Hepatitis E Virus in Cambodia: Prevalence among the General Population and Complete Genome Sequence of Genotype 4.

Hepatitis E virus (HEV) is a growing public health problem in many countries. In this study, we investigated HEV seroprevalence among the general population in the Siem Reap province, Cambodia, and performed HEV genetic analysis with the aim to develop an HEV prevention strategy. This seroepidemiological cross-sectional study conducted from 2010 to 2014 included 868 participants from four different locations in Siem Reap province, Cambodia. They answered questionnaires and provided blood samples for the analysis of hepatitis virus infections. Among the participants (360 men and 508 women; age range, 7-90 years), the prevalence of anti-HEV IgG was 18.4% (95% confidence interval: 15.9-21.0); HEV RNA was detected in two participants (0.23%) and was classified as genotype 3 and 4. Full-length genome of the genotype 4 isolate, CVS-Sie10, was sequenced; it contained 7,222 nucleotides and three ORFs and demonstrated high sequence identity with the swine China isolates swGX40 (95.57%), SS19 (94.37%), and swDQ (91.94%). Multivariate logistic regression analysis revealed that men, elderly people, and house workers were risk groups significantly associated with the positivity for anti-HEV IgG. This is the first report on the detection of HEV genotype 4 in humans in Cambodia and on the complete genome sequence of HEV genotype 4 from this country. Our study demonstrates that new HEV infection cases occur frequently among the general population in Cambodia, and effective preventive measures are required.

Seroprevalence, genotypic distribution and potential risk factors of hepatitis B and C virus infections among adults in Siem Reap, Cambodia.

AIM: We investigated hepatitis B virus (HBV) and hepatitis C virus (HCV) infections among adults in Siem Reap, Cambodia, to consider the prevention strategy in cooperation with the Ministry of Health in Cambodia. METHODS: Serological tests for determining HBV and HCV infections and questionnaires were performed from 2010 to 2012 among the general population in the province of Siem Reap. Multivariate logistic regression analysis was conducted to clarify the factors related to HBV and HCV infections. RESULTS: There were 483 participants, comprising 194 men and 289 women (age range, 18-89 years). The prevalence of hepatitis B surface antigen was not very high at 4.6%, while anti-hepatitis B core (anti-HBc) was high at 38.5%. All HBV DNA samples were classified as genotype C. Anti-HBc showed the trend that the older the age, the higher the positive rate (P = 0.0002). The prevalence of HCV RNA and anti-HCV were 2.3% and 5.8%, respectively. HCV RNA was detected in 39.3% of anti-HCV positive samples and most of them were classified as genotype 6 (54.5%) and 1 (27.3%). Remarkably, in multivariate logistic regression analysis, history of operation and blood transfusion were significantly associated with the positivity for HBV infection and HCV RNA, respectively. CONCLUSION: Our results showed that operation and blood transfusion were potential risk factors for HBV and HCV infection, respectively, and supposed that horizontal HBV transmission may be frequent in adults in Cambodia. Hence, for reducing HBV and HCV infections, it is necessary to improve the safety of blood and medical treatment.

Recovery effects of repeated exposures to normobaric hyperoxia on local muscle fatigue.

Reported recovery effects of hyeroxia are conflicted. This study aimed to identify the effects and the mechanisms of normobaric hyperoxia on the recovery of local muscle fatigue, which is the most commonly encountered form of fatigue both daily and in training and competitions. Twelve male subjects performed 3 × 3 × no less than 30 seconds of isometric quadriceps exercise at 70% of maximum voluntary isometric contraction (MVIC) separated by two 15-minute recovery sessions under 1 of 2 different atmospheric oxygen concentrations, one in normoxia (NOX; 20.9% O2) and another in hyperoxia (HOX; 30.0% O2). To assess the degree of fatigue and recovery, 4 parameters were used; MVIC, endurance time to exhaustion, blood lactate, and perceived exertion measured by a visual analog scale (VAS). Maximum voluntary isometric contraction improved an average by approximately 14% in HOX compared with NOX at the conclusion of the second recovery session. However, this was not associated with changes in other parameters because changes in endurance time, blood lactate, and VAS during the trials were similar. Based on our findings, we conclude that 2 sets of 15-minute recovery session in normobaric hyperoxia are effective for restoring MVIC from local muscle fatigue induced by intermittent intense exercises. For quicker recovery, athletes are recommended to repeat 15-minute recovery process under 30.0% hyperoxia.

Nuclei of the human raphe.

Along the raphe of the brain stem, a series of small neuronal groups can be observed in the medulla oblongata, the pons and the mesencephalon. The neurons located in and adjacent to the raphe are considered to produce mainly serotonin (5-HT). The groups of nuclei containing 5-HT were first reported in experimental animals in the early 1960s. The presence of such nuclei, however, has not yet been brought to light in the human brainstem except the few atlases, although in several neuroanatomy textbooks, extrapolated data are shown in the form of drawings as if they were the data from the human brain. The aim of this study is to present microscopic photos of such raphe nuclei made from serial sections of the human brainstem, and to clarify the differences between findings in human and textbook drawings from animal data.

