Corticotropin-releasing factor immunoreactivity increases in the cerebellar climbing fibers in the novel ataxic mutant mouse, pogo

The ataxic pogo mouse (pogo/pogo) is a novel neurological mutant, which was derived as an inbred strain (KJR/MsKist) from a Korean wild mouse. The pathological manifestations include a difficulty in maintaining a normal posture, the failure of inter-limb coordination and an inability to walk straight. In this study, we examined the distribution of corticotropin-releasing factor (CRF) immunoreactive cerebellar climbing fibres and their projections to tyrosine hydroxylase (TH) immunoreactive Purkinje cells in the cerebellum of the pogo mutant mouse using immunohistochemistry. In the pogo/pogo mouse, a subset of climbing fibres was stained more intensely for CRF than in the control. Moreover, ataxic pogo mouse, neurons of the inferior olivary nucleus projecting climbing fibres were also more intensely stained for CRF than in the control. In the pogo/pogo mouse, TH immunoreactivity was located in the Purkinje cells, whereas no TH expression was found in the control. Double immunostaining for CRF and TH in the pogo/pogo cerebellum revealed that the distribution of TH-immunoreactive Purkinje cells corresponded to terminal fields of CRF-immunoreactive climbing fibres but not to the CRF-immunoreactive mossy fibres. Therefore, we suggest that an increase of CRF level may alter the function of targeted Purkinje cells and that it is related to the ataxic phenotype in the pogo mutant mouse.     

Qualitative and quantitative studies of the inferior olivary complex in the water buffalo (Buballus bubalis)

The shape and neuronal number of the inferior olivary complex (IOC) were determined in the water buffalo (Buballus bubalis). The configuration and interrelations of the IOC compartments were ascertained by investigating serial sections through the whole rostro-caudal extent of the IOC. Nissl-stained celloidin sections of six water buffalo's brainstems were used. The IOC in the water buffalo consisted of three major nuclei and four small cell groups. The medial accessory olivary nucleus (MAO) had the longest rostro-caudal extent as well as the highest number of neurons (98,000 +/- 3,000). Although the total area of the principal olivary nucleus (PO) was smaller than the area of the dorsal accessory olivary nucleus (DAO), the PO had the second largest neuronal number. The total number of neurons on both sides of the IOC was 210,000 +/- 7,000 cells. The average neuronal density was 3,000 cells/mm3. Although the size of the PO relatively increases while the size of MAO decreases with the development of the cerebellar hemispheres, the IOC in most mammals maintains a similar structure except for the higher primates and marsupials. The water buffalo IOC showed morphological similarities to the almost all mammalian IOC including rats as follow; the main part of the MAO consists of three subgroups (a, b and c), the DAO is Boomerang-shaped while the PO is a simple U-shaped structure.     

Pax-7 immunoreactivity in the post-natal chicken central nervous system

In this immunocytochemical study on the constitutive expression of Pax-7 protein in the postnatal chicken brain, Pax-7 showed region and cell type specific expression. In the optic tectum, only cells in grey matter showed positive immunoreactivities (IRs), whereas those in the white matters did not show any IRs. In thalamic nuclei and several pontine nuclei, we also localized Pax-7 positive IRs. On the contrary, in the cerebellum, Pax-7 was mainly localized within the Bergmann glia, whereas Purkinje cells did not show any IRs. In double immunolabelling studies, most of the Pax-7 IRs did not originate from neuroglial cells such as oligodendrocytes, microglia or astrocytes, but from neurons, with the exception of Bergmann glia in the cerebellum. The presence of Pax-7 IRs in the adult chicken brain could suggest that Pax-7 might play a role in maintaining normal physiological function in some postnatal chicken brain cells.     

Progressive encephalomyelopathy and cerebellar degeneration in 10 captive-bred cheetahs

Progressive ataxia, with head tremor, developed in 10 captive-born cheetah cubs under six months of age. The condition was usually preceded by coryza and an ocular discharge. Initially the ataxia and weakness affected the hindquarters, then the forelegs, and head tremor developed later. Significant pathological changes were confined to the central nervous system. There was widespread Wallerian degeneration in the funiculi of the spinal cord (except those in the dorsal columns), in the medulla and in the cerebellum. In the cerebellum there was degeneration of Purkinje cells and of the molecular and granular cell layers. There was chromatolysis in the Purkinje cells, the ventral horn cells of the spinal cord and in the neurons of the lateral vestibular nucleus. The olivary nucleus was necrotic. There were foci of inflammatory cells in the molecular layer of the cerebellum and in the medulla. The cause of the disease remains unknown.     

