A single base transversion in the flanking region of an equine microsatellite locus affects amplification of one allele

The equine dinucleotide microsatellite HMS7 is part of a microsatellite panel utilized in a parentage verification programme at the Veterinary Genetics Laboratory (Davis, California, USA). Apparent non-Mendelian inheritance was noted when a Quarter Horse mare was excluded as the parent of two offspring based on analysis of the HMS7 locus. The mare's DNA type qualified her as a parent of the offspring at an additional 20 microsatellite loci. The three animals appeared homozygous for HMS7 with each possessing an allele different from that of the other two animals. Polymerase chain reaction primers designed to bind outside the published primer-binding sites amplified an additional shared allele in all three horses, which qualified the mare as the dam of the two offspring. Sequencing of this newly detected allele revealed a C to A transversion in one of the published primer-binding regions. Apparent non-Mendelian inheritance at the HMS7 locus has been encountered in an additional 26 Quarter Horse parentage cases. In all instances, the lack of amplification and resultant 'null' allele was shown to be caused by the same transversion.     

A microsatellite map of wheat

Hexaploid bread wheat (Triticum aestivum L. em. Thell) is one of the world's most important crop plants and displays a very low level of intraspecific polymorphism. We report the development of highly polymorphic microsatellite markers using procedures optimized for the large wheat genome. The isolation of microsatellite-containing clones from hypomethylated regions of the wheat genome increased the proportion of useful markers almost twofold. The majority (80%) of primer sets developed are genome-specific and detect only a single locus in one of the three genomes of bread wheat (A, B, or D). Only 20% of the markers detect more than one locus. A total of 279 loci amplified by 230 primer sets were placed onto a genetic framework map composed of RFLPs previously mapped in the reference population of the International Triticeae Mapping Initiative (ITMI) Opata 85 x W7984. Sixty-five microsatellites were mapped at a LOD >2.5, and 214 microsatellites were assigned to the most likely intervals. Ninety-three loci were mapped to the A genome, 115 to the B genome, and 71 to the D genome. The markers are randomly distributed along the linkage map, with clustering in several centromeric regions.     

Laboratory studies in cross-species lung transplantation

The lack of sufficient suitable human donor lungs for the many patients requiring pulmonary transplantation as life-saving therapy for end-stage lung diseases has generated extensive interest in cross-species lung transplantation. Ethical concerns and those of animal rights advocates have prompted studies of nonprimate species as potential solid organ donors for humans. This paper provides an overview of some of the laboratory studies of cross-species pulmonary transplantation performed over the past 20 years and focuses, in particular, on more recent work (from our laboratory and others) in the area of porcine-to-primate pulmonary xenotransplantation.     

Mycobacteriosis caused by Mycobacterium genavense in birds kept in a zoo: 11-year survey

Nucleotide sequences encoding GroEL homologs of intracellular symbionts in three closely interrelated aphids were compared with one another and that for GroEL. It was suggested that in these proteins a particular position is highly susceptible to amino acid substitution, through which the GroEL homologs of symbionts seemed to have acquired a unique function on top of the activity as molecular chaperone. This may represent a rare example of non-neutral evolution of molecule under the positive selection pressure.     

Persistent infection promotes cross-species transmissibility of mouse hepatitis virus

