Body lice of homeless people reveal the presence of several emerging bacterial pathogens in northern Algeria.

BACKGROUND: Human lice, Pediculus humanus, are obligate blood-sucking parasites. Body lice, Pediculus h. humanus, occur in two divergent mitochondrial clades (A and D) each exhibiting a particular geographic distribution. Currently, the body louse is recognized as the only vector for louse-borne diseases. In this study, we aimed to study the genetic diversity of body lice collected from homeless populations in three localities of northern Algeria, and to investigate louse-borne pathogens in these lice. METHODOLOGY/PRINCIPAL FINDINGS: In this study, 524 body lice specimens were collected from 44 homeless people in three localities: Algiers, Tizi Ouzou and Boumerdès located in northern Algeria. Duplex clade specific real-time PCRs (qPCR) and Cytochrome b (cytb) mitochondrial DNA (mtDNA) analysis were performed in order to identify the mitochondrial clade. Screening of louse-borne pathogens bacteria was based on targeting specific genes for each pathogen using qPCR supplemented by sequencing. All body lice belong to clade A. Through amplification and sequencing of the cytb gene we confirmed the presence of three haplotypes: A5, A9 and A63, which is novel. The molecular investigation of the 524 body lice samples revealed the presence of four human pathogens: Bartonella quintana (13.35%), Coxiella burnetii (10.52%), Anaplasma phagocytophilum (0.76%) and Acinetobacter species (A. baumannii, A. johnsonii, A. berezeniae, A. nosocomialis and A. variabilis, in total 46.94%). CONCLUSIONS/SIGNIFICANCE: To the best of our knowledge, our study is the first to show the genetic diversity and presence of several emerging pathogenic bacteria in homeless' body lice from Algeria. We also report for the first time, the presence of several species of Acinetobacter in human body lice. Our results highlight the fact that body lice may be suspected as being a much broader vector of several pathogenic agents than previously thought. Nevertheless, other studies are needed to encourage epidemiological investigations and surveys of louse-associated infections.

Detection of bacterial pathogens including potential new species in human head lice from Mali.

In poor African countries, where no medical and biological facilities are available, the identification of potential emerging pathogens of concern at an early stage is challenging. Head lice, Pediculus humanus capitis, have a short life, feed only on human blood and do not transmit pathogens to their progeny. They are, therefore, a perfect tool for the xenodiagnosis of current or recent human infection. This study assessed the occurrence of bacterial pathogens from head lice collected in two rural villages from Mali, where a high frequency of head lice infestation had previously been reported, using molecular methods. Results show that all 600 head lice, collected from 117 individuals, belonged to clade E, specific to West Africa. Bartonella quintana, the causative agent of trench fever, was identified in three of the 600 (0.5%) head lice studied. Our study also shows, for the first time, the presence of the DNA of two pathogenic bacteria, namely Coxiella burnetii (5.1%) and Rickettsia aeschlimannii (0.6%), detected in human head lice, as well as the DNA of potential new species from the Anaplasma and Ehrlichia genera of unknown pathogenicity. The finding of several Malian head lice infected with B. quintana, C. burnetii, R. aeschlimannii, Anaplasma and Ehrlichia is alarming and highlights the need for active survey programs to define the public health consequences of the detection of these emerging bacterial pathogens in human head lice.

Development of fluorogenic probe-based and high-resolution melting-based polymerase chain reaction assays for the detection and differentiation of Bartonella quintana and Bartonella henselae.

