Culex flavivirus isolates from mosquitoes in Guatemala.

A new strain of Culex flavivirus (family Flaviviridae, genus Flavivirus, CxFV), an insect virus first described in Japan, was isolated from adult Culex quinquefasciatus Say (Diptera: Culicidae) collected in 2006 from Izabal Department on the Caribbean coast of Guatemala. Mosquito pools were assayed for flavivirus RNA by using flavivirus group-specific primers that amplified a 720-bp region of the nonstructural (NS) 5 gene by standard reverse transcriptase-polymerase chain reaction. From 210 pools (1,699 mosquitoes), eight tested positive, and six of these mosquito pools produced virus isolates in Aedes albopictus Skuse C6/36 cells. Nucleotide sequence comparison of the eight flavivirus RNA-positive pools showed that there was 100% identity among them, and phylogenetic analysis of the NS5 and envelope gene regions indicated that they represent a strain of the recently described CxFV from Japan. This is the first report of an insect flavivirus from Central America.

West Nile virus infection and serologic response among persons previously vaccinated against yellow fever and Japanese encephalitis viruses.

It is hypothesized that previous heterologous flaviviral exposure may modulate clinical illness among persons infected with West Nile virus (WNV). Little is known about the serological response in such persons. In summer 2003, a WNV outbreak occurred in Colorado, the location of the Centers for Disease Control and Prevention, Division of Vector-Borne Infectious Diseases (DVBID). DVBID employees, most previously vaccinated with yellow fever virus (YFV) or Japanese encephalitis virus (JEV) vaccines, were studied to determine whether previous vaccination affected symptom development among those subsequently infected with WNV during the outbreak, as well as their serological response. Serum samples collected in December 2003 and previously banked samples were tested using the plaque reduction neutralization test (PRNT) against WNV, Saint Louis encephalitis virus, dengue- 4 virus, JEV, and YFV. Specimens shown to have WNV antibody by PRNT were tested by IgM and IgG enzymelinked immunosorbent assays (ELISAs). Ten (9%) of 113 serosurvey participants had WNV neutralizing antibody titers in December 2003. PRNT titers from previous specimens showed that one of the ten had seroconverted to WNV before 2003. Of the remaining nine participants, seven reported illness in the summer of 2003, two of which were unvaccinated and five previously vaccinated. In the December 2003 specimens, five persons previously unvaccinated or vaccinated only against YFV had a fourfold or greater neutralizing titer with WNV than with other flaviviruses, whereas no persons previously vaccinated against JEV or JEV and YFV showed a similar difference in neutralizing titers. Eight of nine persons infected in 2003 had negative or indeterminate WNV MAC-ELISA results in the December 2003 sample; the ninth person was vaccinated against YFV one month previously, and was also YFV positive by MAC-ELISA. We conclude that previous flaviviral vaccination does not markedly affect the development of WNV fever and that the IgM antibody response in patients without neuroinvasive WNV disease is transient.

Production of a Sindbis/Eastern Equine Encephalitis chimeric virus inactivated cell culture antigen.

Eastern Equine Encephalitis virus (EEEV) is a medically important pathogen that can cause severe encephalitis in humans, with mortality rates ranging from 30 to 80%. Unfortunately there are no antivirals or licensed vaccines available for human use, and laboratory diagnosis is essential to differentiate EEEV infection from other pathogens with similar clinical manifestations. The Arboviral Diseases Branch (ADB) reference laboratory at the CDC Division of Vector-Borne Diseases (DVBD) produces reference antigens used in serological assays such as the EEEV immunoglobulin M antibody-capture enzyme-linked immunosorbent assay (MAC-ELISA). However, EEEV is classified as a HHS select agent and requires biosafety level (BSL) three containment, limiting EEEV antigen production in non-select agent and BSL-2 laboratories. A recombinant Sindbis virus (SINV)/EEEV has been constructed for use under BSL-2 conditions and is not regulated as a select agent. Cell culture production of inactivated EEEV antigen from SINV/EEEV for use in the EEEV MAC-ELISA is reported here. Cell culture conditions and inactivation procedures were analyzed for SINV/EEEV using a recently developed antigen production algorithm, with the MAC-ELISA as the performance indicator.

