Two new species of the Peronospora belbahrii species complex, Pe. choii sp. nov. and Pe. salviae-pratensis sp. nov., and a new host for Pe. salviae-officinalis.

The downy mildew species parasitic to Mentheae are of particular interest, as this tribe of Lamiaceae contains a variety of important medicinal plants and culinary herbs. Over the past two decades, two pathogens, Peronospora belbahrii and Pe. salviae-officinalis have spread globally, impacting basil and common sage production, respectively. In the original circumscription of Pe. belbahrii, the downy mildew of coleus (Plectranthus scutellarioides) was ascribed to this species in the broader sense, but subtle differences in morphological and molecular phylogenetic analyses using two genes suggested that this pathogen would potentially need to be assigned to a species of its own. In the present study, Peronospora species causing downy mildew on members of the Mentheae, including clary sage (Salvia sclarea), meadow sage (S. pratensis), basil (Ocimum basilicum), ground ivy (Glechoma hederacea) and coleus (Plectranthus scutellarioides) were studied using light microscopy and molecular phylogenetic analyses based on six loci (ITS rDNA, cox1, cox2, ef1a, hsp90 and ß-tubulin) to clarify the species boundaries in the Pe. belbahrii species complex. The downy mildew on Salvia pratensis is shown to be distinct from Pe. salviae-officinalis and closely related to Pe. glechomae, and is herein described as a new species, Pe. salviae-pratensis. The downy mildew on S. sclarea was found to be caused by Pe. salviae-officinalis. This is of phytopathological importance, because meadow sage thus does not play a role as inoculum source for common sage in the natural habitat of the former in Europe and Asia, while clary sage probably does. The multi-gene phylogeny revealed that the causal agent of downy mildew on coleus is distinct from Pe. belbahrii on basil, and is herein described as a new taxon, Pe. choii.

Systemic infection with Candida albicans in breast tumor bearing mice: Cytokines dysregulation and induction of regulatory T cells.

OBJECTIVE: The effect of candidemia on immunologic parameters in breast tumor bearing patients is not well studied. Here, we hypothesised that candidemia in the tumor background may change the outcome of immunologic parameters and tumor condition. METHOD: Mice were divided into four groups, including normal, tumor, Candida infected (only Candidiasis) and tumor/Candidiasis groups. Tumor changes were recorded daily after tumor transplantation and induction of candidemia. Splenocytes of mice were harvested, cultured, and stimulated with PHA; afterwards, IL-4, IL-10, IFN-γ, TNF-α and TGF-ß cytokines were assessed using ELISA kits. We also evaluated the population of CD4+CD25+Foxp3+ regulatory T cells in the tumor infiltrated and splenocytes. RESULTS: The results showed that infection with C. albicans decreased the IFN-γ/IL-4 ratio in tumor/candidiasis and candidiasis groups versus their non-infected controls. IL-10, TGF-ß and TNF-α levels increased in the candidiasis group. In addition, Candidemia led to an increase in the Treg population in tumor microenvironment and splenocytes of experimental groups compared with non-infected controls. Finally, candidemia increased tumor growth of tumor/Candidiasis group compared with the tumor group. CONCLUSION: It seems that systemic infection with C. albicans could not only induce regulatory T cells but also result in dysregulation of cytokine network and thereby facilitate tumor growth.

Diagnostic Accuracy of Computed Tomography in Detecting Bone Invasion due to Squamous Cell Carcinoma of Buccal Mucosa.

