The association of Western flower thrips, Frankliniella occidentalis, with a near Erwinia species gut bacteria: transient or permanent?

Associations between insects and gut bacteria are ubiquitous. It is possible to make a distinction between permanent associations (called symbiosis), in which the same type of bacteria is present in more than one generation of the insect, and transient associations. Transient bacteria are ingested together with food but do not settle in the insect gut in such a way that they will be passed on to the next generation. In this study, we describe the permanent association between Western flower thrips (Frankliniella occidentalis), a polyphagous insect species that is a major pest worldwide, and one type of gut bacteria. On the basis of direct microscopic observations and counts of bacteria, it was found that thrips from the populations studied contained large numbers of bacteria in their hindgut. Bacteria were isolated from their host and grown on 10 different agar media. The number of bacteria isolated on agar media equaled the number of direct counts. All isolates had the same colony morphology. On the basis of their 16S rDNA sequence these bacteria were identified as Enterobacteriaceae, closely related to Escherichia coli. Isolates tested with API 20E biochemical tests were Erwinia species. This was the only type of bacteria found in all thrips individuals on any of the 10 different agar media. Universal primers, which would potentially pick up DNA from any bacterium present in the insect, were applied on crude DNA extracts from thrips with bacteria. We only found 16S rDNA sequences similar to those of the isolated thrips gut bacteria. The same type of bacteria was present in all life stages of the thrips and was found to persist in the thrips populations for at least 2 years (more than 50 generations).

Growth and transmission of gut bacteria in the Western flower thrips, Frankliniella occidentalis.

The Western flower thrips (Frankliniella occidentalis), a polyphagous insect with global distribution, has a permanent association with a near Erwinia species TAC bacterium in its hindgut. Since this bacterium is able to grow outside the thrips, it is a facultative symbiont that is not completely dependent on the host. In this study we address the question of how the association is maintained and how bacteria are transmitted to newly hatched thrips larvae. Bacteria are passed on to new thrips via the food source. No evidence was found for vertical transmission from mother to offspring via the egg. Gut bacteria show unlimited growth during the larval (feeding) stages, and in the second instar stage 100% of the larvae become infected with high numbers of bacteria. In the prepupal and pupal stage, the number of bacteria declines, but increases again during the adult phase. A method to rear aposymbiotic (bacteria-free) thrips is described which enables studies on the impact of bacteria on the fitness of thrips.

Protein kinase C prevents oligodendrocyte differentiation: modulation of actin cytoskeleton and cognate polarized membrane traffic.

In a previous study, we showed that activation of protein kinase C (PKC) prevents oligodendrocyte differentiation at the pro-oligodendrocyte stage. The present study was undertaken to identify downstream targets of PKC action in oligodendrocyte progenitor cells. Activation of PKC induced the predominant phosphorylation of an 80-kD protein, identified as myristoylated alanine-rich C-kinase substrate (MARCKS). Upon phosphorylation, MARCKS is translocated from the plasma membrane to the cytosol. Furthermore, PKC activation perturbed the organization of the actin cytoskeleton, causing a redistribution of actin filaments to the submembranous or cortical actin cytoskeleton. As a consequence, transport of a protein traffic marker, the vesicular stomatitis virus glycoprotein, from the trans-Golgi network to the plasma membrane becomes perturbed. The effect of disruption of the actin filament network by cytochalasin D perfectly matched the effect of PKC. These data thus favor the existence of a causal relationship between actin rearrangement and docking and/or fusion of proteins to the plasma membrane. Interestingly, neither in control cells nor in PKC-activated cells did another protein traffic marker, influenza hemagglutinin (HA), reach the cell surface. However, an eminent and specific accumulation of HA just underneath the plasma membrane became apparent upon PKC activation. Yet, this effect could not be simulated by cytochalasin D treatment. Therefore, these observations imply that although MARCKS represents a prominent PKC target site in regulating differentiation, another target involves the differential control of cognate polarized trafficking pathways, which are apparently operating in oligodendrocyte progenitor cells.

Adaptation of Pseudomonas sp. GJ1 to 2-bromoethanol caused by overexpression of an NAD-dependent aldehyde dehydrogenase with low affinity for halogenated aldehydes.