Non-identical neural mechanisms for two types of mental transformation: event-related potentials during mental rotation and mental paper folding.

Reaction times, accuracy and 128-channel event-related potentials (ERPs) were measured from 14 normal, right-handed subjects while they performed two different parity-judgment tasks that require transformations of mental images: a relatively simple task requiring a single transformation (mental letter rotation), and a more complex task involving a coordinated sequence of transformations (mental paper folding). Reaction times increased monotonically with larger angular displacements from the upright (for mental rotation) and with number of squares carried (for mental paper folding). Both the tasks resulted in amplitude modulation of an approximately 420-700 ms latency ERP component at parietal electrodes. Scalp topographies indicated that right parietal cortex was activated during mental rotation, but bilateral parietal regions were activated during mental paper folding. Our results support the notion of a right hemispheric superiority for tasks involving simple, single mental rotations, but indicate greater involvement of the left hemisphere when a more complex sequence of transformations are required. This task-dependent lability of hemispheric function may account for some of the inconsistent results reported by previous neuroimaging and electrophysiological studies.

Evidence of a source of HIV type 1 within the central nervous system by ultraintensive sampling of cerebrospinal fluid and plasma.

Defining the source of HIV-1 RNA in cerebrospinal fluid (CSF) will facilitate studies of treatment efficacy in the brain. Four antiretroviral drug-naive adults underwent two 48-hr ultraintensive CSF sampling procedures, once at baseline and again beginning on day 4 after initiating three-drug therapy with stavudine, lamivudine, and nelfinavir. At baseline, constant CSF HIV-1 RNA concentrations were maintained by daily entry of at least 10(4) to 10(6) HIV-1 RNA copies into CSF. Change from baseline to day 5 ranged from -0.38 to -1.18 log(10) HIV-1 RNA copies/ml in CSF, and from -0.80 to -1.33 log(10) HIV-1 RNA copies/ml in plasma, with no correlation between CSF and plasma changes. There was no evidence of genotypic or phenotypic viral resistance in either CSF or plasma. With regard to pharmacokinetics, mean CSF-to-plasma area-under-the-curve (AUC) ratios were 38.9% for stavudine and 15.3% for lamivudine. Nelfinavir and its active M8 metabolite could not be accurately quantified in CSF, although plasma M8 peak level and AUC(0-8hr) correlated with CSF HIV-1 RNA decline. This study supports the utility of ultraintensive CSF sampling for studying HIV-1 pathogenesis and therapy in the CNS, and provides strong evidence that HIV-1 RNA in CSF arises, at least in part, from a source other than plasma.

Immunoreactive substance P in human saliva. A possible marker of chronic pain.

In all patients and volunteers, the levels of immunoreactive SP measured in saliva were about 100 times higher than the levels measured in plasma. SP per mg protein was consistently lower in both plasma and saliva of chronic pain patients than in healthy volunteers. These findings suggest that a simple noninvasive objective method of determining SP in saliva may become useful in the evaluation and treatment of chronic pain.

Perceptual and motor mechanisms for mental rotation of human hands.

We measured brain potentials from human subjects performing a mental rotation task requiring right-left judgments of misoriented hands, and a control task requiring palm-back judgments of the same stimuli. High-density, 128-channel event-related potentials (ERPs) were recorded from 16 normal, right-handed subjects. There was a main effect of task at five different latencies: 148 ms (occipital), 180 ms (parietal), 388 ms (vertex), 556 ms (central-parietal), and 900 ms (vertex). Source estimations derived from topographic data indicate that frontal brain regions were strongly activated after 300 ms in the control task, but not until about 900 ms in the rotation task. We conclude that the neural computations underlying mental hand rotation may be recruited from relatively early stages of visuo-perceptual analysis; these early computations influence subsequent processing within a parietal-prefrontal system for the integration of perception with action.

