Long-term self-reported health and disability after COVID-19 in public employees.

BACKGROUND: Although the symptomatology has been assessed in multiple studies among persons recovering from coronavirus disease 2019 (COVID-19), less is known regarding long-term general health and disability. We aimed to assess long-term self-reported disability in public employees after predominantly mild COVID-19 in comparison with individuals who had negative COVID-19 polymerase chain reaction (PCR) test results. METHODS: Public employees within Region Västra Götaland were offered tests to identify SARS-CoV-2 infection (n = 56,221) and were invited to complete an online survey that included the World Health Organization Disability Assessment Schedule. Questionnaires were sent out between January 26 and March 5, 2021. A total of 14,222 (25.3%) employees responded, of which 10,194 (18%) were included (women n = 8749, 85.8%). Of these, 7185 (70.5%) participants had a negative PCR test result (controls). A total of 1425 (14%) had a positive PCR result and were categorized in the sub-acute phase (4-12 weeks post COVID-19), and 1584 (15.5%) had a positive PCR test and were categorized in the post COVID-19 phase (> 12 weeks). RESULTS: Fifty-two percent of controls rated disability of varying degrees, versus 73% and 64% of participants in the sub-acute and post COVID-19 phase, respectively. Being "emotionally affected" was the most frequently reported disability in the sub-acute phase, the post COVID-19 phase, as well as in controls. The proportion of participants reporting difficulties for 20-30 days was higher in the sub-acute phase than in the post COVID-19 phase (27.9% vs. 21.8%, p < 0.001) as well as in a comparison between participants in the post COVID-19 phase and controls (21.8% vs 14.2%, p < 0.001). Compared with controls, severe disability was more common in the post COVID-19 phase among both women (15.8% vs. 10.7%,), and men (9.8% vs. 6.8%). CONCLUSIONS: Disability was present in all groups; however, reported disability was greater in the sub-acute phase than in the post COVID-19 phase. The higher levels of disability reported in the COVID-19 patient population may indicate a persisting need for rehabilitation and recovery. In general, women reported a greater degree of disability than men in the sub-acute and post COVID-19 phases.

Lentiviral vector induced insertional haploinsufficiency of Ebf1 causes murine leukemia.

Integrating vectors developed on the basis of various retroviruses have demonstrated therapeutic potential following genetic modification of long-lived hematopoietic stem and progenitor cells. Lentiviral vectors (LV) are assumed to circumvent genotoxic events previously observed with γ-retroviral vectors, due to their integration bias to transcription units in comparison to the γ-retroviral preference for promoter regions and CpG islands. However, recently several studies have revealed the potential for gene activation by LV insertions. Here, we report a murine acute B-lymphoblastic leukemia (B-ALL) triggered by insertional gene inactivation. LV integration occurred into the 8th intron of Ebf1, a major regulator of B-lymphopoiesis. Various aberrant splice variants could be detected that involved splice donor and acceptor sites of the lentiviral construct, inducing downregulation of Ebf1 full-length message. The transcriptome signature was compatible with loss of this major determinant of B-cell differentiation, with partial acquisition of myeloid markers, including Csf1r (macrophage colony-stimulating factor (M-CSF) receptor). This was accompanied by receptor phosphorylation and STAT5 activation, both most likely contributing to leukemic progression. Our results highlight the risk of intragenic vector integration to initiate leukemia by inducing haploinsufficiency of a tumor suppressor gene. We propose to address this risk in future vector design.

European consensus statement on diagnosis and treatment of adult ADHD: The European Network Adult ADHD.

