Amoebal pathogens as emerging causal agents of pneumonia

Despite using modern microbiological diagnostic approaches, the aetiological agents of pneumonia remain unidentified in about 50% of cases. Some bacteria that grow poorly or not at all in axenic media used in routine clinical bacteriology laboratory but which can develop inside amoebae may be the agents of these lower respiratory tract infections (RTIs) of unexplained aetiology. Such amoebae-resisting bacteria, which coevolved with amoebae to resist their microbicidal machinery, may have developed virulence traits that help them survive within human macrophages, i.e. the first line of innate immune defence in the lung. We review here the current evidence for the emerging pathogenic role of various amoebae-resisting microorganisms as agents of RTIs in humans. Specifically, we discuss the emerging pathogenic roles of Legionella-like amoebal pathogens, novel Chlamydiae (Parachlamydia acanthamoebae, Simkania negevensis), waterborne mycobacteria and Bradyrhizobiaceae (Bosea and Afipia spp.

Protochlamydia naegleriophila as Etiologic Agent of Pneumonia

Using ameba coculture, we grew a Naegleria endosymbiont. Phenotypic, genetic, and phylogenetic analyses supported its affiliation as Protochlamydia naegleriophila sp. nov. We then developed a specific diagnostic PCR for Protochlamydia spp. When applied to bronchoalveolar lavages, results of this PCR were positive for 1 patient with pneumonia. Further studies are needed to assess the role of Protochlamydia spp. in pneumonia

Applications of MALDI-TOF mass spectrometry in clinical diagnostic microbiology

Until recently, microbial identification in clinical diagnostic laboratories has mainly relied on conventional phenotypic and gene sequencing identification techniques. The development of matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) devices has revolutionized the routine identification of microorganisms in clinical microbiology laboratories by introducing an easy, rapid, high throughput, low-cost, and efficient identification technique. This technology has been adapted to the constraint of clinical diagnostic laboratories and has the potential to replace and/or complement conventional identification techniques for both bacterial and fungal strains. Using standardized procedures, the resolution of MALDI-TOF MS allows accurate identification at the species level of most Gram-positive and Gram-negative bacterial strains with the exception of a few difficult strains that require more attention and further development of the method. Similarly, the routine identification by MALDI-TOF MS of yeast isolates is reliable and much quicker than conventional techniques. Recent studies have shown that MALDI-TOF MS has also the potential to accurately identify filamentous fungi and dermatophytes, providing that specific standardized procedures are established for these microorganisms. Moreover, MALDI-TOF MS has been used successfully for microbial typing and identification at the subspecies level, demonstrating that this technology is a potential efficient tool for epidemiological studies and for taxonomical classificatio

Distinct Genomic Features Characterize Two Clades of <i>Corynebacterium diphtheriae</i>: Proposal of <i>Corynebacterium diphtheriae</i> Subsp. <i>diphtheriae</i> Subsp. nov. and <i>Corynebacterium diphtheriae</i> Subsp. <i>lausannense</i> Subsp. nov.

&lt;i&gt;Corynebacterium diphtheriae&lt;/i&gt; is the etiological agent of diphtheria, a disease caused by the presence of the diphtheria toxin. However, an increasing number of records report non-toxigenic &lt;i&gt;C. diphtheriae&lt;/i&gt; infections. Here, a &lt;i&gt;C. diphtheriae&lt;/i&gt; strain was recovered from a patient with a past history of bronchiectasis who developed a severe tracheo-bronchitis with multiple whitish lesions of the distal trachea and the mainstem bronchi. Whole-genome sequencing (WGS), performed in parallel with PCR targeting the toxin gene and the Elek test, provided clinically relevant results in a short turnaround time, showing that the isolate was non-toxigenic. A comparative genomic analysis of the new strain (CHUV2995) with 56 other publicly available genomes of &lt;i&gt;C. diphtheriae&lt;/i&gt; revealed that the strains CHUV2995, CCUG 5865 and CMCNS703 share a lower average nucleotide identity (ANI) (95.24 to 95.39%) with the &lt;i&gt;C. diphtheriae&lt;/i&gt; NCTC 11397 &lt;sup&gt;T&lt;/sup&gt; reference genome than all other &lt;i&gt;C. diphtheriae&lt;/i&gt; genomes (&gt;98.15%). Core genome phylogeny confirmed the presence of two monophyletic clades. Based on these findings, we propose here two new &lt;i&gt;C. diphtheriae&lt;/i&gt; subspecies to replace the lineage denomination used in previous multilocus sequence typing studies: &lt;i&gt;C. diphtheriae&lt;/i&gt; subsp. &lt;i&gt;lausannense&lt;/i&gt; subsp. nov. (instead of lineage-2), regrouping strains CHUV2995, CCUG 5865, and CMCNS703, and &lt;i&gt;C. diphtheriae&lt;/i&gt; subsp. &lt;i&gt;diphtheriae&lt;/i&gt; subsp. nov, regrouping all other &lt;i&gt;C. diphtheriae&lt;/i&gt; in the dataset (instead of lineage-1). Interestingly, members of subspecies &lt;i&gt;lausannense&lt;/i&gt; displayed a larger genome size than subspecies &lt;i&gt;diphtheriae&lt;/i&gt; and were enriched in COG categories related to transport and metabolism of lipids (I) and inorganic ion (P). Conversely, they lacked all genes involved in the synthesis of pili (SpaA-type, SpaD-type and SpaH-type), molybdenum cofactor and of the nitrate reductase. Finally, the CHUV2995 genome is particularly enriched in mobility genes and harbors several prophages. The genome encodes a type II-C CRISPR-Cas locus with 2 spacers that lacks &lt;i&gt;csn2&lt;/i&gt; or &lt;i&gt;cas4&lt;/i&gt; , which could hamper the acquisition of new spacers and render strain CHUV2995 more susceptible to bacteriophage infections and gene acquisition through various mechanisms of horizontal gene transfer