Natural products and ring-closing metathesis: synthesis of sterically congested olefins.

Covering: 2002 to August 2017.This review highlights recent RCM reactions towards the synthesis of sterically congested natural products. It offers an insight into various synthetic targets and approaches and provides information on the evolution of catalysts as powerful tools enabling the use of increasingly challenging diene precursors.

Brucellosis of the common vole (Microtus arvalis).

A systemic disease occurred in a wild population of the common vole Microtus arvalis in South Moravia (Czech Republic) during the years 1999-2003. Acute infections were characterized by edema of extremities, occasionally with colliquating abscesses, arthritis, lymphadenitis, perforations of the skin resulting from colliquated abscesses, orchitis, and peritoneal granulomas. From the clinical samples, small Gram-negative coccobacilli were isolated and identified as Ochrobactrum intermedium by API 20NE and colistin sensitivity profiles. However, subsequent rrs (16S rRNA) and recA (recombinase A) gene sequencing analysis of two isolates (CCM 4915=CAPM 6434; CCM 4916=CAPM 6435) identified them as Brucella sp. with sequence identities of 100% to other Brucella spp. Analysis of the omp2a/b genes confirmed the two isolates as Brucella. In AMOS polymerase chain reaction (PCR), a 2000-bp fragment was generated that was not seen in other brucellae. Experimental infection of outbred ICR mice with these isolates resulted in a mortality rate of 50%. Based on the results of the molecular investigations and the mortality observed in experimentally infected mice we conclude that the epizootic was caused by Brucella sp. and not by Ochrobactrum intermedium. The study demonstrates the limitations of commercial biochemical test systems in accurately differentiating among Ochrobactrum and Brucella.

An IL-27/Lag3 axis enhances Foxp3+ regulatory T cell-suppressive function and therapeutic efficacy.

Foxp3-expressing regulatory T cells (Tregs) are central regulators of immune homeostasis and tolerance. As it has been suggested that proper Treg function is compromised under inflammatory conditions, seeking for a pathway that enhances or stabilizes Treg function is a subject of considerable interest. We report that interleukin (IL)-27, an IL-12 family cytokine known to have both pro- and anti-inflammatory roles in T cells, plays a pivotal role in enhancing Treg function to control T cell-induced colitis, a model for inflammatory bowel disease (IBD) in humans. Unlike wild-type (WT) Tregs capable of inhibiting colitogenic T-cell expansion and inflammatory cytokine expression, IL-27R-deficient Tregs were unable to downregulate inflammatory T-cell responses. Tregs stimulated with IL-27 expressed substantially improved suppressive function in vitro and in vivo. IL-27 stimulation of Tregs induced expression of Lag3, a surface molecule implicated in negatively regulating immune responses. Lag3 expression in Tregs was critical to mediate Treg function in suppressing colitogenic responses. Human Tregs also displayed enhanced suppressive function and Lag3 expression following IL-27 stimulation. Collectively, these results highlight a novel function for the IL-27/Lag3 axis in modulating Treg regulation of inflammatory responses in the intestine.

West Nile virus investigations in South Moravia, Czechland.

Seven virus isolates were obtained from 11,334 mosquitoes after the 1997 Morava River flooding in South Moravia (Czech Republic): 6 strains of Tahyna bunyavirus, California antigenic group (5 from Aedes vexans, 1 from Ae. cinereus), and 1 strain of West Nile flavivirus (WNV) from Culex pipiens. In 1999, one isolate of Tahyna virus from Ae. vexans and one isolate of WNV from Cx. pipiens were recovered from a total of 14,354 mosquitoes examined in the same area, whereas no virus was detected there in 1,179 overwintering mosquitoes (mostly Cx. pipiens) in March 2000. The infection rate of mosquitoes with arboviruses was significantly higher in 1997, the year of the flood and an enormously high population density of mosquitoes. Antibodies neutralizing WNV were detected in 13 of 619 (2.1%) hospitalized patients or persons seeking outpatient clinics of the area in 1997. Five of the seroreactors revealed clinical symptoms compatible with West Nile fever: in 2 of them (children), recent infection with WNV was confirmed by a significant increase of antibody titer between acute and convalescent serum samples.

Candidatus Ehrlichia walkerii: a new Ehrlichia detected in Ixodes ricinus tick collected from asymptomatic humans in Northern Italy.

Ixodes ricinus harbors many infectious agents pathogenic for humans. A cause of fever is found in less than 50% of patients exposed to ticks. Investigations on 359 Ixodes ticks removed from asymptomatic patients in Northern Italy revealed the presence of a new ehrlichial agent in 10 ticks. Comparison of the 16S rRNA and the gltA gene sequences showed the organism is most closely related to Ehrlichia ruminantium. We propose this new Ehrlichia be named "candidatus Ehrlichia walkerii."

