Experimental Adaptation of Burkholderia cenocepacia to Onion Medium Reduces Host Range

It is unclear whether adaptation to a new host typically broadens or compromises host range, yet the answer bears on the fate of emergent pathogens and symbionts. We investigated this dynamic using a soil isolate of Burkholderia cenocepacia, a species that normally inhabits the rhizosphere, is related to the onion pathogen B. cepacia, and can infect the lungs of cystic fibrosis patients. We hypothesized that adaptation of B. cenocepacia to a novel host would compromise fitness and virulence in alternative hosts. We modeled adaptation to a specific host by experimentally evolving 12 populations of B. cenocepacia in liquid medium composed of macerated onion tissue for 1,000 generations. The mean fitness of all populations increased by 78% relative to the ancestor, but significant variation among lines was observed. Populations also varied in several phenotypes related to host association, including motility, biofilm formation, and quorum-sensing function. Together, these results suggest that each population adapted by fixing different sets of adaptive mutations. However, this adaptation was consistently accompanied by a loss of pathogenicity to the nematode Caenorhabditis elegans; by 500 generations most populations became unable to kill nematodes. In conclusion, we observed a narrowing of host range as a consequence of prolonged adaptation to an environment simulating a specific host, and we suggest that emergent pathogens may face similar consequences if they become host-restricted.Some emergent pathogens, such as Pseudomonas and Burkholderia species, persist in a wide range of plant and animal hosts, suggesting that the virulence factors needed to infect plants and animals are similar (5, 40). Yet whether adaptation to a new niche tends to compromise niche breadth or, in this case, host range is an open question. Adaptation to a novel host may restrict host range to various degrees, whether by diminishing host-specific virulence traits without affecting host colonization or by reducing the ability to initiate infection in alternative hosts. However, if factors needed to colonize plant and animal hosts are similar, then why are some bacterial populations restricted to a narrow host range while others are not? One explanation for a limited host range may be the result of genetic trade-offs associated with adaptation to a specific host (7, 18). Another explanation may be that prolonged adaptation to a specific host casts a “selective shadow” over unused functions that are relevant to colonizing other hosts but decay by genetic drift (7, 18). To address these possibilities, we quantified the direct and correlated effects of specific host adaptation by the opportunistic pathogen Burkholderia cenocepacia.Members of the Burkholderia cepacia complex (Bcc), which are ubiquitous in the environment, were once used as biocontrol and bioremedial agents but now are banned from these applications because of the potential of some members to cause plant and human disease (39). The type species B. cepacia is well known as a pathogen of the common yellow onion, Allium cepa, in which it causes a characteristic yellow or brown rot. Another species, B. cenocepacia, can also infect onions as well as a range of plants and animals, including humans (2, 6, 26, 36). Bcc bacteria can cause serious infection in the lungs of cystic fibrosis (CF) patients (6, 26). These infections, called “cepacia syndrome,” are highly contagious among CF patients, and infections produce many negative effects on an already poor quality of life, including longer hospital stays, removal from lung transplant lists, blood poisoning, and eventual death (24). B. cenocepacia, one of the two Bcc species most commonly isolated from lung infections, is especially threatening and is associated with more severe cepacia syndrome (35). However, the mechanisms allowing B. cenocepacia to adapt to colonize both human and plant hosts are unclear. Several putative virulence mechanisms have been identified by random mutagenic screens or by knockouts of candidate genes (2, 12, 20, 25, 29, 35, 43, 46), but these mechanisms generally have not been shown to function in host adaptation. One way to directly study adaptation of bacterial populations to susceptible hosts is by experimental evolution, in which bacterial populations evolve in a controlled laboratory setting that enables study of the adaptive process over time (7).We experimentally evolved populations of B. cenocepacia HI2424 to study the extent to which adaptation to the common yellow onion A. cepa affects host range. B. cenocepacia HI2424 is a soil isolate and is classified as part of the PHDC strain lineage, the strain first characterized as responsible for an outbreak of Bcc infections in large treatment centers located in the mid-Atlantic region of the United States (33). We found that adaptation of B. cenocepacia to the onion model was associated with reduced virulence but did not compromise the capacity to colonize (or be consumed by) the nematode Caenorhabditis elegans, and the coincidence of these events suggests that a genetic trade-off (antagonistic pleiotropy) between fitness in onion medium and nematode virulence exists. We also characterized several phenotypes potentially associated with adaptation to the onion or nematode virulence. Most phenotypes varied significantly among replicate populations, suggesting that adaptation to the onion model may follow several different pathways.     

