Prevalence and antimicrobial resistance of Enterococcus spp. isolated from animal feed in Japan

The rising prevalence of antimicrobial resistance (AMR) of bacteria is a global health problem at the human, animal, and environmental interfaces, which necessitates the “One Health” approach. AMR of bacteria in animal feed are a potential cause of the prevalence in livestock; however, the role remains unclear. To date, there is limited research on AMR of bacteria in animal feed in Japan. In this study, a total of 57 complete feed samples and 275 feed ingredient samples were collected between 2018 and 2020. Enterococcus spp. were present in 82.5% of complete feed (47/57 samples), 76.5% of soybean meal (62/81), 49.6% of fish meal (55/111), 33.3% of poultry meal (22/66), and 47.1% of meat and bone meal (8/17) samples. Of 295 isolates, E. faecium (33.2% of total isolates) was the dominant Enterococcus spp., followed by E. faecalis (14.2%), E. hirae (6.4%), E. durans (2.7%), E. casseliflavus (2.4%), and E. gallinarum (1.0%). Of 134 isolates which were tested for antimicrobial susceptibility, resistance to kanamycin was the highest (26.1%), followed by erythromycin (24.6%), tetracycline (6.0%), lincomycin (2.2%), tylosin (1.5%), gentamicin (0.8%), and ciprofloxacin (0.8%). All Enterococcus spp. exhibited susceptibility to ampicillin, vancomycin, and chloramphenicol. Of 33 erythromycin-resistant isolates, only two showed a high minimum inhibitory concentration value (>128 μg/mL) and possessed ermB. These results revealed that overall resistance to antimicrobials is relatively low; however, animal feed is a source of Enterococcus spp. It is essential to elucidate the causative factors related to the prevalence of AMR in animal feed

Ventrolateral Origin of Each Cycle of Rhythmic Activity Generated by the Spinal Cord of the Chick Embryo

BACKGROUND: The mechanisms responsible for generating rhythmic motor activity in the developing spinal cord of the chick embryo are poorly understood. Here we investigate whether the activity of motoneurons occurs before other neuronal populations at the beginning of each cycle of rhythmic discharge. METHODOLOGY/PRINCIPAL FINDINGS: The spatiotemporal organization of neural activity in transverse slices of the lumbosacral cord of the chick embryo (E8-E11) was investigated using intrinsic and voltage-sensitive dye (VSD) imaging. VSD signals accompanying episodes of activity comprised a rhythmic decrease in light transmission that corresponded to each cycle of electrical activity recorded from the ipsilateral ventral root. The rhythmic signals were widely synchronized across the cord face, and the largest signal amplitude was in the ventrolateral region where motoneurons are located. In unstained slices we recorded two classes of intrinsic signal. In the first, an episode of rhythmic activity was accompanied by a slow decrease in light transmission that peaked in the dorsal horn and decayed dorsoventrally. Superimposed on this signal was a much smaller rhythmic increase in transmission that was coincident with each cycle of discharge and whose amplitude and spatial distribution was similar to that of the VSD signals. At the onset of a spontaneously occurring episode and each subsequent cycle, both the intrinsic and VSD signals originated within the lateral motor column and spread medially and then dorsally. By contrast, following a dorsal root stimulus, the optical signals originated within the dorsal horn and traveled ventrally to reach the lateral motor column. CONCLUSIONS/SIGNIFICANCE: These findings suggest that motoneuron activity contributes to the initiation of each cycle of rhythmic activity, and that motoneuron and/or R-interneuron synapses are a plausible site for the activity-dependent synaptic depression that modeling studies have identified as a critical mechanism for cycling within an episode

Hemorrhagic Sudden Onset of Spinal Epidural Angiolipoma

Angiolipomas are relatively rare benign tumors. Spinal angiolipomas that generally induce slow progressive cord compression are most commonly found in the thoracic region. A 49-year-old female with obesity presented with a 1-week history of progressively worsening back pain, paresthesia of lower limbs, and gait disturbance. When thoracic magnetic resonance imaging (MRI) revealed a dorsal epidural mass at the Th5–Th8 level, the patient underwent a laminectomy for gross total excision of the lesion. Both mature fatty tissue and abnormal proliferating vascular elements with thin or expanded walls were observed in the resected tumor. Nonfiltrating spinal angiolipoma was diagnosed and confirmed by pathology. After the operation, sensory loss, numbness, and gait disturbance were improved following the disappearing severe back pain. Following examinations indicated the absence of recurrence within 1 year. The angiolipomas of the spine are rare causes of spinal cord compression that generally induce slow progressive cord compression, but sudden onset or rapid worsening of neurological deterioration is observed in hemorrhagic spinal angiolipoma

Comparison of the spatial distribution of two kinds of intrinsic signal and dye-related signals recorded from the cut, transverse face of the spinal cord.