Pseudomonas sp. GJ1 is able to grow with 2-chloroethanol as the sole carbon and energy source, but not with 2-bromoethanol, which is toxic at low concentrations (1 mM). A mutant that could grow on 2-bromoethanol with a growth rate of 0.034 h-1 at concentrations up to 5 mM was isolated and designated strain GJ1M9. Measurement of enzyme activities showed that mutant and wild-type strains contained a PMS-linked alcohol dehydrogenase that was active with halogenated alcohols and that was threefold overexpressed in the mutant when grown on 2-chloroethanol, but only slightly overproduced when grown on 2-bromoethanol. Both strains also contained an NAD-dependent alcohol dehydrogenase that had no activity with halogenated alcohols. Haloacetate dehalogenase levels were similar in the wild-type and the mutant. Activities of NAD-dependent aldehyde dehydrogenase were only slightly higher in extracts of the mutant grown with 2-bromoethanol than in those of the wild-type grown with 2-chloroethanol. SDS-PAGE, however, showed that this enzyme amounted to more than 50% of the total cellular protein in extracts of the mutant from 2-bromoethanol-grown cells, which was fourfold higher than in extracts of the wild-type strain grown on 2-chloroethanol. The enzyme was purified and shown to be a tetrameric protein consisting of subunits of 55 kDa. The enzyme had low Km values for acetaldehyde and other non-halogenated aldehydes (0.8-4 microM), but much higher Km values for chloroacetaldehyde (1.7 mM) and bromoacetaldehyde (10.5 mM), while V(max) values were similar for halogenated and non-halogenated aldehydes. Cultures that were pregrown on 2-chloroethanol rapidly lost aldehyde dehydrogenase activity after addition of 2-bromoethanol and chloroamphenicol, which indicates that bromoacetaldehyde inactivates the enzyme. To achieve growth with 2-bromoethanol, the high expression of the enzyme thus appears to be necessary in order to compensate for the high Km for bromoacetaldehyde and for inactivation of the enzyme of bromoacetaldehyde.

Isolation of Typical Marine Bacteria by Dilution Culture: Growth, Maintenance, and Characteristics of Isolates under Laboratory Conditions.

Marine bacteria in Resurrection Bay near Seward, Alaska, and in the central North Sea off the Dutch coast were cultured in filtered autoclaved seawater following dilution to extinction. The populations present before dilution varied from 0.11 x 10 to 1.07 x 10 cells per liter. The mean cell volume varied between 0.042 and 0.074 mum, and the mean apparent DNA content of the cells ranged from 2.5 to 4.7 fg of DNA per cell. All three parameters were determined by high-resolution flow cytometry. All 37 strains that were obtained from very high dilutions of Resurrection Bay and North Sea samples represented facultatively oligotrophic bacteria. However, 15 of these isolates were eventually obtained from dilution cultures that could initially be cultured only on very low-nutrient media and that could initially not form visible colonies on any of the agar media tested, indicating that these cultures contained obligately oligotrophic bacteria. It was concluded that the cells in these 15 dilution cultures had adapted to growth under laboratory conditions after several months of nutrient deprivation prior to isolation. From the North Sea experiment, it was concluded that the contribution of facultative oligotrophs and eutrophs to the total population was less than 1% and that while more than half of the population behaved as obligately oligotrophic bacteria upon first cultivation in the dilution culture media, around 50% could not be cultured at all. During one of the Resurrection Bay experiments, 53% of the dilution cultures obtained from samples diluted more than 2.5 x 10 times consisted of such obligate oligotrophs. These cultures invariably harbored a small rod-shaped bacterium with a mean cell volume of 0.05 to 0.06 mum and an apparent DNA content of 1 to 1.5 fg per cell. This cell type had the dimensions of ultramicrobacteria. Isolates of these ultramicrobacterial cultures that were eventually obtained on relatively high-nutrient agar plates were, with respect to cell volume and apparent DNA content, identical to the cells in the initially obligately oligotrophic bacterial dilution culture. Determination of kinetic parameters from one of these small rod-shaped strains revealed a high specific affinity for the uptake of mixed amino acids (a degrees (A), 1,860 liters/g of cells per h), but not for glucose or alanine as the sole source of carbon and energy (a degrees (A), +/- 200 liters/g of cells per h). The ultramicrobial strains obtained are potentially a very important part of picoplankton biomass in the areas investigated.