Edge effects reduce the nesting success of Acadian Flycatchers in a moderately fragmented forest

ABSTRACT.   Forest fragmentation can create negative edge effects that reduce the reproductive success of birds nesting near the forest/nonforest interface, and threaten bird populations deeper in remnant forest habitats. Negative edge effects may be more pronounced in landscapes that are moderately fragmented, particularly where agriculture is the primary land-use fragmenting forests. Information about the extent and strength of edge effects at a site can help guide conservation actions, and determine their effectiveness. We examined edge effects for birds breeding in a nearly contiguous forest fragmented by relatively narrow agricultural corridors in Illinois (USA). We measured rates of nest predation and brood parasitism for Acadian Flycatchers ( Empidonax virescens ) over a continuum of distances from the edge of an agricultural inholding. Nest predation and brood parasitism were highest near the edge and decreased with increasing distance from the edge. Given the cumulative effects of nest predation and brood parasitism on reproductive success, we determined that forest within 600 m of the inholding was sink habitat. We found, however, that deeper forest interior areas currently serve as source habitat, and that conversion of the entire 205 ha agricultural corridor to forest would add 1350 ha of source habitat for Acadian Flycatchers. Such results provide support for a local conservation strategy of forest consolidation and establish baseline measures necessary to determine the relative effectiveness of any subsequent reforestation efforts.     

Potential role for IL-7 in Fas-mediated T cell apoptosis during HIV infection

IL-7 promotes survival of resting T lymphocytes and induces T cell proliferation in lymphopenic conditions. As elevated IL-7 levels occur in HIV-infected individuals in addition to high Fas expression on T cells and increased sensitivity to Fas-induced apoptosis, we analyzed whether IL-7 has a regulatory role in Fas-mediated T cell apoptosis. We show that IL-7 up-regulates Fas expression on naive and memory T cells through a mechanism that involves translocation of Fas molecules from intracellular compartments to the cell membrane. IL-7 induced the association of Fas with the cytoskeletal component ezrin and a polarized Fas expression on the cell surface. The potential role of IL-7 in Fas up-regulation in vivo was verified in IL-7-treated macaques and in HIV-infected or chemotherapy treated patients by the correlation between serum IL-7 levels and Fas expression on T cells. IL-7 treatment primed T cells for Fas-induced apoptosis in vitro and serum IL-7 levels correlated with the sensitivity of T cells to Fas-induced apoptosis in HIV-infected individuals. Our data suggest an important role for IL-7 in Fas-mediated regulation of T cell homeostasis. Elevated IL-7 levels associated with lymphopenic conditions, including HIV-infection, might participate in the increased sensitivity of T cells for activation-induced apoptosis.     

