Nadim Cassir

Culture of previously uncultured members of the human gut microbiota by culturomics

Metagenomics revolutionized the understanding of the relations among the human microbiome, health and diseases, but generated a countless number of sequences that have not been assigned to a known microorganism1. The pure culture of prokaryotes, neglected in recent decades, remains essential to elucidating the role of these organisms2. We recently introduced microbial culturomics, a culturing approach that uses multiple culture conditions and matrix-assisted laser desorption/ionization–time of flight and 16S rRNA for identification2. Here, we have selected the best culture conditions to increase the number of studied samples and have applied new protocols (fresh-sample inoculation; detection of microcolonies and specific cultures of Proteobacteria and microaerophilic and halophilic prokaryotes) to address the weaknesses of the previous studies3–5. We identified 1,057 prokaryotic species, thereby adding 531 species to the human gut repertoire: 146 bacteria known in humans but not in the gut, 187 bacteria and 1 archaea not previously isolated in humans, and 197 potentially new species. Genome sequencing was performed on the new species. By comparing the results of the metagenomic and culturomic analyses, we show that the use of culturomics allows the culture of organisms corresponding to sequences previously not assigned. Altogether, culturomics doubles the number of species isolated at least once from the human gut.

A new strategy to fight antimicrobial resistance: the revival of old antibiotics

The increasing prevalence of hospital and community-acquired infections caused by multidrug-resistant (MDR) bacterial pathogens is limiting the options for effective antibiotic therapy. Moreover, this alarming spread of antimicrobial resistance has not been paralleled by the development of novel antimicrobials. Resistance to the scarce new antibiotics is also emerging. In this context, the rational use of older antibiotics could represent an alternative to the treatment of MDR bacterial pathogens. It would help to optimize the armamentarium of antibiotics in the way to preserve new antibiotics and avoid the prescription of molecules known to favor the spread of resistance (i.e., quinolones). Furthermore, in a global economical perspective, this could represent a useful public health orientation knowing that several of these cheapest “forgotten” antibiotics are not available in many countries. We will review here the successful treatment of MDR bacterial infections with the use of old antibiotics and discuss their place in current practice.

Clostridium butyricum strains and dysbiosis linked to necrotizing enterocolitis in preterm neonates

BACKGROUND Necrotizing enterocolitis (NEC) is the most common and serious gastrointestinal disorder among preterm neonates. We aimed to assess a specific gut microbiota profile associated with NEC. METHODS Stool samples and clinical data were collected from 4 geographically independent neonatal intensive care units, over a 48-month period. Thirty stool samples from preterm neonates with NEC (n = 15) and controls (n = 15) were analyzed by 16S ribosomal RNA pyrosequencing and culture-based methods. The results led us to develop a specific quantitative polymerase chain reaction (qPCR) assay for Clostridium butyricum, and we tested stool samples from preterm neonates with NEC (n = 93) and controls (n = 270). We sequenced the whole genome of 16 C. butyricum strains, analyzed their phylogenetic relatedness, tested their culture supernatants for cytotoxic activity, and searched for secreted toxins. RESULTS Clostridium butyricum was specifically associated with NEC using molecular and culture-based methods (15/15 vs 2/15; P < .0001) or qPCR (odds ratio, 45.4 [95% confidence interval, 26.2-78.6]; P < .0001). Culture supernatants of C. butyricum strains from preterm neonates with NEC (n = 14) exhibited significant cytotoxic activity (P = .008), and we identified in all a homologue of the β-hemolysin toxin gene shared by Brachyspira hyodysenteriae, the etiologic agent of swine dysentery. The corresponding protein was secreted by a NEC-associated C. butyricum strain. CONCLUSIONS NEC was associated with C. butyricum strains and dysbiosis with an oxidized, acid, and poorly diversified gut microbiota. Our findings highlight the plausible toxigenic mechanism involved in the pathogenesis of NEC.

Routine use of Staphylococcus aureus rapid diagnostic test in patients with suspected ventilator-associated pneumonia.

IntroductionIn patients with ventilator-associated pneumonia (VAP), administration of an appropriate empirical antimicrobial treatment is associated with improved outcomes, leading to the prescription of broad-spectrum antibiotics, including a drug active against methicillin resistant Staphylococcus aureus (MRSA). In order to avoid the overuse of antibiotics, the present study aimed to evaluate the technical characteristics of a rapid diagnostic test (Cepheid Xpert assay) in patients with suspected VAP.MethodsFrom June 2011 to June 2012, in patients with suspected VAP, a sample from the bronchialalveolar lavage (BAL) or miniBAL was tested in a point-of-care laboratory for a rapid diagnostic test of methicillin susceptible Staphylococcus aureus (MSSA) and MRSA. Then, the result was compared to the quantitative culture with a threshold at 104 colony-forming units per milliliter for bronchoalveolar lavage and 103 colony-forming units per milliliter for minibronchoalveolar lavage. The study was performed in three intensive care units at two institutions.ResultsFour hundred, twenty-two samples from 328 patients were analyzed. The culture of 6 (1.1%) and 28 (6.5%) samples were positive for MRSA and MSSA. The test was not interpretable in 41 (9.3%) patients. The negative predictive values of the rapid detection test were 99.7% (98.1 to 99.9%) and 99.8% (98.7 to 99.9%) for MSSA and MRSA, respectively.ConclusionThe rapid diagnostic test is reliable in excluding the presence of MSSA and MRSA in the samples of patients with suspected VAP. Its utility should be regarded depending on the prevalence of MRSA.

Clostridium difficile 027 emerging outbreak in Marseille, France.