肉鸡肌纤维与肉质关系研究

选用地方鸡种白耳鸡、泰和鸡、黄羽肉鸡康达尔 12 8cs、石岐杂鸡、快长肉鸡AA鸡cs、蛋用鸡罗曼褐cs和矮脚黄鸡 7个鸡种 (系 ) ,在同样条件下饲养到 13周龄进行品味、肌纤维等性状测定。结果表明 ,地方鸡种白耳鸡、矮脚黄鸡肉质最好 ,石岐杂鸡、康达尔黄鸡 12 8cs次之 ;罗曼褐cs、AA鸡cs、泰和鸡肉质较差。白肌纤维直径大于红肌纤维和中间型纤维直径 ,其所占比例与肉质呈负相关 ,白肌纤维比例愈高 ,肉质愈差。肌纤维直径与鸡的体重大小没有关系。肌纤维密度和直径共同影响肌肉的蛋白质含量 ,且与肉质呈负相关     

Lysosomal storage disease caused by Sida carpinifolia poisoning in goats

A neurologic disease characterized by ataxia, hypermetria, hyperesthesia, and muscle tremors of the head and neck was observed for 2 years in a flock of 28 Anglo-Nubian and Saanen goats on a farm with 5 ha of pasture. Six newborns died during the first week of life, and five abortions were recorded. The predominant plant in the pasture was Sida carpinifolia. The disease was reproduced experimentally in two goats by administration of this plant. Three goats with spontaneous disease and the two experimental animals were euthanatized and necropsied. No significant gross lesions were observed. Fragments of several organs, including the central nervous system, were processed for histopathology. Small fragments of the cerebellar cortex, liver, and pancreas of two spontaneously poisoned goats and two experimentally poisoned goats were processed for electron microscopy. Multiple cytoplasm vacuoles in hepatocytes, acinar pancreatic cells, and neurons, especially Purkinje cells, were the most striking microscopic lesions in the five animals. Ultrastructural changes included membrane-bound vacuoles in hepatocytes, Kupffer cells, acinar pancreatic cells, Purkinje cells, and the small neurons of the granular cell layer of the cerebellum. Paraffin-embedded sections of the cerebellum and pancreas were submitted for lectin histochemical analysis. The vacuoles in different cerebellar and acinar pancreatic cells reacted strongly to the following lectins: Concanavalia ensiformis, Triticum vulgaris, and succinylated Triticum vulgaris. The pattern of staining, analyzed in Purkinje cells and acinar pancreatic cells coincides with results reported for both swainsonine toxicosis and inherited mannosidosis.     

出生前山羊延髓主要灰质核团的形成和发育

对从胚胎期第6周到出生前山羊的延髓主要灰质核团发育的组织学变化进行了系统研究。结果表明：（1）第6周山羊胚胎延髓处在组织发生末期和核团形成初期，是其内部核团构建的关键时期。（2）延髓内不同核团中的神经元发育变化在时间上有较大的差异，在同一核团内神经元胞体发育分化在时间上也有差异，即同一核团内较为成熟的神经元胞体的数量由少到多，细胞质内的尼氏小体也存在由少到多，由小变大的过程。（3）延髓灰质结构形成和神经元发育分化的规律：有些结构发生早，而神经元分化较晚，如下橄榄核、三叉神经脊束核和孤束核；有些结构发生略晚，但其中的神经元胞体分化和发育则较早，如舌下神经核、疑核及延髓的网状结构；有些结构发生早，神经元发育也早，如迷走神经背核。     

鸡小脑突触素的表达与发育研究

应用免疫组织化学SP法，研究了突触素在15、20胚龄、10日龄鸡小脑中的表达。结果表明：突触素免疫反应产物以颗粒、小点、阳性神经元和阳性纤维形式出现；突触素免疫反应产物密度在15、20胚龄、10日龄有3种变化趋势：①分子层、颗粒层、中间核、外侧上核、外侧下核中随胚(日)龄增加保持不变；②蒲肯野氏层随胚(日)龄增加先升高后降低；③小脑内侧核则随胚(日)龄增加逐渐升高。结果揭示：突触素最终合成部位可能在高尔基复合体；20胚龄左右是蒲肯野氏层突触形成的高峰期；小脑各部突触在出壳前后发育具有不均衡性。     

