False positive PCR detection of Tropheryma whipplei in the saliva of healthy people

<p>Abstract</p> <p>Background</p> <p><it>Tropheryma whipplei</it>, the agent of Whipple's disease (WD), has been recently isolated and the genomes of two isolates have been fully sequenced. Previous diagnosis tools for the diagnosis of the disease used sequence analysis of the 16S rRNA gene. Using this target gene, the high percentage of detection of the bacterium in saliva of healthy people was in contrast to the negative results obtained with specific target genes. The aim of our study was to compare previously published primers targeting the 16S rRNA gene to real-time PCR with Taqman* probes targeting specific repeat genes only found in the genome of <it>T. whipplei </it>in a series of 57 saliva from healthy people.</p> <p>Results</p> <p>Although the specific real-time PCR assays with both primers and probes were negative for all the samples, 13 out of 57 samples were positive with different primers previously reported targeting the 16S rRNA gene. Among the positive samples, 8 yielded a 231-bp sequence that was 99.1% identical to that of <it>Actinomyces odontolyticus</it>, 2 yielded a 226-bp that was 99.6% identical to that of <it>A. turicensis</it>, and 3 yielded a 160-bp sequence that was 98.5% identical to that of <it>Capnocytophaga gingivalis</it>. We found that the <it>C. gingivalis </it>and <it>A. odontolyticus </it>16S rRNA sequences obtained in our study share more than 80% homology with the corresponding 16S rRNA sequences of the <it>T. whipplei </it>genomes especially at 5' and 3' end.</p> <p>Conclusion</p> <p>Asymptomatic carriers of <it>T. whipplei </it>in saliva may exist but their prevalence is much lower than those previously reported. Testing the specificity of designed primers is critical to avoid false positive detection of <it>T. whipplei</it>. In atypical case we recommend to test two different specific target genes before concluding.</p

Mechanisms of polymyxin resistance: acquired and intrinsic resistance in bacteria

Polymyxins are polycationic antimicrobial peptides that are currently the last-resort antibiotics for the treatment of multidrug-resistant, Gram-negative bacterial infections. The reintroduction of polymyxins for antimicrobial therapy has been followed by an increase in reports of resistance among Gram-negative bacteria. Some bacteria, such as Klebsiella pneumoniae, Pseudomonas aeruginosa, and Acinetobacter baumannii, develop resistance to polymyxins in a process referred to as acquired resistance, whereas other bacteria, such as Proteus spp., Serratia spp., and Burkholderia spp., are naturally resistant to these drugs. Reports of polymyxin resistance in clinical isolates have recently increased, including acquired and intrinsically resistant pathogens. This increase is considered a serious issue, prompting concern due to the low number of currently available effective antibiotics. This review summarizes current knowledge concerning the different strategies bacteria employ to resist the activities of polymyxins. Gram-negative bacteria employ several strategies to protect themselves from polymyxin antibiotics (polymyxin B and colistin), including a variety of lipopolysaccharide (LPS) modifications, such as modifications of lipid A with phosphoethanolamine and 4-amino-4-deoxy-L-arabinose, in addition to the use of efflux pumps, the formation of capsules and overexpression of the outer membrane protein OprH, which are all effectively regulated at the molecular level. The increased understanding of these mechanisms is extremely vital and timely to facilitate studies of antimicrobial peptides and find new potential drugs targeting clinically relevant Gram-negative bacteria. (Résumé d'auteur

Synergistic activity of antibiotics combined with ivermectin to kill body lice

International audienceIvermectin and doxycycline have been found to be independently effective in killing body lice. In this study, 450 body lice were artificially fed on a Parafilm TM membrane with human blood associated with antibiotics (doxycycline, erythromycin, rifampicin and azithromycin) alone and in combination with ivermectin. Fluorescence in situ hybridisation and spectral deconvolution were performed to evaluate bacterial transcriptional activity following antibiotic intake by the lice. In the first series, a lethal effect of antibiotics on lice was observed compared with the control group at 18 days (log-rank test, P ≤ 10 −3), with a significant difference between groups in the production of nits (P = 0.019, Kruskal–Wallis test). A high lethal effect of ivermectin alone (50 ng/mL) was observed compared with the control group (log-rank test, P ≤ 10 −3). Fluorescence of bacteriocytes in lice treated with 20 ␮g/mL doxycycline was lower than in untreated lice (P < 0.0001, Kruskal–Wallis test). In the second series with antibiotic–ivermectin combinations , a synergistic lethal effect on treated lice (log-rank test, P < 10 −6) was observed compared with the control group at 18 days, associated with a significant decrease in the production of nits (P ≤ 0.001, Kruskal–Wallis test). Additionally, survival of lice in the combination treatment groups compared with ivermectin alone was significant (log-rank test, P = 0.0008). These data demonstrate that the synergistic effect of combinations of antibiotics and ivermectin could be used to achieve complete eradication of lice and to avoid selection of a resistant louse population

Antibiotics as selectors and accelerators 1 of diversity in the mechanisms of resistance: from the resistome to genetic plasticity in the β-lactamases world

