Molecular detection of Rickettsia in ectoparasites (Siphonaptera and Phthiraptera) of domestic and feral pigs from Argentina.

Rickettsioses are distributed among a variety of hematophagous arthropods, and represent an emergent threat. The presence of rickettsial bacteria in ectoparasites collected from pigs from Argentina is still unknown. This study investigated the presence and identity of Rickettsia spp. in fleas, Pulex irritans, and sucking lice, Haematopinus suis, of domestic and feral pigs, Sus scrofa, from Central-Northern Argentina, through the genes gltA and ompB. Rickettsial bacteria were detected in 50% of fleas and 24% of lice. The BLASTn analysis of the ompB gene fragments in P. irritans samples showed identities 99% and 100% with R. felis. Positive samples of H. suis were 99% similar with species from the spotted fever group, future amplifications of a more polymorphic fragment of the ompB gene will allow to corroborate the identity of the Rickettsia species present in these lice samples. The Rickettsia spp. reported in the present study are having eventually been associated with cases of human diseases, and the circulation of these agents in arthropods has already been reported in several countries. Therefore, the identification of circulating pathogenic agents, such as reported in this study, is crucial for development of preventive measures for the control of ectoparasite-borne rickettsiosis diseases. Further studies, using serology techniques, will be allow to explore the ability of pigs as a possible Rickettsia reservoir and its role as part of transmission cycle of Rickettsia spp. in the studied scenarios.

The seal louse (Echinophthirius horridus) in the Dutch Wadden Sea: investigation of vector-borne pathogens.

BACKGROUND: Belonging to the anopluran family Echinophthiriidae, Echinophthirius horridus, the seal louse, has been reported to parasitise a broad range of representatives of phocid seals. So far, only a few studies have focused on the vector function of echinophthiriid lice, and knowledge about their role in pathogen transmission is still scarce. The current study aims to investigate the possible vector role of E. horridus parasitising seals in the Dutch Wadden Sea. METHODS: E. horridus seal lice were collected from 54 harbour seals (Phoca vitulina) and one grey seal (Halichoerus grypus) during their rehabilitation period at the Sealcentre Pieterburen, The Netherlands. DNA was extracted from pooled seal lice of individual seals for molecular detection of the seal heartworm Acanthocheilonema spirocauda, the rickettsial intracellular bacterium Anaplasma phagocytophilum, and the cell wall-less bacteria Mycoplasma spp. using PCR assays. RESULTS: Seal lice from 35% of the harbour seals (19/54) and from the grey seal proved positive for A. spirocauda. The seal heartworm was molecularly characterised and phylogenetically analysed (rDNA, cox1). A nested PCR was developed for the cox1 gene to detect A. spirocauda stages in seal lice. A. phagocytophilum and a Mycoplasma species previously identified from a patient with disseminated 'seal finger' mycoplasmosis were detected for the first time, to our knowledge, in seal lice. CONCLUSIONS: Our findings support the potential vector role of seal lice in the transmission of A. spirocauda and reveal new insights into the spectrum of pathogens occurring in seal lice. Studies on vector competence of E. horridus, especially for bacterial pathogens, are essentially needed in the future as these pathogens might have detrimental effects on the health of seal populations. Furthermore, studies on the vector role of different echinophthiriid species infecting a wide range of pinniped hosts should be conducted to extend the knowledge of vector-borne pathogens.

The genus Rickettsia in Mexico: Current knowledge and perspectives.

The genus Rickettsia encompasses 35 valid species of intracellular, coccobacilli bacteria that can infect several eukaryotic taxa, causing multiple emerging and re-emerging diseases worldwide. This work aimed to gather and summarise the current knowledge about the genus Rickettsia in Mexico, updating the taxonomy of the bacteria and their hosts by including all the records available until 2020, to elucidate host-parasite relationships and determine the geographical distribution of each Rickettsia species present in the country. Until now, 14 species of Rickettsia belonging to four groups have been recorded in Mexico. These species have been associated with 26 arthropod species (14 hard ticks, three soft ticks, two sucking lice, and seven fleas) and 17 mammal species distributed over 30 states in Mexico. This work highlights the high biological inventory of rickettsias for Mexico and reinforces the need to approach the study of this group from a One Health perspective.

First record of natural infection with Anaplasma marginale in sucking lice infesting the water buffalo (Bubalus bubalis) in Mexico.