Auricular injury as a presentation of child abuse.

Child abuse causes serious physical and emotional injury. Those abused, however, are usually unable or unwilling to gain access to agencies that might help them. The otolaryngologist, while usually not the patient's primary physician, may be in the position to detect child abuse. We describe a 3-year-old boy who sustained auricular trauma as the result of child abuse. The importance of physician recognition of an abuse case and the institution of appropriate measures are discussed.

Isolated sphenoid sinus disease.

Isolated sphenoid sinus disease is an unusual entity that is encountered infrequently. With advances in antibiotic therapy and imaging techniques, the nature of isolated sphenoid sinus disease has significantly changed. More tumors and less inflammatory disorders are being encountered. We have retrospectively reviewed the medical records of patients who had undergone sublabial transseptal sphenoidotomy for isolated sphenoid sinus disease at Allegheny General Hospital for Pittsburgh between January 1985 and July 1989. Thirteen patients were identified with isolated sphenoid sinus disease who were successfully managed with sublabial transseptal sphenoidotomy. This approach allows maximal visualization and safety with minimal morbidity.

Metabolic activation of 1,2,4-trichlorobenzene and pentachlorobenzene by rat liver microsomes: a major role for quinone metabolites.

Microsomal metabolism of 1,2,4-[14C]trichlorobenzene (1,2,4-TrCB) and [14C]pentachlorobenzene (PeCB) was studied with special emphasis on the conversion-dependent covalent binding to protein and DNA. 1,2,4-TrCB was metabolized to 2,3,6- and 2,4,5-trichlorophenol, and to a lesser extent to 2,4,6- and 2,3,5-trichlorophenol, and trichlorohydroquinone. About 10% of all metabolites became covalently bound to protein in a rather nonselective way. For 1,2,4-TrCB and PeCB a strong correlation between secondary metabolism to hydroquinones and covalent binding was established. Protein binding was completely inhibited by the addition of ascorbic acid, indicating quinone metabolites as the sole reactive species formed. Both 1,2,4-TrCB and PeCB alkylated DNA, although to a much lesser extent than protein (0.5 and 0.3% of all metabolites, respectively). Nonquinone intermediates, presumably epoxides, were responsible for a minor portion of the observed DNA binding, since complete inhibition by ascorbic acid was not reached. The differential role of cytochrome P450 both in primary and in secondary metabolism was demonstrated by the use of microsomes from rats pretreated with different inducers. Dexamethasone (DEX) microsomes (cytochrome P450IIIA1) showed the highest activity toward these chlorinated benzenes (14 nmol/mg/5 min for 1,2,4-TrCB and 36 nmol/mg/10 min for PeCB, both with regard to the formation of phenols and to the formation of protein-bound metabolites. In addition, DEX microsomes preferentially formed 2,3,6-trichlorophenol, whereas other microsomal suspensions formed 2,4,5-trichlorophenol as the major isomer. The present study clearly demonstrates the high alkylating potency of secondary quinone metabolites derived from chlorinated benzenes and poses a need for reevaluation of the role of epoxides in the observed toxicity of these compounds.

Abscess of the sphenoid sinus after transsphenoidal surgery.

A case of a bacterial abscess developing in the sphenoid sinus 2 weeks after transsphenoidal surgery is presented. Although abscesses within the sella turcica have been reported as rare complications of transsphenoidal surgery, this is the first reported case of the postoperative formation of an abscess of the sphenoid sinus. The patient sought treatment for severe headaches, nausea and vomiting, and marked temperature elevation. A computed tomographic scan demonstrated soft tissue and air within the sphenoid sinus. A regimen of stress doses of hydrocortisone and antibiotics was prescribed, and the patient underwent transsphenoidal drainage of the sphenoid sinus. The sella turcica was not involved. Anaerobic cultures were positive for Fusobacterium necrophorum.

