Human case of bubonic plague resulting from the bite of a wild Gunnison's prairie dog during translocation from a plague-endemic area.

Plague is a zoonotic disease (transmitted mainly by fleas and maintained in nature by rodents) that causes severe acute illness in humans. We present a human plague case who became infected by the bite of a wild Gunnison's prairie dog, and a good practical example of the One Health approach that resulted in a rapid public health response. The exposure occurred while the animal was being transported for relocation to a wildlife refuge after being trapped in a plague enzootic area. This is the first report of a human plague case resulting from the bite of a Gunnison's prairie dog. Additionally, we present an observation of a longer incubation period for plague in captive prairie dogs, leading to a recommendation for a longer quarantine period for prairie dogs during translocation efforts.

Crystallization and quaternary structure analysis of an Lrp-like regulatory protein from the hyperthermophile Pyrococcus furiosus.

The LrpA transcriptional regulator from Pyrococcus furiosus, a member of the leucine-responsive regulatory protein (Lrp) family, has been crystallized by the hanging-drop method of vapour diffusion using ammonium sulfate as the precipitant. The crystals belong to the tetragonal system and are in space group I4(1)22, with unit-cell parameters a = b = 104.5, c = 245.1 A. Consideration of the values of V(M) and possible packing of the molecules within the cell suggest that the asymmetric unit contains a dimer. Examination of the behaviour of the protein on gel-filtration columns and analysis of the self-rotation function suggests that the molecule is an octamer in solution at around pH 5. Determination of the structure of this protein will provide insights into the mechanisms responsible for DNA-protein recognition at high temperature and into how the regulatory properties of the Lrp family are modified by the presence or absence of effector molecules.

Structure of two FREP genes that combine IgSF and fibrinogen domains, with comments on diversity of the FREP gene family in the snail Biomphalaria glabrata.

Upon exposure to infection with digenetic trematodes such as Echinostoma paraensei, the freshwater snail Biomphalaria glabrata produces increased quantities of hemolymph lectins, some of which are unique polypeptides containing both immunoglobulin superfamily (IgSF) and fibrinogen domains. These unusual lectins have been termed fibrinogen-related proteins (FREPs), and recognize and precipitate digenean antigens. We here report 11 distinct FREP-encoding sequences from B. glabrata, and provide the complete genomic sequence for two of the most frequently recovered FREPs. The unique juxtaposition of IgSF and fibrinogen domains, previously known only from incomplete cDNAs, is confirmed. Sequences corresponding to known peptides derived from FREPs from hemolymph were found in one of these genes. Both genes contain four exons, the first encodes a putative signal peptide, the second and third a portion of an IgSF-type loop, and the fourth a fibrinogen domain. Cysteines, postulated to form an intrachain loop, are present in the IgSF domain and are separated from one another by 78 or 79 residues. The IgSF sequences most closely resemble V (variable)-type Ig domains, based on canonical and hydrophobic residues and predicted secondary structure. Some minor differences in genomic fragments isolated for each of the two sequences were noted and may represent allelic variants. The results may be of relevance in understanding the role of B. glabrata in transmission of Schistosoma mansoni, a digenean parasite that infects nearly 100 million people in the tropics.

Crystal structure of the Lrp-like transcriptional regulator from the archaeon Pyrococcus furiosus.

The LrpA protein from the hyperthermophilic archaeon Pyrococcus furiosus belongs to the Lrp/AsnC family of transcriptional regulatory proteins, of which the Escherichia coli leucine-responsive regulatory protein is the archetype. Its crystal structure has been determined at 2.9 A resolution and is the first for a member of the Lrp/AsnC family, as well as one of the first for a transcriptional regulator from a hyperthermophile. The structure consists of an N-terminal domain containing a helix-turn-helix (HtH) DNA-binding motif, and a C-terminal domain of mixed alpha/beta character reminiscent of a number of RNA- and DNA-binding domains. Pyrococcus furiosus LrpA forms a homodimer mainly through interactions between the antiparallel beta-sheets of the C-terminal domain, and further interactions lead to octamer formation. The LrpA structure suggests how the protein might bind and possibly distort its DNA substrate through use of its HtH motifs and control gene expression. A possible location for an effector binding site is proposed by using sequence comparisons with other members of the family coupled to mutational analysis.

