Search

Dridi B, Raoult D, Drancourt M. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry identification of Archaea: towards the universal identification of living organisms. APMIS 2012; 120: 85-91. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) identification of Archaea has been limited to some environmental extremophiles belonging to distant taxa. We developed a specific protocol for MALDI-TOF-MS identification of Archaea and applied it to seven environmental human-associated Methanobrevibacter smithii, Methanobrevibacter oralis, Methanosphaera stadtmanae, and the recently described Methanomassiliicoccus luminyensi Archaea. After mechanical lyse, we observed a unique protein profile for each organisms comprising 7-24 peaks ranging from 3 015 to 10 632 Da with a high quality score of 7.38 ± 1.26. Profiles were reproducible over successive experiments performed at 1, 2, and 3-week growth durations and unambiguously distinguished the Archaea from all of the 3 995 bacterial spectra in the Brüker database. After the incorporation of the determined profiles into a local database, archaeal isolates were blindly identified within 10 min with an identification score of 1.9-2.3. The MALDI-TOF-MS-based clustering of these archaeal organisms was consistent with their 16S rDNA sequence-based phylogeny. These data prove that MALDI-TOF-MS profiling could be used as a first-line technique for the identification of human Archaea. In complement to previous reports for animal cells, Bacteria and giant viruses, MALDI-TOF-MS therefore appears as a universal method for the identification of living unicellular and multicellular organisms. [ABSTRACT FROM AUTHOR]

Rickettsia are best known as strictly intracellular vector-borne bacteria that cause mild to severe diseases in humans and other animals. Recent advances in molecular tools and biological experiments have unveiled a wide diversity of Rickettsia spp. that include species with a broad host range and some species that act as endosymbiotic associates. Molecular phylogenies of Rickettsia spp. contain some ambiguities, such as the position of R. canadensis and relationships within the spotted fever group. In the modern era of genomics, with an ever-increasing number of sequenced genomes, there is enhanced interest in the use of whole-genome sequences to understand pathogenesis and assess evolutionary relationships among rickettsial species. Rickettsia have small genomes (1.1-1.5 Mb) as a result of reductive evolution. These genomes contain split genes, gene remnants and pseudogenes that, owing to the colinearity of some rickettsial genomes, may represent different steps of the genome degradation process. Genomics reveal extreme genome reduction and massive gene loss in highly vertebrate-pathogenic Rickettsia compared to less virulent or endosymbiotic species. Information gleaned from rickettsial genomics challenges traditional concepts of pathogenesis that focused primarily on the acquisition of virulence factors. Another intriguing phenomenon about the reduced rickettsial genomes concerns the large fraction of non-coding DNA and possible functionality of these 'non-coding' sequences, because of the high conservation of these regions. Despite genome streamlining, Rickettsia spp. contain gene families, selfish DNA, repeat palindromic elements and genes encoding eukaryotic-like motifs. These features participate in sequence and functional diversity and may play a crucial role in adaptation to the host cell and pathogenesis. Genome analyses have identified a large fraction of mobile genetic elements, including plasmids, suggesting the possibility of lateral gene transfer in these intracellular bacteria. Phylogenetic analyses have identified several candidates for horizontal gene acquisition among Rickettsia spp. including tra, pat2, and genes encoding for the type IV secretion system and ATP/ADP translocase that may have been acquired from bacteria living in amoebae. Gene loss, gene duplication, DNA repeats and lateral gene transfer all have shaped rickettsial genome evolution. A comprehensive analysis of the entire genome, including genes and non-coding DNA, will help to unlock the mysteries of rickettsial evolution and pathogenesis. [ABSTRACT FROM AUTHOR]

Summary We have developed a matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS)-based identification technique for Planctomycetes organisms, which are used here as bacteria of suitable diversity at genus and species level for testing resolution of the method. Planctomyces maris ATCC 29201, Planctomyces brasiliensis ATCC 49424T, P. brasiliensis ATCC 49425, Planctomyces limnophilus ATCC 43296T, Blastopirellula marina ATCC 49069T, Rhodopirellula baltica DSM 10527T and Gemmata obscuriglobus DSM 5831T were cultured on half-strength marine broth and agar, or alternatively on caulobacter broth and agar. The resulting pellets of organisms (liquid) or colonies (solid agar) were directly applied to a MALDI-TOF plate. This yielded a reproducible, unique protein profiles comprising 23-39 peaks ranging in size from 2403 to 12 091 Da. These peaks were unambiguously distinguished from any of the 3038 bacterial spectra in the Brüker database. Matrix-assisted laser desorption/ionization time-of-flight patterns were similar for isolates grown in solid and in liquid medium, albeit the patterns from solid growth were more easily interpretable. After the incorporation of the herein determined profiles into the Brüker database, Planctomycetes isolates were blindly identified within 10 min, with an identification score in the range of 1.8 to 2.3. Matrix-assisted laser desorption/ionization time-of-flight-based clustering of these Planctomycetes organisms was consistent with 16S rDNA-based phylogeny. However, the incorporation of additional non- Planctomycetes MALDI-TOF profiles in the analysis resulted in inconsequential clustering. In conclusion, MALDI-TOF protein profiling is a new approach for the rapid and accurate identification of cultured environmental organisms, as illustrated in this study through the analysis of Planctomycetes. [ABSTRACT FROM AUTHOR]

Q fever is a worldwide zoonosis caused by Coxiella burnetii. Q fever may be present as an acute or a chronic infection and can be reactivated during subsequent pregnancies. Although its exact prevalence remains unknown, it is likely that the number of cases of Q fever in pregnant women is underestimated. During pregnancy, the illness is likely to be asymptomatic, and diagnosis is based on serology. Acute infection results in appearance of IgM and IgG antibodies mainly directed against the avirulent form of C. burnetii (phase II). Chronic Q fever results in particularly high level of IgG and IgA antibodies directed against both virulent (phase I) and avirulent (phase II) forms of the bacterium. Q fever may result in adverse pregnancy outcome, including spontaneous abortion, intrauterine growth retardation, oligoamnios, intrauterine fetal death (IUFD), and premature delivery. Obstetric complications occur significantly more often as C. burnetii infects the patient at an early stage of her pregnancy. Occurrence of IUFD is correlated with the presence of placental infection by C. burnetii and might be the consequence of direct infection of the fetus. The mother is exposed to the risk of chronic Q fever and endocarditis with potential fatal evolution. Long-term cotrimoxazole therapy prevents from placental infection, IUFD, and maternal chronic Q fever. Such treatment should be used to treat pregnant women with Q fever. Women with previous history of Q fever should have a regular serological follow up. Obstetricians’ knowledge about Q fever must be improved. [ABSTRACT FROM AUTHOR]

