Rapid Detection of Salmonella Microcolonies by Fluorescent Antibody

A microcolony fluorescent-antibody (FA) procedure for detecting salmonellae was compared to the usual direct FA procedure on 304 environmental, food, and feed samples. The microcolony FA test detected all of the specimens found positive by culture, whereas the direct FA missed 3.1% of them. Both FA tests revealed stained organisms in some of the culturally negative specimens. The microcolony FA test has several advantages over the direct FA test: ease of examining the smears, elimination of the fluorescent background material, and increased sensitivity.     

The Laboratory Diagnosis of Whooping Cough by Fluorescent Antibody and by Culture Methods

Attempts were made to hasten the identification of Bordetella pertussis by using the fluorescent antibody (FA) technique and to improve the isolation rate by culture methods. First, a comparison was made between the number of identifications of the organism by growth on Bordet-Gengou medium containing penicillin G and by staining smears of pharyngeal exudate with FA. Of 100 specimens, 29 were positive on culture and 46 were positive by the FA method. Five of the 46 were false positives. Second, 170 specimens were inoculated in quadruplicate on Bordet-Gengou medium containing either penicillin G or methicillin, and on Lacey medium containing either penicillin G or methicillin. In order there were 39, 41, 56 and 57 isolations. The isolation of Bord. pertussis on methicillin-containing media was easier than on penicillin-containing media. When culture and FA methods were compared, the most reliable was found to be the inoculation of specimens on Lacey medium.     

Fluctuating feather asymmetry in relation to corticosterone levels is sex-dependent in Eurasian treecreeper (Certhia familiaris) nestlings

Fluctuating asymmetry (FA) has been widely used as a stress-related phenotypic marker of developmental instability. However, previous studies relating FA to various stressful conditions have produced inconsistent results and we still lack quantitative individual-level evidence that high FA is related to stress in wild vertebrate species. We studied how baseline plasma levels of corticosterone predicted FA of wing and tail feathers in free-living Eurasian treecreeper (Certhia familiaris) nestlings. We found a sex-specific association between corticosterone levels and FA: high corticosterone levels were related to an increased FA in male but not in female nestlings. These results suggest that in treecreepers, FA may correlate with individual stress hormone levels, male developmental trajectory being potentially more sensitive to stress than that of the female.     

Basement membrane material and tigroid background in a fine needle aspirate of clear cell adenocarcinoma of the cervix. A case report

BACKGROUND: Although cytologic findings of clear cell adenocarcinoma of the female genital tract have been reported sporadically, the background on the smear has received little attention. CASE: A 16-year-old female had a large, necrotic mass in the cervix. As cervical brushing cytology and a punch biopsy of the mass could not make a definitive diagnosis, fine needle aspiration cytology was performed from the mass. The smears revealed loose, three-dimensional clusters and sheet arrangements of atypical cells. Dispersed atypical cells were also seen. The atypical cells were large and had abundant, weakly stained cytoplasm and round or oval nuclei with large nucleoli. There were a few tumor cells with clear cytoplasm and distinct cell borders in Papanicolaou-stained smears. The background in Diff-Quik-stained smears revealed a tigroid background and basement membrane material. CONCLUSION: The malignant tumor, revealing both a tigroid background and basement membrane material, seems to have been clear cell adenocarcinoma. Both features are diagnostic clues to clear cell adenocarcinoma in the female genital tract. Our case indicates that aspiration cytology is also an effective method of diagnosing a cervical tumor when the tumor is polypoid and the surface is extensively necrotic.     

Efficacy of laboratory tests for the detection of enterotoxigenic Clostridium perfringens.

Nineteen Clostridium perfringens strains with positive erythemal and ligated intestinal loop reactions, and 22 strains with negative reactions, originating from food-poisoning cases, were tested comparatively using the fluorescent antibody (FA), reversed passive hemagglutination (RPHA), and immunodiffusion (ID) tests. All the biologically positive strains were detected by the three immunological tests used. The FA test detected five additional strains among the biologically negative group which did not react in RPHA or ID tests. Sporulating culture supernatant fluids, after 13 to 17h of growth, were satisfactory for testing for the presence of enterotoxin by the RPHA and ID tests. The FA test was used on cell smears.     