The water buffalo (Bubalus bubalis) is an Asian species of bovine which was introduced in Mexico in 1992, as an alternative for milk and meat production. However, little is known about its associated ectoparasites, in particular about their lice. As a part of a project to identify the diversity and prevalence of Anaplasma in cattle and water buffaloes in the state of Veracruz, Mexico, we visited a ranch in the downtown area in which 42 buffaloes were visually inspected for the presence of ectoparasites. Lice were manually recovered, and morphologically identified using taxonomic keys. They were also molecularly identified by the amplification of a fragment of the cytochrome oxidase C subunit I gene (COX1). As a result, we recovered 20 lice from eight animals. All lice were identified as Haematopinus tuberculatus, and the recovered sequences exhibited a similarity of 99.7% with those of the same species deposited in GenBank. Additionally, we detected the presence of Anaplasma marginale in 70% (14/20) of the analysed samples. Our results represent the first record of this exotic ectoparasite for the country.

First molecular detection of Mycoplasma suis in the pig louse Haematopinus suis (Phthiraptera: Anoplura) from Argentina.

Porcine haemoplasmosis caused by Mycoplasma suis affects the global pig industry with significant economic losses. The main transmission route of M. suis is through the blood and some haematophagous arthropods, like flies and mosquitoes, could be the vectors to this pathogen. However, the presence of M. suis in pig haematophagous ectoparasites in natural conditions has not yet been studied. The most frequent ectoparasite in pigs is the blood-sucking louse Haematopinus suis, an obligate and permanent parasite. Therefore, this work aims to study the occurrence of M. suis in H. suis samples from both domestic and wild pig populations from Argentina; using the 16S rRNA gene. A total of 98 sucking lice, collected from domestic and wild pigs from Buenos Aires Province in central Argentina, were examined. We found M. suis DNA in 15 H. suis samples (15.30%). Positive lice were detected from all studied populations. This is the first report of M. suis presence in H. suis, being also the first detection in a pig ectoparasite species. We conclude that H. suis could serve as a mechanical vector for M. suis. This information not only extends the knowledge about the pathogen spectrum potentially transmitted by H. suis, but may be also useful in epidemiological studies about Mycoplasma.

Molecular Evidence of Rickettsia slovaca in Wild Boar Lice, in Northeastern Algeria.

Using molecular assays, Rickettsia slovaca, the agent of a spotted fever group rickettsia was assessed in 92/325 lice collected on 117 wild boars hunted in the far northeast of Algeria. Overall, 7/92 (7.61%) Haematopinus suis were found positive to R. slovaca. The presence of R. slovaca DNA on H. suis questions about the relationship that may exist between R. slovaca and lice.

Legionella Becoming a Mutualist: Adaptive Processes Shaping the Genome of Symbiont in the Louse Polyplax serrata.

Legionellaceae are intracellular bacteria known as important human pathogens. In the environment, they are mainly found in biofilms associated with amoebas. In contrast to the gammaproteobacterial family Enterobacteriaceae, which established a broad spectrum of symbioses with many insect taxa, the only instance of legionella-like symbiont has been reported from lice of the genus Polyplax. Here, we sequenced the complete genome of this symbiont and compared its main characteristics to other Legionella species and insect symbionts. Based on rigorous multigene phylogenetic analyses, we confirm this bacterium as a member of the genus Legionella and propose the name Candidatus Legionella polyplacis, sp.n. We show that the genome of Ca. Legionella polyplacis underwent massive degeneration, including considerable size reduction (529.746 bp, 484 protein coding genes) and a severe decrease in GC content (23%). We identify several possible constraints underlying the evolution of this bacterium. On one hand, Ca. Legionella polyplacis and the louse symbionts Riesia and Puchtella experienced convergent evolution, perhaps due to adaptation to similar hosts. On the other hand, some metabolic differences are likely to reflect different phylogenetic positions of the symbionts and hence availability of particular metabolic function in the ancestor. This is exemplified by different arrangements of thiamine metabolism in Ca. Legionella polyplacis and Riesia. Finally, horizontal gene transfer is shown to play a significant role in the adaptive and diversification process. Particularly, we show that Ca. L. polyplacis horizontally acquired a complete biotin operon (bioADCHFB) that likely assisted this bacterium when becoming an obligate mutualist.

