Genome sequence of Desulfovibrio sp. A2, a highly copper resistant, sulfate-reducing bacterium isolated from effluents of a zinc smelter at the Urals

Desulfovibrio sp. A2 is an anaerobic gram-negative sulfate-reducing bacterium with remarkable tolerance to copper. It was isolated from wastewater effluents of a zinc smelter at the Urals. Here, we report the 4.2-Mb draft genome sequence of Desulfovibrio sp. A2 and identify potential copper resistance mechanisms.

Genome sequence of Desulfosporosinus sp. OT, an acidophilic sulfate-reducing bacterium from copper mining waste in Norilsk, Northern Siberia

We have sequenced the genome of Desulfosporosinus sp. OT, a Gram-positive, acidophilic sulfate-reducing Firmicute isolated from copper tailing sediment in the Norilsk mining-smelting area in Northern Siberia, Russia. This represents the first sequenced genome of a Desulfosporosinus species. The genome has a size of 5.7 Mb and encodes 6,222 putative proteins.

Classification of organisms previously reported as the SP and Stewart-Letscher groups, with descriptions of Necropsobacter gen. nov. and of Necropsobacter rosorum sp. nov. for organisms of the SP group

To allow classification of bacteria previously reported as the SP group and the Stewart-Letscher group, 35 isolates from rodents (21), rabbits (eight), a dog and humans (five) were phenotypically and genotypically characterized. Comparison of partial rpoB sequences showed that 34 of the isolates were closely related, demonstrating at least 97.4 % similarity. 16S rRNA gene sequence comparison of 20 selected isolates confirmed the monophyly of the SP group and revealed 98.5 %-100 % similarity between isolates. A blast search using the 16S rRNA gene sequences showed that the highest similarity outside the SP group was 95.5 % to an unclassified rat isolate. The single strain, P625, representing the Stewart-Letscher group showed the highest 16S rRNA gene similarity (94.9-95.5 %) to members of the SP group. recN gene sequence analysis of 11 representative strains resulted in similarities of 97-100 % among the SP group strains, which showed 80 % sequence similarity to the Stewart-Letscher group strain. Sequence similarity values based on the recN gene, indicative for whole genome similarity, showed the SP group being clearly separated from established genera, whereas the Stewart-Letscher group strain was associated with the SP group. A new genus, Necropsobacter gen. nov., with only one species, Necropsobacter rosorum sp. nov., is proposed to include all members of the SP group. The new genus can be separated from existing genera of the family Pasteurellaceae by at least three phenotypic characters. The most characteristic properties of the new genus are that haemolysis is not observed on bovine blood agar, positive reactions are observed in the porphyrin test, acid is produced from (+)-L-arabinose, (+)-D-xylose, dulcitol, (+)-D-galactose, (+)-D-mannose, maltose and melibiose, and negative reactions are observed for symbiotic growth, urease, ornithine decarboxylase and indole. Previous publications have documented that both ubiquinones and demethylmenaquinone were produced by the proposed type strain of the new genus, Michel A/76(T), and that the major polyamine of representative strains (type strain not included) of the genus is 1,3-diaminopropane, spermidine is present in moderate amounts and putrescine and spermine are detectable only in minor amounts. The major fatty acids of strain Michel A/76(T) are C(14 : 0), C(16 : 0), C(16:1)omega7c and summed feature C(14 : 0) 3-OH/iso-C(16 : 1) I. This fatty acid profile is typical for members of the family Pasteurellaceae. The G+C content of DNA of strain Michel A/76(T) was estimated to be 52.5 mol% in a previous investigation. The type strain is P709(T) ( = Michel A/76(T) = CCUG 28028(T) = CIP 110147(T) = CCM 7802(T)).

PCR-based diagnosis of Naegleria sp. infection in formalin-fixed and paraffin-embedded brain sections

We developed a real-time PCR which allowed the highly sensitive detection of Naegleria fowleri in histological brain tissue sections from experimentally infected mice. This genus-specific small-subunit (18S) rRNA gene-based PCR can complement conventional (immuno-) histology for the diagnosis of primary amoebic meningoencephalitis in paraffin-embedded brain necropsy specimens that had been fixed in formalin buffered with phosphate-buffered saline.