Bartonella henselae and Bartonella quintana are the major etiological agents of infective endocarditis, which pose a serious threat to human health. To simultaneously detect and differentiate B. henselae and B. quintana, a reliable and fast method to simultaneously detect and differentiate B. henselae and B. quintana is required. In this study, we developed and validated two rapid, highly sensitive and specific, duplex, real-time polymerase chain reaction (PCR) assays-one based on high-resolution melting (HRM) analysis, and the other on TaqMan probes-to simultaneously detect and differentiate B. henselae and B. quintana. The sensitivity of developed assays were found 100 times more sensitive than that of conventional PCR. The specificity of the assays were validated by the absence of any cross reaction with the other Bartonella species, non-Bartonella bacteria and other animals. The results indicate that the duplex HRM-based and TaqMan probe-based assays have high specificity and sensitivity, and good reproducibility for simultaneous the detection of B. henselae and B. quintana. They are cost-effective, sensitive and reliable methods; and are thus suitable for clinical diagnosis, epidemiological surveys, and disease surveillance.

Complete genome sequence of Bartonella quintana, a bacterium isolated from rhesus macaques.

Bartonella quintana is a re-emerging pathogen and the causative agent of a broad spectrum of disease manifestations in humans. The present study reports the complete genome of B. quintana strain RM_11, which was isolated from rhesus macaques.

Bartonella quintana in Ethiopian lice.

Head and clothing lice from Jimma, Ethiopia were investigated for pathogenic bacteria. Genomic DNA from pools of lice was subjected to PCR analysis for Bartonella spp., Borrelia spp. Coxiella burnetii, Rickettsia spp. and Yersinia pestis. All 102 lice pools were negative for the afore mentioned pathogens, with the exception of Bartonella species found among 6 of 65 (9.2%) head lice pools and1 of 33 clothing lice pools. Identification was achieved by sequencing the ribosomal intragenic transcribed spacer region (ITS), revealing all to be Bartonella quintana. Although established as a clothing louse-borne infection, typically causing chronic bacteraemia, trench fever, bacillary angiomatosis and endocarditis, this has only been rarely reported among head lice. The higher numbers of infected head lice pools compared with clothing lice suggests their competence for maintaining this infection within Ethiopia.

Altitude-dependent Bartonella quintana genotype C in head lice, Ethiopia.

To determine the presence of Bartonella quintana in head and body lice from persons in different locations in Ethiopia, we used molecular methods. B. quintana was found in 19 (7%) genotype C head lice and in 76 (18%) genotype A body lice. B. quintana in head lice was positively linked to altitude (p = 0.014).

Skin diseases associated with Bartonella infection: facts and controversies.

The genus Bartonella is composed of a series of species and subspecies. Ten of them are responsible for human infections. The best-identified diseases are cat scratch disease (B henselae and possibly B clarridgeiae), trench fever (B quintana), bacillary angiomatosis (B quintana and B henselae), and the spectrum of verruga peruana, Carrion disease, and Oroya fever (B bacilliformis). Controversies exist about the implication of a few other microorganisms being involved in these diseases. Several other conditions have been associated with the presence of Bartonella spp, but these observations await confirmation.

Bartonella quintana in body lice and head lice from homeless persons, San Francisco, California, USA.

Bartonella quintana is a bacterium that causes trench fever in humans. Past reports have shown Bartonella spp. infections in homeless populations in San Francisco, California, USA. The California Department of Public Health in collaboration with San Francisco Project Homeless Connect initiated a program in 2007 to collect lice from the homeless to test for B. quintana and to educate the homeless and their caregivers on prevention and control of louse-borne disease. During 2007-2008, 33.3% of body lice-infested persons and 25% of head lice-infested persons had lice pools infected with B. quintana strain Fuller. Further work is needed to examine how homeless persons acquire lice and determine the risk for illness to persons infested with B. quintana-infected lice.

Analysis of the first Australian strains of Bartonella quintana reveals unique genotypes.

Bartonella quintana is increasingly recognized as a cause of clinical disease in various geographical locations. We characterized three Australian strains associated with endocarditis, using established molecular-typing techniques, the 16S/23S rRNA intergenic spacer (ITS) region, and multispacer typing (MST). All strains examined demonstrated novel ITS and/or MST genotypes. Further characterization of Australian strains is required to determine whether there is an association between genotype and geographical location.

Isolation of Bartonella quintana from a woman and a cat following putative bite transmission.