Two Bacterial Genera, Sodalis and Rickettsia, Associated with the Seal Louse Proechinophthirus fluctus (Phthiraptera: Anoplura).

UNLABELLED: Roughly 10% to 15% of insect species host heritable symbiotic bacteria known as endosymbionts. The lice parasitizing mammals rely on endosymbionts to provide essential vitamins absent in their blood meals. Here, we describe two bacterial associates from a louse, Proechinophthirus fluctus, which is an obligate ectoparasite of a marine mammal. One of these is a heritable endosymbiont that is not closely related to endosymbionts of other mammalian lice. Rather, it is more closely related to endosymbionts of the genus Sodalis associated with spittlebugs and feather-chewing bird lice. Localization and vertical transmission of this endosymbiont are also more similar to those of bird lice than to those of other mammalian lice. The endosymbiont genome appears to be degrading in symbiosis; however, it is considerably larger than the genomes of other mammalian louse endosymbionts. These patterns suggest the possibility that this Sodalis endosymbiont might be recently acquired, replacing a now-extinct, ancient endosymbiont. From the same lice, we also identified an abundant bacterium belonging to the genus Rickettsia that is closely related to Rickettsia ricketsii, a human pathogen vectored by ticks. No obvious masses of the Rickettsia bacterium were observed in louse tissues, nor did we find any evidence of vertical transmission, so the nature of its association remains unclear. IMPORTANCE: Many insects are host to heritable symbiotic bacteria. These heritable bacteria have been identified from numerous species of parasitic lice. It appears that novel symbioses have formed between lice and bacteria many times, with new bacterial symbionts potentially replacing existing ones. However, little was known about the symbionts of lice parasitizing marine mammals. Here, we identified a heritable bacterial symbiont in lice parasitizing northern fur seals. This bacterial symbiont appears to have been recently acquired by the lice. The findings reported here provide insights into how new symbioses form and how this lifestyle is shaping the symbiont genome.

Identification of different Bartonella species in the cattle tail louse (Haematopinus quadripertusus) and in cattle blood.

Bartonella spp. are worldwide-distributed facultative intracellular bacteria that exhibit an immense genomic diversity across mammal and arthropod hosts. The occurrence of cattle-associated Bartonella species was investigated in the cattle tail louse Haematopinus quadripertusus and in dairy cattle blood from Israel. Lice were collected from cattle from two dairy farms during summer 2011, and both lice and cow blood samples were collected from additional seven farms during the successive winter. The lice were identified morphologically and molecularly using 18S rRNA sequencing. Thereafter, they were screened for Bartonella DNA by conventional and real-time PCR assays using four partial genetic loci (gltA, rpoB, ssrA, and internal transcribed spacer [ITS]). A potentially novel Bartonella variant, closely related to other ruminant bartonellae, was identified in 11 of 13 louse pools collected in summer. In the cattle blood, the prevalence of Bartonella infection was 38%, identified as B. bovis and B. henselae (24 and 12%, respectively). A third genotype, closely related to Bartonella melophagi and Bartonella chomelii (based on the ssrA gene) and to B. bovis (based on the ITS sequence) was identified in a single cow. The relatively high prevalence of these Bartonella species in cattle and the occurrence of phylogenetically diverse Bartonella variants in both cattle and their lice suggest the potential role of this animal system in the generation of Bartonella species diversity.

Detection of Brucella abortus DNA and RNA in different stages of development of the sucking louse Haematopinus tuberculatus.

BACKGROUND: Brucellosis is considered the world's most widespread zoonotic infection. It causes abortion and sterility in livestock leading to serious economic losses and has even more serious medical impact in humans, since it can be a trigger to more than 500,000 infections per year worldwide. The aim of this study was to evaluate the role of Haematopinus tuberculatus, a louse that can parasitize several ruminants, as a new host of brucellosis. Louse specimens were collected from seropositive and seronegative water buffaloes and divided in 3 developmental stages: adults, nymphs and nits. All samples were separately screened for Brucella spp. DNA and RNA detection by Real Time PCR. In particular, primers and probes potentially targeting the 16S rRNA and the Brucella Cell Surface 31 kDalton Protein (bcsp31) genes were used for Real Time PCR and buffalo ß actin was used as a housekeeping gene to quantify host DNA in the sample. A known amount of B. abortus purified DNA was utilized for standard curve preparation and the target DNA amount was divided by the housekeeping gene amount to obtain a normalized target value. A further molecular characterization was performed for Brucella strain typing and genotyping by the Bruce-ladder, AMOS-PCR and MLVA assays. Data were statistically analysed by ANOVA. RESULTS: Brucella abortus DNA and RNA were detected in all developmental stages of the louse, suggesting the presence of viable bacteria. Data obtained by MLVA characterization support this finding, since the strains present in animals and the relative parasites were not always identical, suggesting bacterial replication. Furthermore, the detection of Brucella DNA and RNA in nits samples demonstrate, for the first time, a trans-ovarial transmission of the bacterium into the louse. CONCLUSIONS: These findings identified H. tuberculatus as a new host of brucellosis. Further studies are needed to establish the role of this louse in the epidemiology of the disease, such as vector or reservoir.