A Pilot Study of Short-Duration Sputum Pretreatment Procedures for Optimizing Smear Microscopy for Tuberculosis

Background

Direct sputum smear microscopy for tuberculosis (TB) lacks sensitivity for the detection of acid fast bacilli. Sputum pretreatment procedures may enhance sensitivity. We did a pilot study to compare the diagnostic accuracy and incremental yield of two short-duration (<1 hour) sputum pretreatment procedures to optimize direct smears among patients with suspected TB at a referral hospital in India.

Methodology/Findings

Blinded laboratory comparison of bleach and universal sediment processing (USP) pretreated centrifuged auramine smears to direct Ziehl-Neelsen (ZN) and direct auramine smears and to solid (Loweinstein-Jensen (LJ)) and liquid (BACTEC 460) culture. 178 pulmonary and extrapulmonary TB suspects were prospectively recruited during a one year period. Thirty six (20.2%) were positive by either solid or liquid culture. Direct ZN smear detected 22 of 36 cases and direct auramine smears detected 26 of 36 cases. Bleach and USP centrifugation detected 24 cases each, providing no incremental yield beyond direct smears. When compared to combined culture, pretreated smears were not more sensitive than direct smears (66.6% vs 61.1 (ZN) or 72.2 (auramine)), and were not more specific (92.3% vs 93.0 (ZN) or 97.2 (auramine).

Conclusions/Significance

Short duration sputum pretreatment with bleach and USP centrifugation did not increase yield as compared to direct sputum smears. Further work is needed to confirm this in a larger study and also determine if longer duration pre-treatment might be effective in optimizing smear microscopy for TB.     

The effect of fluctuating asymmetry and leaf nutrients on gall abundance and survivorship

Fluctuating asymmetry (FA) is defined as small and random deviations from perfect symmetry in otherwise bilaterally symmetrical traits. Usually, plants under stress have a higher nutritional quality and are preferred by insect herbivores. Here, we examined the relationship among leaf FA of Bauhinia brevipes (Fabaceae), nutrients and the occurrence of Schizomyia macrocapillata (Cecidomyiidae: Diptera). We evaluated whether: (i) FA was related to S. macropillata abundance and survivorship; (ii) FA correlated with the levels of foliar nutrients (macro- and micronutrients, and organic carbon); and (iii) S. macropillata abundance was related to leaf nutrients. The results showed a positive relationship between FA and gall abundance. S. macrocapillata survivorship was five times higher in B. brevipes individuals with high FA levels, but this difference was not significant. The influence of nutrients in B. brevipes FA was negligible, except for foliar nitrogen, which was 7.3% higher in plants with low FA levels. The abundance of S. macrocapillata was negatively related to nitrogen levels, but gall survivorship was not affected by this nutrient. We found evidence for a causal relationship between FA and gall occurrence, since S. macropillata gall oviposition occurs in young undeveloped leaves, thus mature leaves show an increase in FA. Nonetheless, in this system, neither FA nor nitrogen influenced gall survivorship.     

Fine needle aspiration cytology of breast fibroadenoma. A cytohistologic correlation study of 405 cases.

OBJECTIVE: To evaluate the cytohistologic correlation of breast fibroadenoma (FA) in order to assess the value of cytology in the conservative management of this lesion. STUDY DESIGN: A retrospective analysis of all aspirates diagnosed as FA or fibroadenomatous lesion (n = 1,549) for which a histologic follow-up study was available (n = 362). Forty-three aspirates, including 14 nonrepresentative smears, from histologically proven FAs with a different cytologic report were also included in the study. RESULTS: Cytohistologic agreement was present in 287 of the 362 cytodiagnoses. Lack of correlation was observed in 75 cases. Most diagnostic errors accumulated in the older patient group. The sensitivity of the cytologic diagnosis of FA was 86.9% (90.8% excluding nonrepresentative cases), with a positive predictive value of 79.3%. In 43 cases a histologic diagnosis of FA was given after previous erroneous (n = 29) or nonrepresentative cytodiagnoses (n = 14). The specificity of the cytologic diagnosis of FA reached 93.8%, with a negative predictive value of 96.3% (97.5% excluding nonrepresentative cases). Regarding malignancy, five tumors were diagnosed as FA and were malignant. No false positive diagnoses of malignancy were given, but nine aspirates were included in the category "suspicious for carcinoma." CONCLUSION: FA of the breast remains a diagnostic challenge for the cytopathologist. A considerable amount of benign breast lesions can mimic FA on cytology, and such diagnostic categories as "fibroadenomatous lesion" or "consistent with FA" are associated with low diagnostic accuracy. While the cytologic requisites for entering a program of conservative management of FA are established, it seems that strict diagnostic criteria should be applied even at the expense of diminishing sensitivity.     

