Ovomucoid third domains from 100 avian species: isolation, sequences, and hypervariability of enzyme-inhibitor contact residues

Ovomucoids were isolated from egg whites of 100 avian species and subjected to limited proteolysis. From each an intact, connecting peptide extended third domain was isolated and purified. These were entirely sequenced by single, continuous runs in a sequencer. Of the 106 sequences we report (five polymorphisms and chicken from the preceding paper [Kato, I., Schrode, J., Kohr, W. J., & Laskowski, M., Jr. (1986) Biochemistry (preceding paper in this issue)]), 65 are unique. In all cases except ostrich (which has Ser45), the third domains are either partially or fully glycosylated at Asn45. The majority of the third domain preparations we isolated are carbohydrate-free. Alignment of the sequences shows that their structurally important residues are strongly conserved. On the other hand, those residues that are in contact with the enzyme in turkey ovomucoid third domain complex with Streptomyces griseus proteinase B [Read, R., Fujinaga, M., Sielecki, A. R., & James, M. N. G. (1983) Biochemistry 22, 4420-4433] are not conserved but instead are by far the most variable residues in the molecule. These findings suggest that ovomucoid third domains may be an exception to the widely accepted generalization that in protein evolution the functionally important residues are strongly conserved. Complete proof will require better understanding of the physiological function of ovomucoid third domains. This large set of variants differing from each other in the enzyme-inhibitor contact area and augmented by several high-resolution structure determinations is useful for the study of our sequence to reactivity (inhibitory activity) algorithm. It is also useful for the study of several other protein properties. In the connecting peptide fragment most phasianoid birds have the dipeptide Val4-Ser5, which is absent in most other orders. This dipeptide is often present in only 70-95% of the molecules and appears to arise from ambiguous excision at the 5' end of the F intron of ovomucoid. Connecting peptides from the ovomucoids of cracid birds contain the analogous Val4-Asn5 peptide. In laughing kookaburra ovomucoid third domain we found (in 91% of the molecules) Gln5A, which we interpret as arising from ambiguous intron excision at the 3' end of the F intron.     

Cryptic diversity in a fig wasp community—morphologically differentiated species are sympatric but cryptic species are parapatric

A key debate in ecology centres on the relative importance of niche and neutral processes in determining patterns of community assembly with particular focus on whether ecologically similar species with similar functional traits are able to coexist. Meanwhile, molecular studies are increasingly revealing morphologically indistinguishable cryptic species with presumably similar ecological roles. Determining the geographic distribution of such cryptic species provides opportunities to contrast predictions of niche vs. neutral models. Discovery of sympatric cryptic species increases alpha diversity and supports neutral models, while documentation of allopatric/parapatric cryptic species increases beta diversity and supports niche models. We tested these predictions using morphological and molecular data, coupled with environmental niche modelling analyses, of a fig wasp community along its 2700‐km latitudinal range. Molecular methods increased previous species diversity estimates from eight to eleven species, revealing morphologically cryptic species in each of the four wasp genera studied. Congeneric species pairs that were differentiated by a key morphological functional trait (ovipositor length) coexisted sympatrically over large areas. In contrast, morphologically similar species, with similar ovipositor lengths, typically showed parapatric ranges with very little overlap. Despite parapatric ranges, environmental niche models of cryptic congeneric pairs indicate large regions of potential sympatry, suggesting that competitive processes are important in determining the distributions of ecologically similar species. Niche processes appear to structure this insect community, and cryptic diversity may typically contribute mostly to beta rather than alpha diversity.     

The use of NASBA for the detection of microbial pathogens in food and environmental samples

The isothermal amplification method nucleic acid sequence-based amplification (NASBA), which amplifies RNA, has been reported as useful for the detection of microbial pathogens in food and environmental samples. Methods have been published for Campylobacter spp., Listeria monocytogenes and Salmonella enterica ser. Enteritidis in various foods and for Cryptosporidium parvum in water. Both 16S rRNA and various mRNAs have been used as target molecules for detection; the latter may have advantages in allowing specific detection of viable cells. Most of the methods to detect pathogens in foods have employed enrichment in nutrient medium prior to NASBA, as this can ensure sensitivity of detection and encourage the detection of only viable target cells. Although a relatively recent method, NASBA has the potential for adoption as a diagnostic tool for environmental pathogens.     