Due to improved diagnostic methods and increased interest, the number of representatives of the genus Rickettsia has increased dramatically over the past 20 years, with 25 currently validated species. These arthropod-associated intracellular bacteria are now recognized in all parts of the world. The comparison of the phylogenic organizations obtained from the study of several genes with different functions, and more recently from genomic sequences, provided basis to establish a reliable taxonomy of the bacteria induded in the genus Rickettsia. These data were incorporated into polyphasic consensus guidelines that provide clear recommendations for the taxonomic classification and nomenclature of rickettsiae and rickettsial diseases. [ABSTRACT FROM AUTHOR]

Amoeba-resisting bacteria (ARB) such as Legionella spp. are currently regarded as potential human pathogens living in the environment. To detect ARB from both human and environmental samples, co-culture with amoebae has been demonstrated as an efficient tool. However, using this procedure, mostly water from cooling towers and hospital water supplies have been investigated as the possible reservoir of ARB. In the present study, we studied ARB population in 77 environmental water samples including rivers, fountains, lakes and domestic wells in the south of France. As a result, a total of 244 isolates corresponding to 89 different species of ARB, but not Legionella spp., were identified. Ability to grow within and/or to be lytic for amoebae was revealed for the first time for several human pathogens . Six isolates are likely to be the members of a new or uncharacterized genus/species. An anaerobic bacterium, Clostridium frigidicarnis was demonstrated to be lytic for amoebae. This preliminary work demonstrates that the water environment in the vicinity of humans is a reservoir of ARB, including well-known pathogens for which amoebae and/or water was not recognized earlier as a possible reservoir. [ABSTRACT FROM AUTHOR]

Homeless people are particularly exposed to ectoparasite. The living conditions and the crowded shelters provide ideal conditions for the spread of lice, fleas, ticks, and mites. Body lice have long been recognized as human parasites and although typically prevalent in rural communities in upland areas of countries close to the equator, it is now increasingly encountered in developed countries especially in homeless people or inner city economically deprived population. Fleas are widespread but are not adapted to a specific host and may occasionally bite humans. Most common fleas that parasite humans are the cat, the rat, and the human fleas, Ctenocephalides felis, Xenopsylla cheopis, and Pulex irritans, respectively. Ticks belonging to the family Ixodidae, in particular, the genera Dermacentor, Rhipicephalus, and Ixodes, are frequent parasites in humans. Sarcoptes scabiei var. hominis is a mite ( Arachnida class) responsible for scabies. It is an obligate parasite of human skin. The hematophagic-biting mite, Liponyssoides sanguineus, is a mite of the rat, mouse, and other domestic rodents but can also bite humans. Finally, the incidence of skin disease secondary to infestation with the human bedbug, Cimex lectularius, has increased recently. Bacteria, such as Wolbacchia spp. have been detected in bedbug. The threat posed by the ectoparasite in homeless is not the ectoparasite themselves but the associated infectious diseases that they may transmit to humans. Except for scabies all these ectoparasites are potential vectors for infectious agents. Three louse-borne diseases are known at this time. Trench fever caused by Bartonella quintana ( B. quintana), epidemic typhus caused by Rickettsia prowazekii, and relapsing fever caused by the spirochete Borrelia recurrentis. Fleas transmit plague ( Xenopsylla cheopis and Pulex irritans), murine typhus ( Xenopsylla cheopis), flea-borne spotted rickettsiosis on account of the recently described species Rickettsia felis ( C. felis), and occasionally cat scratch disease on account of Bartonella henselae ( C. felis). The role of fleas as potential vector of B. quintana has recently been suggested. Among the hematophagic-biting mites, L. sanguineus, is responsible for the transmission of Rickettsia akari, the etiologic agent of rickettsialpox. Virtually, no data are available on tick-borne disease in this population. This article will deal with epidemiology, diagnosis, prevention, and treatment of these ectoparasite and the infectious diseases they transmit to the homeless people. [ABSTRACT FROM AUTHOR]

Fenollar F, Raoult D. Molecular genetic methods for the diagnosis of fastidious microorganisms. APMIS 2004;112:785–807.Technological innovations in the detection and identification of microorganisms using molecular techniques such as polymerase chain reaction (PCR) have ushered in a new era with respect to diagnostic microbiology. PCR using universal or specific primers followed by identification of amplified product, mainly by sequencing, has enabled the rapid identification of cultured or uncultured bacteria. Thus, PCR may allow quick diagnosis of infections caused by fastidious pathogens for which culture could be extremely difficult. However, several pitfalls, such as false positives, have been observed with PCR, underlining the necessity to interpret the results obtained with caution. At present, certain improvements in the molecular genetic methods may be helpful for the diagnosis of infectious diseases. Indeed, the recent development of bacterial genome sequencing has provided an important source of potential targets for PCR, allowing rational choice of primers for diagnosis and genotyping. In addition, the development of new techniques such as real-time PCR offers several advantages in comparison to conventional PCR, including speed, simplicity, reproducibility, quantitative capability and low risk of contamination. Herein, we review the general principles of PCR-based diagnosis and molecular genetic methods for the diagnosis of several hard-to-culture bacteria, such asRickettsiaspp.,Ehrlichiaspp.,Coxiella burnetii,Bartonellaspp.,Tropheryma whippleiandYersinia pestis. [ABSTRACT FROM AUTHOR]

Antibiotic resistance in pathogenic bacteria is a major problem for human health. We analyzed metagenomic datasets from ancient and remote samples from diverse environmental sources and observed the presence of all the eleven antibiotic resistance genes (ARG) groups evaluated. Since ancient samples are not subjected to modern effects of antibiotic misuse, they represent a clean model to explore the natural diversity of ARG in the environment. Most sequences showed high divergence compared with known ARG, representing a much larger universe than the currently known and characterized ARGs. We explored whether proteins within the "divergent resistome" may correspond to functional ARG by characterizing a beta-lactamase hit with very low similarity to any known sequence (<45% to best BLAST hit in NCBI). By starting from purely in-silico data, we revived a new family of class B beta-lactamases from ancient medieval samples, which exhibited a very high penicillinase activity. In this work, we explored ancient resistomes and added novel support to previous works showing that the universe of ARG is naturally vast and diverse in microbial communities. Our results bring a new perspective to the exploration of environmental ARG and indicate that this gigantic reservoir represents a natural endless source of emerging resistances. [ABSTRACT FROM AUTHOR]

