGTGTGT-3’), Clostridium clostridioforme reverse (5′-ACCGGCCCGTCAGGGGGAT-3′), rpoB forward (5′-AACATCGGTTTGATCAAC-3′) and rpoB reverse (5′-CGTTGCATGTTGGTACCCAT-3′).…”