Antibiotics and antibiotic resistance determinants, natural molecules closely related to bacterial physiology and consistent with an ancient origin, are not only present in antibiotic-producing bacteria. Throughput sequencing technologies have revealed an unexpected reservoir of antibiotic resistance in the environment. These data suggest that co-evolution between antibiotic and antibiotic resistance genes has occurred since the beginning of time. This evolutionary race has probably been slow because of highly regulated processes and low antibiotic concentrations. Therefore to understand this global problem, a new variable must be introduced, that the antibiotic resistance is a natural event, inherent to life. However, the industrial production of natural and synthetic antibiotics has dramatically accelerated this race, selecting some of the many resistance genes present in nature and contributing to their diversification. One of the best models available to understand the biological impact of selection and diversification are β-lactamases. They constitute the most widespread mechanism of resistance, at least among pathogenic bacteria, with more than 1000 enzymes identified in the literature. In the last years, there has been growing concern about the description, spread, and diversification of β-lactamases with carbapenemase activity and AmpC-type in plasmids. Phylogenies of these enzymes help the understanding of the evolutionary forces driving their selection. Moreover, understanding the adaptive potential of β-lactamases contribute to exploration the evolutionary antagonists trajectories through the design of more efficient synthetic molecules. In this review, we attempt to analyze the antibiotic resistance problem from intrinsic and environmental resistomes to the adaptive potential of resistance genes and the driving forces involved in their diversification, in order to provide a global perspective of the resistance problem

Zoonotic Focus of Plague, Algeria

After an outbreak of human plague, 95 Xenopsylla cheopis fleas from Algeria were tested for Yersinia pestis with PCR methods. Nine fleas were definitively confirmed to be infected with Y. pestis biovar orientalis. Our results demonstrate the persistence of a zoonotic focus of Y. pestis in Algeria

Rickettsia prowazekii and Real-time Polymerase Chain Reaction

This highly standardized and adaptable assay could improve epidemic typhus surveillance

Detection of "Rickettsia sp strain Uilenbergi" and "Rickettsia sp strain Davousti" in Amblyomma tholloni ticks from elephants in Africa

Background: To date, 6 tick-borne rickettsiae pathogenic for humans are known to occur in Africa and 4 of them were first identified in ticks before being recognized as human pathogens. Results: We examined 33 and 5 Amblyomma tholloni ticks from African elephants in the Central African Republic and Gabon, respectively, by PCR amplification and sequencing of a part of gltA and ompA genes of the genus Rickettsia. The partial sequences of gltA and ompA genes detected in tick in Gabon had 99.1% similarity with those of R. heilongjiangensis and 97.1% with those of Rickettsia sp. HL-93 strain, respectively. The partial gltA and ompA gene sequences detected in tick in the Central African Republic were 98.9% and 95.1% similar to those of Rickettsia sp. DnS14 strain and R. massiliae, respectively. Phylogenetic analysis showed Rickettsia sp. detected in Gabon clusters with R. japonica and R. heilongjiangensis in a phylogenetic tree based on the partial gltA and ompA genes. The genotype of the Rickettsia sp. detected in the Central African Republic is close to those of R. massiliae group in the phylogenetic tree based on partial gltA gene sequences, and distantly related to other rickettsiae in the tree based on partial ompA gene. Conclusion: The degrees of similarity of partial gltA and ompA genes with recognized species indicate the rickettsiae detected in this study may be new species although we could only study the partial sequences of 2 genes regarding the amount of DNA that was available. We propose the Rickettsia sp. detected in Gabon be provisionally named "Rickettsia sp. stain Davousti" and Rickettsia sp. detected in the Central African Republic be named "Rickettsia sp. strain Uilenbergi"

Comparison of Real-Time Quantitative PCR and Culture for the Diagnosis of Emerging Rickettsioses

Diagnosis of Rickettsia infection would benefit by use of the more rapid and sensitive method of quantitative real-time PCR than the time-intensive and less sensitive method of culturing Rickettsia species from skin biopsies. We evaluated culture sensitivity compared to PCR according to sampling delay and previous antibiotic treatment. We found that skin biopsies can be positive even when molecular tests were negative, and a negative result using molecular assays did not exclude the diagnosis of Rickettsia spp. infection. Rickettsia africae was the most common species in skin biopsies and R. slovaca was most common in ticks. We found a positive correlation between the number of bacteria copies and the isolation success in skin biopsies and ticks. The probability of isolating Rickettsia spp. was higher in untreated patients and in patients from our hometown. To increase the sensitivity of culture, skin biopsies should be sampled before treatment early in the course of the disease and should be inoculated as soon as possible

Emergence of drug resistant bacteria at the Hajj: A systematic review

Background Hajj is the annual mass gathering of Muslims, and is a reservoir and potential source of bacterial transmission. The emergence of bacterial transmission, including multi-drug resistance (MDR) bacteria, during Hajj has not been systematically assessed. Methods Articles in Pubmed, Scopus, and Google scholar were identified using controlled words relating to antibiotic resistance (AR) at the Hajj from January 2002 to January 2017. Eligible studies were identified by two researchers. AR patterns of bacteria were obtained for each study. Results We included 31 publications involving pilgrims, Hajj workers or local patients attending hospitals in Mecca, Mina, and the Medina area. Most of these publications provided antibiotic susceptibility results. Ten of them used the PCR approach to identify AR genes. MRSA carriage was reported in pilgrims and food handlers at a rate of 20%. Low rates of vancomycin-resistant gram-positive bacteria were reported in pilgrims and patients. The prevalence of third-generation cephalosporin-resistant bacteria was common in the Hajj region. Across all studies, carbapenem-resistant bacteria were detected in fewer than 10% of E.coli isolates tested but up to 100% in K. pneumoniae and A. baumannii. Colistin-resistant Salmonella enterica, including mcr-1 colistin-resistant E.coli and K.pneumoniae were only detected in the pilgrim cohorts. Conclusion This study provides an overview of the prevalence of MDR bacteria at the Hajj. Pilgrims are at high risk of AR bacterial transmission and may carry and transfer these bacteria when returning to their home countries. Thus, pilgrims should be instructed by health care practitioners about hygiene practices aiming at reducing traveler's diarrhea and limited use of antibiotics during travel in order to reduce the risk of MDR bacterial transmission