One century after the first description of rickettsiae as human pathogens, the rickettsiosis remained poorly understood diseases. These microorganisms are indeed characterized by a strictly intracellular location which has, for long, prohibited their detailed study. Within the last ten years, the completion of the genome sequences of several strains allowed gaining a better knowledge about the molecular mechanisms involved in rickettsia pathogenicity. Here, we summarized available data concerning the critical steps of rickettsia-host cell interactions that should contribute to tissue injury and diseases, that is, adhesion, phagosomal escape, motility, and intracellular survival of the bacteria. [ABSTRACT FROM AUTHOR]

Mediterranean spotted fever is the most important tick-borne disease occurring in Southern Europe and North Africa. The first case of this life-threatening zoonosis was reported in 1910. In the 1930s, the role of the brown dog tick, Rhipicephalus sanguineus, and the causative agent, Rickettsia conorii were described. However, basic questions regarding the relationships between the rickettsia and its tick vector are still unresolved, and the life cycle of R. conorii is incompletely known. There is a lack of knowledge associated with the role of Rh. sanguineus in the maintenance and transmission of R. conorii. The infectious rate of Rh. sanguineus ticks with R. conorii has been found low every time it has been tested; usually lower that 1%. The deleterious impact of R. conorii on ticks has been suggested in experimental models as a potential reason to explain a low prevalence in nature. The long-recognized phenomenon known as reactivation has been suggested as a cause of negative effects – that is, the change in temperature and physiology of the tick host induces the agent to emerge from dormancy and attain infectivity with bad effects on ticks. However, naturally infected colonies of ticks have been maintained in laboratory conditions over several generations. We discuss here several aspects that have been recently studied to better understand Rh. sanguineus– R. conorii relationships, including comparison between the fitness of infected and non-infected ticks in laboratory conditions and the role of external factors such as temperature and starvation. [ABSTRACT FROM AUTHOR]

The article discusses the case of a patient who presented with fever, weight loss, diarrhea and adenopathy. He had a history of severe destructive polyarthritis. Duodenal biopsy showed a periodic acid-Schiff-positive macrophages. He was found positive for Tropheryma whipplei in synovial fluid, synovial tissue, and lymph node specimens, and Whipple's disease was confirmed. He was given antiobiotic and his symptoms stopped.

This overview summarize the salient features of advances in the epidemiology, vectors, and clinical and laboratory diagnoses of rickettsiology. Presentations on veterinary rickettsiology highlight the importance of the rickettsiae in animal husbandry. [ABSTRACT FROM AUTHOR]

Atypical pneumonia is a term applied to lower respiratory tract infections that are not characterized by signs and symptoms of lobar consolidation. This article will discuss the epidemiology, clinical manifestations, and laboratory diagnoses of Mycoplasma pneumoniae, Chlamydia sp., Legionella sp., Francisella tularensis, and Coxiella burnetii, which are the agents most commonly associated with atypical pneumonia. Because many of these pathogens are intracellular, diagnosis depends upon serological confirmation. The current serological tests used to identify these agents in the etiologic diagnosis of atypical pneumonia are described. Recently, however, it has become possible to make a diagnosis directly in these cases using DNA or protein microarrays. Here, we describe the development of a new, automated technique for simultaneous testing and detection of several pathogens using a multiplexed serology test. This should prove to be a valuable tool for the rapid determination of patient status, allowing effective and efficient postexposure prophylaxis and treatment. [ABSTRACT FROM AUTHOR]

Rickettsia felis is an obligate intracellular Gram‐negative bacterium which causes flea‐borne spotted fever in humans. In the past decades, R. felis has been detected worldwide in Ctenocephalides felis fleas and various other arthropods. However, due to its shared symptoms with other common vector‐borne diseases, human infection is prone to be underestimated or misdiagnosed, especially in the malaria‐endemic areas including sub‐Saharan Africa, where confirmatory laboratory diagnoses are not usually available. In this study, a 'One Health' approach was adopted to explore potential vector‐borne and zoonotic pathogens in the Democratic Republic of Sao Tome and Principe (DRSTP), an island nation in the Gulf of Guinea. By collaborating with local veterinarians, 1,187 fleas were collected from 95 domestic dogs across the country and later identified as Ct. felis using taxonomic keys. A cytochrome oxidase gene‐based phylogenetic analysis revealed that all collected fleas belonged to a single haplotype and were identical to isolates from Ivory Coast and Brazil that clustered into a clade of tropical distribution. Additional samples of 14 chigoe fleas (Tunga penetrans) were collected from the surrounding environment of the dogs' resting spots. Rickettsia felis infection in fleas was examined by molecular methods targeting the citrate synthase (gltA)‐ and outer membrane protein A (ompA)‐coding genes as well as the R. felis‐specific pRF plasmid. The bacterial DNA was detected in 21.01% (146/695) of cat fleas but none of the chigoe fleas. Microimmunofluorescence assay was then performed to assess pathogen exposure of the residents. Of 240 dried blood spots from participants with dog contacts, 8 (3.33%) exhibited R. felis antibodies. Our findings demonstrated the presence of R. felis in DRSTP. Further extensive epidemiological studies regarding its prevalence and its role in causing febrile illness while the nation is entering pre‐elimination stage of malaria will be carried out. [ABSTRACT FROM AUTHOR]

