Molecular Detection of Tick-Borne Pathogens in Humans with Tick Bites and Erythema Migrans,in the Netherlands

BackgroundTick-borne diseases are the most prevalent vector-borne diseases in Europe. Knowledge on the incidence and clinical presentation of other tick-borne diseases than Lyme borreliosis and tick-borne encephalitis is minimal, despite the high human exposure to these pathogens through tick bites. Using molecular detection techniques, the frequency of tick-borne infections after exposure through tick bites was estimated.MethodsTicks, blood samples and questionnaires on health status were collected from patients that visited their general practitioner with a tick bite or erythema migrans in 2007 and 2008. The presence of several tick-borne pathogens in 314 ticks and 626 blood samples of this cohort were analyzed using PCR-based methods. Using multivariate logistic regression, associations were explored between pathogens detected in blood and self-reported symptoms at enrolment and during a three-month follow-up period.ResultsHalf of the ticks removed from humans tested positive for Borrelia burgdorferi sensu lato, Anaplasma phagocytophilum, Candidatus Neoehrlichia mikurensis, Rickettsia helvetica, Rickettsia monacensis, Borrelia miyamotoi and several Babesia species. Among 92 Borrelia burgdorferi s. l. positive ticks, 33% carried another pathogen from a different genus. In blood of sixteen out of 626 persons with tick bites or erythema migrans, DNA was detected from Candidatus Neoehrlichia mikurensis (n = 7), Anaplasma phagocytophilum (n = 5), Babesia divergens (n = 3), Borrelia miyamotoi (n = 1) and Borrelia burgdorferi s. l. (n = 1). None of these sixteen individuals reported any overt symptoms that would indicate a corresponding illness during the three-month follow-up period. No associations were found between the presence of pathogen DNA in blood and; self-reported symptoms, with pathogen DNA in the corresponding ticks (n = 8), reported tick attachment duration, tick engorgement, or antibiotic treatment at enrolment.ConclusionsBased on molecular detection techniques, the probability of infection with a tick-borne pathogen other than Lyme spirochetes after a tick bite is roughly 2.4%, in the Netherlands. Similarly, among patients with erythema migrans, the probability of a co-infection with another tick-borne pathogen is approximately 2.7%. How often these infections cause disease symptoms or to what extend co-infections affect the course of Lyme borreliosis needs further investigations.     

Tentorial mobility in centipedes (Chilopoda) revisited: 3D reconstruction of the mandibulo-tentorial musculature of Geophilomorpha

Mandibular mechanisms in Geophilomorpha are revised based on three-dimensional reconstructions of the mandibulo-tentorial complex and its muscular equipment in Dicellophilus carniolensis (Placodesmata) and Hydroschendyla submarina (Adesmata). Tentorial structure compares closely in the two species and homologies can be proposed for the 14/17 muscles that attach to the tentorium. Both species retain homologues of muscles that in other Pleurostigmophora are traditionally thought to cause swinging movements of the tentorium that complement the mobility of the mandibles. Although the original set of tentorial muscles is simplified in Geophilomorpha, the arrangement of the preserved homologues conforms to a system of six degrees of freedom of movement, as in non-geophilomorph Pleurostigmophora. A simplification of the mandibular muscles is confirmed for Geophilomorpha, but our results reject absence of muscles that in other Pleurostigmophora primarily support see-saw movements of the mandibles. In the construction of the tentorium, paralabial sclerites seem to be involved in neither Placodesmata nor Adesmata, and we propose their loss in Geophilomorpha as a whole. Current insights on the tentorial skeleton and its musculature permit two alternative conclusions on their transformation in Geophilomorpha: either tentorial mobility is primarily maintained in both Placodesmata and Adesmata (contrary to Manton’s arguments for immobility), or the traditional assumption of the tentorium as being mobile is a misinterpretation for Pleurostigmophora as a whole.     

At the end of the rope: Geophilus hadesi sp. n. – the world’s deepest cave-dwelling centipede (Chilopoda,Geophilomorpha, Geophilidae)

A new geophilomorph centipede, Geophilus hadesi sp. n., is described from caves in the Velebit Mountain, central Croatia. Together with Geophilus persephones Foddai & Minelli, 1999, described from Pierre Saint-Martin cave in France, they are the only two remarkably troglomorphic geophilomorphs hitherto known. The new species apparently belongs to a group of Geophilus species inhabiting mainly Western and Southern Europe, with a uniquely modified pretarsus in the second maxillae. Geophilus hadesi sp. n. shows unusual traits, some of which commonly found in troglobitic arthropods, including exceptionally elongated antennae, trunk segments and leg claws. The species is described upon specimens found in two caves at a depth below -250 m. Another two specimens apparently belonging to the same species have been recorded in another deep vertical cave at -980 m and -1100 m. The latter represents the world’s deepest record of Chilopoda as a whole.     