BACKGROUND: Attention deficit hyperactivity disorder (ADHD) is among the most common psychiatric disorders of childhood that persists into adulthood in the majority of cases. The evidence on persistence poses several difficulties for adult psychiatry considering the lack of expertise for diagnostic assessment, limited treatment options and patient facilities across Europe. METHODS: The European Network Adult ADHD, founded in 2003, aims to increase awareness of this disorder and improve knowledge and patient care for adults with ADHD across Europe. This Consensus Statement is one of the actions taken by the European Network Adult ADHD in order to support the clinician with research evidence and clinical experience from 18 European countries in which ADHD in adults is recognised and treated. RESULTS: Besides information on the genetics and neurobiology of ADHD, three major questions are addressed in this statement: (1) What is the clinical picture of ADHD in adults? (2) How can ADHD in adults be properly diagnosed? (3) How should ADHD in adults be effectively treated? CONCLUSIONS: ADHD often presents as an impairing lifelong condition in adults, yet it is currently underdiagnosed and treated in many European countries, leading to ineffective treatment and higher costs of illness. Expertise in diagnostic assessment and treatment of ADHD in adults must increase in psychiatry. Instruments for screening and diagnosis of ADHD in adults are available and appropriate treatments exist, although more research is needed in this age group.

SPECT and PET of the dopamine transporter in attention-deficit/hyperactivity disorder.

Abnormalities of frontostriatal circuits, which are modulated by dopamine, have been found by brain imaging studies in patients with attention-deficit/hyperactivity disorder (ADHD). With special radiolabeled ligands selective imaging of the dopamine transporter (DAT), which has a key function in dopamine metabolism, can be performed by SPECT and PET. Most of the studies showed a higher DAT availability in untreated patients with ADHD compared with controls. The relationship between DAT availability and a polymorphism of DAT1 gene in patients with ADHD is not clear and the results are controversial. It has been shown that methylphenidate lowers DAT availability very effectively in normal people and in patients with ADHD. First results seem to indicate that nonresponders to methylphenidate among ADHD patients have a low primary DAT availability, whereas patients with a good response to the drug have high DAT. Nicotine seems to lower DAT availability such as stimulant medication; this may explain the high percentage of smokers among patients with ADHD. Zinc is a DAT inhibitor and seems to have a positive therapeutic effect on ADHD symptoms. This article reviews the function and structure of the DAT, the results of DAT imaging with SPECT and PET, the relations between DAT availability and the DAT1 gene polymorphism, the influence of stimulants on DAT and the significance of DAT for therapeutic response, nicotine, zinc and psychotic symptoms in patients with ADHD.

Value of 99mTc-TRODAT-1 SPECT to predict clinical response to methylphenidate treatment in adults with attention deficit hyperactivity disorder.

BACKGROUND: In a previous study, binding of Tc-TRODAT-1 to the dopamine transporter (DAT) was found to be higher in patients with attention deficit hyperactivity disorder (ADHD) as compared to healthy controls. AIM: To determine whether the degree of Tc-TRODAT-1 binding to the striatal DAT may have a predictive role on the response to methylphenidate (MPH) in patients with ADHD. METHODS: Twenty-two adult patients suffering from ADHD underwent a brain SPECT scan with Tc-TRODAT-1. After the scan patients received MPH, individually medicated up to 60 mg.day. Severity of illness was estimated using the Clinical Global Impression (CGI-S) Scale before treatment. Ten weeks after the beginning of MPH treatment the improvement in global symptoms was rated by the Clinical Global Improvement Scale (CGI-I). RESULTS: Before treatment 17/22 patients with ADHD presented with higher striatal DAT binding as compared to age-matched healthy controls (+23.8%; P<0.01). After treatment with MPH a significant improvement of ADHD symptoms was demonstrated by the CGI-I in 16 of these 17 patients (CGI-S before: 4.8; CGI-I after MPH: 1.9; P<0.01). Five patients showed reduced DAT binding prior to therapy (-14.4%; P=0.04); these patients did not respond to MPH therapy (CGI-S before: 4.5; CGI-I after MPH: 4.2; P=0.40). CONCLUSION: Our findings suggest that ADHD patients with primarily elevated binding of Tc-TRODAT-1 to the striatal DAT responded better to therapy with MPH as compared to those with normal or low DAT binding. Consequently, our results - even if obtained on a small collective indicate that measurement of DAT may be an important prognostic predictor for therapy response to MPH.

Striatal dopamine transporter availability and DAT-1 gene in adults with ADHD: no higher DAT availability in patients with homozygosity for the 10-repeat allele.