Objectives: Depicting past epidemics currently relies on DNA‐based detection of pathogens, an approach limited to pathogens with well‐preserved DNA sequences. We used paleoserology as a complementary approach detecting specific antibodies under a mini line‐blot format including positive and negative control antigens. Methods: Mini line blot assay incorporated skim milk as negative control, Staphylococcus aureus as positive control, and antigens prepared from lice‐borne pathogens Rickettsia prowazekii, Borrelia recurrentis, Bartonella quintana, and Yersinia pestis. Paleoserums were extracted from rehydrated dental pulp recovered from buried individuals. Mini line blots observed with the naked eye, were quantified using a scanner and appropriate software. Paleoserology was applied to the indirect detection of lice‐borne pathogens in seven skeletons exhumed from a 16th–17th century suspected military burial site (Auxi‐le‐Château); and 14 civils exhumed from a 5th–13th century burial site (Saint‐Mont). Direct detection of pathogens was performed using quantitative real‐time PCR. Results: In Auxi‐le‐Château, paleoserology yielded 7/7 interpretable paleoserums including 7/7 positives for B. recurrentis including one also positive for B. quintana. In Saint‐Mont, paleoserology yielded 8/14 interpretable paleoserums and none reacted against any of the four pathogens. Antibodies against R. prowazekii and Y. pestis were not detected. The seroprevalence was significantly higher in the military burial site of Auxi‐le‐Château than in the civil burial site of Saint‐Mont. Real‐time PCR detection of B. quintana yielded 5/21 positive (3 at Saint‐Mont and 2 at Auxi‐le‐Château) whereas B. recurrentis was not detected. Conclusions: Paleoserology unmasked an outbreak of relapsing B. recurrentis fever in one 16th – 17th century military garrison, missed by real‐time PCR. Paleoserology offers a new tool for investigating past epidemics, in complement to DNA sequence‐based approaches. [ABSTRACT FROM AUTHOR]

A serological study was carried out in two Senegalese villages located in the Sine‐Saloum region in order to estimate the presence of anti‐leptospiral antibodies in humans and animals, and to identify the predominant serogroups. Seven hundred and forty‐nine serum samples were collected from humans (n = 545), dogs (n = 33), donkeys (n = 20), goats (n = 52), sheep (n = 43) and N'Dama cattle (n = 56), all originated from Dielmo and Ndiop villages. All samples were tested for different serovars of pathogenic Leptospira species by the microscopic agglutination test. Considering titres ≥ 1:100, 7.7% [CI 95:5.5 to 9.9] on the 545 human blood samples tested and 42.2% [CI95:35.4 to 48.9] on the 204 animal blood samples tested were found to be positive to one or more serovars. The results obtained indicate that the Australis serogroup is the most prevalent serogroup in human (67.3%) and cattle (27.3%). Serogroup Icterohaemorhagiae is the most frequent serogroup in goat (55.6%) and donkey (37.5%). Canicola (23.4%), Icterohaemorhagiae (21.1%) and Australis (12.5%) serogroups are the most prevalent serogroups in dogs. This study shows that diverse Leptospira serovars occur in a wide range of wild and domestic mammal species, as well as in humans in Senegal. However, further studies are needed to better understand the complexity of Leptospira epidemiology in Africa, identify the reservoirs of different serogroups and estimate its impact on livestock. Understanding the multi‐host epidemiology of leptospirosis is essential to control and prevent the disease. [ABSTRACT FROM AUTHOR]

Bacterial resistance to antibiotics is a serious medical and public health concern worldwide. Such resistance is conferred by a variety of mechanisms, but the extensive variability in levels of resistance across bacteria is a common finding. Understanding the underlying evolutionary processes governing this functional variation in antibiotic resistance is important as it may allow the development of appropriate strategies to improve treatment options for bacterial infections. The main objective of this study was to examine the functional evolution of β‐lactamases, a common mechanism of enzymatic resistance that inactivates a widely used class of antibiotics. We first obtained β‐lactamase protein sequences and minimal inhibitory concentration (MIC), a measure of antibiotic function, from previously published literature. We then used a molecular phylogenetic framework to examine the evolution of β‐lactamase functional activity. We found that the functional activity of antibiotic resistance mediated by β‐lactamase has evolved in a convergent manner within molecular classes, but is not associated with any single amino acid substitution. This suggests that the dynamics of convergent evolution in this system can vary between the functional and molecular (sequence) levels. Such disassociation may hamper bioinformatic approaches to antibiotic resistance determination and underscore the need for (less efficient but more effective) activity assays as an essential step in evaluating resistance in a given case. [ABSTRACT FROM AUTHOR]

Background: The transfusion of platelet concentrates (PCs) contaminated with bacteria may cause serious, and even fatal, septic reactions in patients. The aim of this study was to compare the VersaTREK with the BACTEC FX automated culture systems for screening bacterial contamination, directly after the delay of 24 hours of preparation to obtain the final pooled buffy coat PCs, to prevent transfusion-transmitted bacterial infections.Study Design and Methods: Seven bacterial strains were each inoculated into five replicate pooled buffy coat PCs at approximately 100 colony-forming units/unit, and 5- or 10-mL samples were inoculated into duplicate aerobic culture bottles. The time and detection rates were compared between BACTEC FX, as a reference method, and VersaTREK.Results: Time to detection was significantly shorter using VersaTREK for most species detected by both systems for the volumes tested. Of 70 VersaTREK cultures, 69 (98.57% detection rate) were positive after 24 hours of incubation with the 5-mL sample. In contrast, the BACTEC FX system detected all positive samples in PCs for the volume of 10 mL, although seven samples were false negatives for the 5-mL volume.Conclusion: The VersaTREK system compared favorably to the BACTEC FX system for 5-mL volumes (p < 0.05) and could be considered a potential method for detecting bacterial contamination in PC samples directly after 24 hours of preparation of the final pooled buffy coat PCs. [ABSTRACT FROM AUTHOR]