Prevalence of Rickettsia spp. in Dermacentor marginatus ticks removed from game pigs (Sus scrofa) in southern France.

During the months of October and November 2000 a total of 70 Dermacentor marginatus ticks were removed from different game pigs Sus scrofa in southern France and investigated by PCR using primers derived from the citrate synthase (gltA) and the outer membrane protein (OmpB) genes of rickettsiae. Based on sequence analysis of 1,150 bp fragment of gltA, Rickettsia slovaca was identified in 11 ticks (15.7%). These results confirm that Rickettsia slovaca, an emerging pathogen is highly prevalent in Dermacentor marginatus ticks in France. Moreover, a new Rickettsia genotype was detected in one specimen (1.4%). The pathogenic role of this new rickettsia has yet to be demonstrated.

Genetic diversity of bacterial agents detected in ticks removed from asymptomatic patients in northeastern Italy.

A total of 360 ticks were removed from 353 asymptomatic subjects in Belluno Province, Italy and surrounding areas, from 1998 to 2001. Ticks were identified as Ixodes ricinus (357), Ixodes hexagonus (1), Rhipicephalus sanguineus (1), and Ixodes ventalloi (1). Tick DNA was investigated by PCR and subsequent sequencing of amplified products to identity associated bacterial agents. Primers targeting different genes of Rickettsia (gltA and OmpA), Borrelia (16S rDNA, rpoB), Francisella (16S rDNA), and all genera members of the Anaplasmataceae (16S rDNA), were used. DNA of bacterial agents was identified in 28 Ixodes ricinus specimens (7.8%). Rickettsia helvetica was detected in 7 ticks. Rickettsia sp. IRS4 and Borrelia afzelii was detected in 4 ticks each. B. garinii and B. valaisiana were identified in one tick each. Anaplasma phagocytophilum, the agent of human granulocytic ehrlichiosis, was identified in 1 specimen of I. ricinus. A new Ehrlichia sp. ("Candidatus Ehrlichia walkerii", sp. nov.) was identified in 10 I. ricinus specimens.

Charting methods to monitor the operational performance of ELISA method for the detection of antibodies against trypanosomes.

Four indirect enzyme-linked immunosorbent assays (ELISAs) for the detection of antibody against trypanosomes using antigen-precoated plates (Trypanosoma congolense and T. vivax) were used in 15 veterinary diagnostic laboratories in Africa and Europe. The study provided data allowing an evaluation of charting methods with respect to the operational performance of each ELISA. Data from standardised internal quality control (IQC) samples were plotted on charts and used as the assay performance indicators with reference to expected upper and lower control limits. Based on unprocessed (optical density) and normalised absorbance values (calculated as a percentage positivity of a control), dispersion of values from the expected data range was estimated plotting the location and deviation of the values. In addition, assay precision was estimated plotting the distribution of coefficients of variation<10% of the IQCs. Binding ratios of controls were calculated to estimate the assay proficiency with respect to the accuracy of assessing that the IQC samples tested positive or negative in the test proper. The graphical analysis of dispersion of absorbance values in combination with assay precision and proficiency criteria was considered fully satisfactory to evaluate the operational performance of the ELISAs and provided useful decision criteria for plate acceptance and rejection. The establishment of standardised and transparent IQC data charting methods for the indirect ELISAs provided an increased measure of confidence to national laboratories with respect to their reports on disease occurrence. Moreover, the relative assay performances between all laboratories were examined using summary data charts with reference to the performance criteria described. The IQC data were also examined using modified Youden plot analysis demonstrating that indirect ELISA methods can be successfully applied at diagnostic laboratories in the tropics for monitoring trypanosomosis control programmes.

First evidence of Anaplasma platys in Rhipicephalus sanguineus (Acari: Ixodida) collected from dogs in Africa.

A total of 27 ticks, comprising Rhipicephalus sanguineus (Latreille) (n = 21), Haemaphysalis leachi (Andouin) (n = 4) and Haemaphysalis paraleachi (Camicas, Hoogstraal & El Kammah) (n = 2) were recovered from two clinically healthy female dogs in the Democratic Republic of the Congo. DNA of Anaplasma platys was detected in a female R. sanguineus, using primers derived from the 16S rRNA gene, which amplify members of the family Anaplasmataceae . Anaplasma platys DNA was also detected in the blood of one of the dogs. Phylogenetic analysis based on partial sequences of the 16S rRNA, the gltA and the groEL genes ranged the detected agent within the Anaplasma clade. This is the first reported detection of A. platys in ticks in Africa. This finding raises the question of the possible involvement of R. sanguineus in A. platys infection of dogs.

Disruption of the Wolbachia surface protein gene wspB by a transposable element in mosquitoes of the Culex pipiens complex (Diptera, Culicidae).