OBJECTIVE: To determine the diagnostic accuracy of contrast enhanced computed tomography (CECT) of head and neck in detecting adjacent bone invasion due to squamous cell carcinoma of buccal mucosa using histopathology as gold standard. STUDY DESIGN: Cross sectional / descriptive study. PLACE AND DURATION OF STUDY: Department of Radiology, Dow Medical College, Civil Hospital, Karachi, from September 2015 to October 2016. METHODOLOGY: A total of 116 patients clinically suspected of oral cavity squamous cell carcinoma (OCSCC) involving buccal mucosa was included. After taking proper history, all these patients referred to Radiology Department for contrast enhanced contiguous thin slice sections of 3-5 mm the primary site of the lesion prior to surgery. Reconstruction of all images done by using bone algorithm. The diagnostic accuracy of CECT was analysed against the gold standard of histopathology. Statistical analyses were carried out by using SPSS 21. Mean ±SD were calculated for quantitative variable like age. For qualitative variables, frequencies and percentages were calculated. Sensitivity, specificity, positive predictive value, negative predictive value, and diagnostic accuracy were computed for CECT, taking histopathology as gold standard. RESULTS: Eighty-three male (71%) and 33 female (28%) patients. Mean age was 46.66±10.77 years. Mean duration of sign and symptoms was 3.47 ±1.54 months. Sensitivity, specificity, PPV, NPV and accuracy were 69.2%, 78.4%, and 80.4%, 66.7% and 73.3%, respectively. CONCLUSION: CECT is a reliable technique for detecting adjacent bone invasion due to squamous cell carcinoma of buccal mucosa by using 3-5 mm thin-section CT reconstructed with bone algorithm.

Schwannoma of Nasal Inferior Turbinate in Young Male: A Rare Occurrence.

A12-year old boy presented to the ENT Department of Abbasi Shaheed Hospital, Karachi, with a one year history of progressively increasing unilateral right sided nasal obstruction. He denied any other symptoms like that of rhinorrhea, epistaxis, facial pain, headache and any history of nasal trauma. After routine clinical examination, CT scan nose/PNS with contrast was advised and subsequently biopsy was done that revealed the mass to be as schwannoma. This was then removed via a lateral rhinotomy approach and the postoperative specimen again revealed the same. The boy was followed for 1-year and no recurrence is seen. He is still in the follow-up phase.

First Report of Plant-Parasitic Nematode Meloidoderita salina in the Netherlands.

After the description of the root-parasitic nematode Meloidoderita salina from a tidal salt marsh in France (1), an additional sampling was carried out to search for the presence of this unusual nematode in a tidal salt marsh area close to Sint-Annaland, Zeeland Province, the Netherlands. In August and October 2012, a total of 25 soil and root samples were collected from the halophytic plants Atriplex portulacoides L. (so far the only known host for this nematode species), A. littoralis L., A. prostrata Boucher ex DC., Limonium vulgare Mill., Salicornia europaea L., Aster tripolium L., and Plantago maritima L. All these halophytes grow in a cohesive muddy soil type within the salt marsh, except A. littoralis and A. prostrata, which grow in the litter tidal zones on the edges of this area. Nematodes from roots and soil were extracted by centrifugal flotation (2) and Oostenbrink's cotton-wool filter methods (4), respectively. Additionally, roots were used for direct observation of females and young cystoids with a dissecting microscope. Finally, all stages were compared morphologically with available type material (1). Root and soil samples demonstrated that only nematodes isolated from A. portulacoides, A. littoralis, and A. prostrata contained all life stages of the genus Meloidoderita, while on the roots of L. vulgare, S. europaea, A. tripolium, and P. maritima, no Meloidoderita was observed. The soil samples included males, cystoids, and second-stage juveniles (J2) in low densities (<20 nematodes/100 ml), while swollen females and young cystoids were observed on root samples. All stages (n = 10 per life stage) fit morphologically with the recently described M. salina. Females were swollen with an oval to pear shaped body with a small posterior protuberance, irregular and twisted neck, oval and backwardly sloping stylet knobs, a prominent secretory-excretory (S-E) pore with cuticular lobes, and a swollen uterus with a thick hyaline wall. Males were without stylet, strongly sclerotized S-E duct, and tail tapering to rounded terminus ending in one or two ventrally terminal mucron. J2s had a well-developed stylet and rounded knobs set off from shaft and conical tail slightly curved ventrally and tapering to a finely pointed terminus with a finger-like projection. Cystoids showing the unique sub-cuticular hexagonal beaded pattern (1). J2s were also used for molecular analysis. DNA was extracted by incubating individual J2s in a lysis buffer as described in (3). Two primer combinations were used to amplify the small subunit ribosomal DNA (SSU rDNA) from a 100-times-diluted crude lysate (two overlapping fragments, [3]). The resulting (nearly) full-length SSU-rDNA sequences (GenBank KF751617 and KF751618) showed >99% identity with M. salina sequences from nematodes collected in the aforementioned tidal salt marsh in France (FJ969126 and FJ969127). To our knowledge, this is the first report of M. salina in the Netherlands. Moreover, this is the first record of M. salina parasitizing A. littoralis and A. prostrata. Although these Atriplex species are used for human consumption, the effect of M. salina on the host is unknown so far. References: (1) S. Ashrafi et al. Zookeys 249:1, 2012. (2) W. A. Coolen. Pages 317-329 in: Root-knot nematodes (Meloidogyne species). Systematics, biology and control. Academic Press, New York, 1979. (3) Holterman et al. Mol. Biol. Evol. 23:1792, 2006. (4) M. Oostenbrink. Pages 85-102 in: Nematology. University of North Carolina Press, Chapel Hill, 1960.

