Colicin Typing as an Epidemiological Tool in the Investigation of Outbreaks of Shigella sonnei

Shigella sonnei has become the most frequently reported cause of shigellosis in the United States. Since Shigella subgroup D has no other serotypes, colicin production has been used as a basis for differentiating and identifying epidemiologically related strains. The results of colicin typing 115 cultures of S. sonnei from eight outbreaks of shigellosis occurring in widely separated regions of the United States support the usefulness of this technique. In each outbreak, the cultures were either of the same colicin type or were uniformly untypable. Unrelated cases yielded a variety of types. Definitions of the relative frequencies and geographic distributions of the various strains of S. sonnei in the United States await an accumulation of experience with the method.     

Comparison of the Deoxyribonucleic Acid Molecular Weights and Homologies of Plasmids Conferring Linked Resistance to Streptomycin and Sulfonamides

Bacterial strains showing linked resistance to streptomycin (Sm) and sulfonamides (Su) were chosen representing a wide taxonomic and geographical range. Their SmSu resistances were transferred to Escherichia coli K-12 and then plasmid deoxyribonucleic acid (DNA) was isolated by ethidium bromide CsCl centrifugation. The plasmid DNA was examined by electron microscopy and analyzed by sedimentation through 5 to 20% neutral sucrose gradients. Plasmid DNA from strains having transmissible SmSu resistance consisted of two or three molecular species, one of which had a molecular mass of about 5.7 Mdal (10(6) daltons), the others varying between 20 to 60 Mdal. By using transformation or F' mobilization, we isolated the SmSu-resistance determinant from any fellow resident plasmids in each strain and again isolated the plasmid DNA. Cosedimentation of each of these with a differently labeled reference plasmid DNA (R300B) showed 9 out of 12 of the plasmids to have a molecular mass not significantly different from the reference (5.7 Mdal); two others were 6.3 and 9.2 Mdal, but PB165 consisted of three plasmids of 7.4, 14.7, and 21.4 Mdal. Three separate isolations of the SmSu determinant from PB165 gave the same three plasmids, which we conclude may be monomer, dimer, and trimer, respectively. DNA-DNA hybridizations at 75 C demonstrated 80 to 93% homology between reference R300B DNA and each isolated SmSu plasmid DNA, except for the 9.2-Mdal plasmid which had 45% homology and PB165 which had 35%. All the SmSu plasmids were present as multiple copies (about 10) per chromosome. The conjugative plasmid of R300 (present as 1.3 copies per chromosome) has been shown to have negligible effect on the number of copies of its accompanying SmSu plasmid R300B. We conclude that the SmSu plasmids are closely related and probably have a common evolutionary origin.     

Plasmid RP4, with Escherichia coli DNA inserted in vitro, mediates chromosomal transfer.

The wide host-range plasmid RP4 is generally poor at mobilizing chromosomal markers. Insertion in vitro of Escherichia coli chromosomal DNA into its HindIII site (recognized by loss of its kanamycin resistance marker) or its EcoRI site generated RP4-primes that efficiently mobilized chromosomal markers. These RP4-primes behaved analogously to F-primes, each had a particular transfer origin and direction of marker mobilization, and this was abolished by a recA allele in the donor. Insertion sequences on the RP4-primes do not appear to be necessary for mobilization, it seems very unlikely that each of the RP4-primes generated contained an insertion sequence. The construction and use of such plasmids should facilitate inter- and intrageneric genetic analyses and manipulations.     

Zur Feinstruktur einer elektrischen Synapse

Summary Free-running, naked axons (diameter 2000 to 7000 Å) can be found in the lumen of the pineal organ. Their axoplasm contains microtubules, mitochondria as well as synaptic (diameter 350 to 450 Å) and granulated vesicles (diameter 500 to 1500 Å). In Pleurodeles waltlii, the axons in the pineal lumen form synapses on the free, apical surface of the pineal ependyma which is supplied with microvilli. In addition to usual cytoplasmic elements the innervated ependymal cells contain myeloid bodies and accumulations of glycogen granules. Without forming synapses these axons pass by and occasionally contact the inner and/or outer segments of the pinealocytes. The synapses found on the pineal ependymal cells furnish evidence of a neuronal control of these glial elements.The nerve fibers of the pineal lumen are being compared with known CSF contacting axons; they resemble one another in their ultrastructure and synaptic connections. Therefore and since in amphibians the pineal lumen communicates with the 3rd ventricle, the axons of the pineal lumen are considered to represent CSF contacting axons and to belong to the so-called CSF contacting axon system of the brain.In addition, the pineal CSF contacting axons are being compared with the following nerve fibers and terminals found in the pineal tissue: 1) axons containing large, granulated vesicles (diameter 1300 to 1500 Å) and terminating on the dendrites of nerve cells situated among the basal processes of the pinealocytes; 2) the synaptic ribbons-containing pinealocyte processes forming likewise synapses on the nerve cells; 3) the neurohormonal, synaptic semidesmosomes of pinealocytic processes on the lamina basalis separating the connective tissue spaces of the pia mater from the proper nervous tissue of the pineal organ; 4) the perivasal, autonomic nerve fibers of the pial septa. Though granulated vesicles of various diameters are present in all these terminals the greatest morphological similarity is found between the pineal CSF contacting axons and those nerve fibers containing large, granulated vesicles and forming axo-dendritic synapses on the pineal nerve cells. A similar nature and origin of both axons are suggested.