<p>A–C. Distribution of intrinsic (A), dye-related (B) and filtered intrinsic (C) signals during an episode of rhythmic activity. The signals in A and C were obtained in an unstained slice which was subsequently stained with the voltage sensitive dye NK2671 to provide the VSD signals shown in B. The data were acquired at an interval of 0.42 ms. The signals in A were unfiltered and those in B were low pass filtered at 100Hz. The signals shown in D were obtained by bandpass (0.1–10 Hz) filtering the signals in A. Data in A is from a complete episode, while the signals shown in B and C were from 3 cycles from within the episode. Note that the dye signals and the filtered intrinsic signals exhibit a similar distribution of rhythmicity with the highest amplitude in the ventrolateral part of the cord. DR-dorsal root; VR-ventral root. D. The upper two traces compare the time course of the slow electrical activity (black trace) with the intrinsic optical signal recorded over the lateral motor column (red trace). Note that the peak of the optical signal is delayed with respect to the peak of the slow electrical signal. The lower traces are the cycle-averaged electrical (grey-integrated neurogram; black-low pass filtered slow potential) and optical (red-lateral motor column optical signal) responses generated from the cycles identified by the asterisks. The average was triggered from the peak of the slow potential as indicated by the vertical dotted grey lines. Data obtained from an E11 embryo.</p

Structure of an episode of activity.

<p>Electrical activity recorded from a ventral root during a single episode of rhythmic activity. The duration of the complete episode is demarcated by the grey box. The onset of the individual cycles is defined by the dotted grey lines. 2 cycles are identified at the end of the episode. Data from an E8 embryo. R VR DC-Right Ventral Root DC recording; R VR AC-Right Ventral Root AC recording high pass filtered to show the discharge pattern.</p

Timing of voltage-sensitive dye signals recorded from different regions of the cord at the onset of individual cycles.

<p>A. Electrical and optical recordings from an E10 embryo during an episode initiated by a dorsal root stimulus. The optical recording is from a single diode over the later motor column ipsilateral to the neural recordings. The dotted grey lines demarcate the onset of discharge in each cycle of activity. B. Electrical (black) and optical recordings from 3 different regions of the cord. As in previous figures, the red traces were averaged from diodes over the lateral motor column, the green traces from over the intermediate region and the blue traces from over the dorsal region. The diodes and their locations are shown in panel D. The neurogram is the DC trace low pass filtered between DC-20Hz. The episode was triggered by a single stimulus to the ipsilateral dorsal root at the time marked by the arrow (S). The numbered cycles correspond to those in panel A. The initial part of the first cycle (demarcated by a gray box) has been blown up in the inset to show the timing of the earliest optical and electrical activity. C. Quantification of the timing of optical activity in each cycle with respect to the onset of the electrical activity recorded from the ventral root. The onset delay of the optical signals with respect to the onset of the electrical signal is plotted for each cycle for each of the three regions. The numbers beside the plots correspond to the numbered cycles shown in A and B. Note that the dorsal optical activity precedes that of the electrical activity in the first cycle. Moreover the first cycle exhibits a dorsoventral sequence of activation but all of the other cycles are activated ventrodorsally. D. Location of the diodes whose signals were averaged to produce the traces shown in panels B and E. The diode signals are superimposed over a stained section in which the motoneurons were labeled with DiO and the dorsal root afferents were labeled with DiI. E. Cycle-triggered, averaged DC ventral root potential (VR DC-smoothed with a 20 point moving average) and integrated ventral root discharge (VR INT) together with the optical responses from the three different cord regions ipsilateral to the earliest ventral root activity. The last three cycles (3–5 in A) were averaged for these records. As before, each trace was averaged from several adjacent diodes (shown in D) and was normalized to its peak amplitude. Data were obtained at a sampling interval of 1.011 ms. F. Montage showing the pseudocolored diode array signals superimposed on the outline of the cord slice. Each image is the average of 45 frames obtained at the times indicated by the numbered regions over the traces in E. The first panel in the sequence (Anti) is an image generated during antidromic stimulation of the ventral root to identify the location of the lateral motor column. The arrow in image 3 shows the initial activity within the lateral motor column. The last image in the series (pre) was averaged from the frames delimited by the red rectangle marked pre in panel A. This image shows the spatial distribution of activity just before the last cycle.</p

Optical signals originate ventrolaterally at the onset of a spontaneous episode.