Safety of Brucella abortus strain RB51 in deer mice

Brucella abortus strain RB51 is an approved brucellosis vaccine for use in cattle that may have potential as an oral vaccine for use in elk (Cervus elaphus) and/or bison (Bison bison). This study was designed to determine effects of strain RB51 on deer mice (Peromyscus maniculatus), a nontarget species that could have access to treated baits in a field situation. In February 1994, 90 mice were orally dosed or intraperitoneally injected with 1 x 10(8) colony forming units strain RB51 and 77 controls were similarly dosed with sterile saline. At weekly intervals through early April 1994, 4 to 6 mice from each group were euthanized, gross necropsies performed, spleens and uteruses cultured, and tissues examined histologically. All orally inoculated mice cleared the infection by 6 wk post-inoculation (PI). While most of the injected mice cleared the infection by 7 wk PI, a few required 9 wk. There were minimal adverse effects attributable to strain RB51. Apparently, strain RB51 would not negatively impact P. maniculatus populations if it were used in a field situation. Also, deer mice appear to be able to clear the vaccine in 6 to 9 wk, thus the probability of these mice transmitting the vaccine to other animals is low.     

Environment and position of first bud to break on apple shoots affects lateral outgrowth

A study was conducted to determine which bud (terminal or lateral) breaks first, and thereby exerts primigenic dominance, on ‘Granny Smith’ and ‘Golden Delicious’, 1-year-old apple (Malus × domestica Borkh.) shoots grown in two locations in the Western Cape, South Africa, with differing degrees of chilling. Primigenic dominance of laterals was more common in a warm area than a cool area, and more common in ‘Granny Smith’ than ‘Golden Delicious’. Laterals rarely broke before the terminal in ‘Golden Delicious’, and so differences in lateral development due to position of first bud to break were only analyzed in ‘Granny Smith’ shoots from this point on in the study. In ‘Granny Smith’, lateral budbreak and growth was influenced by the position of the first bud to break on the shoot, but did not differ between locations. On ‘Granny Smith’ shoots with primigenic dominance of the terminal, lateral budbreak and growth was suppressed, in accordance with the typical ‘delayed foliation’ commonly observed in warm winter climates. However, when at least one lateral broke before the terminal, lateral budbreak and growth were similar to previous observations in cold winter areas.     

Using biogeographical history to inform conservation: the case of Preble's meadow jumping mouse

The last Pleistocene deglaciation shaped temperate and boreal communities in North America. Rapid northward expansion into high latitudes created distinctive spatial genetic patterns within species that include closely related groups of populations that are now widely spread across latitudes, while longitudinally adjacent populations, especially those near the southern periphery, often are distinctive due to long‐term disjunction. Across a spatial expanse that includes both recently colonized and long‐occupied regions, we analysed molecular variation in zapodid rodents to explore how past climate shifts influenced diversification in this group. By combining molecular analyses with species distribution modelling and tests of ecological interchangeability, we show that the lineage including the Preble's meadow jumping mouse (Zapus hudsonius preblei), a US federally listed taxon of conservation concern, is not restricted to the southern Rocky Mountains. Rather, populations along the Front Range are part of a single lineage that is ecologically indistinct and extends to the far north. Of the 21 lineages identified, this Northern lineage has the largest geographical range and low measures of intralineage genetic differentiation, consistent with recent northward expansion. Comprehensive sampling combined with coalescent‐based analyses and niche modelling leads to a radically different view of geographical structure within jumping mice and indicates the need to re‐evaluate their taxonomy and management. This analysis highlights a premise in conservation biology that biogeographical history should play a central role in establishing conservation priorities.     

Enzymatic Synthesis of 2′,5′-Dideoxv Purine Nucleosides and Related Compounds

Abstract

A wide range of 2′,5′-dideoxy-nucleosides, including 6- substituted purine, pyrazolo[3,4-d]pyrimidine and 1-deazapurine derivatives, has been enzymatically prepared using purine nucleoside phosphorylase. Specificity towards cleavage by bacterial versus mammalian purine nucleoside phosphorylase was evaluated.