The key steroidogenic enzyme 3β-hydroxysteroid dehydrogenase in <Emphasis Type="Italic">Taenia solium</Emphasis> and <Emphasis Type="Italic">Taenia crassiceps</Emphasis> (WFU)

Larval and adult stages of Taenia solium and Taenia crassiceps WFU strain were analyzed by histochemical and biochemical methods to determine the existence of steroid pathways. The presence of the key enzyme 3β-hydroxisteroid-dehydrogenase (3β-HSD) was examined in frozen sections of cysticerci obtained from mice and segments of tapeworms obtained from the intestine of hamsters. 3β-HSD activity was detected by nitroblue-tetrazolium products after incubation with dehydroepiandrosterone, androstendiol, or pregnenolone. Tapeworm tissues exhibited 3β-HSD activity in the subtegumentary areas of the neck and immature proglottids following incubation with androstendiol, as well as surrounding the testes in mature proglottids. T. solium cysticerci exhibited 3β-HSD activity in the subtegumentary tissues. The synthesis of steroid hormones involving the activity of 3β-HSD was studied in cysticerci or tapeworms incubated in the presence of tritiated steroid precursors. The culture media were analyzed by thin layer chromatography and showed synthesis of androstendiol, testosterone, and 17β-estradiol by cysticerci, androstendiol, and 17β-estradiol by tapeworms. The results strongly suggest the activity of 3β-HSD in taeniid parasites that have at least a part of the enzymatic chain required for androgen and estrogen synthesis and that the enzymes are present in the larval stage and from the early strobilar stages to the mature proglottids.     

<Emphasis Type="Italic">Phlebotomus</Emphasis> (<Emphasis Type="Italic">Euphlebotomus</Emphasis>) <Emphasis Type="Italic">mascomai</Emphasis> n. sp. (Diptera–Psychodidae)

A new species of sandfly is described from limestone caves in Thailand. The inclusion of this species in the subgenus Euphlebotomus is justified on the basis of characters of the male genitalia (paramere, basal lobe). The male–female gathering in the same taxon is based on ecological (cavernicolous species), morphological (length of male genital filaments and female spermathecal ducts) and molecular (homology of cytochrome b mt DNA sequences) criteria. A differential diagnosis between Phlebotomus mascomai n. sp. and P. argentipes Annandale & Brunetti, the vector of Leishmania donovani (Laveran & Mesnil) in India, is proposed based on several morphological characters like antennal formula and genitalia.     

The <Emphasis Type="Italic">rpl5</Emphasis>–<Emphasis Type="Italic">rps14</Emphasis> mitochondrial region: a hot spot for DNA rearrangements in <Emphasis Type="Italic">Solanum</Emphasis> spp. somatic hybrids

Southern analysis with rpl5 and rps14 mtDNA gene probes of Solanum tuberosum, S. commersonii and a sample of somatic hybrids detected polymorphisms between parents and the appearance of a novel restriction fragment in various hybrids. In one of them, detailed mtDNA analyses revealed various configurations of the rpl5–rps14 region present at different stoichiometries. Multiple inter-parental recombination events across homologous sequences were assumed to have caused these rearrangements. Sequence similarity searches detected one sequence putatively involved in the recombination upstream of the rpl5 gene. The presence of a second recombinogenic sequence was inferred. We propose two models to explain the mechanism responsible for obtaining the different rpl5–rps14 arrangements shown after somatic hybridization. Variability in the rpl5–rps14 region observed in both the parental species and their somatic hybrids suggests this region is a hot spot for mtDNA rearrangements in Solanum spp.Contribution no. 39 from the Institute of Plant Genetics, Research Division of Portici.Communicated by A. Brennicke     

<Emphasis Type="Italic">RNR4</Emphasis> mutant alleles <Emphasis Type="Italic">pso3-1</Emphasis> and <Emphasis Type="Italic">rnr4Δ</Emphasis> block induced mutation in <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis>