Abstract: Dogs are competent reservoir hosts of several zoonotic agents, including Filariidae nematodes and Anaplasmataceae family bacteria. The latter family unites human and veterinary pathogens (Anaplasma, Ehrlichia and Neorickettsia bacteria) with Wolbachia, some of which are obligatory endosymbionts of pathogenic filarial nematodes. The epidemiology of Anaplasmataceae and Filariidae species infecting dogs living in kennels in New Caledonia was studied. 64 EDTA blood samples were screened for the presence of Anaplasmataceae and filarial nematodes. Molecular study was conducted using primers and probe targeting the of 23S rRNA long fragment of Anaplasmataceae species. Next, all blood sample was screened for the presence of Filariidae species targeting the primers and probe targeting the COI gene, as well as primers targeting the COI and 5S rRNA genes of all filarial worms. Anaplasma platys was identified in 8/64 (12.5, 95% confidence interval [CI]: 4.4–20.6%) and Wolbachia endosymbiont of Dirofilaria immitis in 8/64 (12.5%, CI: 4.4–20.6%). Filariidae species investigation was performed and showed that 11/64 (17.2%, CI: 7.9–26.4%) dogs were infected with D. immitis, whereas, 2/64 (3.1%, CI: 0.0–7.3%) were infected with Acanthocheilonema reconditum. Finally, we checked the occurrence of co‐infection between Anaplasmataceae and Filariidae species. Co‐occurrence with Wolbachia endosymbiont of D. immitis was observed in seven dogs, one dog was co‐infected with A. platys and A. reconditum and another was co‐infected with Wolbachia endosymbiont of D. immitis and A. reconditum. These results are the first report of Anaplasmataceae and Filariidae occurring in dogs in New Caledonia. [ABSTRACT FROM AUTHOR]

Background: Platelets (PLTs) are especially affected by the risk of bacterial contamination. Matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) is an accurate method for the routine identification of bacterial isolates in microbiology laboratories. We directly applied the MALDI-TOF method to bacterial detection in PLTs. In this study, we evaluated the sensitivity, specificity, and speed of a direct MALDI-TOF approach compared to the conventional method BACTEC.Study Design and Methods: Eight bacteria associated with PLT contamination, cited by the ISBT on transfusion-transmitted infectious diseases, were spiked into PLTs for a final concentration of approximately 100 CFU/bag (n = 5 for each strain). The PLTs were then agitated for 24 hours. One milliliter of PLTs was incubated in a shaker incubator for 8 hours at 37°C with 1 mL of trypticase soy broth (TSB). The spectra were analyzed using the MALDI Biotyper software. As a control, 8 mL of PLTs incubated into BACTEC bottles and a positive bottle were subcultured to ensure identification of bacterial growth.Results: Regardless of the strain of PLTs tested, MALDI-TOF analysis made detection and early identification possible at 8 hours. Analysis by BACTEC of PLTs infected with Escherichia coli, Bacillus cereus, and Providencia stuartii made early identification possible. For the remaining bacteria, the detection time by BACTEC was significantly longer than 8 hours.Conclusion: We demonstrated the possibility of detecting bacteria in PLTs using a standardized culture step in TSB with MALDI-TOF, regardless of the strain, with the same specificity and analytical sensitivity and with a time to results of 12 hours. This direct method presented rapid and reliable results. [ABSTRACT FROM AUTHOR]

M. genitalium is a reemerging microorganism, responsible for sexually transmissible infections ( STIs), with prevalence which varies depending on the country and population group studied. We report here M. genitalium prevalence among the specimens received for STI diagnosis in our routine microbiological laboratory in the university hospital in Marseille, France. We tested 4 624 samples from 3 793 patients using qPCR for M. genitalium, C . trachomatis, N. gonorrheae, T. pallidum. Of these samples, 528 (13.6%) patients were tested positive for at least one pathogen and 126 (3.3%) were positive for M. genitalium. M. genitalium is the second most prevalent micro-organism detected in women after C. trachomatis (10.4%) and the third most prevalent in men after C. trachomatis (5.1%) and N. gonorrhoeae (4.4%). We observed no significant differences between the prevalence of M. genitalium in vaginal, urethral and urine specimens (p = 0.9). Prevalence of M. genitalium is significantly higher in patients aged between 10-30 years (4.1%) compared to those aged between 30 and 50 years (2.7%) (p = 0.02, RR = 1.54 [1.06-2.24]) and patients over 50 years of age (1.1%) (p = 0.003, RR= 3.98 [1.47-10.8]). M. genitalium is a common agent of STI, therefore we suggest that this micro-organism should be systematically tested during chronic, recurrent, or antibiotic resistant genital infections and in populations at high-risk of STIs. [ABSTRACT FROM AUTHOR]

Central nervous system Whipple's disease ( CNS- WD) with clinically isolated neurological involvement is a rare condition fatal without an early diagnosis. We aimed to present clinical and neuropathological features of three cases of pre- or post-mortem polymerase chain reaction confirmed CNS- WD with distinct clinical presentation, outcome and pathological findings. One patient had an acute onset with spinal and brainstem involvement and died without CNS- WD diagnosis after 14 weeks. Neuropathology showed extensive inflammatory and necrotizing lesions with abundant foamy periodic-acid- Schiff ( PAS)+ macrophages. The second patient had a subacute evolution with late CNS- WD diagnosis and death occurring 18 months after onset despite antibiotic treatment. Brain examination showed inflammatory lesions in the brainstem, thalamus and cerebellum, and abundant foamy PAS+ macrophages. The third case was diagnosed within 4 weeks of onset and treated with an excellent response. He died after a disease-free period of 24 months of unrelated causes. Neuropathology showed cystic residual lesions devoid of microorganisms without inflammatory reaction. CNS- WD may have an acute or subacute course with variable response to treatment. Accordingly, subjacent lesions may be those of a severe acute necrotizing encephalitic process or subacute inflammatory lesions involving diencephalic, brainstem, cerebellar and spinal regions. Chronic, cavitary brain lesions may be sequelae of a successful treatment. Early diagnosis should allow appropriate treatment and improve prognosis. [ABSTRACT FROM AUTHOR]

Bartonella quintana is transmitted by the infected faeces of body lice. Recently, this bacterium was detected in cat fleas ( Ctenocephalides felis) and in two humans with chronic adenopathy whose only risk factor was contact with cat fleas. In this study, a total of 960 C. felis were divided into 12 groups (2 control groups and 10 infected groups) each containing 80 fleas. The fleas were fed B. quintana-inoculated human blood at different dilutions (≈3.6 × 104 − 8.4 × 109 bacteria) for 4 days via an artificial membrane. Subsequently, all flea groups were fed uninfected blood until day 13 postinfection (dpi). On day 3 pi, B. quintana was detected with two specific genes by quantitative PCR in 60-100% of randomly chosen fleas per dilution: 52% (26/50) in the infected fleas in Trial 1 and 90% (45/50) of the fleas in Trial 2. B. quintana was also identified by molecular and culture assays in flea faeces. The average number of B. quintana as determined by q PCR decreased until the 11th dpi and was absent in both trials at the 13th dpi. Bacteria were localized only in the flea gastrointestinal gut by specific immunohistochemistry. Our results indicate that cat fleas can acquire B. quintana by feeding and release viable organisms into their faeces. Therefore, fleas may play a role as vectors of trench fever or other clinical manifestations that are caused by B. quintana. However, the biological role of C. felis in the transmission of B. quintana under natural conditions is yet to be defined. [ABSTRACT FROM AUTHOR]