Lymphocyte subset distribution in apparently normal and single intradermal test-positive water buffaloes analyzed by flow cytometry

Monoclonal antibodies (mAbs) against bovine lymphocyte cell surface antigens namely, MHC Class I, MHC class II (DP, DQ and DR), CD3, CD4, CD8, gamma delta TCR, WC1N1 and WC1N2, were tested for their reactivity on apparently normal buffalo mononuclear cells prepared from spleen, lymph nodes and peripheral blood. All the mAbs cross-reacted with the buffalo mononuclear cells. The mean (+/-SD) CD4:CD8 cell ratio in the peripheral blood of apparently normal buffaloes was 1.08+/-0.049 while in the spleen and lymph nodes it was 0.90+/-0.080 and 1.81+/-0.430, respectively. The lymphocyte subsets in the buffaloes positive for tuberculosis by the single intra dermal (SID) test was found to be altered; the CD4 cells were reduced while the CD8 and gamma delta cells were increased. The mean CD4:CD8 ratio in the SID positive buffaloes was 0.36+/-0.010.     

Naturally occurring parvovirus-associated feline hypogranular cerebellar hypoplasia-- A comparison to experimentally-induced lesions using immunohistology

Three cases of feline cerebellar hypoplasia are presented. At the time of examination, the ages of the cats ranged from 2 months to 1 year. Necropsy revealed cerebellar and pons hypoplasia. Polymerase chain reaction for parvoviral deoxyribonucleic acid was positive in cerebellar tissue. Cell-specific immunolabeling was used to characterize the lesions, which were characterized into 2 types. In type 1 lesions, the cortex was nearly agranular, with an extremely thin molecular layer; the Purkinje cells were randomly placed and oriented, and their stunted main dendrite produced a thorn-covered atrophic dendritic tree; the basket cell axons ran randomly and had dysmorphic endings; and myelinated fibers were severely reduced in folia axes. In type 2 lesions, the cortex was hypogranular; the Purkinje cells were linearly organized, but their main dendrite extended too far in the molecular layer before giving up smooth, bent secondary dendrites; many basket cells were located along the cerebellar surface, and their axons ran at right angle to the surface; myelinated fibers were moderately reduced. Defects in climbing fiber synapse translocation and elimination were evident in both types of lesion. This immunohistologic study allowed a comparison between lesions in these spontaneous cerebellar hypoplasia cases with those documented when using silver impregnation studies after perinatal experimental cerebellar damage. Such a comparison is consistent with viral infection that occurs before birth in all 3 cases. Progress in parvovirus biology knowledge suggests that viral NS1 protein cytotoxicity might explain degenerative changes in the Purkinje cells that were present, in addition to the development defect.     

Comparison of cytologic and histologic evaluations of the conjunctiva in the normal equine eye

OBJECTIVE: To describe the cells observed in conjunctival brush cytology (CBC) from normal horses and compare these findings with conjunctival structural histology so as to understand which cells are recovered from CBC. METHODS: This study was divided into three parts. (1) Conjunctival brush smears were collected from 20 healthy horses on both eyes and a differential count on 300 cells was carried out on May Grünwald-Giemsa (MGG) smears. (2) A similar protocol was used for whole eyes from five horses obtained rapidly after death from a slaughterhouse. The eyes were then assessed for conjunctival histology. (3) Cytobrush smears were collected from five healthy horses. Smears were examined after MGG or periodic acid Schiff (PAS) staining. RESULTS: The differential cell count showed a majority of deep and intermediate epithelial cells with very few superficial and goblet cells in both eyes. A stratified columnar to cuboidal epithelium was observed on nearly the whole surface of the conjunctiva. A stratified squamous type was observed at the palpebral and bulbar edges. Areas with highest mucus cell indices were found from the nasal to the temporal edge of the equine inferior conjunctiva in the upper palpebral segment near the fornix and in a part of the nasal fornix. In MGG smears no mucus cells were identified; however, they were numerous in PAS smears (22.6% +/- 11) and were mostly cylindrical cells (42.5% +/- 14.4 PAS positive). CONCLUSIONS: Cytobrush smears in the healthy horse are characterized by a majority of polyhedral and cylindrical cells and a few squamous cells. The cylindrical cells may be mucous cells and probably originate from the main stratified columnar to cuboidal epithelium.     