Evaluation of the spinal cord white matter.

Morphological findings or evaluations of the nervous system have traditionally concentrated on cell somata; evaluations of the white matter have not been put forward up to now. This study was conducted to evaluate the white matter in the spinal cord with the LPH discriminative staining method which was proposed by Goto. Thanks to the minimum shrinkage ratio (10 ± 0% in length) which this technique allows, it is possible to evaluate the sizes of nerve axons, and to compare the arrangement of nerve fibers in various parts of the spinal white matter. As the axonal sizes reflect nerve conduction velocities, we would like to emphasize that these sizes or the differences in the arrangement of axons may be important for a better understanding of neurosymptomatology.

Morphological evaluation of the human pyramidal tract: tapering of axons.

It is generally accepted that nerve fiber conduction velocity is directly proportional to the fiber diameter under the condition, based on a supposition, that the transverse area of axons is normally maintained constant. Using an ideal preparation method for the purpose of axonal discrimination, we examined 43 human spinal cords after making transverse sections at the cervical, thoracic, lumbar and sacral levels and found a tapering of the axons in the lateral pyramidal tract, as the cross-sectional area of the pyramidal axons showed a definite decrease from the cervical to the sacral levels. Our results contradict the supposition that the transverse area of axons is normally maintained constant, which has for a long time been believed to be true without any evidence for it.

Morphometric and functional correlation of human neuronal somata: pyramidal motor, special sensory and general sensory systems.

Using an ideal tissue preparation method, we found a definite correlation between various human neuronal somata from the view point of accurate morphometry and functional evaluations. We believe this study may be of value, or even indispensable in the correct understanding of neurological symptomatology and phenomenology.

Development of the human parvocellular red nucleus. A morphological study.

Morphology of the human parvocellular red nucleus (RNp) was investigated in 14 fetuses aged from 12 to 39 weeks of gestation (WG). The brains were processed into celloidin-embedded serial sections. At 12 WG, the anlage of RNp was observed as an ovoid mass of immature neurons clustering into some groups. Lobular appearance in cross-sectional images was conspicuous during the early stages (12-23 WG), particularly at rostral levels. The fasciculus retroflexus of Meynert was seen as a prominent bundle of fibers surrounded by the most rostral part of RNp. Two types of neurons were identified: large and small neurons. Large neurons were earlier observed at 16 WG, and had a polygonal or multipolar perikaryon with abundant Nissl bodies from 28 WG onwards. Small neurons later appeared among large neurons at 21 WG, and had a triangular or ovoid perikaryon with scanty Nissl bodies. The volume of RNp showed an exponential increase with age during 20-39 WG. The mean of neuronal perikaryonal areas showed a linear increase with age in both types during 16-39 WG, although the degree of change was much greater in large neurons than small neurons. The current study has clearly demonstrated the presence of two neuronal populations and their differential growth in developing human RNp.

Comparison of preparation methods for the evaluation of nerve axons.

We have conducted a study to compare various preparation methods, including a certain number of new methods, to find which ones are best suited to the morphological evaluation of nerve fibers in the pyramidal tract of the human medulla oblongata. Our main concern was to find fixation and staining methods that would minimize errors, especially regarding the tissue shrinkage ratio and the ease of staining. From the two fixation methods we examined, the most satisfactory was the secondary chromic acid fixation (which gave the best overall results when followed by nitrocellulose embedding), as it gives the lowest shrinkage ratio with the narrowest range (10 +/- 0%). Among the ten staining methods we tested, we found that the most suitable for morphological evaluation were the discriminative staining methods (Luxol fast blue-Periodic acid-Schiff-Hematoxylin stain, Masson-Goldner-Goto method and modified Hematoxylin-Eosin stain) and the silver impregnation methods (Luxol fast blue-Silver impregnation and Luxol fast blue-Silver impregnation-Periodic acid-Schiff-Hematoxylin).

Morphometric evaluation of the aging process in various human nerve fibers.

We carried out a morphometric comparison of tissue sections from the human spinal cord, medulla oblongata, cranial nerves, autonomic nerves and spinal nerves with the help of a very accurate method that includes two-step fixation, nitrocellulose embedding and discriminative staining. We conducted morphometric evaluations to compare various axonal areas between different individuals using a combination of an image analyzer and a high power microscope. Our study showed a negative correlation between age and axonal area in all nerves except the greater splanchnic nerve. We believe that such data would not have been discovered without the use of our sophisticated electronic equipment together with the special preparation method we employed. Recourse to such methodology will allow more precise study of neurohistology and lead to better understanding of the aging process in human beings.

Morphological evaluation of the human pyramidal tract: gender and age differences.