Occurrence of Leishmania sp. in cutaneous lesions of horses in Central Europe

The present report describes a novel etiological agent of cutaneous leishmaniasis in horses that, at least for some cases, sporadically appeared as autochthonous infections in geographically distant regions of Germany and Switzerland. The infection was initially diagnosed upon clinical and immunohistological findings. Subsequent comparative sequence analysis of diagnostic PCR products from the internal transcribed spacer 1 (ITS1) of ssrRNA classified the respective isolates as neither Old World nor New World Leishmania species. However, four isolates subjected to molecular analyses all exhibited a close phylogenetic relationship to Leishmania sp. siamensis, an organism recently identified in a visceral leishmaniasis patient from Thailand. Future investigations will demonstrate if this form of leishmaniasis represents an emerging, and perhaps zoonotic, disease of European, or even global, importance.

Sensitivity of pelagic calcification to ocean acidification

Ocean acidification might reduce the ability of calcifying plankton to produce and maintain their shells of calcite, or of aragonite, the more soluble form of CaCO3. In addition to possibly large biological impacts, reduced CaCO3 production corresponds to a negative feedback on atmospheric CO2. In order to explore the sensitivity of the ocean carbon cycle to increasing concentrations of atmospheric CO2, we use the new biogeochemical Bern3D/PISCES model. The model reproduces the large scale distributions of biogeochemical tracers. With a range of sensitivity studies, we explore the effect of (i) using different parameterizations of CaCO3 production fitted to available laboratory and field experiments, of (ii) letting calcite and aragonite be produced by auto- and heterotrophic plankton groups, and of (iii) using carbon emissions from the range of the most recent IPCC Representative Concentration Pathways (RCP). Under a high-emission scenario, the CaCO3 production of all the model versions decreases from ~1 Pg C yr−1 to between 0.36 and 0.82 Pg C yr−1 by the year 2100. The changes in CaCO3 production and dissolution resulting from ocean acidification provide only a small feedback on atmospheric CO2 of −1 to −11 ppm by the year 2100, despite the wide range of parameterizations, model versions and scenarios included in our study. A potential upper limit of the CO2-calcification/dissolution feedback of −30 ppm by the year 2100 is computed by setting calcification to zero after 2000 in a high 21st century emission scenario. The similarity of feedback estimates yielded by the model version with calcite produced by nanophytoplankton and the one with calcite, respectively aragonite produced by mesozooplankton suggests that expending biogeochemical models to calcifying zooplankton might not be needed to simulate biogeochemical impacts on the marine carbonate cycle. The changes in saturation state confirm previous studies indicating that future anthropogenic CO2 emissions may lead to irreversible changes in ΩA for several centuries. Furthermore, due to the long-term changes in the deep ocean, the ratio of open water CaCO3 dissolution to production stabilizes by the year 2500 at a value that is 30–50% higher than at pre-industrial times when carbon emissions are set to zero after 2100.

A survey on intestinal parasites of livestock guardian dogs and herding dogs in Switzerland

The present study describes the occurrence of intestinal parasite infections in livestock guardian dogs and herding dogs. A total of 71 guardian dogs (more than half of the total number of guardian dogs in Switzerland) and 21 herding dogs were coprologically examined, using a combined sedimentation-flotation method. In 21 (23 %) of the dogs intestinal parasites were detected, and 8 (8.7 %) of these dogs shed either sporocysts of Sarcocystis sp. (n = 6) or taeniid eggs (n = 2). The evaluation of questionnaires providing data on age, origin and deworming schemes of the dogs completed the study.

Truncated recombinant human SP-D attenuates emphysema and type II cell changes in SP-D deficient mice

BACKGROUND: Surfactant protein D (SP-D) deficient mice develop emphysema-like pathology associated with focal accumulations of foamy alveolar macrophages, an excess of surfactant phospholipids in the alveolar space and both hypertrophy and hyperplasia of alveolar type II cells. These findings are associated with a chronic inflammatory state. Treatment of SP-D deficient mice with a truncated recombinant fragment of human SP-D (rfhSP-D) has been shown to decrease the lipidosis and alveolar macrophage accumulation as well as production of proinflammatory chemokines. The aim of this study was to investigate if rfhSP-D treatment reduces the structural abnormalities in parenchymal architecture and type II cells characteristic of SP-D deficiency. METHODS: SP-D knock-out mice, aged 3 weeks, 6 weeks and 9 weeks were treated with rfhSP-D for 9, 6 and 3 weeks, respectively. All mice were sacrificed at age 12 weeks and compared to both PBS treated SP-D deficient and wild-type groups. Lung structure was quantified by design-based stereology at the light and electron microscopic level. Emphasis was put on quantification of emphysema, type II cell changes and intracellular surfactant. Data were analysed with two sided non-parametric Mann-Whitney U-test. MAIN RESULTS: After 3 weeks of treatment, alveolar number was higher and mean alveolar size was smaller compared to saline-treated SP-D knock-out controls. There was no significant difference concerning these indices of pulmonary emphysema within rfhSP-D treated groups. Type II cell number and size were smaller as a consequence of treatment. The total volume of lamellar bodies per type II cell and per lung was smaller after 6 weeks of treatment. CONCLUSION: Treatment of SP-D deficient mice with rfhSP-D leads to a reduction in the degree of emphysema and a correction of type II cell hyperplasia and hypertrophy. This supports the concept that rfhSP-D might become a therapeutic option in diseases that are characterized by decreased SP-D levels in the lung.