Determination of the minimum anesthetic concentration and cardiovascular dose response for sevoflurane in chickens during controlled ventilation

OBJECTIVE: To determine the minimum anesthetic concentration for sevoflurane and effects of various multiples of minimum anesthetic concentration on arterial pressure and heart rate during controlled ventilation in chickens. STUDY DESIGN: Prospective experimental study. ANIMALS: Seven healthy chickens, 6 to 8 months old, weighing 1.6 to 3.4 kg. METHODS: A rebreathing, semiclosed anesthetic circuit was used. Anesthesia was induced by mask with sevoflurane in oxygen. Each chicken was endotracheally intubated, then controlled ventilation was started and the end-tidal CO2 partial pressure was maintained at 30 to 40 mm Hg. Body temperature was maintained at 39.5 degrees to 41.0 degrees C. The inspired and end-tidal sevoflurane concentration were monitored with a multigas monitor. Minimum anesthetic concentration was determined as the minimal end-tidal sevoflurane concentration which prevented gross purposeful movement in response to clamping a toe for 1 minute. After the determination, the cardiovascular effects of sevoflurane at 1.0, 1.5, and 2.0 times the minimum anesthetic concentration were determined. RESULTS: The minimum anesthetic concentration for sevoflurane was 2.21% + 0.32% (mean +/- SD). Mean arterial pressure and heart rate at minimum anesthetic concentration were 84 +/- 13 mm Hg and 150 +/- 58 beats/min, respectively. There was a dose-dependent decrease in arterial pressure. The heart rate did not change significantly over the range 1 to 2 x minimum anesthetic concentration. No cardiac arrhythmias developed throughout the experiments. CONCLUSIONS AND CLINICAL RELEVANCE: The minimum anesthetic concentration for sevoflurane in chickens was within the range of minimum alveolar concentration reported in mammals. When the concentration of sevoflurane is increased during controlled ventilation in chickens, decrease in arterial pressure should be expected.     

The comparison of the collagen fiber contents and hepatic stellate cell distribution in male and female chicken livers

The difference in collagen fiber content, morphological properties and hepatic stellate cell (HSC) distribution was investigated in the liver of both sexes in chicken. Collagen fiber specimens were obtained by maceration treatment with NaOH solution. HSCs were detected using desmin‐specific immunohistochemistry. The ratio of liver weight to body weight was larger in the female than the male chickens. Collagen fiber content, the numerical density of HSCs and the percentage area displaying desmin immunopositivity were not different between the right and left lobes of the liver, in both male and female chickens. However, all of these parameters were larger in the males than the females. In the light microscopic observation, many HSCs in the male had large and elongated cytoplasmic processes. Conversely, HSCs with poorly developed cytoplasmic processes were frequently observed in females. Liver tissue is structurally stronger in male chickens than females and the activity and density of HSCs may be related to the collagen fiber content in chicken liver.     

Immunohistochemical distribution of alpha B-crystallin in the cerebellum of dogs infected with canine distemper virus

The cerebella of 12 dogs infected with canine distemper virus (CDV) and those of three normal dogs were examined. The avidin-biotin-peroxidase complex technique was used to detect alphaB-crystallin (alphaB-c) immunoreactivity and immunolocalisation of the CDV antigen. CDV antigens, immunopositive astrocytes, oligodendrocytes and granular neurons were seen in both the white and grey matter of the infected dogs. In the controls, alphaB-c immunopositive glial cells were seen in the white matter and around the Purkinje cells. In dogs with distemper, alphaB-c immunoreactivity was not observed in some of the glial cells around the Purkinje cells. A significant negative correlation of P < 0.01 level was found between areas of severe demyelination and the number of alphaB-c immunopositive cells in dogs infected with CDV. Such correlation was not observed between mild and moderate demyelinating areas and alphaB-c immunostaining. The alphaB-crystallin/ total number of cells ratio was found to be significant in severely affected demyelinating areas (P < 0.05). These data indicate that there was a relationship between the degrees of CDV associated with demyelination and the level of alphaB-c expression in the glial cells.     