Primary and secondary healing in infected wounds. An experimental study.

The beneficial effects of delayed closure of contaminated wounds are recognized, but not well defined. Rats with abdominal incisions were infected with Staphylococcus aureus or Escherichia coli and then subgrouped for primary or secondary healing. Noninfected rats served as controls. Between seven and 119 days later, the wounds were subjected to breaking-strength determination, hydroxyproline analysis, and light and scanning electron microscopy. In the controls, secondary closure gave stronger wounds than primary closure. In those infected with S aureus the wounds were stronger than in the primary-closure control group, regardless of the closure method. Of the E coli-infected wounds those primarily closed were weaker than those secondarily closed. Secondary closure gave infected wounds fewer complications. Biochemical and microscopic examination did not explain these findings.

High-density mapping in an N400 paradigm: evidence for bilateral temporal lobe generators.

OBJECTIVE: The aim of this study was to obtain a more detailed description of N400 scalp topography than has previously been reported. METHODS: High-density (128 channel) visual event-related potentials were measured in an N400 paradigm using semantically incongruous sentence endings. RESULTS: The stimuli elicited an N400 with a centroparietal scalp distribution. In addition, P400s with similar timing and functional characteristics were observed at non-standard recording locations inferior to the temporal lobes. CONCLUSIONS: The data are consistent with intracranial evidence for bilateral activation of anterior medial temporal lobe structures. These structures are oriented such that the positive regions of their scalp fields lie largely outside of the area sampled by standard electrode montages. P400s at other non-standard scalp locations, including infraorbital and infraoccipital sites, may reflect volume conduction from the same generators, or activation of non-temporal lobe generators.

The use of controlled-release oxycodone for the treatment of chronic cancer pain: a randomized, double-blind study.

To compare the effectiveness and safety of controlled-release (CR) oxycodone tablets with immediate-release (IR) oxycodone in patients with chronic cancer pain, a multicenter, randomized, double-blind, parallel-group study was performed in 111 patients with cancer pain. Patients were treated with 6 to 12 tablets or capsules of fixed-combination opioid/nonopioid analgesics per day at study entry. Patients received 30 mg of CR oxycodone tablets every 12 hr or 15 mg of IR oxycodone four times daily for 5 days. No titration or supplemental analgesic medications were permitted. The mean (+/- SE) baseline pain intensity (0 = none, 1 = slight, 2 = moderate, 3 = severe) was 1.5 +/- 0.1 for the CR oxycodone-treated group and 1.3 +/- 0.1 for the group given IR oxycodone (P > 0.05). The 5-day mean pain intensity was 1.4 +/- 0.1 and 1.1 +/- 0.1 for the CR and IR groups, respectively (P > 0.05). Discontinuation rates were equivalent (33%). There was no significant difference between treatment groups in the incidence of adverse events. This study demonstrates that cancer pain patients given 6 to 12 tablets or capsules of fixed-dose combination analgesics can be equally well treated with CR oxycodone administered every 12 hr or IR oxycodone four times daily at the same total daily dose. CR oxycodone offers the benefits of twice daily dosing.

In vitro method for detecting precipitation of parenteral formulations after injection.

Many injectable formulations currently on the market, including diazepam and alprazolam, utilize one or more cosolvents to solubilize the active constituents. On injections into an aqueous medium, some of these components tend to precipitate. A simple procedure is described for measuring the degree of precipitation that occurs when a solubilized drug is injected. This in vitro technique was used to show that alprazolam injection shows less precipitation than diazepam injection under all tested conditions, and that the precipitation observed with diazepam can be controlled by ensuring that the formulation is injected very slowly. This simple technique also can be used during preformulation development to evaluate the relative potential for precipitation of various formulations.