Lice outbreak in buffaloes: evidence of Anaplasma marginale transmission by sucking lice Haematopinus tuberculatus.

Lice infestations are commonly seen in buffaloes, causing damage directly to the animal, i.e., itching, skin lesions, and anemia. In addition, these insects can also be vectors for infectious diseases. The present study describes an outbreak of lice in buffaloes as well as evidence for Haematopinus tuberculatus acting as a vector of anaplasmosis. Lice and blood were collected from 4 young buffaloes (2- to 4-mo-old) and a molecular analysis for the presence of Anaplasma marginale was conducted. DNA of A. marginale was detected in the blood of all 4 animals. Twelve lice were collected and separated in 4 groups, with 3 insects each, to comprise a pool of samples. After DNA extraction and molecular analysis, a positive PCR for A. marginale was found in all pooled samples. These results identify sucking lice as potential vectors of anaplasmosis. However, additional studies are necessary to fully evaluate the vector potential of H. tuberculatus for A. maginale transmission.

Evidence of antibodies to spotted fever group rickettsiae in small mammals and quail from Mississippi.

Rickettsia parkeri is a recently recognized human pathogen primarily associated with the Gulf Coast tick Amblyomma maculatum, with immature stages of this tick reported from wild vertebrates. To better understand the role of vertebrates in the natural history of this bacterium, we evaluated small mammals and ground-dwelling birds for evidence of infection with R. parkeri or exposure to the organism. We sampled small mammals (n=39) and passerines (n=47) in both north-central and southeast Mississippi, while northern bobwhite (Colinus virginianus) samples (n=31) were obtained from farms in central Mississippi. Blood from all sampled animals was tested using polymerase chain reaction (PCR) for spotted fever group rickettsiae (SFGR), and for antibodies to SFGR using R. parkeri antigen. Ectoparasite samples were removed from animals and included mites, lice, fleas, and immature ticks. Of 39 small mammal samples collected, 7 were positive for antibodies to SFGR; none tested positive by PCR for DNA of SFGR. Of 47 passerine blood samples collected, none were positive for DNA of SFGR by PCR, nor did any show serological evidence of exposure. Finally, none of 31 northern bobwhite samples tested were positive for SFGR DNA, while 7 were seropositive for rickettsial antibodies. Detection of seropositive rodents and quail suggests a role for these host species in the natural history of SFGR, possibly including R. parkeri, but the extent of their role has not yet been elucidated.

Intestinal endocellular symbiotic bacterium of the macaque louse Pedicinus obtusus: Distinct endosymbiont origins in anthropoid primate lice and the old world monkey louse.

A symbiotic bacterium of the macaque louse, Pedicinus obtusus, was characterized. The symbiont constituted a gammaproteobacterial lineage distinct from the symbionts of anthropoid primate lice, localized in the midgut epithelium and the ovaries and exhibiting AT-biased genes and accelerated molecular evolution. The designation "Candidatus Puchtella pedicinophila" was proposed for it.

On the ubiquity and phylogeny of Wolbachia in lice.

Wolbachia are intracellular bacteria that occur in an estimated 20% of arthropod species. They are of broad interest because they profoundly affect the reproductive fitness of diverse host taxa. Here we document the apparent ubiquity and diversity of Wolbachia in the insect orders Anoplura (sucking lice) and Mallophaga (chewing lice), by detecting single or multiple infections in each of 25 tested populations of lice, representing 19 species from 15 genera spanning eight taxonomic families. Phylogenetic analyses indicate a high diversity of Wolbachia in lice, as evidenced by the identification of 39 unique strains. Some of these strains are apparently unique to lice, whereas others are similar to strains that infect other insect taxa. Wolbachia are transmitted from infected females to their offspring via egg cytoplasm, such that similar species of lice are predicted to have similar strains of Wolbachia. This predicted pattern is not supported in the current study and may reflect multiple events of recent horizontal transmission between host species. At present, there is no known mechanism that would allow for this latter mode of transmission to and within species of lice.