Parasite-responsive IgSF members in the snail Biomphalaria glabrata: characterization of novel genes with tandemly arranged IgSF domains and a fibrinogen domain.

Two novel genes of the immunoglobulin superfamily (IgSF), FREP3 and FREP7, are reported from the snail Biomphalaria glabrata, a prominent intermediate host of the human parasite Schistosoma mansoni. They resemble other B. glabrata genes that encode fibrinogen-related proteins (FREPs), but differ in that they encode proteins with two tandemly arranged IgSF domains followed by a C-terminal fibrinogen domain. FREPs are hemolymph proteins that increase in abundance following exposure to a digenetic trematode, Echinostoma paraensei, and that bind to and precipitate parasite antigens. Within each gene, the two IgSF-coding regions are dissimilar from one another: the N-terminal IgSF1 domain is encoded by a single exon whereas the downstream IgSF2 domain is encoded by three exons. For both FREPs 3 and 7, the IgSF2 domain belongs to the variable (V) type, whereas the IgSF1 domain is not easily classified with respect to IgSF type. The fibrinogen-encoding region in both genes is relatively conserved and lacks introns. FREP3 exhibits extensive variation in the IgSF1 region. A ratio of nonsynonymous versus synonymous substitutions of 2.56 suggests that this region is under positive selection. A genomic fragment identifiable as FREP7 but lacking an exon was also found, further suggestive of variability within FREP IgSF-encoding regions. Insofar as FREPs are hypothesized to function in nonself recognition, the identification of additional novel FREP genes as part of a growing gene family in B. glabrata is of interest. Such genes, particularly given their variable nature, serve as a model to study the complexity of invertebrate defense responses.

Analysis of messages expressed by Echinostoma paraensei miracidia and sporocysts, obtained by random EST sequencing.

A lambdaZAP Express cDNA library was constructed with mRNA obtained from immature miracidia within eggs, hatched miracidia, and sporocysts of Echinostoma paraensei. This cDNA library was amplified and 213 expressed sequence tag (EST) sequences (averaging 466 nucleotides in length) were obtained. The mean percentage of unresolved bases within the EST sequences was 0.4%, ranging from 0 to 4.6%. The 213 ESTs represent 151 unique messages. BLAST (version 2.0.8) analysis disclosed that 64 unique E. paraensei messages (42.4%) had significant similarities (BLAST score < or =e-5), at deduced amino acid or nucleotide levels, with known sequences in the nonredundant GenBank databases or the dbEST database (NCBI). The remainder, 57.6% of the unique EST-encoded messages, scored nonsignificant hits. Most of the E. paraensei messages that could be assigned a cellular role based on sequence similarities were involved in gene/protein expression. Several ESTs scored highest similarities with sequences obtained from trematode species. A total of 22,560 nucleotides present in open reading frames from ESTs that aligned with known sequences was used to determine codon usage for E. paraensei. Analysis of a subset of eight ESTs that contained full-length open reading frames did not reveal a bias in codon usage. Also, EST sequences were found to contain 3' untranslated regions with an average length of 69.9 +/- 88.4 nucleotides (n = 46). The EST sequences were submitted to GenBank/dbEST, adding to the 51 available Echinostoma-derived sequences, to provide reference information for both phylogenetic analysis and study of general trematode biology.

Expressed sequences from conidial, mycelial, and sexual stages of Neurospora crassa.

In the Neurospora Genome Project at the University of New Mexico, expressed sequence tags (ESTs) corresponding to three stages of the life cycle of the filamentous fungus Neurospora crassa are being analyzed. The results of a pilot project to identify expressed genes and determine their patterns of expression are presented. 1,865 partial complementary DNA (cDNA) sequences for 1,409 clones were determined using single-pass sequencing. Contig analysis allowed the identification of 838 unique ESTs and 156 ESTs present in multiple cDNA clones. For about 34% of the sequences, highly or moderately significant matches to sequences (of known and unknown function) in the NCBI database were detected. Approximately 56% of the ESTs showed no similarity to previously identified genes. Among genes with assigned function, about 43.3% were involved in metabolism, 32.9% in protein synthesis and 8.4% in RNA synthesis. Fewer were involved in defense (6%), cell signalling (3.4%), cell structure (3.4%) and cell division (2.6%).