Assessment of stress during periodontal surgery with intravenous sedation and with local anesthesia only.

Ten patients with moderate to advanced periodontal disease were subjected to two similar periodontal surgical procedures. Each patient received either intravenous conscious sedation with local anesthesia or local anesthesia only. The stress-reducing effects of a conscious sedation regimen consisting of pentobarbital, meperidine, and diazepam were evaluated in these patients. Stress was evaluated by monitoring changes in serum cortisol, human growth hormone, and vital signs. Blood samples were obtained at 15- to 30-minute intervals throughout each procedure and were evaluated for serum cortisol and growth hormone. The conscious sedation group had significantly lower serum cortisol levels and lower systolic blood pressure, indicating that the patients having periodontal surgery with conscious sedation experienced reduced stress. Physiologic stability was maintained for each patient, indicating that this conscious sedation regimen can be used to reduce measurable parameters of stress that patients develop during periodontal surgery.

An adaptable optimal controller for electric wheelchairs.

A microcomputer-based optimal control system for electric wheelchairs is presented. As the weight of a wheelchair is generally small compared to that of a user, the user's actual weight can have a tremendous impact on the dynamics of the wheelchair. The velocity feedback controller presented here was designed to give an optimal response for all users by incorporating a measure of the individual user's weight into the control algorithm. This allows optimal control of the motor velocities while assuring the constant stability of the system. Control of the system is managed by a modified proportional-integral-derivative (PID) controller, and the adaptability is handled by use of variable-structure control. The factors and considerations involved in such a system are identified, as are the advantages and disadvantages of the particular control strategy used. Experimental results are presented.

Automated grammatical tagging of child language samples.

Recent studies of the automated grammatical categorization ("tagging") of words using probabilistic methods have reported substantial levels of accuracy-over 95% agreement with manual tagging for words from a variety of texts. However, the texts with which this method has been tested were written by adults and edited by publishers. The present study examined the accuracy with which such methods could tag transcribed conversational language samples from 30 normally developing children. On a word-by-word basis, automated accuracy levels ranged from 92.9% to 97.4%, averaging 95.1%. Accuracy at correctly tagging whole utterances was lower, ranging from 60.5% to 90.3%, with an average of 77.7%. Probabilistic methods of coding language samples hold potential as a viable tool for child language research. Further study and improvement of automated grammatical tagging is warranted and necessary before widespread use can be made of this technology.

Results of grid keratotomy, superficial keratectomy and debridement for the management of persistent corneal erosions in 92 dogs.

Ninety-two cases of persistent corneal erosions in dogs were analyzed for breed, gender, age and which eye was affected. The results of the treatment of 92 persistent corneal erosions in dogs by superficial keratectomy (SK), grid keratotomy (GK), or debridement with a sterile dry cotton swab are presented. These techniques gave better rates of healing than have been previously reported. All cases of persistent corneal erosions healed in this study. However, it must be noted that three cases treated with debridement only failed to heal after several treatments and were eventually treated with SK. After one procedure 80 out of 92 (87%) had healed. After one procedure, 63% of cases treated with debridement healed, 100% of cases treated with SK healed, and 85% of cases treated with GK healed. At the first postoperative visit, 88% (21/24 cases) of ulcers treated by SK had healed, and 75% (39/52 cases) of ulcers treated by GK had healed. Only 25% of the persistent corneal erosions had healed at the first visit after debridement. All 24 cases of persistent corneal erosions treated with SK healed after one treatment in a mean +/- SD of 9.3 +/- 3.9 days (median of 7 days). Fifty-two cases were managed with GK; 44 (83%) of these healed with one procedure and eight cases required a second GK procedure to resolve. A mean +/- SD of 13.4 +/- 5.1 days (median of 11.5 days) following GK was required for the persistent corneal erosions to heal. Nineteen cases were initially managed by debridement with a dry cotton swab under local anesthesia. Sixteen out of these 19 debridement cases healed (giving an overall healing rate of 84%) in a mean +/- SD time of 23.4 +/- 11.1 days (median 21.5). There were three cases that did not heal with debridement. These cases were debrided at 10-20 day intervals for 30-60 days, and were then treated with SK. Two of these cases healed within 7 days, the other case required 18 days to heal. Sixty-three per cent of persistent corneal erosions treated with debridement healed after one procedure; however, only four out of 19 cases (21%) were healed at the first revisit. Complications were rare: corneal edema occurred in two cases following multiple GK, and excessive granulation tissue in one case was managed with a SK. There was the occurrence of an ulcer adjacent to the surgery site in four cases, two cases following GK and two cases following SK.