Using a preparation method composed of secondary chromic acid fixation, nitrocellulose embedding and Luxol fast blue-PAS-hematoxylin stain (a discriminative staining method) for the purpose of axonal discrimination, we examined 43 human spinal cords (31 males and 12 females) at the C5 level to find the differences due to gender and aging in axons of the lateral corticospinal tract. These results can be of great help in understanding the motor functions in relation to gender differences and the aging process.

Morphometric development of the posterior funicular nucleus in the human medulla oblongata.

Using serial sections of 11 human brains of fetuses 18-40 weeks of gestation (WG), 2-month-old infant and 63-year-old adult, we conducted the quantitative analysis of the development of the posterior funicular nucleus (PFN) in the medulla oblongata. The results suggest the following: (1) The cells of the human cuneatus lateralis nucleus are easily distinguished from those of the gracilis nucleus and cuneatus medialis nucleus because the neuronal somatic areas are the largest; the smallest neuronal areas are in the cuneatus medialis nucleus. (2) The development of the PFN occurs at least in four stages characterized by the degree of maturation of the neurons: (a) immature neuronal stage before 20 WG, (b) preparatory stage between (a) and (c), (c) accelerated development stage from 30 to 40 WG, and (d) postnatal stage. The development of the human posterior funicular nucleus accelerates after 30 WG, and postnatal qualitative maturation also occurs. Developmental differences of the three nuclei of the PFN in the medulla oblongata are presented and discussed from the morphological and morphometric point of view.

Morphometric evaluations of the human nervous system.

It is very important to evaluate and accurately understand the various conditions of the human nervous system. In this review article, we introduce several morphometric reports that are proven to be accurate from the view point of various errors (range of tissue shrinkage ratios, microscopic multiple counting, artifacts of microscopic structures, etc.). We review the following aspects of the selected reports: methodology, developmental research, neuronal differences, gender differences, aging process and miscellaneous (nerve fibers, unmyelinated fibers, in relation to neuropathology, clinical image-analysis and immunohistochemistry).

Development of the human magnocellular red nucleus: a morphological study.

The development of the human magnocellular red nucleus (RNm) was studied in 20 fetuses at 12-39 weeks of gestation (WG). With microscopic observation on serial sections of the brain, we measured the profile area of a neuronal cell body. At 12WG, several islands of immature cells of the RNm appeared dorsal to the parvocellular red nucleus (RNp). At 16WG, the RNm was detected ventral to the RNp as a cluster of semilunar shape, consisting of basophilic neurons of various sizes. During 18-23WG, the neurons were dispersed dorsal to the RNp. They were isolated or aggregated as small clusters among the myelinated oculomotor nerve roots. Twenty-eight WG onwards, the neurons were widely distributed ventrolateral to the superior cerebellar peduncle and around the caudal pole of the RNp. Measurement of the profile area revealed that the average size of overall neurons increased almost linearly with the gestational age, and that two populations (large and small neurons) were clearly distinguished on the histogram from 33WG onwards. The relative position of the RNm to the RNp may vary among the individuals, especially in earlier fetal stage. This study suggests that the differentiation and maturation of neuronal cytoarchitecture of the RNm may gradually and monotonously progress during the later half of gestation.

Morphometric comparison of human nerve cells: pyramidal motor system.

We compared morphological and morphometric data on various motor neurons in the human pyramidal system using the modified Klüver-Barrera staining method with extremely minimized shrinkage ratio and an image-analyzer. We classified motor neurons in the human pyramidal system into three groups according to the measurement data. This report may be of interest to better understand the process of nerve conduction in the human pyramidal system.

Morphometric evaluation of posterior funiculus nerve fibers in relation to aging.

We morphologically evaluated the size of axons in the posterior funiculus in different age groups and examined the changes due to aging. In the past, such studies have been conducted at the cervical spinal cord (C6) level, and a decrease in the size and number of axons due to aging has been noted. The current study was conducted at the lower lumbar spinal cord (L2) level.

Morphological characteristics of nerve cells in the human general sensory system.

We conducted a study to compare morphological characteristics of various nerve cells in the sensory system. This sort of evaluation is indispensable for a better understanding of the nervous system in relation to clinical physiology and neurology.

Morphometric comparison of human spinal cord anterior horn cells between the cervical and lumbar levels.

Morphometric comparisons of anterior horn cells (AHCs) were conducted between cervical and lumbar spinal cords in 29 cadavers. The sizes of the AHCs were always larger at the lumbar level than at cervical level irrespective of the age of the spicemens, but a decrease in size with age was observed at both levels. The correlation coefficient shows a larger decrease in numbers and volumes at lumbar level than at cervical level.

Morphometric comparison of human nerve cells: special sensory system.

We compared morphometric data on various nerve cells in the human special sensory system using the modified Klüver-Barrera staining method with an extremely minimized shrinkage ratio and an image-analyzer. According to the measurement data of cell-body sizes, we classified nerve cells of the various nerve nuclei in the special sensory system into three groups. These data are of interest to better understand the process of nerve conduction in the special sensory system.