Persistent infection with mouse hepatitis virus (MHV) strain A59 in murine DBT (delayed brain tumor) cells resulted in the emergence of host range variants, designated V51A and V51B, at 210 days postinfection. These host range mutants replicated efficiently in normally nonpermissive Chinese hamster ovary (CHO), in human hepatocarcinoma (HepG2), and to a lesser extent in human breast carcinoma (MCF7) cell lines. Little if any replication was noted in baby hamster kidney (BHK), green African monkey kidney (COS-7), feline kidney (CRFK), and swine testicular (ST) cell lines. By fluorescent antibody (FA) staining, persistent viruses V10B and V30B, isolated at days 38 and 119 days postinfection, also demonstrated very low levels of replication in human HepG2 cells. These data suggest that persistence may rapidly select for host range expansion of animal viruses. Pretreatment of HepG2 cells with a polyclonal antibody directed against human carcinoembryonic antigens (CEA) or with some monoclonal antibodies (Col-1, Col-4, Col-12, and Col-14) that bind human CEA significantly inhibited V51B infection. Under identical conditions, little or no blockade was evident with other monoclonal antibodies (kat4c or Col-6) which also bind the human CEA glycoproteins. In addition, an antibody (EDDA) directed against irrelevant antigens did not block V51B replication. Pretreatment with the Col-4 and Col-14 antibodies did not block Sindbis virus replication in HepG2 cells or MHV infection in DBT cells, suggesting that one or more CEA glycoproteins likely functioned as receptors for V51B entry into human cell lines. To test this hypothesis, the human biliary glycoprotein (Bgp) and CEA genes were cloned and expressed in normally nonpermissive BHK cell lines by using noncytopathic Sindbis virus replicons (pSinRep19). By growth curves and FA staining, human CEA and to a much lesser extent human Bgp functioned as receptors for V51B entry. Furthermore, V51B replication was blocked with polyclonal antiserum directed against human CEA and Bgp. Under identical conditions, the parental MHV strain A59 failed to replicate in BHK cells expressing human Bgp or CEA. These data suggest that MHV persistence may promote virus cross-species transmissibility by selecting for virus variants that recognize phylogenetic homologues of the normal receptor.     

A microsatellite genetic linkage map of human chromosome 13

We have characterized 21 polymorphic (CA)n microsatellites for the development of a genetic map of chromosome 13. Fifteen markers were isolated from a flow-sorted chromosome 13 library, four CA repeats were derived from NotI-containing cosmid clones, and two polymorphic markers were described previously (J. L. Weber, A. E. Kwitek, and P. E. May, 1990, Nucleic Acids Res. 18: 4638; L. Warnich, I. Groenwald, L. Laubscher, and A. E. Retief, 1991, Am. J. Hum. Genet. 49(Suppl.): 372 (Abstract)). Regional localization for all of the markers was performed by amplification of DNA from five somatic cell hybrids containing different deletions of chromosome 13. Genetic markers were shown to be distributed throughout 6 of the 11 resolvable chromosomal subregions. Using data from nine families provided by the Centre d'Etude du Polymorphisme Humain (CEPH), a framework map of 12 of these 21 markers was developed. Six of the 12 markers form three pairs, with each two members of a pair being tightly linked, such that nine systems of markers can be distinguished. The average heterozygosity of these 12 markers is 0.75. The total length of the sex-averaged map is 65.4 cM (Kosambi), with an average distance of 8.2 cM between systems of markers (eight intervals). Seven remaining markers were placed provisionally into the framework map.     

A role for thyroid hormones in the regulation of diet-induced thermogenesis in birds

The possible involvement of thyroid hormones in avian diet-induced thermogenesis (DIT) was investigated in two lines of cockerels divergently selected for high (R-) or low (R+) food efficiency. For a given body weight, R+ cockerels exhibited a higher food intake than R- cockerels (+49 to +76%) and increased DIT (+25%). Plasma thyroxine (T4) level did not differ between lines whatever the feeding status of the birds. Plasma 3,5,3'-triiodothyronine (T3) level was lower in fasted R+ than in fasted R- cockerels while the opposite was observed after a meal. Iopanic acid injections reduced both plasma T3 concentrations and heat production to the same levels in both lines. Hepatic 5'-deiodinase activity measured with an exogenous sulfhydryl group (dithiothreitol) did not differ between lines, but when the sulfhydryl group was omitted, the activity was higher in R+ than in R- birds (90 v. 42 pmol T3/min per liver). T3-binding capacity of isolated hepatic nuclei was higher (+76%) in R+ than in R- birds. Long-term or acute pair-feeding of R+ cockerels to the level of R- controls did not alter these results. The present results suggest that T3, mainly originating from peripheral conversion of T4 to T3, is involved in DIT in the R+ line. Availability of endogenous sulfhydryl groups appears to play an important part in the modulation of hepatic deiodinase activity. The higher concentration of nuclear T3 receptors may further increase the effects of the hormone, suggesting a major role of thyroid hormones associated with catecholamines in the stimulation of avian DIT. The underlying thermogenic mechanisms remain to be elucidated.     