In 29 adults with attention deficit hyperactivity disorder (ADHD) striatal dopamine transporter (DAT) availability was assessed by [(99m)Tc]TRODAT-1 SPECT and correlated with 3' VNTR polymorphism of the DAT gene on chromosome 5p15.3. Seventeen patients showed homozygosity for the 10-repeat allele, two homozygosity of the 9 allele and 10 were heterozygous (9-10). No statistically significant difference in DAT availability was found between patients with 10-10 carriers (DAT 1.28 +/- 0.34) and with at least one 9 allele (DAT 1.31 +/- 0.27); when smokers were excluded, DAT availability was 1.38 +/- 0.28 in the 10-10 carriers (n = 12) and 1.42 +/- 0.19 in the 9-10 and 9-9 carriers (n = 7). In conclusion, no higher striatal DAT was found in patients with homozygosity of the 10 allele of the DAT gene in this study. These results differ from a study in 11 Korean children with ADHD, in which 10-10 carriers showed higher DAT availability in [(123)I]IPT SPECT. Discrepancies may be explained by differences in patient's age, ethnical differences, different imaging techniques or the limited number of patients included in both studies.

ADHD in adolescence and adulthood, with a special focus on the dopamine transporter and nicotine.

The persistence of attention deficit hyperactivity disorder (ADHD) into adolescence and adulthood has now been accepted as a clinical entity. The rate of prevalence among adults is assumed to be from 2% to 4%. With increasing age, a symptom change has to be considered; disturbance of attention becomes more prominent, whereas hyperactivity often diminishes or changes to inactivity. Neuroimaging studies show a high striatal dopamine transporter (DAT) availability in most adults with ADHD; this can be reduced by stimulants. Nicotine seems to have a stimulant-like action on the DAT. In most adults with ADHD, therapy has to be multimodal, combining psychotherapy and medication. Methylphenidate is the first-line drug in adult ADHD; further options are amphetamine and noradrenaline reuptake inhibitors. Nonresponders to methylphenidate seem to have no elevated DAT availability prior to therapy. Combination with other psychiatric disorders occurs frequently in adults with ADHD; in these patients additional pharmacological treatment with special regard to the comorbid disease is recommended.

Influence of striatal dopamine transporter availability on the response to methylphenidate in adult patients with ADHD.

In this study, we investigated whether availability of striatal dopamine transporter (DAT) may have an influence on the response of adult patients with attention deficit hyperactivity disorder (ADHD) on methylphenidate (MPH). In 18 non-smoking and non-medicated adult patients with ADHD, availability of DAT was measured with [(99m)Tc] TRODAT-1 SPECT. Then, the patients received methylphenidate (MPH), individually titrated up to 60 mg per day. Ten weeks later, clinical improvement was rated by Clinical Global Impressions scale. In all, 6 patients were classified as non-responders, and 12 responded to MPH. From the non-responders, 5 presented with a DAT availability below that of normal controls of the same age, whereas in the group of responders all patients had elevated DAT availability. There was a significant negative correlation between values for global clinical improvement and striatal DAT availability. In conclusion, ADHD patients with low DAT availability seem not to respond to therapy with MPH.

The dopamine transporter and neuroimaging in attention deficit hyperactivity disorder.

There is evidence that abnormalities within the dopamine system in the brain play a major role in the pathophysiology of attention deficit hyperactivity disorder (ADHD). For instance, dopaminergic psychostimulants, the drugs of first choice in ADHD, interact directly with the dopamine transporter (DAT). Molecular genetic studies suggest involvement of a polymorphism of the DAT gene in ADHD. More recent imaging studies show abnormalities in various brain structures, but particularly in striatal regions. In the current paper we review recent studies in this area. First in vivo measurements of DAT with single photon emission computed tomography (SPECT) in ADHD patients revealed an elevation of striatal DAT density. No differences in DAT density between the left and right side and between putamen and caudate nucleus have been found in [99mTc]TRODAT-1 SPECT of ADHD patients. Patients with ADHD and with a history of nicotine abuse both displayed lower values of DAT density in [99mTc]TRODAT-1 SPECT than non-smokers with ADHD. DAT seem to be elevated in non-smoking ADHD patients suffering from the purely inattentive subtype of ADHD as well as in those with the combined or purely hyperactive/impulsive subtype.

