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Neuron navigator 1 (Nav1) is a cytoskeleton-associated protein expressed during brain development that is necessary for proper neuritogenesis, but the underlying mechanisms are poorly understood. Here we show that Nav1 is present in elongating axon tracts during mouse brain embryogenesis. We found that depletion of Nav1 in cultured neurons disrupts growth cone morphology and neurotrophin-stimulated neuritogenesis. In addition to regulating both F-actin and microtubule properties, Nav1 promotes actin-rich membrane ruffles in the growth cone and promotes macropinocytosis at those membrane ruffles, including internalization of the TrkB receptor for the neurotrophin brain-derived neurotropic factor (BDNF). Growth cone macropinocytosis is important for downstream signaling, neurite targeting, and membrane recycling, implicating Nav1 in one or more of these processes. Depletion of Nav1 also induces transient membrane blebbing via disruption of signaling in the Rho GTPase signaling pathway, supporting the novel role of Nav1 in dynamic actin-based membrane regulation at the cell periphery. These data demonstrate that Nav1 works at the interface of microtubules, actin, and plasma membrane to organize the cell periphery and promote uptake of growth and guidance cues to facilitate neural morphogenesis during development.

Science is a core curricular area of instruction for all students and the federal mandates of the Individuals with Disabilities Education Act (2004) and No Child Left Behind (2001) require that students with disabilities are educated in the least restrictive environment and have access to general education science content, based upon rigorous standards. While, most students with visual impairments are educated in the general science classroom, few studies have been done to determine whether appropriate accommodations and modifications are being made in those classrooms to meet the specialized needs of these students. A 35 question survey instrument was disseminated to teachers of the visually impaired through a Visual Impairments Listserve and Facebook group to help determine what pedagogical practices, accommodations, modifications, adaptive equipment and instructional practices are being used to educate students with visual impairments in the United States and Canada. This study helped inform how students with visual impairments are being educated in the science classroom.

This article presents the results of a study on the use of 3-D printed models in a science classroom for students with visual impairments and examines whether the use of these models impacts student conceptual understanding and misconceptions related to geosciences concepts, specifically plate tectonics. Data were collected one week prior to instruction, one week after instruction and throughout the 3-week instructional period. Results showed that students with visual impairments held many of the same misconceptions about plate tectonics as students who are typically sighted. All students in this study had fewer misconceptions after the instructional period than they held before instruction began; however, both the 3D group and the TG group continued to hold approximately equal numbers of misconceptions. The adaptations and hands-on experiences in this 3-week curriculum proved effective for students with visual impairments; helping them understand the unifying theory of plate tectonics. Some unique misconceptions held by the students with visual impairments in this research study include plates floating on the ocean, earthquakes moving with the plates, and volcanoes working together with the plates to cause earthquakes. There was a difference between students who had low vision and those with light perception only. The study helps to shed light on the use of 3-D printed models in the science classroom and their effectiveness at helping students with visual impairments learn important geoscience concepts.

Syndromic sensorineural hearing loss is multigenic and associated with malformations of the ear and other organ systems. Herein we describe a child admitted to the NIH Undiagnosed Diseases Program with global developmental delay, sensorineural hearing loss, gastrointestinal abnormalities, and absent salivation. Next-generation sequencing revealed a uniparental isodisomy in chromosome 5, and a 22 kb homozygous deletion in SLC12A2, which encodes for sodium, potassium, and chloride transporter in the basolateral membrane of secretory epithelia. Functional studies using patient-derived fibroblasts showed truncated SLC12A2 transcripts and markedly reduced protein abundance when compared with control. Loss of Slc12a2 in mice has been shown to lead to deafness, abnormal neuronal growth and migration, severe gastrointestinal abnormalities, and absent salivation. Together with the described phenotype of the Slc12a2-knockout mouse model, our results suggest that the absence of functional SLC12A2 causes a new genetic syndrome and is crucial for the development of auditory, neurologic, and gastrointestinal tissues.