Fibrous dysplasia of mandible.

An elderly male presented to the ENT, Outpatient Department at Abbasi Shaheed Hospital (ASH) with a mass on the left side of the face and lower jaw for 8 - 9 years. It was of a tennis ball size. No treatment was sought by him during those years. A diagnosis of benign fibro-osseous lesion of the mandible was made. Later the histopathology proved the same. he first occurrence in a male gender and involvement of the mandible is uncommon.

An unusual foreign body in trachea.

We report a very unusual case of foreign body (FB) in the tracheo-bronchial tract of a male child, who presented to us in the Emergency Department at our tertiary care centre with the complaint of FB in throat, cough bouts and choking sensation with pain and difficulty in breathing for whom an emergency rigid endoscopy was carried out after appropriate investigations.

Varicocelectomy: semen parameters and protamine deficiency.

Different methods have been used to evaluate the beneficial effect of varicocelectomy; these include semen parameters and pregnancy rate. Because of high biological variability of semen parameters, sperm functional tests have been considered as an efficient end point in assessment of fertility. Therefore, the aim of this study was to evaluate the effect of varicocelectomy on semen parameters and sperm protamine deficiency in 192 patients. The results of the present study show that all the three semen parameters and percentage of sperms with normal protamine content have improved post-surgery. The cumulative pregnancy rate was 34.6%. Comparing the results of the semen parameters and protamine content between patients whose partner became pregnant to those who did not benefit from varicocelectomy before and 6 months after surgery, show that patients may benefit from varicocelectomy that had higher initial semen density and better sperm morphology prior to surgery. Detailed analyses of sperm morphology, along with aforementioned results reveal that the factors which account for pregnancy difference are: (i) improvement in early events of spermatogenesis, possibly during spermatocytogenesis and reduction division; and (ii) late spermiogenesis events. Thus, it can be suggested that patients with low initial sperm count may benefit more from assisted reproductive techniques or varicocelectomy followed by assisted reproduction.

Wilting of date palm branches by Fusarium oxysporum in south of Iran and its control managements with soil solarization method.

Wilting of some branches in nurseries and orchards of date palm were studied in south of Iran including Ahvaz and Abadan cities in 2005-2006 years. Different infected plants were visited and samples showing symptoms including wilting or death of branches collected from various areas and transferred to laboratory. Samples were cultured in common media (PDA) and different fungi were studied and identified. The most frequently isolated pathogen was Fusarium oxysporum which caused wilting of some branches of date palm seedling or trees in studied areas. Results showed that the disease caused main losses where date palm cuttings were cultured in infected soils, previously cropped to susceptible plants. Since chemical control was not managed the disease, soil disinfestations by soil solarization method was carried in Ahvaz as the warmer climate in studied areas to control the pathogen. Application of this method reduced population density of the pathogen from 1800 CFU -g/soil to 600 after 5 week. This method was simple, effective, non negative side and economic which can be used in nearly warm areas.

Smokeless tobacco-induced lamellar body abnormalities.