Experience of applying plant and animal fluctuating asymmetry in assessment of environmental quality in terrestrial ecosystems: Results of 20-year studies of wildlife and anthropogenically transformed territories

The value of fluctuating asymmetry (FA) of trees and small mammals is considered as an indicator of developmental instability. The urban territories with recreation zones, sites with increased radiation background, and the areas affected by mining facilities have been analyzed as anthropogenically transformed areas. The following tree and small mammalian species are the objects of the study: Siberian silver birch (Betula platyphylla), spreading birch (B. divaricata), dwarf birch (B. exilis), shrub alder (Duschekia fruticosa), basket willow (Salix viminalis), northern red-backed vole (Clethrionomys rutilus), tundra vole (Microtus oeconomus), Laxmann’s shrew (Sorex caecutiens), and tundra shrew (S. tundrensis). In total, 60000 leaves have been measured and 3500 rodent sculls have been examined. The issues of selection of bioindicator species and the factors distorting the results of environmental quality assessment according to developmental instability are discussed. A statistically significant increase in FA of plants is recorded in urban territories as compared with the wildlife biotopes. A positive correlation of FA value with transport load, concentration of heavy metals in soil, and atmospheric air pollution is observed. The proximity to a roadway and the state of road pavement influence the FA value. The main source of pollution in the administrative center (Yakutsk) is motor vehicles; correspondingly, large streets and crossroads with a high transport load are the most polluted sites. As for the industrial center (city of Mirnyi), pollution spreads from the areas affected by mining to the residential area. The main pollution source in a small administrative center (city of Aldan) is the federal highway. It is recommended to concurrently use the FA characteristics of plant and animals in the area affected by the mining industry. Characteristic of this industry is withdrawal of large areas of natural landscapes; typically, transformed plots alternate weakly affected plots. The specific features of the effects of diamond, gold, and coalmining industries, as well as oil and gas field development, are discussed, which should be taken into account in sampling for bioindication. The problem of bioindication in the areas completely devoid of soil and plant cover can be solved by biotesting via assessment of bunching onion seed germination rate and the rate of mitotic aberrations. An increase in the FA level in trees and small mammals is observed in the areas transformed by the mining industry displaying a chronically increased radiation background and oil-polluted as a result of technogenic accidents. The use of FA index makes it possible to estimate the environmental quality and to compare the state of territories located under different natural and climatic conditions and influenced by multicomponent anthropogenic impacts differing in their strength and genesis, which interferes with their comparison.     

Antigenic similarity among Anabaena azollae separated from different species of Azolla

Direct fluorescent antibody (FA) reaction results of 5 FAs against symbiotic Anabaena azollae indicated that all the A. azollae freshly separated from 32 specimens of Azolla collected worldwide (belonging to 6 different species) shared identical and highly specific antigens. None of these FAs exhibited cross-reaction with any of the free-living blue-green algae tested. FA absorption results confirmed these results and also indicate the existence of cross-reactive antigens between Azolla leaves and the surfaces of A. azollae. Antibodies made against free-living A. azollae did not cross-react with any of the symbiotic A. azollae indicating either: (i) these isolates are not true isolates, or (ii) their antigenic properties were altered during isolation and culturing. Such possibilities and their implications are discussed.     

Excreted urine,bladder urine,and the delay of sexual maturation in female house mice

A series of experiments tested effects of excreted and bladder urine of group- and singly-caged female house mice (Mus musculus) on sexual maturation of young female mice caged alone or in groups of eight. Puberty was determined by vaginal smears. A maturation-delaying pherompne was present in bladder urine of all female mice and excreted urine of group-caged females, but not in excreted urine from singly-caged females. Test mice treated with bladder urine homogenized with urethras from singly-caged females matured at the same age as controls. Apparently the urethras or associated glands of singly-caged females produce a substance which deactivates the maturation-delaying pheromone contained in bladder urine.     