Aminoacyl-tRNA synthetases (ARSs) function to transfer amino acids to cognate tRNA molecules, which are required for protein translation. To date, biallelic mutations in 31 ARS genes are known to cause recessive, early-onset severe multi-organ diseases. VARS encodes the only known valine cytoplasmic-localized aminoacyl-tRNA synthetase. Here, we report seven patients from five unrelated families with five different biallelic missense variants in VARS. Subjects present with a range of global developmental delay, epileptic encephalopathy and primary or progressive microcephaly. Longitudinal assessment demonstrates progressive cortical atrophy and white matter volume loss. Variants map to the VARS tRNA binding domain and adjacent to the anticodon domain, and disrupt highly conserved residues. Patient primary cells show intact VARS protein but reduced enzymatic activity, suggesting partial loss of function. The implication of VARS in pediatric neurodegeneration broadens the spectrum of human diseases due to mutations in tRNA synthetase genes.

Domestic cats are the natural reservoir of Bartonella henselae, B. clarridgeiae and B. koehlerae. To determine the role of wild felids in the epidemiology of Bartonella infections, blood was collected from 14 free-ranging California mountain lions (Puma concolor) and 19 bobcats (Lynx rufus). Bartonella spp. were isolated from four (29%) mountain lions and seven (37%) bobcats. These isolates were characterized using growth characteristics, biochemical reactions, molecular techniques, including PCR-RFLP of selected genes or interspacer region, pulsed-field gel electrophoresis (PFGE), partial sequencing of several genes, and DNA-DNA hybridization. Two isolates were identical to B. henselae genotype II. All other isolates were distinguished from B. henselae and B. koehlerae by PCR-RFLP of the gltA gene using endonucleases HhaI, TaqI and AciI, with the latter two discriminating between the mountain lion and the bobcat isolates. These two novel isolates displayed specific PFGE profiles distinct from B. henselae, B. koehlerae and B. clarridgeiae. Sequences of amplified gene fragments from the three mountain lion and six bobcat isolates were closely related to, but distinct from, B. henselae and B. koehlerae. Finally, DNA-DNA hybridization studies demonstrated that the mountain lion and bobcat strains are most closely related to B. koehlerae. We propose naming the mountain lion isolates B. koehlerae subsp. boulouisii subsp. nov. (type strain: L-42-94), and the bobcat isolates B. koehlerae subsp. bothieri subsp. nov. (type strain: L-17-96), and to emend B. koehlerae as B. koehlerae subsp. koehlerae. The mode of transmission and the zoonotic potential of these new Bartonella subspecies remain to be determined.

Domestic cats are the natural reservoir of Bartonella henselae, B. clarridgeiae and B. koehlerae. To determine the role of wild felids in the epidemiology of Bartonella infections, blood was collected from 14 free-ranging California mountain lions (Puma concolor) and 19 bobcats (Lynx rufus). Bartonella spp. were isolated from four (29%) mountain lions and seven (37%) bobcats. These isolates were characterized using growth characteristics, biochemical reactions, molecular techniques, including PCR-RFLP of selected genes or interspacer region, pulsed-field gel electrophoresis (PFGE), partial sequencing of several genes, and DNA-DNA hybridization. Two isolates were identical to B. henselae genotype II. All other isolates were distinguished from B. henselae and B. koehlerae by PCR-RFLP of the gltA gene using endonucleases HhaI, TaqI and AciI, with the latter two discriminating between the mountain lion and the bobcat isolates. These two novel isolates displayed specific PFGE profiles distinct from B. henselae, B. koehlerae and B. clarridgeiae. Sequences of amplified gene fragments from the three mountain lion and six bobcat isolates were closely related to, but distinct from, B. henselae and B. koehlerae. Finally, DNA-DNA hybridization studies demonstrated that the mountain lion and bobcat strains are most closely related to B. koehlerae. We propose naming the mountain lion isolates B. koehlerae subsp. boulouisii subsp. nov. (type strain: L-42-94), and the bobcat isolates B. koehlerae subsp. bothieri subsp. nov. (type strain: L-17-96), and to emend B. koehlerae as B. koehlerae subsp. koehlerae. The mode of transmission and the zoonotic potential of these new Bartonella subspecies remain to be determined.