Effects of inflammation and aqueous tear film deficiency on conjunctival morphology and ocular mucus composition in cats.

An experimental model of keratoconjunctivitis sicca (KCS) was produced by removing the lacrimal gland and the gland of the third eyelid from the left eye of 6 cats. The right eye of each cat was left intact and used as a control. After 2 weeks, cats were euthanatized and the central portion of the upper eyelid from both eyes of each cat was excised. Histologic sections were stained with either hematoxylin and eosin or with a battery of biotinylated lectins including concanavalin A (conA), soybean agglutinin (SBA), wheat germ agglutinin (WGA), succinylated wheat germ agglutinin (S-WGA), Ulex europaeus agglutinin I (UEA), Dolichos biflorus agglutinin (DBA), Ricinus communis agglutinin (RCA), peanut agglutinin (PNA), and PNA pretreated with neuraminidase. Consistent differences in histologic features were not observed between conjunctivas with KCS and control conjunctivas. A variable degree of mononuclear cell infiltration of the substantia propria was observed in control conjunctivas and those with KCS. In both groups, conjunctival goblet cell density decreased and epithelial stratification increased as the degree of submucosal inflammatory cell infiltration increased. Lectin binding sites for DBA, WGA, S-WGA, UEA, PNA, and PNA pretreated with neuraminidase were detected on conjunctival goblet cells of conjunctivas with KCS and control conjunctivas. The mucus/glycocalyx layer of conjunctival epithelial cells in both groups of conjunctivas bound lectins RCA, WGA, UEA, and conA, but inconsistently bound S-WGA. In both groups, DBA principally bound to the mucus layer overlying normal epithelium, whereas PNA pretreated with neuraminidase consistently bound to the mucus layer of stratified epithelial surfaces free of goblet cells. Binding of SBA to goblet cells and the mucus/glycocalyx layer was variable.

Intraocular Cuterebra in a cat.

A cat had sudden onset of unilateral corneal edema, fibrin in the anterior chamber, uveal inflammation, and retinal dysfunction. A dead dipteran fly larva was removed surgically from the anterior chamber and was identified as an early instar of a Cuterebra sp.

Infrared laser diode irradiation has no behavioral or biochemical effect on pain in the sciatic nerve ligation-induced mononeuropathy in rat.