<p>Both intrinsic (A) and dye-related (B) optical signals originate over the motor column at the onset of a spontaneous episode. Each optical signal was spatially averaged from the diodes shown in the schematic of the cord above the traces and then normalized to its peak amplitude. The electrical activity was simultaneously recorded from the ipsilateral ventral root (VR). DC-unfiltered. AC-band pass filtered from 20-312Hz. The arrows delineate the initial slow rise of the depolarization and the onset of discharge (dotted gray line) in the ventral root. Data in A were obtained from an E9 embryo, while those in B were from an E10 embryo. C. Spread of dye-related activity at the beginning of a spontaneously occurring episode. The data were averaged from two E10 embryos and synchronized to the onset of the ventral root discharge (dotted gray line). The smoothed DC ventral root potential (VR DC) and the integrated ventral root discharge (VR INT-25ms integration time) are displayed together with the optical responses from three different cord regions (ventral/lateral motor column-red; intermediate-green; dorsal-blue). Each record was averaged from several adjacent diodes (shown in panel A of the montage illustrated in D) and normalized to its peak amplitude. Data were obtained at a sampling interval of 0.64ms. The gray lines over the traces in C, delineate the frames that were averaged to produce the pseudocolored montage. D. Montage of the pseudocolored diode array signals superimposed on the outline of the transverse face of the cord slice. The number in each panel corresponds to the numbered intervals over the electrical and optical traces. The first image (Anti) in the sequence is a pseudocolored image of the optical signals during antidromic stimulation of the ipsilateral ventral root to identify the location of the lateral motor column. The colored regions on the image identify the location of the diodes whose signals were averaged to produce the optical records in C. The second image (1, Gain×4) was obtained before the onset of discharge (vertical dotted gray line in C) and was averaged from 105 frames (67ms/frame) is displayed at 4×the gain of the remaining images. All remaining images (2–8) were averaged from 45 frames (29ms/frame). The arrow shows the earliest detectable activity occurs over the lateral motor column.</p

Summary of the timing of activity in different cycles within episodes evoked by a stimulus to the dorsal root.

<p>A. Sequence of activity for the first cycle in 5 experiments in which the dorsal optical activity preceded the onset of the electrical activity. B. Average delays of the optical signals measured from the onset of the ipsilateral electrical activity for 7 experiments. Time 0 corresponds to the onset of slow electrical activity recorded from the ipsilateral ventral root. The color code is the same as in the previous figures. Data are mean±SEM.</p

Antidromic stimulation of the ventral roots to identify the location of the motor nucleus.

<p>A. The optical responses recorded from the photodiode array are shown superimposed over a confocal transverse section of the slice in which the recordings were made. After the optical experiment, the slice was fixed and motoneurons were labeled retrogradely with DiI (green) applied to the ventral root and primary afferents were labeled anterogradely with DiO (red) applied to the dorsal root. Each optical signal is the response to a single antidromic stimulus averaged from a train of stimuli (8 stimuli at 20 Hz) and recorded in the presence of a cholinergic (mecamylamine 50 µM) and a GABAA (bicuculline 50 µM) antagonist to block the synaptic inputs from R-interneurons onto motoneurons (33). The largest antidromic responses coincided with the location of motoneuron cell bodies. B. Antidromically-evoked optical responses from another cord in which it was possible to detect antidromic spikes adjacent to the central canal at a location corresponding to that of preganglionic neurons (red traces), in addition to those over the motor nucleus (green traces). DR-dorsal root; VR-Ventral Root. The colored traces in panel B have been averaged and are displayed on an expanded scale in C. It can be seen that the antidromic responses near the central canal are smaller and have a longer latency that those recorded from the motor nucleus. D. The slow antidromic responses recorded over motoneurons are depressed in the presence of the nicotinic cholinergic antagonist mecamylamine and the GABAA receptor antagonist bicuculline to block the recurrent synaptic input from R-interneurons to motoneurons. The upper trace (control) was obtained under control conditions and the lower trace (bicuculline and mecamylamine) in the presence of the drugs. The time of the ventral root stimuli (stim.) are marked by the dots beneath the traces. E. Spectrum of optical responses recorded over motoneurons and evoked by ventral root stimulation under control conditions. The signals were obtained at three different illumination wavelengths in response to a train of ventral root stimuli (stim.). Note the reversal of the signals at 580 nm and the absence of a significant signal at 630 nm, the isosbestic point of the dye. The times of the ventral root stimuli (stim.) are indicated beneath the traces. In this, and all other figures, the vertical calibration arrows indicate the direction of increased light transmission (decreased light absorption). Data in A from an E11 embryo and in B, C and D from an E10 embryo. Data in E from an E11 embryo.</p