Ectoparasites of gray squirrels in two different habitats and screening of selected ectoparasites for bartonellae.

Gray squirrels, Sciurus carolinensis, were livetrapped in 2 different habitat types, woodland (67 squirrels) and parkland (53 squirrels), in southeastern Georgia. Ectoparasites were recovered from anesthetized squirrels and compared between hosts from the 2 habitats. Because of the absence of low vegetation in parkland habitats, it was hypothesized that the ectoparasite fauna, especially ticks and chiggers, would be more diverse on woodland squirrels. The results were generally in agreement with this hypothesis. Seventeen species of ectoparasites were recovered from woodland squirrels, compared with 6 species from parkland squirrels. Five species of ticks and 3 species of chiggers parasitized the woodland squirrels compared with no ticks or chiggers on the parkland squirrels. Significantly higher infestation prevalences were recorded on woodland compared with parkland squirrels for the flea Orchopeas howardi, the tick Amblyomma americanum, and the mesostigmatid mite Androlaelaps fahrenholzi. The mean intensity for O. howardi also was significantly higher on woodland than on parkland squirrels. Because a new strain of Bartonella sp. was isolated recently from S. carolinensis in Georgia, selected ectoparasites from this study were screened for bartonellae by polymerase chain reaction (PCR). Some of the fleas and lice, but none of the mites tested, were PCR positive, suggesting that fleas, or lice, or both, might be vectors of bartonellae between squirrels. Six distinct strains of Bartonella sp. were detected, 2 in fleas and 4 in lice.

The sex ratio distortion in the human head louse is conserved over time.

BACKGROUND: At the turn of the 19th century the first observations of a female-biased sex ratio in broods and populations of the head louse, Pediculus humanus capitis, had been reported. A study by Buxton in 1940 on the sex ratio of lice on prisoners in Ceylon is still today the subject of reanalyses. This sex ratio distortion had been detected in ten different countries. In the last sixty years no new data have been collected, especially on scalp infestations under economically and socially more developed conditions. RESULTS: Here we report a female bias of head lice in a survey of 480 school children in Argentina. This bias is independent of the intensity of the pediculosis, which makes local mate competition highly unlikely as the source of the aberrant sex ratio; however, other possible adaptive mechanisms cannot be discounted. These lice as well as lice from pupils in Britain were carrying several strains of the endosymbiotic bacterium Wolbachia pipientis, one of the most wide spread intracellular sex ratio distorters. Similar Wolbachia strains are also present in the pig louse, Haematopinus suis, suggesting that this endosymbiont might have a marked influence on the biology of the whole order. The presence of a related obligate nutritional bacterium in lice prevents the investigation of a causal link between sex ratio and endosymbionts. CONCLUSIONS: Regardless of its origin, this sex ratio distortion in head lice that has been reported world wide, is stable over time and is a remarkable deviation from the stability of frequency-dependent selection of Fisher's sex ratio. A female bias first reported in 1898 is still present over a hundred years and a thousand generations later.

The neglected saliva: medically important toxins in the saliva of human lice.

Although there has been a great deal of research effort within the last two decades on identifying the active components of the saliva of blood-sucking ticks, mosquitoes, biting flies, fleas and bugs, essentially neglected have been the human lice. Despite initial reports in the early part of this century suggestive of vasodilatory, anticoagulant and immunosuppressive properties of the saliva, for the next 50 years there were no biochemical studies on the active principles. Very recently, anatomical and biochemical studies have begun to characterize the bioactive molecules in lice saliva. The louse stocks a salivary vasodilator in excess over what is needed for a single bite, and injects similar amounts at each successive bite. The vasodilator in lice saliva appears to have different pharmacological properties than peroxidative, oxidative and maxidilan types of vasodilators reported from other blood-sucking insects. Possible anticoagulant activities have also been characterized. This belated, but welcome, interest comes at a time of resurgence of lice-born disease in certain parts of Africa, and of resistance to chemical control in Europe and North America.