Until now, Planctomycetes bacteria were considered as environmental organisms. Nevertheless, some studies detected Planctomycetes DNA from human gut. We therefore explored the human gut Planctomycetes content. Planctomycetes-specific PCR primers were designed to amplify a 240-bp 16S rRNA gene fragment in human stool specimens from individuals in France and in Senegal and from endocarditis patients receiving antibiotics in France. PCR products were then cloned and sequenced. PCR detection revealed a significantly higher prevalence (1.8% vs 0.4%, p = 0.05) and higher diversity (62 vs 6 phylotypes, p = 0.02) of Planctomycetes 16S rRNA gene in stool specimens collected in Senegal than in France. Also, stool specimens from endocarditis patients exhibited non-significantly higher prevalence (0.6% vs 0.4%) and the ratio of phylotypes by positive patient (3 vs 1.5) than those collected from untreated French individuals. Gemmata sp. related sequences were found in 6/12 individuals. Planctomycetes organisms are a part of the human digestive tract microbiota. Their diversity varied by environment including the geographical origin of the individual and antibiotics treatment. [ABSTRACT FROM AUTHOR]

Several bacteria, viruses, and parasites cause diarrhea as coinfecting pathogens. We designed a DNA microarray comprising 60-bp probes spotted 194 times for the multiplex detection of 33 enteropathogenic bacteria and seven enteropathogenic viruses, and the archaeon Methanobrevibacter smithii was used as an internal positive control. Nine pathogen-free stool specimens were used as negative controls. One of these control specimens was further spiked with Salmonella enterica as a positive control. The microarray was then tested with 40 pathological stool specimens, comprising S. enterica (n = 30), Campylobacter jejuni (n = 4), pathogenic Escherichia coli (n = 2), and adenovirus (n = 4). M. smithii was detected in 47/49 (95.9%) specimens, no pathogen was detected in negative controls and S. enterica was identified in the S. enterica-spiked positive control. The overall specificity was 100% and the overall sensitivity was 97.5% because one S. enterica sample was missed by the microarray. The multiplexed detection of C. jejuni spiked into an adenovirus-positive stool sample gave positive results, with fluorescence values of 14.3 and 9.1, respectively. These data indicate that using the protocol developed in this article, the DNA array allows for the multiplexed detection of some enteropathogens in stool samples. [ABSTRACT FROM AUTHOR]

Giant viruses of the Megavirales order have been recently isolated from aquatic environments and have long been neglected because they are removed from samples during viral purification for viral metagenomic studies. Due to bacterial overgrowth and susceptibility to high concentrations of antibiotics, isolation by amoeba co-culture has a low efficiency and is highly time-consuming. Thus, few environments have been exhaustively investigated to date, although the ubiquitous distribution of the Acanthamoeba sp. suggests that these viruses could also be ubiquitous. In this work, we have implemented a high-throughput method to detect amoebae lysis on agar plates that allows the testing of hundreds of samples in a few days. Using this procedure, a total of 11 new Marseilleviridae strains and four new Mimiviridae strains, including a virus infected with a virophage, were isolated from 1000 environmental samples from Tunisia. Of these, four corresponded to new genotypic variants. These isolates are the first African environmental isolates identified from these two families, and several samples were obtained from a hypersaline aquatic environment. These results demonstrate that this technique can be used for the evaluation and characterization of large collections of giant viruses to provide insight into understanding their ecology. [ABSTRACT FROM AUTHOR]

Carbapenem-resistant Acinetobacter baumannii infections are a worldwide endemic nosocomial threat. Between December 2010 and April 2011, an increase of carbapenem-resistant A. baumannii infections occurred in several Marseille University Hospitals. The aim of this study was to investigate the increase of carbapenem-resistant A. baumannii infections and to characterize the mechanisms of carbapenem resistance. The increase was detected by a homemade computer surveillance program, known as EPIMIC, that monitors antibiotic resistance profiles on a weekly basis. During this period, positive samples of carbapenem-resistant A. baumannii were retrieved from patients hospitalized in different units. Genotyping of the isolates was performed using pulsed-field gel electrophoresis ( PFGE) and multi-locus sequence typing ( MLST), and carbapenemase gene analyses were performed to detect the presence of carbapenemases and to determine the relationships of the isolates. Carbapenem-resistant A. baumannii were isolated in a total of 11 patients who were hospitalized in different hospitals units. We identified the presence of the blaOXA23-like carbapenemase-encoding gene in all of the isolates and found four major PFGE groups and different MLST groups. These results demonstrate a current evolution in the A. baumannii epidemiology in Marseille with a switch from an epidemic situation to an endemic situation and with several circulating clones. [ABSTRACT FROM AUTHOR]

A 16S rDNA sequence-based investigation of methanogenic Archaea in the human stools found Methanobrevibacter smithii in 99.2% and Methanosphaera stadtmanae in 32.6%. The recently described Methanomassiliicoccus luminyensis found by others to be representative of a new order of methanogenic Archaea was found in 4% of stool specimens. The prevalence of M. luminyensis significantly increased with age, contrary to M. smithii and M. stadtmanae. [ABSTRACT FROM AUTHOR]

A correction to the article "Rickettsioses: From Genome to Proteome, Pathobiology, and Rickettsiae As an International Threat," by Karim E. Hechemy and colleagues that was published in a previous issue of the journal is presented.