Trypanosoma cruzi prevalence and epidemiologic trends in lemurs on St. Catherines Island, Georgia

Lemurs on St. Catherines Island, Georgia were tested for Trypanosoma cruzi infection to develop a better understanding of the epizootiology of the parasite in nonhuman primates in the southeastern United States. Fifty-six ring-tailed (Lemur catta), blue-eyed black (Eulemur macaco flavifrons), and black-and-white ruffed (Varecia variegata variegata) lemurs were tested by hemoculture and serology to determine the prevalence of T. cruzi in the population. Of those tested 3 (5%) were identified as culture positive and 25 (44.6%) as seropositive. When hemoculture results were compared with those from a similar study performed in 1997, prevalence remained unchanged. Genetic characterization of the 3 culture isolates indicated they belong to the T. cruzi IIa group, which is identical to strains previously isolated from raccoons on the island. Despite the occurrence of T. cruzi in the population, there was no evidence that the health of the lemurs was compromised as a result of infection. Based upon prevalence and available breeding records we speculate that both vertical and vector-mediated transmission play significant roles in the epidemiology of T. cruzi on the island. This also represents the first report of autochthonous infection in blue-eyed black and black-and-white ruffed lemurs.     

Evidence for enhanced eNOS function in coronary microvessels during the second window of protection

Nitric oxide (NO) derived from endothelial NO synthase (NOS) (eNOS) has been identified as a trigger for the second window of protection (SWOP), but its role as a mediator during the SWOP is a matter of debate. Eighteen mongrel dogs were chronically instrumented to measure left ventricular function, coronary blood flow, and wall thickening. Myocardial preconditioning was induced by 10 min coronary artery occlusion. After 24 h of reperfusion (during the SWOP), the hearts were excised. Coronary microvessels were isolated and incubated in presence of 1) the endothelium-dependent agonists carbachol and bradykinin, 2) the calcium ionophore A23187, and 3) the angiotensin-converting enzyme (ACE) inhibitors enalaprilat and ramiprilat. Nitrite, a metabolite of NO, was measured. Under baseline conditions, nitrite production in microvessels from SWOP was 30% higher than that from normal (96 +/- 4 vs. 74 +/- 3 pmol/mg, P < 0.01, respectively). Nitrite production in response to carbachol, bradykinin, and A23187 was also enhanced in microvessels from SWOP (P < 0.05). These enhanced responses were abolished by N(G)-nitro-l-arginine methyl ester (l-NAME) or the endothelial receptor-specific antagonists atropine and HOE-140. The level of eNOS protein in the SWOP myocardium was twofold higher than that in the non-SWOP myocardium. Nitrite production in response to the ACE inhibitors was greater in microvessels from SWOP. These effects were blocked by l-NAME, HOE-140, or dichloroisocoumarin (which inhibits kinin formation). We found that a brief ischemic episode induced delayed, enhanced NO production in coronary microvessels and an upregulation of eNOS protein. These findings suggest that eNOS is a mediator during the SWOP. The ability of ACE inhibitors to enhance NO release during the SWOP points to an additional clinical application for these drugs.     

Fundamental training for individuals involved in the care and use of laboratory animals: a review and update of the 1991 NRC Core Training Module

Public trust demands that individuals who do research, testing, or teaching with animals use humane, ethical, and scientifically sound methods. Furthermore, the Animal Welfare Act and the Public Health Service Policy require research institutions to provide basic training and to ensure that anyone who cares for and/or works with laboratory animals has the appropriate training or experience relevant to their job responsibilities. Institutions accredited by the Association for Assessment and Accreditation of Laboratory Animal Care International must also provide training programs and ensure the qualifications of personnel. The primary goal of this training is to provide individuals with basic knowledge and to reinforce attitudes and behaviors that help to ensure humane animal care and use. This article provides an overview of the core training module outline and content from the 1991 report of the Institute for Laboratory Animal Research, Education and Training in the Care and Use of Laboratory Animals: A Guide for Developing Institutional Programs, as well as pertinent updates for introducing personnel to information regarding the care and use of laboratory animals. Both mandatory and suggested training topics are reviewed, including relevant regulations and standards, ethical considerations, humane methods of animal experimentation and maintenance, and other pertinent topics. Although the fundamental training course content and delivery will vary depending on the nature and complexity of an institution's animal care and use program, this basic training provides the foundation for more in-depth training programs and supports humane and ethical animal care and use.