The PSO3 gene of Saccharomyces cerevisiae was molecularly cloned by complementing the cold-sensitivity phenotype of a pso3-1 mutant and was found to be allelic to RNR4, encoding one of the two DNA damage-inducible small subunits of the ribonucleotide reductase (RNR) complex. Compared to a rnr4Δ mutant that allows only very little mutation induction at very low doses of 254_nm ultraviolet light (UVC), the pso3-1 mutant allele confers leakiness in that it permits some DNA damage-induced mutagenesis at low doses of UVC. Similarly, the pso3 mutant is slightly less sensitive to UVC than an rnr4Δ mutant. Cloning and sequencing of the RNR4 locus of the pso3-1 mutant revealed that its intermediate phenotype is attributable to a G → A transition at nucleotide 352, leading to replacement of glycine by arginine [G118R] in the mutant’s protein. Both RNR4 mutant alleles confer significantly less sensitivity to UVC than mutant alleles of non-UVC-mutable REV3, indicating that, apart from nucleotide excision repair, RAD6-dependent error-free DNA repair may still be functional. The phenotype of a strongly reduced UVC-induced mutagenesis for rnr4 mutant alleles has not yet been described; it suggests the importance of this gene for a fully functional RNR providing correct amounts of DNA precursor molecules, thereby, allowing translesion synthesis (error-prone) of UVC-damaged DNA. Stationary phase cells of the rnr4Δ mutant, but not of the original pso3-1 mutant, are swollen with a fourfold to eightfold increase in volume. The central role of RNR in DNA precursor metabolism and its complex regulation allow for several modes of suppression that may influence the phenotypes of RNR4 mutants, especially those containing the leaky pso3-1 mutant allele.     

Comparative genomic sequence analysis of novel <Emphasis Type="Italic">Helicoverpa</Emphasis> <Emphasis Type="Italic">armigera</Emphasis> nucleopolyhedrovirus (NPV) isolated from Kenya and three other previously sequenced <Emphasis Type="Italic">Helicoverpa</Emphasis> spp. NPVs

A newly cloned Helicoverpa armigera nucleopolyhedrovirus (HearNPV) from Kenya, HearNPV-NNg1, has a higher insecticidal activity than HearNPV-G4, which also exhibits lower insecticidal activity than HearNPV-C1. In the search for genes and/or nucleotide sequences that might be involved in the observed virulence differences among Helicoverpa spp. NPVs, the entire genome of NNg1 was sequenced and compared with previously sequenced genomes of G4, C1 and Helicoverpa zea single-nucleocapsid NPV (Hz). The NNg1 genome was 132,425 bp in length, with a total of 143 putative open reading frames (ORFs), and shared high levels of overall amino acid and nucleotide sequence identities with G4, C1 and Hz. Three NNg1 ORFs, ORF5, ORF100 and ORF124, which were shared with C1, were absent in G4 and Hz, while NNg1 and C1 were missing a homologue of G4/Hz ORF5. Another three ORFs, ORF60 (bro-b), ORF119 and ORF120, and one direct repeat sequence (dr) were unique to NNg1. Relative to the overall nucleotide sequence identity, lower sequence identities were observed between NNg1 hrs and the homologous hrs in the other three Helicoverpa spp. NPVs, despite containing the same number of hrs located at essentially the same positions on the genomes. Differences were also observed between NNg1 and each of the other three Helicoverpa spp. NPVs in the diversity of bro genes encoded on the genomes. These results indicate several putative genes and nucleotide sequences that may be responsible for the virulence differences observed among Helicoverpa spp., yet the specific genes and/or nucleotide sequences responsible have not been identified.     

Schizogonic stages of<Emphasis Type="Italic"> Haemoproteus</Emphasis> from Wenyon’s Baghdad sparrows are also found in<Emphasis Type="Italic"> Passer domesticus biblicus</Emphasis> in Israel

Schizont bodies reminiscent of those described by Wenyon from Baghdad sparrows were found in the liver and lungs of an Israeli house sparrow (Passer domesticus biblicus Hartert 1904) infected with Haemoproteus passeris Kruse, 1890. All observed schizonts were composed of packed assemblages of walled compartments, each holding a differentiating schizogonic body. The schizogonic bodies in the various compartments demonstrated sequential stages in the differentiation process from a compact multinucleate cytoplasmic mass to massive formation of multiple merozoites. Young non-differentiated schizont assemblages reached 0.2×0.25 mm in size and the fully differentiated ones, containing merozoites, were double or triple that. Among species of Haemopterus, division to compartmented and single-plasmodium schizonts could not be associated with any recognized generic or infrageneric division. Moreover, in some species of Haemoproteus, both types of schizogony co-existed.     