Hereditary striatonigral and cerebello-olivary degeneration of the Kerry blue terrier. I. Gross and light microscopic central nervous system lesions

The character and progression of gross and light microscopic central nervous system lesions associated with a hereditary neurodegenerative disease with juvenile onset are described in ten affected Kerry blue terriers. The central nervous system lesions were progressive and, although there was some variability, followed a relatively well-defined temporal course. Degeneration of Purkinje cells in the cerebellar cortex was evident at the onset of clinical signs (approximately 4.5 months of age). After two weeks to one month of clinical illness, retrograde transsynaptic neuronal degeneration occurred in the olivary nucleus. Degeneration of both large and small neurons in the caudate nucleus began approximately two to three months after the onset of clinical signs, and by seven to eight months of clinical illness, the caudate nucleus was reduced to numerous microcystic cavities and almost was devoid of neurons except for a narrow subependymal zone and the tail of the nucleus. Neuronal depletion in the pars reticularis of the substantia nigra, which was evident after five to seven months of clinical illness, was attributed to an anterograde transsynaptic mechanism of neuronal degeneration. The disease in the Kerry blue terrier is compared with similar neurodegenerative diseases in man. The pathogenesis may involve altered neurotransmitter systems in the cerebellar cortex and caudate nucleus.     

Morphologic and morphometric study of the facial nerve in clinically normal adult dogs.

A morphologic and morphometric study was carried out on the facial nerve: to determine normal histologic data in myelinated fibers of clinically normal young adult dogs; to establish reference values for mean fiber diameter, and to delineate the relative diameter frequency distribution curve. Few degenerative changes were observed in single teased-nerve fibers and semithin cross-sectional nerve preparations. Statistical difference was not observed between left and right facial nerves. The distribution of fiber diameters in the facial nerve was unimodal. Mean (+/- SD) fiber diameter of the facial nerve was 3.92 +/- 1.18 microns. Approximately 89% of fibers in the facial nerve had diameter between 3 and 6 microns. Fiber diameter ranged from 2 to 12 microns; however, less than 1% of fibers had diameter greater than 8 microns.     

Calcium-Binding Proteins and Neurotransmitters in the Stomach Myenteric Plexus of the Korean Native Goat

The expression of calbindin D-28k (CB), calretinin (CR), substance P (SP) and calcitonin gene-related peptide (CGRP) in the stomach myenteric plexus of the Korean native goat stomach was investigated by immunohistochemistry. The results demonstrated the presence of nerve fibers and cell bodies immunoreactive (IR) to CB, CR, SP and CGRP. In tissues of rumen, reticulum, omasum and abomasum, some distinct neuronal populations could be distinguished according to their morphologic and neuronal chemical properties: Dogiel type I cells which have irregular lamellar dendrites and a single axon, Dogiel type II cells which have large ovoid cell bodies and several long axon-like processes, and small filamentous interneurons. CB-, CR-, SP- and CGRP-IR neurons and fibers were observed in the myenteric plexus of stomach, and varicose nerve fiber immunostained to SP and CGRP also were found in the muscle layer. In myenteric plexus of the stomach, CB- and SP-positive neurons were characterized by Dogiel type II and CR-IR neurons were classified Dogiel type I with lamellar dendrites, and immunoreactivity of CGRP was very weak in the somata. SP- and CGRP-IR nerve fibers formed dense networks within the myenteric ganglia. SP-IR cell bodies and their fibers were found in the myenteric plexus, and the immunoreactivity and number of cell bodies were more than CB-, CR-, and CGRP-IR neurons. These results suggest that SP, CGRP, CB and CR in the myenteric neurons of Korean native goat stomach may have play an important role in the dynamic movement.
(Support contributed by: Korean Research Foundation 2003-015-E00195).     