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Zusammenfassung Im Lumen des Pinealorgans können frei verlaufende, nackte Axone (Durchmesser 2000–7000 Å) beobachtet werden. Ihr Axoplasma enthält Mikrotubuli, Mitochondrien, synaptische (Durchmesser 350–450 Å) und granulierte Vesikel (Durchmesser 500–1500 Å). Bei Pleurodeles waltlii bilden die im Lumen des Pinealorgans verlaufenden Axone Synapsen auf der freien, apikalen Oberfläche der pinealen Ependymzellen. In den innervierten Ependymzellen kommen neben sonstigen Zytoplasmabestandteilen Myeloidkörper und Anhäufungen von Glykogengranula vor. Die Axone verlaufen am Innen- und Außenglied der Pinealozyten vorbei, können diese berühren, bilden aber dort keine Synapsen. Die auf den pinealen Ependymzellen nachgewiesenen Synapsen beweisen eine neuronale Kontrolle dieser Gliaelemente.Die Nervenfasern des pinealen Lumens wurden mit bekannten Liquorkontaktaxonen verglichen. Sie ähneln einander in ihrer Ultrastruktur und ihren synaptischen Verbindungen. Aus diesem Grunde und da bei den Amphibien das pineale Lumen mit dem 3. Ventrikel kommuniziert, werden die Axone des pinealen Lumens als Liquorkontaktaxone und als Glied des sogenannten Liquorkontakt-Axonsystems des Gehirns angesehen.Ferner wurden die pinealen Liquorkontaktaxone mit folgenden Nervenfasern und Endigungen verglichen, die im pinealen Gewebe vorkommen: 1) Axone, die große, granulierte Vesikel (Durchmesser 1300–1500 Å) enthalten und an den Dendriten von Nervenzellen endigen, welche zwischen den basalen Fortsätzen der Pinealozyten liegen; 2) Pinealozytenfortsätze, die synaptische Bänder enthalten und ebenfalls an diesen Neuronen Synapsen bilden; 3) die neurohormonalen, synaptischen Semidesmosomen von Pinealozytenfortsätzen an der Lamina basalis, die die bindegewebigen Räume der Pia mater vom eigentlichen Nervengewebe des Pinealorgans begrenzt: 4) die perivasalen, autonomen Nervenfasern der pialen Septen. Obwohl granulierte Vesikel verschiedener Durchmesser in allen diesen Terminalen vorhanden sind, stellten wir die größte, morphologische Ähnlichkeit zwischen den pinealen Liquorkontaktaxonen und denjenigen Nervenfasern fest, die große, granulierte Vesikel aufweisen und an den pinealen Neuronen axo-dendritische Synapsen bilden. Eine ähnliche Natur und Herkunft beider Axone werden angenommen.

     

Initiation and regulation of oöcyte growth by the brain and corpora allata of the cockroach,Nauphoeta cinerea

Oöcytes of Nauphoeta cinerea begin to grow about 2 days after adult ecdysis. This growth can be totally prevented by surgical ablation of the corpora allata (CA) before day 2 and restored in a dose-related manner by injection of juvenile hormone (JH). Ovarian maturation can also be prevented in some animals by brain extirpation within a day of adult ecdysis but proceeds normally in others, indicating that the brain acts to promote oöcyte lengthening very soon after emergence. Wounding (mouthparts removal) blocks ovarian maturation in a similar percentage of animals, indicating that the ‘wounding effect’ is mediated by the brain. However, those wounded animals which do develop oöcytes do not reach normal values by autopsy on day 7, suggesting a temporary inhibition of the CA. Because the effects of head ligation, which effectively removes the brain as well as the CA, can be reversed by only a little more JH than the effects of allatectomy, we conclude that the primary rôle of the brain is to control the CA. We suggest that both stimulatory neurohumoral material and inhibitory nervous transmission may be utilized for this control.     

The release of attack and escape behavior by vibratory stimuli in a wandering spider (Cupiennim salei keys.)