Expression of the <Emphasis Type="Italic">Acremonium chrysogenum</Emphasis><Emphasis Type="Italic">cefT</Emphasis> gene in <Emphasis Type="Italic">Penicillum chrysogenum</Emphasis> indicates that it encodes an hydrophilic β-lactam transporter

The Acremonium chryrsogenum cefT gene encoding a membrane protein of the major facilitator superfamily implicated in the cephalosporin biosynthesis in A. chrysogenum was introduced into Penicillium chrysogenum Wisconsin 54-1255 (a benzylpenicillin producer), P. chrysogenum npe6 pyrG (-) (a derivative of Wisconsin 54-1255 lacking a functional penDE gene) and P. chrysogenum TA98 (a deacetylcephalosporin producer containing the cefD1, cefD2, cefEF and cefG genes from A. chrysogenum). RT-PCR analysis revealed that the cefT gene was expressed in P. chrysogenum strains. HPLC analysis of the culture broths of the TA98 transformants showed an increase in the secretion of deacetylcephalosporin C and hydrophilic penicillins (isopenicillin N and penicillin N). P. chrysogenum Wisconsin 54-1255 strain transformed with cefT showed increased secretion of the isopenicillin N intermediate and a drastic decrease in the benzylpenicillin production. Southern and northern blot analysis indicated that the untransformed P. chrysogenum strains contain an endogenous gene similar to cefT that may be involved in the well-known secretion of the isopenicillin N intermediate. In summary, the cefT transporter is a hydrophilic beta-lactam transporter that is involved in the secretion of hydrophilic beta-lactams containing alpha-aminoadipic acid side chain (isopenicillin N, penicillin N and deacetylcephalosporin C).     

Suppression of <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis><Emphasis Type="Italic">rad27</Emphasis> null mutant phenotypes by the 5′ nuclease domain of <Emphasis Type="Italic">Escherichia coli</Emphasis> DNA polymerase I

RNA primer removal from Okazaki fragments during lagging-strand replication and the excision of damaged DNA bases requires the action of structure-specific nucleases, such as the mammalian flap endonuclease 1 (FEN-1). This nuclease contains two conserved motifs enriched with acidic amino acid residues that are important for catalytic function. Similar motifs have been identified in nucleases found in viruses, archebacteria, eubacteria, and in eukaryotes ranging from yeast to humans. Unique among these proteins, the putative FEN-1 homologue in Escherichia coli is contained within the N-terminal region of the DNA polymerase I (PolN). To demonstrate that the cellular functions of FEN-1 reside in PolN, we cloned and expressed the amino terminal domain (323 amino acid residues) of PolI in a Saccharomyces cerevisiae strain lacking the FEN-1 homologue RAD27. Overexpression of PolN suppressed, to varying degrees, phenotypes associated with a rad27 null strain. These include temperature sensitivity, Okazaki fragment processing, a mutator phenotype, a G2/M cell cycle arrest, minichromosome loss, and methyl methane sulfonate sensitivity. We purified Rad27 and PolN proteins in order to determine whether differences in their intrinsic nuclease activities or interaction with proliferating cell nuclear antigen (PCNA) could explain the partial suppression of some phenotypes. We found that the in vitro nuclease activities of Rad27 were more potent than those of PolN and the activity of Rad27, but not PolN, was stimulated by PCNA. We conclude that the N-terminal nuclease domain of E. coli polymerase I encodes a functional homologue of FEN-1.     

Cavidine Ameliorates Lipopolysaccharide-Induced Acute Lung Injury <Emphasis Type="Italic">via</Emphasis> NF-κB Signaling Pathway <Emphasis Type="Italic">in vivo</Emphasis> and <Emphasis Type="Italic">in vitro</Emphasis>

Acute lung injury (ALI) is characterized by widespread inflammation in the lungs and alveolar-capillary destruction, causing high morbidity and mortality. Cavidine, isolated from Corydalis impatiens, have been exhibited to have potent anti-inflammatory effects in previous studies. The purpose of this study was to evaluate the protective effect of cavidine on lipopolysaccharide (LPS)-induced ALI and to enunciate the underlying in vivo and in vitro mechanisms. Mice were intraperitoneally administrated with cavidine (1, 3, or 10 mg/kg) at 1 and 12 h, prior to the induction of ALI by intranasal administration of LPS (30 mg/kg). Blood samples, lung tissues, and bronchoalveolar lavage fluid (BALF) were harvested after LPS challenge. Furthermore, we used LPS-induced lung epithelial cells A549 to examine the mechanism of cavidine to lung injury. The results showed that pretreatment with cavidine significantly decreased lung wet-to-dry weight (W/D) ratio, reduced pro-inflammatory cytokine levels including TNF-α and IL-6 in BALF and serum from LPS-stimulated mice, and attenuated lung histopathological changes. In addition, western blot results showed that cavidine inhibited the phosphorylation of nuclear factor-kappaB (NF-κB) p65 and IκBα induced by LPS. In conclusion, our results demonstrate that cavidine protects against LPS-induced acute lung injury in mice via inhibiting of pro-inflammatory cytokine TNF-α and IL-6 production and NF-κB signaling pathway activation. Taken together, cavidine may be useful for the prevention and treatment of pulmonary inflammatory diseases, such as ALI.     