Historical and anthropological data suggest that skeletons excavated from an 11th to 15th century mass grave in Bondy, France, may be those of victims of the Great Plague. Using high-throughput real-time PCR investigation of the dental pulp collected from 14 teeth from five such skeletons, we detected Bartonella quintana DNA in three individuals and Yersinia pestis DNA in two individuals. DNA from five other deadly pathogens was not found. Suicide PCR genotyping confirmed Y. pestis DNA belonging to the Orientalis biotype. One individual had co-infection. These data suggest a plague epidemic in a population already infected by the body louse-transmitted B. quintana or a body louse-driven transmission of the plague that drove a medieval epidemic in inland Europe. Am J Phys Anthropol, 2011. © 2011 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]

Coxiella burnetii, the agent of Q fever, is an obligate intracellular bacterium that is considered a potential biological weapon of category B. C. burnetii survives within myeloid cells by subverting receptor-mediated phagocytosis and preventing phagosome maturation. The intracellular fate of C. burnetii also depends on the functional state of myeloid cells. This review describes the mechanisms used by C. burnetii to circumvent uptake and trafficking events, and the role of cytokines on C. burnetii survival in myeloid cells. [ABSTRACT FROM AUTHOR]

During our more than 20 years of monitoring, we have used epidemiological, field, and experimental methods for characterization of natural foci of tick-borne rickettsioses in Russia. The main results were obtained through genetic methods (PCR sequence) at the Université de la Mediterranée (Marseille, France). We describe considerable heterogeneity of tick-borne α1-proteobacteria: 16 microorganisms the of the order Rickettsiales were detected in Russia and Kazakhstan. R. sibirica–caused North Asiatic tick-borne rickettsiosis is the main tick-borne rickettsiosis in Russia, with wide distribution in Siberia and the Russian Far East and high epidemic activity of natural foci of different landscape types. Our results show circulation of different pathogenic rickettsiae in the same endemic territories. In the Far East region, R. sibirica R. sibirica, R. sibirica BJ-90, and R. heilongjiangensis were detected; in the Altay and Krasnojarsk regions, R. sibirica R. sibirica and R. heilongjiangensis; and in the Kurgan district of West Siberia, R. sibirica R. sibirica and R. slovaca. The roles of more than 15 new genotypes of α1-proteobacteria in infectious disease in Russia and Kazakhstan are in need of further study. [ABSTRACT FROM AUTHOR]

We recently reported the first documented cases of a new rickettsial disease caused by Rickettsia heilongjiangensis in the Russian Far East (Far Eastern tick-borne rickettsiosis). Here we report the amplification of DNA of R. heilongjiangensis from both the skin biopsy of an acutely ill patient and the tick removed from him prior to the disease development. The tick has been identified as Haemaphysalis spp. The clinical picture was that of a spotted fever group rickettsiosis and a seroconversion was noted with R. heilongjiangensis antigen . Screening testing of both species of Haemaphysalis ticks inhabiting Russian Far Eastern regions showed that up to 28.13% of H. concinnae and 4.48% of H. japonica douglasii ticks harbor R. heilongjiangensis. It has been concluded that H. concinnae may serve as the main vector for the transmission of R. heilongjiangensis. H. japonica douglasii ticks harbor several varieties of rickettsiae. DNA of “ Candidatus Rickettsia tarasevichiae,” previously found in Ixodes persulcatus ticks, was amplified from one male tick. Two sequenced complete gltA genes belong to the novel spotted fever group rickettsial species provisionally called here “ Candidatus ” variants Hjd54 and Hjd61. The rate of infection has been found to be not higher than 1.5%. [ABSTRACT FROM AUTHOR]

A series of sudden unexpected cardiac deaths among Swedish elite orienteerers in the 1980s have resulted from the combination of infectious diseases and physical exercise. Studies in the late 1990s have pointed to Chlamydia and Barontella, which both had a high seroprevalence among Swedish elite orienteerers. We conducted a case–control study aimed to elucidate the serologic prevalence of rickettsial diseases among Danish elite orienteerers. Ticks are known as vectors for some rickettsial diseases. None of the orienteerers had a positive antibody titer against any of the tested Rickettsia despite a very high frequency of tick bites in this group. [ABSTRACT FROM AUTHOR]

The presumptive cases of Mediterranean spotted fever have been identified in 1993 and since that time, its frequency has steadily increased. The prospective study, in summer 2004, was conducted in order to present the descriptive clinic and epidemiology, to identify more severe forms, the presence of the multiple eschars, and different rickettsial strains caused the disease in our region. In Oran, the cases were diagnosed clinically. In Marseille, serum specimens were tested by IFA using the panel of eight rickettsial antigen; Western blot and cross-adsorption studies were also performed in order to confirm the diagnosis. Ninety-three patients clinically diagnosed were recorded from July 3 to October 28, 2004. Eighty percent were male, the mean age was 44.3 years, 90% were exposed to dog and 32% reported tick bites. Clinical signs were as follow: presence of underlying disease (44%), sudden onset (78%), fever (100%), loss of weight (63%), conjunctivitis (43%), and a tache noire was noticed in 70%. Interestingly, two patients had two and three eschars, respectively. The rash was maculopapular (palm and sole) and purpuric in nine cases. Doxycycline was the most antibiotic (91%) with favourable outcome in 91% of the cases. Malignant form with death is reported for three patients (3.2%). Among the 93 patients, 104 serum from 65 patients were tested (serums of others patients were lost or ticket not found on tube. Sixty-three patients out of 65 had a positive serology by IFA with cross-reactive antibodies especially between R. conorii, R. felis and/or R. typhi. Two others negative serology were: one precocious serum and second from the patient, which presented symptoms of MSF and tested two serums, Western blot and cross-adsorption. [ABSTRACT FROM AUTHOR]

Rhipicephalus turanicus tick colony infected in the laboratory with Rickettsia massiliae showed that the rickettsia is transovarially and transdatially tramsmitted. The expression of rOmpB did not change with temperature or the stages of the tick life cycle. In contrast, rOmpA was less expressed during the larval stage. [ABSTRACT FROM AUTHOR]

Ticks collected in Northern Algeria between May 2001 and November 2003 were tested by PCR for the presence of Rickettsia spp. DNA using primer amplifying gltA and OmpA genes. Three different spotted fever group rickettsias were amplified from these ticks: R. Conorii subsp. P. conorii strain Malish in Rhipicephalus sanguineus, R. aeschlimannii in Hyalomma marginatum, and R. massiliae in Rhipicephalus turanicus. Our results confirm the presence of R. conorii in ticks in Algeria and provide the first detection of R. aeschlimannii and R. massiliae in Algeria. [ABSTRACT FROM AUTHOR]