Cloning and characterization of plasmid-encoded genes for the degradation of 1,2-dichloro-, 1,4-dichloro-, and 1,2,4-trichlorobenzene of Pseudomonas sp. strain P51.

Pseudomonas sp. strain P51 is able to use 1,2-dichlorobenzene, 1,4-dichlorobenzene, and 1,2,4-trichlorobenzene as sole carbon and energy sources. Two gene clusters involved in the degradation of these compounds were identified on a catabolic plasmid, pP51, with a size of 110 kb by using hybridization. They were further characterized by cloning in Escherichia coli, Pseudomonas putida KT2442, and Alcaligenes eutrophus JMP222. Expression studies in these organisms showed that the upper-pathway genes (tcbA and tcbB) code for the conversion of 1,2-dichlorobenzene and 1,2,4-trichlorobenzene to 3,4-dichlorocatechol and 3,4,6-trichlorocatechol, respectively, by means of a dioxygenase system and a dehydrogenase. The lower-pathway genes have the order tcbC-tcbD-tcbE and encode a catechol 1,2-dioxygenase II, a cycloisomerase II, and a hydrolase II, respectively. The combined action of these enzymes degrades 3,4-dichlorocatechol and 3,4,6-trichlorocatechol to a chloromaleylacetic acid. The release of one chlorine atom from 3,4-dichlorocatechol takes place during lactonization of 2,3-dichloromuconic acid.

Jejunal interposition for repair of stricture or fistula after laryngectomy.

Complications following total laryngectomy may include pharyngocutaneous fistula or pharyngeal stricture. Traditional techniques of repair of fistula and stricture with local or regional flaps lead to a high rate of failure. In this study, we report 18 patients treated by secondary jejunal interposition (JI) to rehabilitate swallowing following recalcitrant postlaryngectomy stricture or fistula. All patients had undergone total laryngectomy with or without partial pharyngectomy for treatment of squamous cell carcinoma of the larynx (8) or hypopharynx (10). Four were stage II; 5, stage III; and 9, stage IV. Thirteen patients (72%) regained swallowing function. Complications of secondary JI included perioperative death (2), flap loss (1), and persistent fistulas (3). Jejunal interposition may be the best modality in the rehabilitation of swallowing in patients with persistent fistula or stricture that fails to respond to traditional management.

Neutropenic enterocolitis. A new complication of head and neck cancer chemotherapy.

Neutropenic enterocolitis is a recognized complication of immunosuppression or chemotherapy for leukemia. It presents as severe abdominal pain and tenderness, fever, and diarrhea associated with granulocytopenia. Gastrointestinal symptoms associated with chemotherapy for head and neck neoplasms include nausea and emesis, but not acute abdominal distress. We present, to our knowledge, the first case of neutropenic enterocolitis in a patient receiving cisplatin and fluorouracil chemotherapy for metastatic head and neck cancer.

A new method to predict safe resection of the internal carotid artery.

A patent internal carotid artery (ICA) is essential in most patients. Management of skull base lesions often requires translocation, balloon embolization, or resection of this vessel. Preoperative tests to assess the availability of collateral flow have not been uniformly accurate. A new test that significantly increases the safety of surgical removal of the ICA is described. One hundred thirty-six patients were studied with temporary balloon occlusion (TBO) of the ICA and determination of stable xenon-enhanced computed tomography cerebral blood flow (Xe/CT CBF) measurements. Eleven patients failed TBO and were determined to be at very high risk of stroke with loss of the ICA. Ninety-six of the patients were predicted to be at minimal risk with permanent loss of the ICA by Xe/CT CBF studies. Twenty-one patients in this group had either permanent balloon occlusion (PBO) or surgical resection of the ICA with no permanent neurologic sequelae. Our studies show that the combination of preoperative TBO and Xe/CT CBF studies significantly increases the safety of ICA resection.

Reconstruction of the cervical esophagus: free jejunal transfer versus gastric pull-up.