First report of <Emphasis Type="Italic">Cryptosporidium</Emphasis><Emphasis Type="Italic">parvum</Emphasis> ‘ferret’ genotype in American mink (<Emphasis Type="Italic">Mustela vison</Emphasis> Shreber 1777)

A total of 51 faecal samples from wild and farmed mink were analysed by a direct immunofluorescence antibody test. Cryptosporidium oocysts were identified in eight, apparently healthy, farmed American mink (Mustela vison). The isolates were identified as Cryptosporidium parvum ‘ferret’ genotype by PCR-RFLP and sequencing analysis of a 341-base-pair fragment of the Cryptosporidium oocyst wall protein (COWP) gene. This is the first report of Cryptosporidium in American mink. Genbank accession numbers of the sequences used in this study C. parvum, ‘bovine’ genotype AF266273; C. hominis, AF266265; C. parvum, ‘mouse’ genotype AF266268, C. wrairi, U35027; C. parvum, ‘ferret’ genotype AF266267; C. meleagridis, AF266266; C. parvum, ‘marsupial’ genotype AF266269; C. parvum, ‘pig’ genotype AF266270; C. canis, AF266274; C. felis, AF266263; C. baileyi, AF266276; C. serpentis, AF266275; C. andersoni, AF266262 and C. muris, AF161579     

Effects of Periodic Body Acceleration on the <Emphasis Type="Italic">In Vivo</Emphasis> Vasoactive Response to N-<Emphasis Type="Italic">w</Emphasis>-nitro–L-arginine and the <Emphasis Type="Italic">In Vitro</Emphasis> Nitric Oxide Production

Periodic acceleration (pGz), a novel method of ventilatory support, is achieved using a platform that moves cyclically in the headward–footward direction. PGz has been shown to increase vascular shear stress and regional blood flows, as well as decrease pulmonary and systemic vascular resistances. PGz also increases nitric oxide (NO) production. This study was undertaken to determine the effects of pGz on the NO inhibiting effects of N-w-nitro–L-arginine (L-NAME) in vivo, and to determine if increased NO production due to pGz could be reproduced in vitro with isolated arteries. Pigs were assigned to conventional ventilation (CV), or pGz, with no additional breathing assistance. L-NAME was infused in cumulative doses of 1, 3, 10, 30, and 100 mg/kg. Cardiac output decreased in both groups by 50%. There was also a dose-dependent increase in blood pressure, pulmonary artery pressure, and vascular resistances. However, pGz attenuated the vascular response of L-NAME. Isolated porcine aortas exposed to nonpulsatile, pulsatile, and pulsatile flow plus pGz exhibited an increase in nitrites with the addition of pulsatile flow (300%, relative to steady flow), and a further increase with pGz (1000%, relative to steady flow). It has been determined that pGz, a novel method of increasing shear stress on the vascular endothelium, attenuates the vasoactive response to L-NAME. The in vitro experiments demonstrated that increases in NO production in vivo could be reproduced in vitro, which provides the opportunity to investigate the mechanisms of cardiovascular pGz effects. © 2003 Biomedical Engineering Society. PAC2003: 8719Uv, 8719Rr, 8780-y     

Genetic regulation of arrested development in nematodes: are <Emphasis Type="Italic">age</Emphasis>-1 and <Emphasis Type="Italic">daf</Emphasis>-gene orthologs present in <Emphasis Type="Italic">Dictyocaulus viviparus</Emphasis>?

In opposite to the free-living soil nematode Caenorhabditis elegans, the genetic regulation of hypobiosis or inhibited or arrested development in parasitic nematodes is completely unknown. In C. elegans, the daf-genes or the age-1 gene are of major importance in signaling pathways regulating arrested development. To investigate if orthologs of these genes are present in the bovine lungworm Dictyocaulus viviparus, a PCR analysis with gene-specific primer combinations was performed. No orthologs of the age-1 or daf-genes could be identified in D. viviparus. The possible differences in the role of the daf-genes concerning arrested development in parasitic and free-living nematodes will be discussed.     

