Conservation genomics of an endangered montane amphibian reveals low population structure, low genomic diversity and selection pressure from disease.

Wildlife diseases are a major global threat to biodiversity. Boreal toads (Anaxyrus [Bufo] boreas) are a state-endangered species in the southern Rocky Mountains of Colorado and New Mexico, and a species of concern in Wyoming, largely due to lethal skin infections caused by the amphibian chytrid fungus Batrachochytrium dendrobatidis (Bd). We performed conservation and landscape genomic analyses using single nucleotide polymorphisms from double-digest, restriction site-associated DNA sequencing in combination with the development of the first boreal toad (and first North American toad) reference genome to investigate population structure, genomic diversity, landscape connectivity and adaptive divergence. Genomic diversity (π = 0.00034-0.00040) and effective population sizes (Ne = 8.9-38.4) were low, likely due to post-Pleistocene founder effects and Bd-related population crashes over the last three decades. Population structure was also low, likely due to formerly high connectivity among a higher density of geographically proximate populations. Boreal toad gene flow was facilitated by low precipitation, cold minimum temperatures, less tree canopy, low heat load and less urbanization. We found >8X more putatively adaptive loci related to Bd intensity than to all other environmental factors combined, and evidence for genes under selection related to immune response, heart development and regulation and skin function. These data suggest boreal toads in habitats with Bd have experienced stronger selection pressure from disease than from other, broad-scale environmental variations. These findings can be used by managers to conserve and recover the species through actions including reintroduction and supplementation of populations that have declined due to Bd.

Framework for a proactive parent support group: the Syracuse cystic fibrosis model.

Many support groups are organized and run according to goals and by-laws set by facilitators. This article introduces a model in which all goals and by-laws are created by the parents based on their group's specific needs. It also describes a "proactive" approach in which the group members identify projects or problems in their community and collectively form a plan in an attempt to improve these situations. This model is based on the real-life experience of a cystic fibrosis parent support group in Syracuse, New York. Lessons learned from this model can be applied to other parent support groups.

Emergency care of children in shelters.

Children living in homeless shelters often lack the health care resources usually available to other children. They are often more acutely and chronically ill than domiciled children and frequently use the emergency department (ED) as their point of entry into the health care system. To identify differences in health status, we surveyed sheltered children and domiciled controls during a nine-month period in our ED. One hundred sixty-two families completed a self-administered questionnaire during the study period: 54 homeless and 108 age-matched controls. Mean patient age was 3.4 years, mean maternal age was 27 years in both groups, and average time spent in shelters was 7.8 months. Shelter families had more children, more single mothers, and higher rates of unemployment and uninsurance than did control families. Shelter children showed greater frequencies of immunization delay, lack of TB testing, and lack of a regular health care site and higher rates of medical admissions from the ED. These data show that children in shelters have limited personal, financial, and medical resources and suggest that there are significant disparities in health status. These patients need to be identified when they present to the ED in order to meet subtle, as well as obvious, health needs.

Comparison of two culture approaches, blind passage and dual observation, for detecting Chlamydia trachomatis in various prevalence populations.

Chlamydia trachomatis diagnosis in our laboratory consisted of dual inoculation of shell vials and detection of inclusions by using fluorescein-conjugated monoclonal antiserum; the second culture vial was conventionally used for blind passage when the first vial was negative. We compared the increase in positivity using blind passage with that of a strategy utilizing observation of two stained monolayers (dual observation) without blind passage, in an effort to reduce the reporting time and labor associated with the conventional approach. A total of 4,329 specimens were obtained from an obstetrics and gynecology (OB-GYN) clinic (2,563 specimens) and the sexually transmitted disease clinic (1,766 specimens). These specimens were used to compare the two strategies. Blind passage of 1,269 initially culture-negative specimens from the OB-GYN clinic resulted in an additional 6 positive chlamydial diagnoses. In comparison, a similar number of specimens (1,294) from the OB-GYN clinic collected subsequently to the first group were tested by dual observation. There were five additional positive findings. A similar evaluation of specimens from the sexually transmitted disease clinic was performed. Blind passage of 313 initially culture-negative specimens yielded 3 additional positive diagnoses, whereas dual observation of 1,435 similar specimens resulted in 9 positive diagnoses. On the basis of analysis of 4,332 specimens, sensitivity of dual observation is comparable to that of blind passage; labor, cost, and reporting time of dual observation are reduced in comparison to those of blind passage.

Evaluation of a visual, rapid, membrane enzyme immunoassay for the detection of herpes simplex virus antigen.

We evaluated a 12-min, direct, monoclonal antibody-based enzyme immunoassay (EIA) (SureCell; Kodak, Rochester, N.Y.) which aids in the detection of herpes simplex virus infection; the assay system is also approved for culture confirmation. The test was evaluated from direct clinical samples and compared with conventional culture methodology by using a single swab. A total of 265 specimens from 180 female cervical-urogenital sites, 62 male urogenital sites, 4 rectal sites, 3 skin sites, 6 oral sites, and 10 colposcopy sites were collected on Dacron or cotton swabs and placed in viral transport medium (VTM). Within 6 h of receipt, 0.2 ml of the vortexed VTM was inoculated into each of two replicate cell cultures. Cell monolayers were observed daily for ten days, and cytopathic effect was confirmed by using an indirect immunoperoxidase reagent. The procedure for the SureCell assay conformed to the manufacturer's recommendations. When conventional culture was compared with EIA results, the overall sensitivity, specificity, positive predictive value, negative predictive value, and agreement were 64.4, 98.9, 96.7, 84.4, and 87.2%, respectively. Variables affecting the EIA sensitivity are the stage of the lesion and conventional culture methodologies. A review of culture results for 32 EIA false-negative tests indicated that 15 were detected after 48 h of incubation. Cytopathic effect observed at 48-, 72-, and 96-h cutoffs altered the sensitivity for the EIA. To ensure detection of SureCell herpes simplex virus-negative specimens, it is recommended that an unused aliquot of VTM be tested in cell culture.

Case report and seeded blood culture study of Brucella bacteremia.

Diagnosis of brucellosis requires prompt detection and identification of the coccobacillus for appropriate patient management, as the organism is associated with a potentially severe outcome. In a recent experience, an 18-year-old migrant farm worker presented at a local hospital with nonspecific symptoms. A significant Brucella titer of 2,560 was followed by the recovery of a gram-negative coccobacillus, subsequently identified as Brucella abortus, from subcultured 5-day-old BACTEC NR730 negative blood cultures. The organism proved to be susceptible to a variety of antimicrobial agents and resistant to nitrofurantoin. The patient was administered antimicrobial therapy for Brucella spp. consisting of tetracycline and streptomycin for 21 days. During the course of therapy the patient experienced defervescence and was discharged with the recommendation for periodic follow-up examinations. Seeded culture studies of this isolate with fresh human blood and target inocula of 5 and 500 CFU/ml indicated that the larger (500-CFU/ml) inoculum produced positive instrument detection within 2 days, whereas the smaller (5-CFU/ml) inoculum required 5.5 to 7.5 days for detection, depending on the medium used. These findings underscore the potential for Brucella bacteremia to escape instrument detection given a low bacterial inoculum.