A total of 395 adult ixodid ticks from three genera ( Dermacentor, Haemaphysalis, and Ixodes) collected from the Urals to the Far East of Russia were tested by PCR and sequencing for the presence of spotted fever rickettsiae, anaplasmae, and ehrlichiae. Four, pathogens recognized in humans were detected in ticks: Rickettsia sibirica, R. heilongjiangensis, R. helvetica, and Anaplasma phagocytophilum. In addition, rickettsiae and ehrlichiae of unknown pathogenicity were detected, including Rickettsia sp. RpA4, Rickettsia sp. DnS14, Rickettsia sp. DnS28, “ Candidatus R. tarasevichiae,” a rickettsia closely related to R. helvetica, A. bovis, Ehrlichia muris, “ Ehrlichia-like”“Schotti variant,” and bacterium “Montezuma.” Our findings indicated the distribution of rickettsiae and ehrlichiae in hard ticks in Russia. [ABSTRACT FROM AUTHOR]

The Rickettsia sibirica species is composed of isolates that are genotypically close but can be classified within two distinct serotypes, that is, R. sibirica sensu stricto and R. sibirica mongolitimonae (incorrectly named R. mongolotimonae). We investigated the possibility of classifying rickettsiae closely related to R. sibirica as R. sibirica subspecies, as proposed by the ad hoc Committee on Reconciliation of Approaches to Bacterial Systematics. For this, we first estimated the genotypic variability by using multilocus sequence typing (MLST), including the sequencing of five genes, and multispacer typing (MST) using three intergenic spacers, of five isolates and three tick amplicons of R. sibirica sensu stricto and six isolates of R. sibirica mongolotimonae. Then, we selected a representative of each MLST genotype and used mouse serotyping to estimate their degree of taxonomic relatedness. Among the 14 isolates or tick amplicons studied, 2 MLST genotypes were identified: (i) the R. sibirica sensu stricto type; and (ii) the R. sibirica mongolitimonae type. Representatives of the two MLST types were classified within three MST types and into two serotypes. Therefore, as isolates within the R. sibirica species are genotypically homogeneous but show MST genotypic, serotypic, and epidemio-clinical dissimilarities, we propose to modify the nomenclature of the R. sibirica species through the creation of subspecies. We propose the names R. sibirica subsp. sibirica subsp. nov. (type strain = 2-4-6, ATCC VR-541T), and R. sibirica subsp. mongolitimonae subsp. nov. (type strain = HA-91, ATCC VR-1526T). The description of R. sibirica is emended to accommodate the two subspecies. [ABSTRACT FROM AUTHOR]

The presence of R. prowazekii in lice using R. prowazekii-specific qPCR cassettes showed the changes of the rickettsial burden in the lice post-infection. [ABSTRACT FROM AUTHOR]

In 2004 between the months of May–November, 11 patients with spotted fever group (SFG) rickettsioses were admitted to the Trakya University Hospital in Edirne, Turkey. SFG rickettsioses were diagnosed clinically. Before treatment, punch biopsy from skin lesions, especially from the eschar, was performed. Serum specimens were tested by IFA using a panel of nine rickettsial antigens, including SFG rickettsiae and R. typhi. Western blotting and standard PCR were also performed. The average age of the 11 patients (4 male and 7 female) was 51 years. All the patients had high fever; 10 (91%) had maculopapular rash; 8 (73%) had rash in the palms or on the soles. Five patients had a unique eschar; two had double eschars (64%). Two patients presented with multiple organ failure and one of them died. All the patients had significant antibody titers against SFG rickettsiae. PCR experiments of skin biopsies were positive in six (60%) of 10 skin biopsy samples and DNA sequencing of the positive PCR products gave 100% homology with Rickettsia conorii Malish 7 for opmA and gltA. Trakya Region in an endemic area for rickettsioses. In this series, three patients presented with life-threatening diseases and one of them died. This patient was the first fatal case (2.8%). Atypic and serous life-threatening presentations of rickettsioses must be kept in mind for the differential diagnosis of febrile disease in Turkey. [ABSTRACT FROM AUTHOR]

The article presents a case study of an 18-month-old boy with intermittent painful limp who was at the time of his birth followed by physical examination which resulted in no fever, mild tenderness and swelling of the left knee. The boy showed signs of lytic lesion in the epiphysis with no chest abnormality. The article discusses limp infection and a empty area was deducted through Magnetic resonance imaging.

To the best of our knowledge, we report the first case of pre‐transplant unrecognized disseminated Coxiella burnetii infection, unmasked in the post‐transplant period leading to both heart and kidney allograft dysfunction. A 59 year old man with a history of simultaneous heart‐kidney transplantation due to end stage heart failure from severe aortic regurgitation (AR) and cryoglobulinemic immune complex mediated concentric necrotizing glomerulonephritis (GN), presents with a history of intermittent fevers and fatigue. Prior to transplantation he was treated for multiple episodes of culture negative endocarditis requiring bio‐prosthetic valve replacement. Evaluation of fever included a transesophageal echocardiogram (TEE) that revealed a large hyperechoic mass on the anterior mitral leaflet with perforation, severe mitral regurgitation and moderate AR. Blood cultures were negative at that time. Owing to development of allograft mitral and aortic valve insufficiency, he underwent allograft bio‐prosthetic mitral valve (MV) replacement and aortic valvuloplasty 2 years following his transplantation. Pathologic examination of the allograft mitral valve demonstrated fibrinopurulent exudate with degenerating bacterial organisms, consistent with vegetation and myxoid degenerative changes. Due to a high suspicion for native heart C. burnetii prosthetic valve endocarditis prior to transplantation, we re‐evaluated the native explanted heart histopathology, as well as the explanted allograft MV. Cardiac allograft and native MV were positive for C. burnetii by real‐time PCR. C. burnetii serology was consistent with persistent infection as well. [ABSTRACT FROM AUTHOR]

Blood culture–negative endocarditis is due to fastidious bacteria, including Coxiella burnetii and Bartonella spp. Diagnosis of such infection relies on serology and microimmunofluorescence is therefore the reference method. We developed a multiplex serology test featuring automatic incubation and reading and incorporating internal controls. Preliminary results indicate that this new serologic test is valuable for the rapid, automated serological diagnosis of blood culture–negative endocarditis. [ABSTRACT FROM AUTHOR]

A foreword to the journal "Annals of the New York Academy of Sciences" is presented.