A man with newly diagnosed AIDS presented with months of back pain and fever. Computed tomography (CT) results demonstrated aortitis with periaortic tissue thickening. DNA amplification of biopsy tissue revealed Bartonella quintana, and Bartonella serologies were subsequently noted to be positive. The patient improved with prolonged doxycycline and rifabutin treatment. This case illustrates how molecular techniques are increasingly important in diagnosing Bartonella infections.

Bartonella quintana is a vector-borne bacterial pathogen that causes fatal disease in humans. During the infectious cycle, B. quintana transitions from the hemin-restricted human bloodstream to the hemin-rich body louse vector. Because extracytoplasmic function (ECF) sigma factors often regulate adaptation to environmental changes, we hypothesized that a previously unstudied B. quintana ECF sigma factor, RpoE, is involved in the transition from the human host to the body louse vector. The genomic context of B. quintana rpoE identified it as a member of the ECF15 family of sigma factors found only in alphaproteobacteria. ECF15 sigma factors are believed to be the master regulators of the general stress response in alphaproteobacteria. In this study, we examined the B. quintana RpoE response to two stressors that are encountered in the body louse vector environment, a decreased temperature and an increased hemin concentration. We determined that the expression of rpoE is significantly upregulated at the body louse (28°C) versus the human host (37°C) temperature. rpoE expression also was upregulated when B. quintana was exposed to high hemin concentrations. In vitro and in vivo analyses demonstrated that RpoE function is regulated by a mechanism involving the anti-sigma factor NepR and the response regulator PhyR. The ΔrpoE ΔnepR mutant strain of B. quintana established that RpoE-mediated transcription is important in mediating the tolerance of B. quintana to high hemin concentrations. We present the first analysis of an ECF15 sigma factor in a vector-borne human pathogen and conclude that RpoE has a role in the adaptation of B. quintana to the hemin-rich arthropod vector environment.

Rasamsonia argillacea (formerly known as Geosmithia argillacea) is a fungus recently recognized as a pathogen of immunocompromised patients. Here we report the first case of Rasamsonia infection in an immunocompetent host, presenting as a pulmonary and aortic graft infection. Its morphological similarity to nonpathogenic Penicillium species delayed the diagnosis and initiation of appropriate treatment.

BackgroundBartonella henselae is the zoonotic agent of cat scratch disease and causes potentially fatal infections in immunocompromised patients. Understanding the complex interactions between the host's immune system and bacterial pathogens is central to the field of infectious diseases and to the development of effective diagnostics and vaccines.MethodologyWe report the development of a microarray comprised of proteins expressed from 96% (1433/1493) of the predicted ORFs encoded by the genome of the zoonotic pathogen Bartonella henselae. The array was probed with a collection of 62 uninfected, 62 infected, and 8 “specific-pathogen free” naïve cat sera, to profile the antibody repertoire elicited during natural Bartonella henselae infection.ConclusionsWe found that 7.3% of the B. henselae proteins on the microarray were seroreactive and that seroreactivity was not evenly distributed between predicted protein function or subcellular localization. Membrane proteins were significantly most likely to be seroreactive, although only 23% of the membrane proteins were reactive. Conversely, we found that proteins involved in amino acid transport and metabolism were significantly underrepresented and did not contain any seroreactive antigens. Of all seroreactive antigens, 52 were differentially reactive with sera from infected cats, and 53 were equally reactive with sera from infected and uninfected cats. Thirteen of the seroreactive antigens were found to be differentially seroreactive between B. henselae type I and type II. Based on these results, we developed a classifier algorithm that was capable of accurately discerning 93% of the infected animals using the microarray platform. The seroreactivity and diagnostic potential of these antigens was then validated on an immunostrip platform, which correctly identified 98% of the infected cats. Our protein microarray platform provides a high-throughput, comprehensive analysis of the feline humoral immune response to natural infection with the alpha-proteobacterium B. henselae at an antigen-specific, sera-specific, and genome-wide level. Furthermore, these results provide novel insight and utility in diagnostics, vaccine development, and understanding of host-pathogen interaction.