Summary The wandering spiderCupiennius salei responds to vibration of the substrate either with predatory behavior (approach) or with a startle reaction or escape behavior (withdrawal) (Fig. 3). The effects of different parameters of the signal in releasing this behavior were studied by applying various artificial stimuli to a spider standing on a vibrating platform with one or more legs. Receptors sensitive to substrate vibration and the trichobothria, which respond to airborne vibration, together determine the response. Spiders without trichobothria: The type of response to vertical vibrations isfrequency-dependent (Fig. 4a), with predatory reactions predominant at low frequencies (3–4 Hz), and withdrawal reactions at high frequencies (350–460 Hz). Whereas approach is most likely to occur at an intermediate, frequency-dependentamplitude, the probability of withdrawal increases continuously with increasing amplitude (Fig. 6). With sine wave stimuli the lowest threshold amplitude for approach is 9 m peak-to-peak (550 Hz, range tested 1–550 Hz) whereas that for withdrawal is 17 m (800 Hz, range tested 1–800 Hz). The threshold for approach is lower by 6–8 dB whenband-limited noise is used, and the probability of an approach response increases as the bandwidth is expanded. The threshold curve for withdrawal, however, is the same in all cases (Fig. 4b and 5). The spider is capable of both frequency and amplitude discrimination.The metatarsal and pretarsal slit sense organs contribute to these responses as is shown by increased thresholds following their destruction (Fig- 7). Intact animals, with functional trichobothria as well as slit sense organs: They have lower thresholds for withdrawal (by ca. 10 dB; Fig. 9) and shorter reaction times than do spiders without trichobothria. Unlike animals without trichobothria the amplitude thresholds of intact animals to bandlimited noise are ca. 7.5 dB lower than those to sine wave stimuli. The approach threshold is the same as that of spiders without trichobothria. According to direct observation the trichobothria are deflected by airborne sound generated by the substrate motion; the deflection angle increases with both amplitude and frequency of substrate vibration (Fig. 10).There is acentral nervous interaction between the signals from the trichobothria and the slit sense organs with the following basic properties: when both of the two receptor systems receive either a prey-like stimulus or a stimulus eliciting withdrawal their effects add, but when the trichobothria receive stimuli unlike prey they inhibit the approach reaction that would otherwise be triggered by substrate vibration.     

Structure-activity relationship in the enzymatic hydrolysis of dipeptide aryl amides by dipeptidyl peptidase IV

Quantitative structure activity analysis of the substrate types Ala-Ala-AR and Ala-Pro-AR containing different substituents in the aryl ring showed that the rate-limiting step in the hydrolysis of the alanine substrates by dipeptidyl peptidase IV occurs in th acylation reaction (kcat approximately k2). Probably, the tetrahedral intermediate of the acylation process has a real life time. The positive q-value of the Hammett-equation in k'cat suggests that the N-atom of the arylamide is charged more negatively in the transition state TI not equal to than in the original state TI. The analysis of the quantitative conformation activity relationship (QCAR) gives information on the steric situation in the tetrahedral intermediate of the acylation step near the transition state. The rate limiting step in the hydrolysis of the substrates of the proline type occurs in the deacylation reaction.     

In vitro biosynthesis, core glycosylation and membrane integration of opsin

A membrane-integrated , core-glycosylated form of bovine opsin was synthesized in vitro when bovine retina mRNA was translated in a wheat germ cell-free system supplemented with dog pancreas microsomal vesicles; glycosylation and integration of opsin into membranes were coupled to translation. Proteolysis with themolysin was used to probe the orientation of opsin within the dog pancreas microsomal membrane, and to compare it with that of opsin in rod cell disk membranes isolated from bovine retina. Intact microsomal or disk vesicles were required for production of discrete, membrane-associated thermolysin fragments of opsin; no discrete opsin fragments were detected when membranes were incubated with thermolysin in the presence of the nonionic detergent, Triton X-100. The major opsin fragments produced by themosylin treatment of intact microsomal vesicles resembled those from disk vesicles in their size, oligosaccharide content, and order of appearance. In each case, the first cleavage of opsin took place at the COOH-terminus, generating a glycosylated fragment, O’, which was only slightly smaller than intact opsin. Both the microsomal and disk membrane forms of O’ were next cleaved internally; glycosylated fragments of similar sizes in both cases were detected which were derived from the NH(2)-terminal portion of O’. Several smaller NH(2)-terminal fragments of opsin were detected only in thermolysin-treated microsomal membranes, and not in disk membranes. The data suggest that the topology of opsin integrated into dog pancreas microsomal vesicles is similar to that in rod cell disk vesicles, although not identical. In each case, the glycosylated NH(2)-terminal region of opsin is located within the lumen of the vesicle, while discrete COOH-terminal and internal segments of opsin apparently emerge at the outer, cytoplasmic face of the membrane. Thus, opsin in the heterologous microsomal membrane, like its counterpart in the native disk membrane, may cross the bilayer at least three times. The internal domain of the polypeptide that emerges at the outer membrane surface is apparently more highly exposed in the case of opsin in microsomal membranes, evidenced by the additional internal thermolysin cleavage sites detected.