We report here the detection of Bartonella quintana, after putative bite transmission, in pre-enrichment blood cultures from a woman and from two feral barn cats. Prospective molecular epidemiological studies are necessary to characterize the risk of human Bartonella quintana infection following cat bites.

Bartonella quintana characteristics and clinical management.

Bartonella quintana, a pathogen that is restricted to human hosts and louse vectors, was first characterized as the agent of trench fever. The disease was described in 1915 on the basis of natural and experimental infections in soldiers. It is now recognized as a reemerging pathogen among homeless populations in cities in the United States and Europe and is responsible for a wide spectrum of conditions, including chronic bacteremia, endocarditis, and bacillary angiomatosis. Diagnosis is based on serologic analysis, culture, and molecular biology. Recent characterization of its genome allowed the development of modern diagnosis and typing methods. Guidelines for the treatment of B. quintana infections are presented.

Evidence for louse-transmitted diseases in soldiers of Napoleon's Grand Army in Vilnius.

BACKGROUND: Many soldiers in Napoleon's Grand Army died of infectious diseases during its retreat from Russia. Because soldiers were commonly infested with body lice, it has been speculated that louse-borne infectious diseases, such as epidemic typhus (caused by Rickettsia prowazekii), were common. METHODS: We investigated this possibility during recent excavations of a mass grave of Napoleon's soldiers in Vilnius, Lithuania. Segments of 5 body lice, identified morphologically and by polymerase chain reaction (PCR) amplification and sequencing, were found in earth from the grave that also contained fragments of soldiers' uniforms. RESULTS: DNA of Bartonella quintana (the agent of trench fever) was identified by PCR and sequencing in 3 of the lice. Similarly, PCR and sequencing of dental pulp from the remains of 35 soldiers revealed DNA of B. quintana in 7 soldiers and DNA of R. prowazekii in 3 other soldiers. CONCLUSIONS: Our results show that louse-borne infectious diseases affected nearly one-third of Napoleon's soldiers buried in Vilnius and indicate that these diseases might have been a major factor in the French retreat from Russia.

Bartonella quintana and Rickettsia felis in Gabon.

We detected Rickettsia felis DNA in Ctenocephalides felis and Bartonella quintana DNA in 3 Pulex irritans fleas taken from a pet Cercopithecus cephus monkey in Gabon, sub-Saharan Africa. This is the first report of B. quintana in the human flea.

Multispacer typing technique for sequence-based typing of Bartonella quintana.

Bartonella quintana is a worldwide fastidious bacterium of the Alphaproteobacteria responsible for bacillary angiomatosis, trench fever, chronic lymphadenopathy, and culture-negative endocarditis. The recent genome sequencing of a B. quintana isolate allowed us to propose a genome-wide sequence-based typing method. To ensure sequence discrimination based on highly polymorphic areas, we amplified and sequenced 34 spacers in a large collection of B. quintana isolates. Six of these exhibited polymorphisms and allowed the characterization of 4 genotypes. However, the strain variants suggested by the noncoding sequences did not correlate with the results of pulsed-field gel electrophoresis (PFGE), which suggested a higher degree of variability. Modification of the PFGE profile of one isolate after nine subcultures confirmed that rearrangement frequencies are high in this species, making PFGE unreliable for epidemiological purposes. The low extent of sequence heterogeneity in the species suggests a recent emergence of this bacterium as a human pathogen. Direct typing of natural samples allowed the identification of a fifth genotype in the DNA extracted from a human body louse collected in Burundi. We have named the typing technique herein described multispacer typing.

Bartonella quintana in cynomolgus monkey (Macaca fascicularis).

We identified a Bartonella quintana strain by polymerase chain reaction amplification, cloning, and sequencing of DNA extracted from lysed erythrocytes and cultured colonies grown from peripheral blood collected from a captive-bred cynomolgus monkey (Macaca fascicularis). This report describes naturally acquired B. quintana infection in a nonhuman primate.

Molecular characterization of first human Bartonella strain isolated in Italy.