Morphologic and morphometric studies of the intrinsic laryngeal muscles in clinically normal adult dogs

To establish normal histologic and histochemical data and to determine reference values for fiber type proportions (percentages, mean fiber diameters, atrophy and hypertrophy factors, and variability coefficients), a histochemical study was carried out on intrinsic muscles of the larynx (cricothyroid, cricoarytenoid lateralis, cricoarytenoid dorsalis, and thyroarytenoid muscles) of clinically normal dogs. Using myosin adenosine triphosphatase stain under acidic preincubation (pH 4.3) conditions, 3 histochemical fiber types--1, 2A, and 2C--were recognized. The percentage of type-2C fibers varied from 1 to 2% in thyroarytenoid muscles to approximately 10% in cricoarytenoid lateralis muscles. There was no significant difference in mean diameter between left- and right-side specimens of each muscle for type 1 vs type 2. The largest fibers (mean +/- SD) of both types were observed in the cricothyroid muscles (type 1, 38.19 +/- 7.76 microns; type 2, 43.25 +/- 8.66 microns), and the smallest fibers were found in the thyroarytenoid muscles (type 1, 29.38 +/- 5.12 microns; type 2, 33.84 +/- 6.20 microns). Respective mean diameters of fiber types from cricoarytenoid dorsalis (type 1, 32.05 +/- 5.69 microns; type 2, 38.95 +/- 7.75 microns) and cricoarytenoid lateralis (type 1, 33.75 +/- 5.98 microns; type 2, 37.09 +/- 7.01 microns) muscles were similar. The histographic distribution of fiber type diameters was unimodal in all muscles. In each muscle, the mean fiber diameter of type-2 fibers was greater than that of type-1 fibers.(ABSTRACT TRUNCATED AT 250 WORDS)     

鸡海马结构向小脑各叶的直接投射——HRP逆行追踪法研究

采用HRP法逆行追踪鸡海马结构向小脑各叶投射的起始神经元.将50%HRP溶液分别引入鸡小脑Ⅳ、Ⅴ、Ⅵ、Ⅶ、Ⅷ、Ⅸ各叶,对端脑及间脑、脑干及小脑进行冰冻切片,TMB呈色,观察脑内标记细胞出现的位置.结果发现,除第Ⅸ叶外,注射小脑各叶双侧端脑海马结构的旁海马内侧区(APHm)出现大量标记细胞.在注射Ⅳ、Ⅴ、Ⅵ后偶尔在内侧隔核出现少量逆标细胞.随着注射点由Ⅳ叶到Ⅸ叶的后移,在APHm出现标记细胞的可能性渐小,而小脑前核,包括延髓大细胞网状核、下橄榄核、内外侧桥核、中脑的内侧螺旋核等标记的可能性相应地增大.隔核的标记细胞主要定位于外侧隔核.本研究结果表明,鸡存在海马向小脑的直接投射,这种投射主要终止于小脑的前叶、中叶.因为哺乳动物,鸡的隔核与海马有密切的联系.     

The immunologists' debt to the chicken

The immune system of the chicken is an invaluable model for studying basic immunology and has made seminal contributions to fundamental immunological principles. Graft versus host responses and the key role of lymphocytes in adaptive immunity were first described in work with chicken embryos and chickens. 2. Most notably, the bursa of Fabricius provided the first substantive evidence that there are two major lineages of lymphocytes. Bursa-derived lymphocytes, or B cells, make antibodies while thymus-derived, or T cells, are involved in cell-mediated immune responses. 3. Gene conversion, the mechanism used by the chicken to produce its antibody repertoire, was first described in the chicken and requires the unique environment of the bursa. Subsequently it has been shown that some mammals also use gene conversion. 4. The chicken's Major Histocompatibility Complex (MHC), the first non-mammalian MHC to be sequenced, is minimal, compact and some 20-fold smaller than that of mammals. Uniquely, the chicken MHC is strongly associated with resistance to infectious diseases. 5. The first attenuated vaccine was developed by Louis Pasteur against a chicken pathogen, fowl cholera, and the first vaccine against a natural occurring cancer agent, Marek's disease virus, was developed for the chicken. 6. Vaccination of chick embryos on the 18th d of incubation, another breakthrough using chickens, provides protection early after hatching. In ovo vaccination now is widely practised by the poultry industry. 7. Evidence that widespread and intensive vaccination can lead to increased virulence with some pathogens, such as Marek's disease virus and infectious bursal disease virus, was first described with chicken populations. It warns of the need to develop mo resustainable vaccination strategies in future and provides useful lessons for other species, including in the human population. 8. Recombinant DNA technologies now provide the opportunity for the rational design of new vaccines. Such vaccines could contain the protective immunogenic elements from several pathogens and immunomodulatory molecules to direct and enhance immune responses so providing improved protection. The important thing will be to design vaccines that are sustainable and do not drive pathogens to ever-increasing virulence.