OBJECTIVES: To compare the morphological changes and quantitative distribution of lamellar bodies (Lb) (membrane coating granules) in the hamster cheek pouch epithelium with smokeless tobacco (ST). MATERIALS AND METHODS: Archives of experimental material from previously published studies [S. Ashrafi, A. Das, R. Worawongvasu, B. Mehdinejad and J. Waterhouse (1992) Scanning Microscopy6: 183] were utilized. Animals in experimental group received most ST (snuff) in their right pouch, 5 days weekly, for 24 months, while no snuff was given to control group. After 24 months, the epithelial tissues were processed for electron microscopic study. Volume densities of Lb were assessed by morphometry. MAIN OUTCOME MEASURES: Densities of Lb in the two groups, experimental vs control. RESULTS: In the control, Lb extruded their contents into the intercellular spaces of upper granular layers and in between the last granular cell layers and keratin layers to form a permeability barrier. Conversely, in the smokeless tobacco-treated epithelium, the majority of the Lb that were formed remained inside and accumulated within the granular cells, without extruding their contents into the intercellular spaces to form a lipid compound permeability barrier. CONCLUSIONS: Commercial alkaline ST may have contributed to the abnormal accumulation of Lb in the granular cell layer and affected the extrusion process of Lb to form an incomplete permeability barrier in the oral epithelium.

Mitochondrial volume densities in the smokeless tobacco-treated hamster cheek pouch epithelium.

OBJECTIVES: To compare the morphological changes and quantitative distribution of mitochondria in the hamster cheek pouch (HCP) epithelium treated with smokeless tobacco (ST). MATERIALS AND METHODS: Archives of experimental material from previously published studies (Ashrafi et al., 1992) were utilized. Animals in experimental group received moist ST (snuff) in their right pouch, 5 days weekly for 24 months, while no snuff was given to control group. After 24 months, the epithelial tissues were processed for electron microscopy study. Volume densities of mitochondria were assessed by morphometry. MAIN OUTCOME MEASURES: Mitochondrial volume densities in the two groups, experimental vs control. RESULTS: In both control and experimental groups mitochondria were concentrated between the nucleus and basal cell plasma membrane. A decrease in the mean mitochondrial volume density (Vvmit) was observed from the basal layer to the more superficial layers in both groups. The experimental HCP displayed more mitochondria than control, and the granular epithelial cell layer in experimental group showed significantly a higher mean Vvmit than the control group (P = 0.03). It was concluded that greater numbers of mitochondria were retained in ST-treated granular cells of the hyperplastic epithelia than in the normal epithelium.

Zinc deficiency produces time-related ultrastructural changes in rat cheek epithelium.

The subject of this study was whether the ultrastructural changes in cheek epithelium of zinc-deficient rats are time related. Weanling male Sprague Dawley rats were fed a zinc-deficient diet containing 0.4 ppm zinc (ZD) ad libitum and controls were pair-fed zinc adequate diet containing 40 ppm zinc. After 9, 18, and 27 days of zinc deficiency, specimens from cheek epithelium of both groups were processed for transmission electron microscopy. Partial conversion of the orthokeratinized cheek epithelium to parakeratinized was seen as early 9 days. An electron-lucent band surrounding the nucleus was observed in ZD cells. Mitochondria, tonofilaments, keratohyalin granules and ribosomes seemed to be increased with the increase in time of zinc deficiency. There was a thickening of the stratum corneum as well as hyperplasia and widening of the intercellular spaces of the spinous layer cells. Retention of a few membrane coating granules (MCGs) in the parakeratinized layer was seen after 9 days. Parakeratinization was further increased after 18 days of zinc deficiency, and the number of MCG profiles also increased. The epithelium was fully parakeratinized following 27 days of zinc deficiency, and the number of MCG profiles was increased. It was concluded that zinc deficiency affected cell proliferation and differentiation of the epithelium as early as 9 days, and caused a delay in loss of nuclei and MCGs in parakeratinized cells.

Scanning electron microscopy of cyclosporine-induced gingival overgrowth.