Use of enteric grafts is a popular method for reconstruction of the cervical esophagus and hypopharynx. Free jejunal transfer (FJT) and gastric pull-up (GP) are the most popular methods used. This discussion is a retrospective review of our experience with 50 cases of free jejunal transfer and 15 cases of gastric pull-up. The graft survival rate was 94 percent (47 of 50) for free jejunal transfer and 87 percent (13 of 15) for gastric pull-up. Successful swallowing was achieved in 88 percent (44 of 50) of free jejunal transfers and 87 percent (13 of 15) of gastric pull-ups. Patients with free jejunal transfers were able to swallow and leave the hospital sooner: 10.6 versus 16.0 days and 22.3 versus 29.0 days, respectively. Fistulas occurred in 16 percent (8 of 50) of free jejunal transfers, most of which (6 of 8) healed spontaneously. Fistulas occurred in 20 percent (3 of 15) of gastric pull-ups, only one of which healed spontaneously. Stricture was the most common late complication for free jejunal transfers, 22 percent (11 of 50), whereas reflux was most common in gastric pull-ups, 20 percent (3 of 15). In patients with advanced cancer, extensive esophageal resection into the chest is often required, and gastric pull-up seems to be an easier and more direct form of reconstruction. In limited resection of the hypopharynx and esophagus, especially with proximal lesions, free jejunal transfer is simpler and avoids mediastinal dissection. This concept as well as other advantages and disadvantages of both techniques will be discussed.

Paraganglioma of the thyroid.

We present a case of paraganglioma of the thyroid. The preoperative evaluation, surgical management, and histopathology are described. Four similar cases that have been reported are summarized, and a hypothesis for the origin of a paraganglioma within the thyroid capsule is proposed.

Hypopharyngeal reconstruction: a comparison of two alternatives.

Gastric pull-up or free jejunal interposition was used for reconstruction after total laryngopharyngectomy in 31 patients. Complications and functional outcomes of the two methods are compared. Primary swallowing was achieved in 86% of patients after gastric pull-up and in 82% of patients after jejunal interposition. Patients who underwent jejunal interposition were able to swallow sooner and had a shorter hospital stay than patients who underwent gastric pull-up. Esophageal tumor recurrence after jejunal interposition was not observed. Hepatic failure occurred in two gastric pull-up patients, leading to perioperative death in one. Flap necrosis occurred in two jejunal interposition patients and one gastric pull-up patient. Two additional fistulas occurred in jejunal interposition patients as a result of microvascular complications. Stricture developed in four jejunal interposition patients, requiring revision surgery in two. Minor complications were more common in the gastric pull-up group. Long-term speech and swallowing function are compared. Our current choice of jejunal interposition or gastric pull-up for reconstruction after total laryngopharyngectomy primarily depends on the location of the tumor.

Nasopharyngeal teratoma involving the temporal bone.

Teratoma is the most common nasopharyngeal tumor in neonates. The tumor is usually limited to the oropharynx and is manifest by stridor and respiratory distress. Complete excision affects a cure. An unusual case of nasopharyngeal teratoma extending into the temporal bone, causing facial paralysis and conductive hearing loss, in addition to stridor, is presented. Stridor was relieved by transoral partial excision shortly after birth. Complete removal of the tumor by way of a subtemporal and infratemporal fossa approach was performed at 14 months of age. The defect was filled with a rectus abdominis muscle graft with microvascular anastomoses. The facial nerve was reconstructed secondarily with a sural nerve graft. The patient has no recurrence tumor and has good facial function at 30 months. This case demonstrates the first known case of facial paralysis due to nasopharyngeal teratoma. The surgical approach for tumor removal: lateral infratemporal fossa dissection, and the method of reconstruction: free rectus abdominis muscle flap with microvascular anastomoses, had so far not been described in a patient this young.

Elective resection of the internal carotid artery without reconstruction.

Curability of skull base tumors is related to the ability to achieve a complete resection. Resection of the internal carotid artery with the tumor puts the patient at risk for catastrophic cerebral injury. Autogenous vein grafting is not always technically or physiologically possible. We present eight patients with tumors eroding the cranial base who underwent safe resection or occlusion of the internal carotid artery as predicted by three tests: 1. carotid arteriography, 2. temporary balloon occlusion of the internal carotid artery, and 3. xenon computerized tomography cerebral blood flow mapping. No patient suffered permanent central nervous system injury.