Stimulant-like action of nicotine on striatal dopamine transporter in the brain of adults with attention deficit hyperactivity disorder.

Eleven adult patients with attention deficit hyperactivity disorder (ADHD) without medication, consuming 7-40 cigarettes per day, showed statistically significant lower values for striatal dopamine transporter (DAT) measured by [99mTc]TRODAT-1 SPECT compared to 11 non-smoking drug-naive patients with ADHD, matched for sex and age, despite higher ADHD scores for the smokers. Because stimulants have been shown to reduce primarily elevated DAT density in adults with ADHD, it can be suggested that nicotine acts in a similar way on striatal DAT as do stimulants.

[Development of prospective diapause-germs (Bombyx mori L.)in vitro without dormancy : III. Their competence and interference inLYS-media, without extraembryonic storage material]. / Die Entwicklung prospektiver Diapause-Keime (Bombyx mori L.)in vitro ohne Dormanz : III. Ihre Kompetenz und Interferenz in LYS-Medien ohne extraembryonales Depotmaterial.

In a foreign protein medium (LTS), naked prospective diapause-germs have a tendency to develop into fully segmented germ bands (stage of dormancy): however, deposition of chorion, serosa or yolk will stimulate themvia the medium to pass through organogenesis without delay (non-dormancy =Nd). The question remained if a germ from a non-diapausing egg would stimulate the uninterrupted development of such a test-germ (Tk). Using 2 different or 2 to 3 equal germs as well asTk alone (control) in hanging drops, experimental evidence was obtained permitting conclusions not only as to the competence and sensitivity of the embryonic reaction system, but also as to factors of the extraembryonicNd-action systems in eggs with and without predetermined diapause regulatory mechanism. 1. Control explants (1 prospective diapause-Tk inLYS) have aNd-level of 10% (= average ofNd-rates in operative stages). This difference from former controls (0%Nd) can be explained by the longer operative procedure in the cups when several test-germs were used. Seven minutes after opening an egg one can reckon on aNd-stimulationvia LYS medium from egg-residues in the operation cup. While transfering the test-germs, yolk-derived substances related to theNd-stimulation can be brought into the hanging drop. The difference betweenNd-controls and the results ofNd-stimulation have to be statistically significant. 2. In experiments with two different germs in LYS-droplets no influence of a non-diapause germ (polyvoltine) on a prospective diapause germ (univoltine) could be noted. Accordingly, the naturally prospectiveNd-germs, have a tendency to develop to dormant germs only, as do the 2Tk and 1Tk controls. In combination with artificially prospectiveNd-germs (univoltine, HCl-treated) no stimulation of the test germ occcurs; nor after explantation with an older germ-band or a young embryo. 3. Combination with two prospective diapause germs of equal age in a drop, having the same minor contamination ofNd-factors in the operation-cup, will result in a threefold increase of theNd-level as compared to the 1Tk-controls. The competence of the embryonic reaction system ranges from a coatlike germ-anlage to a germ with thoracic segmentation. In the dish-like germ-anlage at diapause onsetin ovo maximalNd-rate (70%) is foundin vitro, i.e. highest sensibility towardsNd-stimulatory factors. Also at this operative stage, bothTk of a drop, most frequently developed beyond the dormancy stage. 4. Experiments with three prospective diapause germs of equal age in a drop, having the same minor contamination ofNd-factors in the operation-cup, show a sevenfold increase of theNd-level as compared to controls, showing an almost similar stage-range in competence and stage-specificity in sensibility. The increase of theNd-stimulation rate in relation with the number of test germs in the drop corresponds to a morphogenetic crowding-effect, the influence of the test germs on each other being called "interference". 5. Experiments with LYS-media which was slightly contaminated with bacteria and then refiltered had a 42%Nd-level in the 1Tk experiments. This significant difference from the 10% control level can be related to a sensitivity increasing factor or to aNd-stimulating "masterkey"-effect of the bacterial products in addition to the yolk factors already present. The 65%Nd-level in the 2Tk experiments clearly indicates "interference". 6. Interference was also shown inexperiments with artificially prospective non-diapause germ of the same age. Test-germs isolated immediately after the HCl-treatment had a very highNd-rate (78% in 2Tk and 100% in 3Tk). Interference also occurred in germ-bands that were removed at a later stage, i.e.in ovo prior to organogenesis (2Tk, 60%Nd-rate instead of 1Tk, 20%Nd-rate). Consequently, interference may be caused by a factor promoting an overall increase of tissue activity. 7. Critical evaluation of the methods and discussion of the resultsin vitro permit the following conclusions as to the developmentin ovo. Up to organogenesis, the continuous development of the germ-anlage (non-dormancy), is stimulated by an extra-embryonic system. In particular, the yolk-cell system contains factors, without which all germ-bands (including nondiapausal eggs fromBombyx and presumably other insect eggs) can only pass autonomously through the already programmed and/or determined gastrulation and segmentation. In pre-determined egg-diapause, an extra-embryonic dormancy regulatory mechanism inactivates thisNd-action system. Maternal hormonal depots may influence the continuous stimulation of cell division and the information-transfer for the programming of organogenesis. The activation of enzyme-systems inNd-stimulation can be imitatedin vitro by bacterial contamination and by interference from test germs of the same age, and may be appear as increased sensibility of the test germs.