Author(s): Jean-Christophe Lagier, MD, PhD; Gregory Dubourg, PharmD; Nadim Cassir, MD; Pierre-Edouard Fournier, MD, PhD; Philippe Colson, PharmD, PhD; Hervé Richet, MD, PhD; Philippe Brouqui, MD, PhD; Didier Raoult, MD, PhD Source: Infection Control and Hospital Epidemiology, Vol. 34, No. 12 (December 2013), pp. 1339-1341 Published by: The University of Chicago Press on behalf of The Society for Healthcare Epidemiology of America Stable URL: http://www.jstor.org/stable/10.1086/673995 . Accessed: 28/06/2014 14:14

Risk factors for surgical site infections after neurosurgery: A focus on the postoperative period

BACKGROUND Surgical site infection (SSI) after neurosurgery has potentially devastating consequences. METHODS A prospective cohort study was conducted over a period of 24 months in a university center. All adult patients undergoing neurosurgical procedures, with exception of open skull fractures, were included. Multivariate logistic regression analysis was used to identify independent risk factors. RESULTS We included 949 patients. Among them, 43 were diagnosed with SSI (4.5%). A significant reduction in postneurosurgical SSI from 5.8% in 2009 to 3.0% in 2010 (P = .04) was observed. During that period, an active surveillance with regular feedback was established. The most common microorganisms isolated from SSI were Staphylococcus aureus (23%), Enterobacteriaceae (21%), and Propionibacterium acnes (12%). We identified the following independent risk factors for SSI postcranial surgery: intensive care unit (ICU) length of stay ≥7 days (odds ratio [OR] = 6.1; 95% confidence interval [CI], 1.7-21.7), duration of drainage ≥3 days (OR = 3.3; 95% CI, 1.1-11), and cerebrospinal fluid leakage (OR = 5.6; 95% CI, 1.1-30). For SSIs postspinal surgery, we identified the following: ICU length of stay ≥7 days (OR = 7.2; 95% CI, 1.6-32.1), coinfection (OR = 9.9; 95% CI, 2.2-43.4), and duration of drainage ≥3 days (OR = 5.7; 95% CI, 1.5-22). CONCLUSION Active surveillance with regular feedback proved effective in reducing SSI rates. The postoperative period is associated with overlooked risk factors for neurosurgical SSI. Infection control measures targeting this period are therefore promising.

Robinsoniella peoriensis infection following surgery for scoliosis: a case report

IntroductionRobinsoniella peoriensis was recently identified as a Gram-positive, spore-forming, anaerobic bacillus originally isolated from swine manure storage pits. Seven isolates have been subsequently reported from human sources.Case presentationWe report the case of an infection caused by R. peoriensis in a 45-year-old Caucasian woman after posterior instrumentation correction of idiopathic thoracolumbar scoliosis. The identification was made by culture of samples inoculated onto blood agar and chocolate agar and was confirmed by 16 S ribosomal ribonucleic acid gene sequencing.ConclusionsWe discuss similar cases suggesting that R. peoriensis is responsible for health care-associated infections with the colonic flora as a potential source of infection.

Clostridium butyricum: from beneficial to a new emerging pathogen.

Clostridium butyricum, a strictly anaerobic spore-forming bacillus, is a common human and animal gut commensal bacterium, and is also frequently found in the environment. Whereas non-toxigenic strains are currently used as probiotics in Asia, other strains have been implicated in pathological conditions, such as botulism in infants or necrotizing enterocolitis in preterm neonates. In terms of the latter, within the same species, different strains have antagonist effects on the intestinal mucosa. In particular, short-chain fatty acids, which are products of carbohydrate fermentation, have a dose-dependent paradoxical effect. Moreover, toxin genes have been identified by genome sequencing in pathological strains. Asymptomatic carriage of these strains has also been reported. Herein, we provide an overview of the implications of C. butyricum for human health, from the beneficial to the pathogenic. We focus on pathogenic strains associated with the occurrence of necrotizing enterocolitis. We also discuss the need to use complementary microbiological methods, including culture, in order to better assess gut bacterial diversity and identify new emergent enteropathogens at the strain level.

Outbreak of adenovirus type 1 severe pneumonia in a French intensive care unit, September–October 2012

We herein describe and analyse the first outbreak of severe pneumonia caused by human adenovirus type1 (HAdV C type 1), which included immunocompetent patients in an intensive care unit (ICU) of Marseille, France, and occurred between September and October 2012. Seven successive patients were diagnosed by HAdV specific real-time polymerase chain reaction with a positive bronchoalveolar lavage. After the collection of nasopharyngeal swabs from healthcare workers, three nurses working night shifts tested positive for HAdV C including one that had exhibited respiratory signs while working one week before the outbreak. She was the most likely source of the outbreak. Our findings suggest that HAdV-1 could be considered as a possible cause of severe pneumonia even in immunocompetent patients with a potential to cause outbreaks in ICUs. HAdV rapid identification and typing is needed to curtail the spread of this pathogen. Reinforcing hand hygiene with antiseptics with demonstrated activity against non-enveloped viruses and ensuring that HCWs with febrile respiratory symptoms avoid direct patient contact are critical measures to prevent transmission of HAdV in healthcare settings.

Infectious Diseases of High Consequence and Personal Protective Equipment: A Didactic Method to Assess the Risk of Contamination.

Infectious Diseases of High Consequence and Personal Protective Equipment: A Didactic Method to Assess the Risk of Contamination Nadim Cassir;Sophia Boudjema;Valerie Roux;Pauline Reynier;Philippe Brouqui; Infection Control & Hospital Epidemiology