Overgrown human gingival specimens were examined histologically and by scanning electron microscopy (SEM) to study structural changes caused by cyclosporine. The biopsy specimens were from organ transplant recipients receiving cyclosporine to suppress the rejection of the transplanted organ. The epithelium of the overgrown gingiva was thickened, acanthotic and parakeratotic. Retepegs were anastomosing and extending into connective tissue. The SEM examination of the outer surface of the attached gingival showed loss of cellular attachments and cells were exfoliating. The normal honeycomb structure formed by interconnecting microvilli surrounding the pits was distorted. Outer gingival cell surface showed numerous round, ovoid and dome-like structures instead of parallel, reticular or fingerprint-like microridges. It was concluded that cyclosporine not only caused hyperplasia but also changed the structure of the outer epithelial cell surface.

In situ hybridization and monoclonal antibody analysis of plasma membrane Ca-pump mRNA and protein in submandibular glands of rabbit, rat and man.

The degree of supersaturation of saliva with calcium (Ca) is related to the mineral phase of enamel in erupted teeth, the incidence of caries, and the formation of calculus. The mechanisms for regulating salivary Ca concentration are therefore of relevance to dentistry. Sections of rabbit, rat and human submandibular gland (SMG) were processed for immuno-histochemistry with a specific anti-plasma membrane Ca-pump antibody, 5F10. Western blots confirm that the molecular weight of the proteins identified by our antibody (135 kDa) is consistent with an appropriate molecular weight for PMCA antigen (135-150 kDa). Tissue sections were also processed for in situ hybridization to study the distribution of the PMCA mRNA isoforms. In mammals, the PMCA1 gene is reported to code for a PMCA protein with a role in maintaining the intracellular Ca levels in both epithelial and non-epithelial cells. Other genes including the PMCA2 and PMCA4 genes may code for PMCA proteins specific to Ca transporting tissues. Our studies demonstrate cytoplasmic labeling of PMCA mRNA with hPMCA-1 and hPMCA-4 specific cDNA probes in humans, and rPMCA-1 and rPMCA-2 specific oligonucleotide probes in rats. Labeling of PMCA protein and all mRNA isoforms was found in the cytoplasm of the interlobular and intralobular ducts (except for intercalated ducts). The demonstrated presence of PMCA in SMGs of rabbit, rat, and man, may suggest a role for PMCA in the regulation of intracellular Ca and in a mechanism for regulating and maintaining the high concentration of Ca in salvia.

Time-related changes induced by zinc-deficient diet in the concentration of rat cheek epithelial membrane-coating granules.

Weanling male Sprague-Dawley rats were fed a diet containing 0.4 parts/10(6) zinc and controls were fed an identical diet supplemented with 40 parts/10(6) zinc. After 9, 18 and 27 days of zinc deficiency, specimens were excised from cheek epithelium and processed for transmission electron microscopy to study the concentration of membrane-coating granules (MCG). Their concentration was increased in the granular-cell layers of the zinc-deficient epithelium and became significantly greater after 18 and 27 days than 9 days of deficiency. MCGs appeared in the parakeratinized layers of zinc-deficient epithelium and their concentration became significantly greater after 27 days in comparison with 9 and 18 days of deficiency. Thus the intracellular retention of MCGs was increased in the granular and parakeratinized layers with the increase in time of zinc deficiency.

Migration of epithelium during phenytoin-dependent gingival overgrowth in mice.

A small cavity was made in the mesiopalatal area of the maxillary first molar adjacent to the gingiva. Mice were maintained on 40 mg/kg phenytoin (or on diluent for control) by daily intraperitoneal injections. After 9 weeks, light microscopic observations revealed that in experimental mice, epithelial cells migrated towards the cavity and covered it. In controls, epithelial cell migration towards the cavity did not occur. For scanning electron microscopic (SEM) studies, specimens were fixed in 4% glutaraldehyde in 0.1 M phosphate buffer, pH 7.2, for 2 hours, dehydrated, critical point dried and coated with gold. The surface of the outer gingival epithelium of experimental and of control mice showed a honeycomb arrangement of the microridges suggesting their keratinized nature. Epithelial cells lining the cavity showed well marked macroridges along their borders. Parallel microridges were observed on the upper surface of these cells suggesting that they were non-keratinized. It was concluded that the migrating epithelial cells, that covered the cavity during phenytoin-dependent gingival overgrowth, were of the non-keratinized type.

The effect of lasers on dental hard tissues.