[Development of prospective diapause-germs (Bombyx mori L.) in vitro without dormancy : II. Medium with foreign proteins (LYS) and extraembryonic deposits from different phases of egg diapause]. / Die Entwicklung prospektiver Diapause-Keime (Bombyx mori L.) in vitro ohne Dormanz : II. Fremdeiweißmedium (LYS) mit extraembryonalem Depotmaterial aus verschiedenen Phasen der Eidiapause.

When extraembryonic egg material is placed apart from prospective diapause germ anlagen or early germ bands, it will stimulate their development without dormancy via the medium. This method of a bipartial systemin vitro (21° C) helps to analyze the regulatory mechanisms involved in the egg diapause ofBombyx mori. 1. In preliminary experiments, a culture medium free of egg extracts but containing foreign proteins (LYS) proved useful, since 100% of the test germs reached dormancy in the absence of stored egg material. Mitoses decrease and morphogenesis decelerates until the stage of the fully segmented germ-band is reached, which means the end of the prediapausal period. 2. When eggs were opened they developedin vitro without egg diapause. One may assume that the access of free oxygen activates some regulatory mechanisms permitting development without dormancy (nondormancy=Nd). In addition, the separate deposit of chorion, serosa and yolk cells (CDS Depot) will in any case prevent dormancy. Thus, the factors responsible for egg diapause must be sought in the extraembryonic egg system. A direct contact between the extraembryonic action-system and the embryonic reactionsystem is not a prerequisite. TheLYS medium without deposits offers sufficient oxygen to the test germ. Therefore the prospective diapause germ possesses a tendency to dormancy, according to its reaction norm. The potency to stimulateNd was tested with various depotmaterials (C, D, S, CS, DS, CDS) removed from eggs during prediapause (21° C), diapause (3° C) and post-diapause (returned to 21° C for 4 days). Each material produced a specificNd rate.In vitro, the test germs can progress in their organogenesis optimally to the stage of a small larva. The means of a collective effect in development are determined and related to one of the nine possibledegrees of organogenesis. 3. In comparison to serosa and chorion, yolk material has the highest mean both inNd rate (68 %,n=219) and degree of organogenesis. Surprisingly, cell-free chorionic material prevents dormancy development in 55% (n=296). As compared to theD Depot, the combination ofDS elicits a higherNd rate (79%,n=234), which is only surpassed by theCDS combination (100%,n=76). In comparison toS Depot(44%,n=294) theNd rate of aCS Depot reaches only 37% (n=161), presumably due to a restriction of the experiments to young material only. Probes, tested separately according to germ anlage or germ band, showed that there was no influence of the operational age of the test germ on theNd rate. 4. However, theNd-stimulating potency ofC, D, S, CS andDS depends on the operational age of the donator egg. Yolk material starts out having a highNd effect, decreasing with pre-diapausal age and staying relatively high in diapausal age. Similar changes are observed in the combination of yolk and serosa. TheNd rate of chorion starts low, increases steeply with the operational age and remains rather uniform. TheNd rate of serosa increases steeply in the stage critical to the beginning of egg diapause (dish-like germ anlage), decreases after pre-diapause and increases again after the minimal period for diapause (3 months at 3° C). HigherNd rates are observed whenS, D, andDS Depot were returned to 21° C for 4 days.D Depot has the maximal potency favouring organogenesis at the dish-like germ anlage stage. 