A survey of Blastocystis in domestic chickens

The prevalence and morphology of Blastocystis in fresh faecal material from 227 domestic chickens was investigated. A very high prevalence of infection (approximately 95%) was found in chickens from four of the five commercial farms studied. Extremely high numbers of Blastocystis were found in the majority of samples. Blastocystis cells showed considerable variation in size, ranging from approx. 3 microm to approx. 120 microm in diameter. This size range is more extreme than those previously recognised for the organism from chickens. All chickens from one farm appeared free of Blastocystis infection. Most Blastocystis cells appeared to be the vacuolar form, although the shape of the cells and the appearance of the central vacuole contents varied considerably within and among faecal samples. Nuclei showed "spots" of electron-opaque material, generally arranged as a band within the nuclei. Multiple individual cysts within a single outer fibrillar layer were found in addition to single cysts without an encompassing fibrillar layer.     

A highly conserved microsatellite in the dystrophin gene of diverse mammalian species

Corticobasal degeneration (C.B.D.) is a neurodegenerative disorder characterized mainly by an asymmetrical a kineto-rigid syndrome associated with fronto-parietal cortical signs, particularly apraxia. Conventional imaging even magnetic resonance imaging (M.R.I.) has often been considered as poorly contributive for the diagnosis of C.B.D. We retrospectively studied routinely performed M.R.I. scans of 15 patients presenting a clinical and metabolic (P.E.T/S.P.E.C.T.) syndrome characteristic of probable C.B.D. M.R.I. scans were assessed by 3 investigators, not aware of the clinically most affected side, taking into account M.R.I. technical parameters. We quantified, on each side, the cortical atrophy (frontal, parietal and temporal) and the white matter changes, by using the semi-quantified method of Victoroff et al. (1994). Abnormalities were considered if observed by at least 2 of the 3 investigators. Abnormalities were then correlated with the side initially and most severely affected. The most contributive findings were the asymmetric parietal atrophy (clinically correlated in 93 p. 100 of cases), asymmetric frontal atrophy (clinically correlated in 60 p. 100) and asymmetric dilatation of the lateral ventricles (clinically correlated in 60 p. 100). 80 p. 100 of affected subjects displayed at least 2 of these M.R.I. abnormalities. These results are in accordance with the metabolic and pathologic features of C.B.D. This study demonstrates that M.R.I. evaluation of the cortical atrophy asymmetry may contribute to the diagnosis of C.B.D.     

Tuberculosis in captive exotic birds

Avian tuberculosis was studied clinically and pathologically in 137 affected birds from the National Zoological Park during a 7-year period (1969-1975). Twelve of 22 orders exhibited were affected by the disease, and the highest annual mortality was 4% (in 1975). Antemortem diagnosis of early cases of the disease, based on tuberculin testing, and serologic, hematologic, and radiographic studies, was inconsistent and often not conclusive. Pathologically, the diseases primarily involved digestive organs and spleen. There was a spectrum of lesions consisting of nodules of large foamy histiocytes packed with acid-fast bacilli to giant cell-containing granulomas that were often caseous but not cavitated or calcified. Amyloidosis was seen in approximately 20% of the cases. Mycobacterium avium serotype 1 was isolated from 30 tuberculous birds cultured. There was no sex predilection, and most of the affected birds were adults ranging from 1 to 10 years of age.     

Water conservation in xerophilic birds

Adaptation of birds to dry climates involves decrease in relative evaporative water loss (mechanism unknown); decrease in GFR; and high urine osmolality in the dehydrated state in conjunction with cloacal resorption parameters, which allows the urine to enter the cloaca without a further water loss. The net result of the adaptations is such that the best adapted birds, such as the budgerygah (12) and the zebra finch (13) fed dry seeds alone can live without water. Similar features have been observed also in the best adapted desert rodents (27). Finally, birds tolerate a 7 to 13% increase in plasma osmolality (9). Tucker (28) has calculated that the budgerygah has fuel for 14 h flight; we have calculated that it also has water for 14 h (12) at neutral temperature and humidity. This is better than the Jumbo Jet, which can only fly for 12 h.     