There is a quest to find a method to remove diseased and healthy dental hard tissues without the negative stimuli associated with dental handpieces. Today, lasers are being considered as a potential replacement. This report evaluates effects of three lasers on dentin and pulpal tissues. The Er:YAG laser appears to have a lesser thermal effect.

Expression of plasma membrane Ca++ pump epitopes parallels the progression of enamel and dentin mineralization in rat incisor.

We investigated the expression of Ca++ pump epitopes during enamel and dentin mineralization in the rat incisor. Secretory and maturation ameloblasts were studied as well as odontoblasts, using a monoclonal antibody (5F10) against human erythrocyte plasma membrane Ca++, Mg(++)-ATPase. A progressive increase in staining intensity in ameloblasts and the odontoblasts was observed beginning with the onset of mineralization. The mainly membrane-related labeling of ameloblasts showed variable intensity depending on the stage of enamel formation, whereas that of the odontoblasts showed even intensity during continued dentinogenesis. Staining of papillary cells was evident only during enamel maturation. Western blot analysis of freeze-dried ameloblasts was also used to determine the molecular weight of the Ca++ pump epitopes as well as the distribution and relative concentration of epitopes at each stage. An immunoreactive band of MW 140 KD and lower molecular weight bands that are more intense in late than in early maturation were demonstrated. Our studies suggest that the expression of plasma membrane Ca++ pump parallels the progression of mineralization in rat incisor enamel and dentin.

A light, transmission and scanning electron microscope study of snuff-treated hamster cheek pouch epithelium.

The effects of smokeless tobacco (snuff) on hamster cheek mucosa were studied by light microscopy, transmission (TEM) and scanning electron microscopy (SEM). Two grams of commercially available smokeless tobacco were placed into the blind end of the right cheek pouch of each experimental animal, once a day and five days a week for 24 months. The control animals did not receive smokeless tobacco. After 24 months treatment with smokeless tobacco, hamster cheek mucosal epithelium lost its translucency and had become whitish in color. By light microscopy hyperorthokeratosis, prominent granular cell layers with increased keratohyalin granules and hyperplasia were seen. At the ultrastructural level, wider intercellular spaces filled with microvilli, numerous shorter desmosomes, many thin tonofilament bundles, increased number of mitochondria, membrane coating granules and keratohyalin granules were seen in snuff-treated epithelium. The changes in the surface of the epithelium as seen by SEM were the development of an irregular arrangement of the microridges and the disappearance of the normal honeycomb pattern. The microridges were irregular, widened and surrounded the irregular elongated pits. Some smooth areas without microridges and pits were also seen. The long-term histological, TEM and SEM changes induced by smokeless tobacco treatment of the epithelium are well correlated with each other and were similar to those reported in human leukoplakia without dyskeratosis. They imply changes of pathological response resulting from topically applied snuff.

Cytochemical localization of calcium and Ca2+,Mg2(+)-adenosine triphosphatase in colchicine-altered rat incisor ameloblasts.

Colchicine is known to affect secretory, transport, and degradative functions of ameloblasts. The effects of colchicine on membrane-associated calcium and Ca2+,Mg2(+)-ATPase in secretory and maturation ameloblasts were investigated cytochemically. The pyroantimonate (PPA) method was used for localizing calcium and a modified Wachstein-Meisel medium was used to localize Ca2+,Mg2(+)-ATPase. Sections representing secretory and early maturation stages were examined by transmission electron microscopy. Morphological changes induced by colchicine included dislocated organelles and other well-established reactions to such anti-microtubule drugs. Calcium pyroantimonate (Ca-PA) deposits in most ameloblast types were markedly reduced, with the greater reduction occurring in those cells more severely altered morphologically. However, the cell membranes of both control and experimental smooth-ended maturation ameloblasts were essentially devoid of Ca-PA. The normal distribution and intensity of Ca2+,Mg2(+)-ATPase was not affected by colchicine. Because the observed reduction of membrane-associated calcium is apparently not mediated by Ca2+,Mg2(+)-ATPase in this case, other aspects of the calcium regulating system of ameloblasts are apparently targeted by colchicine.