First description of natural <Emphasis Type="Italic">Echinococcus multilocularis</Emphasis> infections in chinchilla (<Emphasis Type="Italic">Chinchilla laniger</Emphasis>) and Prevost’s squirrel (<Emphasis Type="Italic">Callosciurus prevostii borneoensis</Emphasis>)

This report describes for the first time the occurrence of alveolar echinococcosis in two exotic rodent species in Europe. A pet chinchilla (Chinchilla laniger) was euthanized due to a painful enlargement of the abdominal cavity, and a Prevost's squirrel (Callosciurus prevostii borneoensis) was found dead in the enclosure of a zoo. At necropsy, extended liver lesions consisting of small vesicles and cysts were observed in the livers of both animals. Histological examination revealed that these cysts were composed of an outer, homogenous, eosinophilic layer and an inner, cellular germinal layer. The cysts from both animals contained numerous protoscolices. The morphological diagnosis of Echinococcus multilocularis metacestode infections was confirmed by molecular means.     

<Emphasis Type="Italic">Ex vivo</Emphasis> production of interferon-γ, tumor necrosis factor-α, and interleukin-6 by mouse macrophages during infection with <Emphasis Type="Italic">M. bovis</Emphasis> and <Emphasis Type="Italic">M. tuberculosis</Emphasis> H37Rv

Ex vivo production of IFN-γ, TNF-α, and IL-6 by mouse peritoneal macrophages was studied during successive infection with the vaccine strain M. bovis BCG and virulent strain M. tuberculosis H37Rv. The increase in the concentrations of TNF-α and IL-6 did not depend on the sequence of macrophage infection with the vaccine or virulent strain, but was related to the presence of the vaccine strain M. bovis BCG in the medium. IFN-γ production depended on infection of macrophages with the virulent strain M. tuberculosis H37Rv. The concentration of IFN-γ was maximum during primary infection with the virulent strain and did not increase after successive infection with the virulent and vaccine strain. __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 144, No. 11, pp. 552–555, November, 2007     

Is <Emphasis Type="Italic">Duhuo Jisheng Tang</Emphasis> containing <Emphasis Type="Italic">Xixin</Emphasis> safe? A four-week safety study

Background  

Though the nephrotoxicity and carcinogenicity of aristolochic acid (AA) are known, its safety in clinical usage is not clear. This study aims to evaluate the safety of Duhuo Jisheng Tang (DJT) in a four-week study to treat osteoarthritis (OA) of the knee.     

Impact of <Emphasis Type="Italic">Bacillus sphaericus</Emphasis> exposure on <Emphasis Type="Italic">Anopheles dirus</Emphasis>’s fecundity and resistance development

Mosquitoes are important vectors of many infectious diseases. Bacillus sphaericus (Bs) is an ideal larvicide and has attracted more and more attention, recently. However, the fundamental research of its application is very limited, especially on the subsequent impact of Bs exposure on mosquito’s fecundity and resistance emergence. Through bioassay, LC50 and LC95 of Bs in killing Anopheles dirus larvae were determined as 9.793 ± 1.878 IU/L and 62.4 ± 6.438 IU/L at 48 h posttreatment, 7.128 ± 0.913 IU/L and 34.385 ± 12.547 IU/L at 72 h post treatment, respectively. After being treated with a sub-lethal dose of Bs, gravidity, oviposition, hatch, pupation, and eclosion of the surviving mosquitoes were counted and analyzed to elucidate the subsequent effects of Bs exposure on the reproductive capacity of A. dirus. The result interestingly showed that the exposure of Bs significantly reduced the oviposition ability of the surviving A. dirus, without effect on egg formation/gravidity, hatch, pupation, and eclosion. The surviving mosquitoes were also maintained routinely for generations to test the sustained effect of Bs exposure on the fecundity of the offsprings. After conventional breeding for generations, the capacity of egg laying totally recovered. To explore the rules of resistance development, bioassays were performed after treatment twice with a sub-lethal dose of Bs on two continuous generations of A. dirus larvae. The killing efficacies between the Bs treated group and control group were compared. The results showed that LC50 and LC95 increased by 4.35- and 7.37-folds after treatment with the sub-lethal dose of Bs on two consecutive generations, respectively. The results indicated that A. dirus was sensitive to Bs, which could reduce oviposition of the surviving A. dirus. The subsequent effect might help to further decrease the mosquito population. However, a sub-lethal dose of Bs exposure could easily cause resistance development. Our study provides a dose standard and reference for the rational use of Bs, which will be helpful for mosquito control.