The aim of this study was to evaluate the effect of acute and repeated (5 days) treatment with various types of infrared (IR) diode lasers and probes (single- vs cluster-beam) on the pain response in rats with peripheral mononeuropathy produced by sciatic nerve ligation. Male Sprague-Dawley rats were anesthetized with sodium pentobarbital, and the mid-thigh was surgically exposed to reveal the sciatic nerve, around which four ligatures were loosely tied. On postoperative day 5, the skin over the sciatic nerve lesion was subjected to a 30-min daily local exposure from a 904-nm IR diode laser (700 Hz, average output power 10 mW) with a single-beam probe, a 830-nm IR diode laser (700 Hz) with either a single-beam (average output power 50 mW) or cluster-beam probe (average output power 15 mW), or placebo for 5 consecutive days. Two pain responses (foot-withdrawal time and the hind-paw elevation time) were measured on both sides using the radiant heat method on days 5 and 9. In addition, cold allodynia was measured on day 9 of treatment by placing the rats on a chilled metal plate (4 degrees C) and measuring the duration of elevation of either of the hind paws. On day 9, the animals were sacrificed for collection of the samples of brain and lumbar spinal cord for the determination of the tissue concentrations of dynorphin A1-8-like immunoactivity (DYN) using specific radioimmunoassay (RIA). The hind-paw withdrawal and elevation times on the right side in all groups subjected to the various methods of IR laser irradiation did not differ significantly as compared with the placebo-treated group when measured on days 5 and 9 after surgery. No statistically significant differences in withdrawal response and elevation time of the unaffected left hind paw were noted either. The measurement of cold allodynia similarly failed to reveal any effect in laser-treated groups versus placebo. The RIA analysis found that tissue concentrations of DYN were significantly elevated in the spinal cord ipsilaterally to the ligation side, as compared with the contralateral side, in all rats with sciatic nerve ligation. All modalities of IR diode laser treatment did not produce any significant difference in the brain and spinal cord level of DYN on postoperative day 9 in all treatment groups. It is concluded that repeated IR diode laser treatment did not reduce hyperalgesia induced by sciatic nerve ligation in rats.

Neuromagnetic responses associated with perceptual segregation of pitch.

In recent EEG investigations [Johnson, 2003] [Hautus, 2005], we described a novel late negative ERP component associated with binaural processing of auditory pitch based solely on interaural timing differences ("dichotic pitch"), an acoustic phenomenon that is closely analogous to visual perception of stereoscopic depth based on retinal disparities. The present study extends this research with neuromagnetic recordings of auditory evoked fields (AEFs) elicited by dichotically-embedded pitches. Eight healthy adult subjects listened to control stimuli consisting of 500 ms segments of broadband acoustic noise presented identically to both ears via earphones, and dichotic pitch stimuli created by introducing a dichotic delay to a narrow frequency region of the same noise segments and resulting in a perception of a pitch lateralized to the left or right of auditory space. Auditory-evoked fields (AEFs) were recorded using a 151 channel whole-head MEG system. Comparison of control and dichotic-pitch AEFs showed reliable amplitude differences during a time window of 150-350 ms. AEFs over the left hemisphere showed larger effects for contralateral than ipsilateral pitches, while the right hemisphere showed no differences for differently lateralized sources. The results indicate a relatively late stage of neural processing of binaurally-derived cues for the perceptual segregation of concurrent sound sources and support a right-hemisphere dominance for the processing of sound-source localization.

Modulation of neuromagnetic oscillatory activity during the observation of oro-facial movements.

A number of MEG/EEG studies have shown modulation of endogenous sensorimotor (mu and beta) rhythms during the observation of hand movements. These modulations are similar to patterns that occur during execution of movement and it has been hypothesised that the neural substrates of these rhythms may play a role in action representation and understanding the actions of others. In this experiment we wished to determine whether similar responses would be obtained during the observation of oro-facial movements. Neuromagnetic recordings (151 channels, CTF Systems) were obtained from six healthy subjects while they (1) observed a video of an experimenter making oro-facial movements (2) imitated the same movements and (3) observed hand movements. Source scanning using synthetic aperture magnetometry (SAM) was used to find changes in source power between these active conditions compared to pre-stimulus control conditions where no movement occurred. SAM images were created with 5 mm resolution in the beta (15-35 Hz) and mu (8-15 Hz) bands and showed source power decreases over parietal, occipital and sensorimotor areas. Time-frequency analysis of virtual SAM sensors from sensorimotor areas showed event-related desynchronisation of mu and beta bands following the onset of movement in all three conditions. These data demonstrate comparable activations of visuomotor mechanisms during observation or imitation of mouth movements and during observation of hand movements. These results support the notion that sensorimotor mechanisms play a role in achieving a representation of the oro-facial gestures of others.