A technique for eliminating allele specific amplification failure during DNA amplification of heterozygous cells for preimplantation diagnosis

Advances in techniques of molecular biology have made possible the amplification of specific genes from single cells. This has a major clinical application in preimplantation diagnosis of monogenic disorders. However, the incidence of allele specific amplification failure (allele drop out) in heterozygous single cells can lead to misdiagnosis and the transfer of affected embryos. Few studies have been done to investigate the actual cause of allele drop out, although some investigators have succeeded in reducing but not eliminating it. Here we report the efficiency of amplifying both alleles in heterozygous cells lysed according to two different protocols. A total of 177 heterozygous cells from carriers of cystic fibrosis (CF) and haemoglobin C (HbC) were lysed using two different lysis buffers. Interestingly none of the cells that were lysed with sodium dodecyl sulphate/proteinase K showed any example of allele specific amplification failure whereas in those lysed by KOH/dithiothreitol it was present in 17.6 and 4.7% of the CF and HbC cells respectively. Our results suggest that the phenomenon of allele specific amplification failure is at least in part dependent on the lysis buffer used.     

A histologic survey of trabeculectomy

Thirty-two tissue blocks removed during trabeculectomy were examined histologically, 16 by both scanning electron microscopy and light microscopy, and 16 by light microscopy alone. Trabecular meshwork was identified in 27 specimens (84.3%) and Schlemm's canal in 17 (53%), each found with equal frequency by either technique. Ciliary muscle was identified in eight of 32 of specimens (25%) examined by light microscopy, but was present in 12 of 16 specimens (75%) examined by scanning electron microscopy. The identification of these specific anatomical structures did not correlate with the postoperative outcome. These findings support the contention that trabeculectomy is a filtering procedure.     

元宇宙技术综述

元宇宙作为一个新兴的概念受到了产业界、学术界、媒体界及公众的广泛关注,国内外众多公司也纷纷在“元宇宙”领域布局,但布局的背后离不开强大的技术支持。本文从技术维度深入剖析元宇宙。首先,从科学与技术的角度论述元宇宙的概念及内涵,并总结了目前从业者、专家以及学者对元宇宙的不同见解。其次,概述了元宇宙相关关键技术,包括网络及运算技术（5G、6G、物联网、云计算、雾计算及边缘计算）、管理技术（能耗管理、资源管理、会话管理及时空一致性管理）、虚实对象连接、建模与管理技术（X联网、身份建模、社会计算、去中心化管理技术）、虚实空间交互与融合技术（拓展现实、电子游戏技术、脑机接口技术）等。元宇宙不宜称为新技术,而是IT新技术的综合运用。技术的进步与发展,将为元宇宙的实现和应用奠定坚实的基础,同时元宇宙的发展也会促进现有技术的升级换代。最后,展望了元宇宙发展面临的诸多挑战,以清晰认识、理智决策、稳步探索元宇宙。      

A survey of audit activity in general practice

BACKGROUND: Since 1991, all general practices have been encouraged to undertake clinical audit. Audit groups report that participation is high, and some local surveys have been undertaken, but no detailed national survey has been reported. AIM: To determine audit activities in general practices and the perceptions of general practitioners (GPs) regarding the future of clinical audit in primary care. METHOD: A questionnaire on audit activities was sent to 707 practices from 18 medical audit advisory group areas. The audit groups had been ranked by annual funding from 1992 to 1995. Six groups were selected at random from the top, middle, and lowest thirds of this rank order. RESULTS: A total of 428 (60.5%) usable responses were received. Overall, 346 (85%) responders reported 125.7 audits from the previous year with a median of three audits per practice. There was no correlation between the number of audits reported and the funding per GP for the medical audit advisory group. Of 997 audits described in detail, changes were reported as 'not needed' in 220 (22%), 'not made' in 142 (14%), 'made' in 439 (44%), and 'made and remeasured' in 196 (20%). Thus, 635 (64%) audits were reported to have led to changes. Some 853 (81%) of the topics identified were on clinical care. Responders made 242 (42%) positive comments on the future of clinical audit in primary care, and 152 (26%) negative views were recorded. CONCLUSION: The level of audit activity in general practice is reasonably high, and most of the audits result in change. The number of audits per practice seems to be independent of the level of funding that the medical audit advisory group has received. Although there is room for improvement in the levels of effective audit activity in general practice, continued support by the professionally led audit groups could enable all practices to undertake effective audit that leads to improvement in patient care.