Role of protein translocation pathways across the endoplasmic reticulum in Trypanosoma brucei

The translocation of secretory and membrane proteins across the endoplasmic reticulum (ER) membrane is mediated by co-translational (via the signal recognition particle (SRP)) and post-translational mechanisms. In this study, we investigated the relative contributions of these two pathways in trypanosomes. A homologue of SEC71, which functions in the post-translocation chaperone pathway in yeast, was identified and silenced by RNA interference. This factor is essential for parasite viability. In SEC71-silenced cells, signal peptide (SP)-containing proteins traversed the ER, but several were mislocalized, whereas polytopic membrane protein biogenesis was unaffected. Surprisingly trypanosomes can interchangeably utilize two of the pathways to translocate SP-containing proteins except for glycosylphosphatidylinositol-anchored proteins, whose level was reduced in SEC71-silenced cells but not in cells depleted for SRP68, an SRP-binding protein. Entry of SP-containing proteins to the ER was significantly blocked only in cells co-silenced for the two translocation pathways (SEC71 and SRP68). SEC63, a factor essential for both translocation pathways in yeast, was identified and silenced by RNA interference. SEC63 silencing affected entry to the ER of both SP-containing proteins and polytopic membrane proteins, suggesting that, as in yeast, this factor is essential for both translocation pathways in vivo. This study suggests that, unlike bacteria or other eukaryotes, trypanosomes are generally promiscuous in their choice of mechanism for translocating SP-containing proteins to the ER, although the SRP-independent pathway is favored for glycosylphosphatidylinositol-anchored proteins, which are the most abundant surface proteins in these parasites.

Characterization of a stem cell population in lung cancer A549 cells

We isolated a stem cell subpopulation from human lung cancer A549 cells using FACS/Hoechst 33342. This side population (SP), which comprised 24% of the total cell population, totally disappeared after treatment with the selective ABCG 2 inhibitor fumitremorgin C. In a repopulation study, isolated SP and non-SP cells were each able to generate a heterogeneous population of SP and non-SP cells, but this repopulation occurred more rapidly in SP cells than non-SP. An MTT assay and cell cycle distribution analysis reveal a similar profile between SP and non-SP groups. However, in the presence of doxorubicin (DOX) and methotrexate (MTX), SP cells showed significantly lower Annexin V staining when compared to non-SP cells. Taken together, these results demonstrate that SP cells have an active regeneration capacity and high anti-apoptotic activity compared with non-SP cells. Furthermore, our GeneChip data revealed a heightened mRNA expression of ABCG2 and ABCC2 in SP cells. Overall these data explain why the SP of A549 has a unique ability to resist DOX and MTX treatments. Therefore, we suggest that the expression of the ABCG2 transporter plays an important role in the multidrug resistance phenotype of A549 SP cells.

Questing Dermacentor reticulatus harbouring Babesia canis DNA associated with outbreaks of canine babesiosis in the Swiss Midlands

In 2011 and 2012, outbreaks of clinical canine babesiosis were observed in 2 areas of the Swiss Midlands that had no history of this disease so far. In one area, cases of canine babesiosis occurred over 2 consecutive tick seasons. The outbreaks involved 29 dogs, 4 of which died. All dogs were infected with large Babesia sp. as diagnosed in Giemsa-stained blood smears and/or PCR. These were identified as B. canis (formerly known as B. canis canis) by subsequent partial sequencing of the 18S rRNA gene of Babesia sp. Interestingly, the sequence indicated either a genotype with heterogeneity in the ssrRNA gene copies or double infection with different B. canis isolates. None of the dogs had a recent travel history, but one had frequently travelled to Hungary and had suffered twice from clinical babesiosis 18 and 24 months prior to the outbreak in autumn 2011. Retrospective sequencing of a stored blood DNA sample of this dog revealed B. canis, with an identical sequence to the Babesia involved in the outbreaks. For the first time in Switzerland, the partial 18S rRNA gene of B. canis could be amplified from DNA isolated from 19 out of 23 adult Dermacentor reticulatus ticks flagged in the same area. The sequence was identical to that found in the dogs. Furthermore, one affected dog carried a female D. reticulatus tick harbouring B. canis DNA. Our findings illustrate that, under favourable biogeographic and climatic conditions, the life-cycle of B. canis can relatively rapidly establish itself in previously non-endemic areas. Canine babesiosis should therefore always be a differential diagnosis when dogs with typical clinical signs are presented, regardless of known endemic areas.