Bartonella rochalimae was first isolated from the blood of a human who traveled to Peru and was exposed to multiple insect bites. Foxes and dogs are likely natural reservoirs for this bacterium. We report the results of experimental inoculation of two dogs, five cats and six guinea pigs with the only human isolate of this new Bartonella species. Both dogs became bacteremic for 5-7 weeks, with a peak of 10(3)-10(4) colony forming units (CFU)/mL blood. Three cats had low bacteremia levels (< 200 CFU/mL) of 6-8 weeks' duration. One cat that remained seronegative had two bacterial colonies isolated at a single culture time point. A fifth cat never became bacteremic, but seroconverted. None of the guinea pigs became bacteremic, but five seroconverted. These results suggest that dogs could be a reservoir of this strain of B. rochalimae, in contrast to cats and guinea pigs.

Multiple locus variable number tandem repeat analysis was performed on 178 Bartonella henselae isolates from 9 countries; 99 profiles were distributed into 2 groups. Human isolates/strains were placed into the second group. Genotype I and II isolates shared no common profile. All genotype I isolates clustered within group B. The evolutive implications are discussed.

The first case of canine endocarditis caused by "Bartonella rochalimae" is reported. By PCR-restriction fragment length polymorphism, sequence, and phylogenetic analyses, Bartonella isolates from a dog with endocarditis, 22 gray foxes, and three dogs, described as B. clarridgeiae like, were confirmed to belong to the new species "B. rochalimae," suggesting canids as the natural reservoir.

Bartonella spp. are facultative intracellular bacteria that cause characteristic hostrestricted hemotropic infections in mammals and are typically transmitted by blood-sucking arthropods. In the mammalian reservoir, these bacteria initially infect a yet unrecognized primary niche, which seeds organisms into the blood stream leading to the establishment of a long-lasting intra-erythrocytic bacteremia as the hall-mark of infection. Bacterial type IV secretion systems, which are supra-molecular transporters ancestrally related to bacterial conjugation systems, represent crucial pathogenicity factors that have contributed to a radial expansion of the Bartonella lineage in nature by facilitating adaptation to unique mammalian hosts. On the molecular level, the type IV secretion system VirB/VirD4 is known to translocate a cocktail of different effector proteins into host cells, which subvert multiple cellular functions to the benefit of the infecting pathogen. Furthermore, bacterial adhesins mediate a critical, early step in the pathogenesis of the bartonellae by binding to extracellular matrix components of host cells, which leads to firm bacterial adhesion to the cell surface as a prerequisite for the efficient translocation of type IV secretion effector proteins. The best-studied adhesins in bartonellae are the orthologous trimeric autotransporter adhesins, BadA in Bartonella henselae and the Vomp family in Bartonella quintana. Genetic diversity and strain variability also appear to enhance the ability of bartonellae to invade not only specific reservoir hosts, but also accidental hosts, as shown for B. henselae. Bartonellae have been identified in many different blood-sucking arthropods, in which they are typically found to cause extracellular infections of the mid-gut epithelium. Adaptation to specific vectors and reservoirs seems to be a common strategy of bartonellae for transmission and host diversity. However, knowledge regarding arthropod specificity/restriction, the mode of transmission, and the bacterial factors involved in arthropod infection and transmission is still limited.