The aim of this study was to characterize a Bartonella strain (BA-1) isolated from a blood culture of an Italian, human immunodeficiency virus-positive patient with bacillary angiomatosis. We analyzed the isolate using molecular biology methods such as whole-cell fatty acid analysis, PCR-restriction fragment length polymorphism analysis, type-specific 16S rRNA PCRs, sequence analysis of the 16S rRNA, pulsed-field gel electrophoresis, and arbitrarily primed PCR. The BA-1 isolate turned out to be a Bartonella quintana strain, similar but not identical to B. quintana Oklahoma, which was used as a control strain.

Detection and culture of Bartonella quintana, Serratia marcescens, and Acinetobacter spp. from decontaminated human body lice.

As part of a survey for trench fever among homeless people in Marseilles, France, we attempted isolation of Bartonella quintana from body lice. A decontamination protocol of immersion in 70% ethanol with 0.2% iodine was devised and was tested with a laboratory colony of body lice. Lice which had been experimentally contaminated with either Escherichia coli, Staphylococcus epidermidis, or Acinetobacter spp. were successfully decontaminated, and this process did not prevent the culture of B. quintana from these lice. One hundred sixty-one lice obtained from homeless patients were studied by the protocol. B. quintana was isolated on axenic medium from 15 of 161 body lice and was detected in 41 of 161 lice by PCR. Acinetobacter spp. and Serratia marcescens were also isolated from body lice. The sensitivities of PCR and culture of B. quintana were 98 and 36%, respectively. These procedures may be useful for epidemiologic studies of trench fever and for the recovery of strains for characterization and comparison.

Use of rpoB gene analysis for detection and identification of Bartonella species.

Identification of Bartonella species is of increasing importance as the number of infections in which these bacteria are involved increases. To date, these gram-negative bacilli have been identified by various serological, biochemical, and genotypic methods. However, the development of alternative tools is required, principally to circumvent a major risk of contamination during sample manipulation. The aim of our study was to investigate the possible identification of various Bartonella species by comparison of RNA polymerase beta-subunit gene (rpoB) sequences. This approach has previously been shown to be useful for the identification of members of the family Enterobacteriaceae (C. M. Mollet, M. Drancourt, and D. Raoult, Mol. Microbiol. 26:1005-1011, 1997). Following PCR amplification with specific oligonucleotides, a 825-bp region of the rpoB gene was sequenced from 13 distinct Bartonella strains. Analysis of these sequences allowed selection of three restriction enzymes (ApoI, AluI, and AflIII) useful for discerning the different strains by PCR-restriction fragment length polymorphism (PCR-RFLP) analysis. To confirm the potential value of such an approach for identification of Bartonella, the rpoB PCR was then applied to 94 clinical samples, and the results obtained were identical to those obtained by our reference PCR method. Twenty-four isolates were also adequately identified by PCR-RFLP analysis. In all cases, our results were in accordance with those of the reference method. Moreover, conserved regions of DNA were chosen as suitable primer targets for PCR amplification of a 439-bp fragment which can be easily sequenced.

Determination of the rpoB gene sequences of Bartonella henselae and Bartonella quintana for phylogenic analysis.

Using the Genome Walker procedure, which allows PCR amplification of genomic DNA using a single gene-specific primer and direct automated sequencing methodology, we obtained the nucleotide sequence of the RNA polymerase beta subunit (rpoB) from Bartonella henselae and Bartonella quintana. A phylogenetic tree constructed from these data and other rpoB sequences available in GenBank is, in part, consistent with those previously derived from 16S rRNA gene sequences and confirms the position of Bartonella within the alpha subdivision of Proteobacteria. In fact, this analysis showed that rpoB data are similar to 16S rRNA data for the alpha, beta and gamma subdivisions of Proteobacteria. In contrast, concerning other bacteria included in our study, the topologies of phylogenetic trees were different. Based on the bootstrap values derived from rpoB phylogenic analysis, we believe that this molecule should contribute to better understanding the evolutionary process.