5. The following subjects are discussed: the results of Chino (1957, 1958) on glycogen metabolismin ovo, the findings of Okada (1971) on the development of de-chorionized eggs under paraffin oil and our ownin ovo observations on the ultrastructural changes in the chorion, the mitotic activity before and after diapause and the distribution of glycogen in germ, yolk cells and serosa. These facts can be utilized to formulate a concept of the physiological phases of egg diapausein ovo: Egg diapause begins during a critical stage of the germ anlage with a reaction between serosa and chorionic material, which reduces the rate of oxygen consumption. Under these conditions, glycogen is metabolized into sorbitol and glyoerol. The physiologicalprophase of egg diapause is terminated, when the germ-band reaches dormancy. Diapause begins (e.g. at 3° C) with themesophase, during which the metabolism of glycogen continues decreasingly. Now glycogen is found only in the germ.Metaphase may begin with the re-uptake of oxygen, which starts the re-synthesis of glycogen from sorbitol and glycerol via oxydation and phosphorylation. However, the exposure to cold (3° C) will inhibit mitosis in the dormant germ band. In thetelophase of egg diapause, after terminated resynthesis, the dormant germ can remain in quiescence. When exposed to 21° C during the embryonic post-diapause period, it consumes the stored glycogen. If the high temperature starts prematurely during the mesophase, no embryo will hatch. However, when high temperatures set in during the metaphase, glycogen resynthesis and glycogen-breakdown in embryogenesis will compete and thus the hatching rate will be low. 6. Assuming that in the depot experimentsin vitro at 21° C and with free access of oxygen, glycogen metabolism can be considered one parameter of theNd rate, a satisfactory explanation of our experiments can be offered. With aCDS Depot,Nd stimulatory mechanism will always work satisfactorily, assuming a considerable resynthesis of glycogen of previously cold-exposed depot material.Nd rate ofD Depot will first follow the glycogen parameterin ovo; when removed from diapause, it may be capable of the resynthesis of glycogen. This will also explain the correspondingNd rates of theDS Depots. There is no correlation between theNd rates ofC Depots and the glycogen parameterin ovo. S Depots acquire a dependency on the glycogen parameter, which is independent of exposure to high temperature and oxygenin vitro. Further investigations on the glycogen metabolim of the depot materialin vitro are necessary to clarify these hypotheses. 7. The observations on the physiological phases inBombyx may also hold true for egg diapause of other insects. Various experiments with eggs of other strains ofBombyx with different reaction norms may substantiate our present conclusions. The enzymatic basis of the regulatory mechanisms with special regard to chorion should receive further clarification.

[Development of prospective diapause-germs (Bombyx mori L.)in vitro without dormancy : I. Extracts of diapause or nondiapause eggs in defined stages with or without a yolk depot]. / Die Entwicklung prospektiver Diapause-Keime (Bombyx mori L.)in vitro ohne Dormanz : I. Extrakte aus Diapause- oder Nondiapause-Eiern bestimmten Stadiums ohne oder mit Dotterdepot.