Culex pipiens quinquefasciatus Say and Culex pipiens pipiens Linnaeus are sibling species incriminated as important vectors of emerging and re-emerging infectious diseases worldwide. The two forms differ little morphologically and are differentiated mainly based upon ecological, behavioural, physiological and genetic traits. Within the North American zone of sympatry, populations of Cx. p. quinquefasciatus and Cx. p. pipiens undergo extensive introgression and hybrid forms have been reported in nature. Both Cx. p. quinquefasciatus and Cx. p. pipiens are infected with the endosymbiotic bacteria Wolbachia pipientis. Here, we report the presence of a transposable element belonging to the IS256 family (IS256wPip) associated with Wolbachia in both Cx. p. quinquefasciatus and Cx. p. pipiens populations. Using reverse transcriptase PCR and sequence analysis, we show that IS256wPip has disrupted the wspB locus, a paralogue of the Wolbachia outer membrane protein (wspA) gene. The inactivation of the wspB appears to be specific to Cx. p. quinquefasciatus and to hybrids of the two forms, and was not observed in the surveyed Cx. p. pipiens mosquitoes. Our results support the hypothesis of a different origin of North American Cx. p. quinquefasciatus and Cx. p. pipiens populations. The flux of mobile genetic elements in the Wolbachia wPip genome could explain the high level of crossing types observed among different Culex populations. The insertion of IS256wPip into wspB may comprise a genetic candidate for discriminating Wolbachia symbionts in Culex.

Molecular discrimination of Wolbachia in the Culex pipiens complex: evidence for variable bacteriophage hyperparasitism.

The medically important members of the Culex pipiens species complex provide an enigma for systematists, evolutionary biologists, and vector biologists. The species complex is composed of forms that differ in their ecology, behaviour, physiology and vector competence. Cytoplasmic incompatibility (CI) caused by endosymbiotic Wolbachia bacteria is thought to play an important role in restricting gene flow and the evolution of the Culex complex. Here we describe the first molecular marker useful for discriminating between Wolbachia infections in Culex. A putative bacteriophage locus (orf7) varies between Culex forms in copy number and sequence. We provide evidence that the orf7 loci are strictly associated with Wolbachia and are maternally inherited.

[Trypanosomiasis and trypanocidal treatments in Ndama calves in a glossina infected area (the ranch of Madina-Diassa in Mali)]. / Trypanosomose et traitements trypanocides chez le veau Ndama en milieu fortement infesté de glossines (ranch de Madina-Diassa au Mali).

This work aims at contributing to the knowledge of trypanosomiasis epidemiology in calves of trypanotolerant breeds and at defining an appropriate treatment to improve the survival of such calves in a tsetse infested area. The first study was a parasitological survey of 100 calves from the day of birth to the age of one year. According to the results of this survey, the period from birth to three months is a "critical" moment in the life of the calves, due to a high infection rate and mortality related to trypanosomiasis. The purpose of the second study was to investigate the possible interference of early trypanocidal treatments with the further expression of trypanotolerance. For this purpose three groups of over one-year old animals were established. The groups had different trypanosomiasis history due to the different treatments they had undergone during their first year of life. All the animals had been exposed to trypanosomiasis without treatment and followed up parasitologically and clinically during the second year. The results showed no interference of early trypanocidal treatments (including preventive ones) with the expression of resistance in potentially trypanotolerant animals.

[Epidemiology of trypanosomiasis caused by T. evansi in camels in mali: results of parasitological and clinical survey]. / Epidémiologie de la trypanosomose à T. evansi chez le dromadaire au Mali: résultats d'enquêtes parasitologiques et cliniques.

An epidemiological study of trypanosomiasis was conducted in the rearing areas of dromedary camels in Mali. According to the parasitological and clinical surveys performed, the overall infection rates were 9.5% (29/305) in Western Sahel (region I) and 4.5% (28/627) in the areas of Tombouctou and Gao (region II). The proportion of contaminated herds was 55% in region I and 68% in region II and in some herds the infection rate exceeded 50%. The surveys showed a trend for increasing parasitological prevalence with age. While it was almost nonexistent in young camels less than one year old, it increased with age and reached a maximum in 2 to 5-year old camels. The authors showed that the infection has a significantly negative effect on PCV and on the overall status of the animals, confirming the pathogenicity of Trypanosoma evansi in dromedary camels. This trypanosome is almost the only species detected in the dromedary camel in Mali and it does not seem to cause infections in other animals reared in the same environment.

Isolation of Borrelia afzelii from overwintering Culex pipiens biotype molestus mosquitoes.

During the years 1995-1996, a total of 1,743 overwintering Culex pipiens biotype molestus female mosquitoes were tested for the presence of spirochetes in several localities in South Moravia, Czech Republic.The spirochetes were observed in 5% of the mosquitoes investigated. One of the five isolated strains of spirochetes (BR-84) was identified as Borrelia afzelii. The potential role of mosquitoes in the ecology and epidemiology of Lyme disease (LD) borreliae should be further investigated.