Roof rats (Rattus rattus) damage an estimated 5 to 10% of the developing nut crop in Hawaiian macadamia (Macadamia integrifolia) orchards. Relevant aspects of roof rat biology in macadamia orchards have and continue to be studied with the ultimate goal of developing an ecologically sound and cost-effective integrated pest management plan. The field component of a two-year study of roof rat populations in macadamia orchards has recently been completed. The goal of this study is to clarify the relationship between roof rat seasonal abundance, macadamia flowering, and nut production on five orchards in three regions on the island of Hawaii. The authors herein present preliminary results from selected aspects of this research. This and other completed studies on rat feeding locations and the effect of simulated rat damage during different stages of nut development will aid in the determination of critical points in the crop cycle when rats cause significant economic damage and control of damage is warranted. This paper is intended to be an overview of research leading to the development of a realistic integrated pest management plan for roof rats in Hawaiian macadamia orchards.

Black rats (Rattus rattus) cause extensive damage in Hawaiian macadamia (Macadamia integrifolia) orchards. In a previous study, extensive and persistent snap trapping significantly reduced rat populations and depredations on developing macadamia nuts, but had little effect on subsequent yields of mature nuts. This suggested that macadamia trees may compensate for rat damage, and that commonly used indices based on rodent activity and proportion of nuts damaged may overestimate the impact of rodent depredations and exaggerate the effectiveness of control measures. To clarify the effects of rat feeding on developing macadamia nuts, two levels of damage at two times during nut development and evaluated yields of mature nuts were simulated. Both number of nuts per raceme (P = 0.0001) and total weight of mature kernels per raceme (P = 0.0001), but not mean weight per mature kernel (P = 0.90), varied among treatments. Both number of nuts and total weight of kernels decreased (P < 0.05) with increasing damage. Time during nut development that damage was simulated had no apparent effect (P > 0.05) on yields. These results indicate that racemes did not compensate for damage by retaining other nuts on the same raceme that might otherwise have dropped prematurely. A variance component analysis was also conducted to determine how best to sample the orchard in a practical fashion while minimizing potential sources of bias and retaining sensitivity for distinguishing among treatment effects. All of the random variability in the number of nuts per raceme and total weight of nuts per raceme, and > 93% of the variability in mean weight per mature nut were due to variability between racemes on a tree. Thus, blocking was not needed to control for variability among the different areas in the orchard; sampling fewer trees and concentrating available resources on measuring more racemes per tree would have provided a more sensitive comparison of treatments. Focusing on entire branches or trees instead of racemes as experimental units might have provided a more realistic model for investigating compensatory mechanisms in macadamia trees.

We conducted a series of preliminary feeding trials with Norway rats (Rattus norvegicus), roof rats (R. rattus), and Polynesian rats (R. exulans) to examine the effects of carbon disulfide (CS2 on consumption of nontoxic foods. We formulated CS2 at target concentrations of 10 ppm in deionized water, and of 50 ppm to 100,000 ppm in a starch xanthate matrix. However, we did not analyze actual concentrations of CS2 in the test foods or measure its rate of volatilization, and thus cannot verify the levels of CS2 the rats were exposed to CS2 diluted in water and applied directly to food had no apparent effect on consumption by any of the species. Formulations with 50 ppm CS2 in starch xanthate influenced food choice by Norway rats and roof rats in one test, but not in another. Concentrations ≥ 10,000 ppm repelled Norway rats. CS2-starch xanthate had little effect on consumption by Polynesian rats. Further testing is needed to develop effective formulations and delivery methods for utilizing CS2 as a bait additive.

Various frightening stimuli, primarily visual and acoustic, have been used to prevent or alleviate damage by depredating mammals (e.g., deer (Odocoileus spp.), raccoons (Procyon lotor), tree squirrels (Sciurus spp.), coyotes (Canis latrans). Frightening methods are most appropriate for use where a crop or situation needs protection from pest mammals for only a period of a few days or weeks. The ability of animals to habituate to such stimuli limits their long-term usefulness. Against nocturnal species, various types of lights and noisemakers are the most useful. Combining acoustic and visual stimuli can enhance effectiveness, while varying the techniques used, the placement of frightening devices, and/or the timing sequence can delay habituation. Other types of physical frightening stimuli are also reviewed.