Genetic diversity among Mycoplasma species bovine group 7: clonal isolates from an outbreak of polyarthritis, mastitis, and abortion in dairy cattle

A comprehensive genetic analysis of 60 Mycoplasma sp. bovine group 7 isolates from different geographic origins and epidemiological settings is presented. Twenty-four isolates were recovered from the joints of calves during sporadic episodes of polyarthritis in geographically distinct regions of Queensland and New South Wales, Australia, including two clones of the type strain PG5O. A further three Australian isolates were also recovered from the tympanic bulla, retropharyngeal lymph node and the lung and another three isolates had unconfirmed histories. Six isolates originated from Germany, Portugal, Nigeria, and France. Twenty-four epidemiologically related isolates of Mycoplasma sp. bovine group 7 were recovered from multiple tissue sites and body fluids of infected calves with polyarthritis, mastitic milk, and from the stomach contents, lung and liver from aborted foetuses in three large, centrally managed dairy herds in New South Wales, Australia. Restriction endonuclease analysis (REA) of genomic DNA differentiated 29 Cfol profiles among these 60 isolates and grouped all 24 epidemiologically related isolates in a defined pattern showing a clonal origin. Three isolates of this clonal cluster were recovered from mastitic milk and the synovial exudate of clinically-affected calves and appeared sporadically for periods up to 18 months after the initial outbreak of polyarthritis indicating a persistent, close association of the organism with cattle in these herds. The Cfol profile representative of the clonal cluster was distinguishable from profiles of isolates recovered from multiple, unrelated cases of polyarthritis in Queensland and New South Wales and from other countries. All 24 isolates from the clonal cluster possessed a plasmid (pBG7AU) with a molecular size of 1022 bp. DNA sequence analysis of pBG7AU identified two open reading frames sharing 81 and 99% DNA sequence similarity with hypothetical replication control proteins A and B respectively, previously described in plasmid pADB201 isolated from M. mycoides subspecies mycoides. Other isolates of bovine group 7, epidemiologically unrelated to the clonal cluster, including two clones of the type strain PG5O, possessed a similar-sized plasmid. These data confirm that Mycoplasma sp. bovine group 7 is capable of migrating to, and multiplying within, different tissue sites within a single animal and among different animals within a herd.

Molecular and serological diagnosis of Bartonella infection in 61 dogs from the United States

Molecular diagnosis of canine bartonellosis can be extremely challenging and often requires the use of an enrichment culture approach followed by PCR amplification of bacterial DNA. HYPOTHESES: (1) The use of enrichment culture with PCR will increase molecular detection of bacteremia and will expand the diversity of Bartonella species detected. (2) Serological testing for Bartonella henselae and Bartonella vinsonii subsp. berkhoffii does not correlate with documentation of bacteremia. ANIMALS: Between 2003 and 2009, 924 samples from 663 dogs were submitted to the North Carolina State University, College of Veterinary Medicine, Vector Borne Diseases Diagnostic Laboratory for diagnostic testing with the Bartonella α-Proteobacteria growth medium (BAPGM) platform. Test results and medical records of those dogs were retrospectively reviewed. METHODS: PCR amplification of Bartonella sp. DNA after extraction from patient samples was compared with PCR after BAPGM enrichment culture. Indirect immunofluorescent antibody assays, used to detect B. henselae and B. vinsonii subsp. berkhoffii antibodies, were compared with PCR. RESULTS: Sixty-one of 663 dogs were culture positive or had Bartonella DNA detected by PCR, including B. henselae (30/61), B. vinsonii subsp. berkhoffii (17/61), Bartonella koehlerae (7/61), Bartonella volans-like (2/61), and Bartonella bovis (2/61). Coinfection with more than 1 Bartonella sp. was documented in 9/61 dogs. BAPGM culture was required for PCR detection in 32/61 cases. Only 7/19 and 4/10 infected dogs tested by IFA were B. henselae and B. vinsonii subsp. berkhoffii seroreactive, respectively. CONCLUSIONS AND CLINICAL IMPORTANCE: Dogs were most often infected with B. henselae or B. vinsonii subsp. berkhoffii based on PCR and enrichment culture, coinfection was documented, and various Bartonella species were identified. Most infected dogs did not have detectable Bartonella antibodies.