Species limits and phylogeography of Newportia (Scolopendromorpha) and implications for widespread morphospecies

The genus Newportia Gervais, 1847, includes some 60 nominal species distributed in the Caribbean islands and from Mexico to central South America. Modern keys to species and subspecies are available, greatly facilitating identification, but some species are based on few specimens and have incomplete documentation of taxonomically-informative characters. In order to explore genetic variability and evolutionary relationships within geographically-widespread morphospecies, specimens of Newportia (Newportia) stolli (Pocock, 1896) and Newportia (Newportia) divergens Chamberlin, 1922, two nominal species distinguished principally by differences in suture patterns on T1, were sequenced for mitochondrial 16S rRNA and cytochrome c oxidase subunit I (COI) genes from populations in southern Mexico, Guatemala, Honduras and Brazil. Newportia (Newportia) stolli is paraphyletic with respect to Newportia (Newportia) divergens within a clade from Guatemala, Honduras, and Chiapas (Mexico), most trees being consistent with a single loss of a connection between the anterior transverse suture on T1, whereas specimens of “Newportia (Newportia) stolli” from Brazil are not closely allied to those from the Mesomerican type area. The widespread morphospecies Newportia (Newportia) monticola Pocock, 1890, was sequenced for the same loci from populations in Costa Rica, Colombia and Brazil, finding that specimens from these areas do not unite as a monophyletic group. Samples of Newportia (Newportia) oreina Chamberlin, 1915, from different regions of Mexico form geographic clusters that resolve as each other’s closest relatives. These results suggest that some widespread species of Newportia may be taxa of convenience more so than natural groupings. In several cases geographic proximity fits the phylogeny better than taxonomy, suggesting that non-monophyletic species do not result from use of inappropriate molecular markers. Molecular identification is possible for specimens missing taxonomically informative morphological characters, notably damaged specimens that lack the ultimate leg pair, a protocol that may also apply to other taxonomically difficult genera that are prone to damage (such as Cryptops).     

Homozygous STIL Mutation Causes Holoprosencephaly and Microcephaly in Two Siblings

Holoprosencephaly (HPE) is a frequent congenital malformation of the brain characterized by impaired forebrain cleavage and midline facial anomalies. Heterozygous mutations in 14 genes have been identified in HPE patients that account for only 30% of HPE cases, suggesting the existence of other HPE genes. Data from homozygosity mapping and whole-exome sequencing in a consanguineous Turkish family were combined to identify a homozygous missense mutation (c.2150G>A; p.Gly717Glu) in STIL, common to the two affected children. STIL has a role in centriole formation and has previously been described in rare cases of microcephaly. Rescue experiments in U2OS cells showed that the STIL p.Gly717Glu mutation was not able to fully restore the centriole duplication failure following depletion of endogenous STIL protein indicating the deleterious role of the mutation. In situ hybridization experiments using chick embryos demonstrated that expression of Stil was in accordance with a function during early patterning of the forebrain. It is only the second time that a STIL homozygous mutation causing a recessive form of HPE was reported. This result also supports the genetic heterogeneity of HPE and increases the panel of genes to be tested for HPE diagnosis.     

Mutagenesis using Tn917 in Bacillus licheniformis

Abstract Two plasmid vectors containing Tn 917 have been transformed into protoplasts of Bacillus licheniformis . Replication of the vectors was inhibited at high temperature. The transporon integrated efficiently into the chromosome and auxotroph formation was high (9% of transposed colonies). Most of these were argG or argH indicating a hot-spot region for integration of Tn 917 into the chromosome. However, about 0.5% of the mutants had other auxotrophies.     

Time Constants and Electrotonic Length of Membrane Cylinders and Neurons

A theoretical basis is provided for the estimation of the electrotonic length of a membrane cylinder, or the effective electrotonic length of a whole neuron, from electrophysiological experiments. It depends upon the several time constants present in passive decay of membrane potential from an initially nonuniform distribution over the length. In addition to the well known passive membrane time constant, τ_m = R_mC_m, observed in the decay of a uniform membrane potential, there exist many smaller time constants that govern rapid equalization of membrane potential over the length. These time constants are present also in the transient response to a current step applied across the membrane at one location, such as the neuron soma. Similar time constants are derived when a lumped soma is coupled to one or more cylinders representing one or more dendritic trees. Different time constants are derived when a voltage clamp is applied at one location; the effects of both leaky and short-circuited termination are also derived. All of these time constants are demonstrated as consequences of mathematical boundary value problems. These results not only provide a basis for estimating electrotonic length, L = [unk]/λ, but also provide a new basis for estimating the steady-state ratio, ρ, of cylinder input conductance to soma membrane conductance.     

Adding mitochondrial sequence data (16S rRNA and cytochrome c oxidase subunit I) to the phylogeny of centipedes (Myriapoda: Chilopoda): an analysis of morphology and four molecular loci

Relationships within Chilopoda (centipedes) are assessed based on 222 morphological characters, complete 18S rRNA sequences for 70 chilopod terminals, the D3 region of 28S rRNA for 65 terminals, 16S rRNA sequences for 54 terminals and cytochrome c oxidase subunit I sequences for 45 terminals. Morphological and molecular data for seven orders of Diplopoda are used to root cladograms for Chilopoda. Analyses use direct character optimization for 15 gap and substitution models. The Pleurostigmophora and Epimorpha s.l. hypotheses are largely stable to parameter variation for the combined data; the latter clade is formalized as the new taxon Phylactometria. The combined data include parameter sets that support either the monophyly of Epimorpha s.str. (=Scolopendromorpha + Geophilomorpha) or Craterostigmus + Geophilomorpha; the former derives its support from morphology and the nuclear ribosomal genes. Monophyly of Lithobiomorpha and the sister group relationship between Lithobiidae and Henicopidae are stable for morphological and combined data, and are also resolved for the molecular data for 14 of 15 parameter sets. The fundamental split in Scolopendromorpha is between Cryptopidae and Scolopendridae sensu Attems. Blind scolopendromorphs unite as a clade in most molecular and combined analyses, including those that minimize incongruence between data partitions. Geophilomorpha divides into Placodesmata and Adesmata under nine of 15 explored parameter sets.