1. Naked germ-anlagen and germ-bands during segmentation from univoltine strains ofBombyx mori L. were used without HCl-treatment for culturein vitro. With improved methods germs could develop without their prospective diapause until they had finished organogenesis and if kept in hanging drops nearly one month. 5 culture mediums contained defined extracts: (1) eggs with prospective diapause germs, (2) eggs with nondiapausing germs (3) eggs with germbands in eudiapause, (4) eggs with embryos after diapause and (5) eggs with nondiapausing embryos. In every case explanted germs developed at least to fully segmented germbands (stage of dormancyin ovo) but often further. 2. In culture medium (1), (2), (4), (5) 64-73% of germ-anlagen develop to embryos with articulate limbs and open backs, but in medium (3) only 55%. On the contrary germbands develop to this stage and to shortened embryos in medium (1) only to 38%, in (2) to 5%, in (3) to 12% but in (4) to 75% and in (5) even to 84%. In culture medium (3) there occur abnormal differentiations. 3. A store of yolk or of yolk and serosa separately from the tested germ allows development in many more cases without dormancy and to small larvae with closed backs, with eye pigmentation and muscle contractions. Embryos with appendage formation grow to 100% in culture medium (4) with depositum. Therefore it is the most efficient medium for experiments with fragmented parts of a germ. 4. The different rate of development without dormancy with germbands in culture medium (1) to (5) points to their competence for determining factors of diapause. One day diapausing germbands inovo arein vitro with yolk stores able to develop further. But 2 days old germs in dormancy also with stores cannot any further develop. The germband is definitively determined to eudiapause. 5. Also the material in the stores can develop. Yolkcells may aggregate and will be enclosed by the serosa. Together they form bubbles which pulsate and yolk inside. This material could include factors for diapause, but also for basal metabolism and for syntheses of cell differentiation. Possibly it couldclean also the drop. 6. The results invite discussions aboutin vitro methods, about the ability of the explanted germ for formation and about factors determining dormancy or organogenesis. The selected hypothesis requires experiments with a medium without egg-extract but with stores from extraembryonal egg-materials and it needs investigations of regulation in cell differentiation.

Morphogenetic movements and segmentation of posterior egg fragmentsin vitro : Calliphora erythrocephala meig., diptera.

Eggs ofCalliphora were operated (I) at the undifferentiated blastoderm stage, (II) after blastoderm differentiation and in the stages showing border furrows and the beginnings of gastrulation, and (III) during forward extension of the posterior germ band region along the dorsal egg surface. The operation consisted of cutting off up to 1/3 of the egg anteriorly and removing the posterior egg fragment from the vitelline membrane for culture in a hanging drop. As control experiments for the "explanted" egg fragments, anteriorly pricked eggs and corresponding posterior fragments not removed from the vitelline membrane were cultured in hanging drops. Different technical prerequisites for the sterile culture ofCalliphora embryo fragmentsin vitro were tested. Development of the living embryo fragments was followed, and representative embryos fixed and microscopically examined for comparison with normally developing embryos. In explants I cell formation in the blastoderm did not reach completion and the blastoderm nuclei did not divide further. Slight contractions of parts of the preblastoderm did occur, but no morphogenetic movements. -In control fragments cell formation mostly proceeded further, but gastrulation was abortive except in large fragments. In explants II, irregular immigration of mesoderm cells was possible in fragments explanted before mesoderm groove initiation, but the explants did not develop further. Invagination of mesoderm through a groove, followed by segmentation of the germ band and some differentiation, was possible only in explants operated after initiation of groove formation. The germ band did not extend antero-dorsally, but became folded laterally, these folds foreshadowing the intersegmental boundaries formed later. Organogenesis and histological differentiation were in many respects abnormal. - In contrast, mesoderm invagination and initiation of germ band extension were possiblein control fragments operated before mesoderm groove initiation, although abnormalities in gastrulation were common. Explants III, despite cessation of germ band extension after explantation, often underwent further development resulting in histologically differentiated partial embryos, showing various abnormalities in organogenesis. The dorso-lateral furrows often persisted until segmentation and may correspond to primitive intersegmental furrows. The hind-gut and part of the posterior mid-gut-rudiment often evaginated during germ band contraction. Only when avitelline membrane was present did the germ band continue to extend antero-dorsally and, although this extension was rarely complete, further development with normal organogenesis and histological differentiation took place.Of the three germ layers, the ectoderm most closely approached normal differentiation in both explants and control fragments, with differentiation of nerve ganglia, tracheae, salivary glands, and a hind-gut epithelium with Malpighian tubules. Differentiation of the mesoderm in the explants was confined to muscle fibres situated near the ganglia. The endoderm remained undifferentiated in explants but formed a mid-gut epithelium in some control fragments, when splanchnic mesoderm was also present.The bearings of these results on morphogenetic problems in the insect egg is discussed.