Direct Immunofluorescence Test for the Diagnosis of Genital Herpesvirus Infections

Herpetic lesions of the genitalia may be confused clinically with other ulcerative, genital lesions. Direct immunofluorescence (FA) provides a rapid method of diagnosis, and the utility of this method for the diagnosis of genital ulcers was examined. One hundred and ten patients with genital lesions were examined by darkfield for syphilis and by FA and culture for herpes simplex virus (HSV) infections. Satisfactory samples were obtained from 102 patients, of which 81 were clinically suspected cases of HSV. Acetone-fixed slides of scrapings of ulcerative lesions were stained with conjugated antiserum prepared in rabbits against HSV type 2. HSV was isolated from 73% of specimens of suspected herpetic lesions, and 77% of these specimens were positive by FA. Nine percent were positive by FA only and these were not thought to represent false positives. Five percent were positive by culture only. A comparison of clinical diagnoses with laboratory findings revealed that 4% of the cases were misdiagnosed when only the clinical evaluation was considered. The data suggest that the inclusion of a diagnostic FA test for HSV along with the darkfield examination may be useful for differentiating the etiological agents of ulcerative, genital lesions.     

Mastopathic fibroadenoma of the breast: a pitfall of aspiration cytology

OBJECTIVE: To assess the correlation between cytological diagnoses and histological subtypes of fibroadenoma (FA) and to clarify the cytological features of a specific group of FA displaying variable features similar to fibrocystic disease (mastopathic type, MFA), and to evaluate the significance of this subtype in cytological diagnosis. METHODS: A review of 141 cases of histologically proven FA was performed. We re-classified them into four subtypes according to Kinoshita's criteria [Jpn J Breast Cancer6 (1991) 377] and further selected 92 cases for which both fine needle aspiration (FNA) smears and histological specimens were available. Among them, 18 cases of MFA and their cytological smears were selected for the present study. RESULTS: There was significant correlation between MFA and cytological diagnosis of 'indeterminate' or 'suspicious for malignancy' (P < 0.01). Although no false-positive diagnosis was experienced in our series, 56% of the MFAs (10/18) had cytological diagnoses of indeterminate or were included in the category 'suspicious for malignancy'. Smears from MFA revealed high cellularity (9/18 smears had more than 10 epithelial clusters each composed of more than 50 cells), presence of cellular discohesiveness (13/18, 72.2%), but only mild nuclear atypia (5/18, 27.8%). Anisonucleosis was present in fewer than half the cases and no apparent condensed chromatin was identified. CONCLUSION: We highlight the significance of subclassification of MFA in aspiration cytology of breast. MFA had a significantly higher chance of falling into the 'suspicious for malignancy' or 'indeterminate' diagnostic category in aspiration cytology. It might be a diagnostic challenge for cytopathologist to identify this subtype of FA in FNA smears.     

Cycles of mating behaviour, oestrogen and progesterone in the thick-tailed bushbaby (Galago crassicaudatus crassicaudatus) under laboratory conditions

Vaginal smears and blood samples were taken throughout the reproductive cycle of female Galago c. crassicaudatus. Blood plasma was assayed for oestradiol and progesterone, and vaginal smears were initially classified dioestrus or vaginal oestrus. During vaginal oestrus the females were tested daily for sexual receptivity by being placed with a male. Those days on which the male achieved intromission were reclassified as behavioural oestrus. During dioestrus the females were tested weekly with males. Female receptivity increased and then declined across a 6-day period of behavioural oestrus during the 44-day cycle. Fully cornified smears were characteristic of the period of maximal receptivity and oestradiol secretion. The luteal phase lasted 24 days with a plasma progesterone peak midway through dioestrus.     

Evaluation of Fluorescein Isothiocyanate-labeled Whole Antiserum in the Immunofluorescent Identification of Microorganisms

Portions of a whole antiserum to Histoplasma capsulatum were reacted with amounts of fluorescein isothiocyanate (FITC) that ranged from 50 to 400 mug/mg of protein. Portions of the globulin from the same antiserum were reacted with amounts of FITC that ranged from 12.5 to 50 mug of FITC per mg of protein. The globulin conjugates (postlabeled globulins), the whole serum conjugates, and the globulins from the whole serum conjugates (prelabeled globulins) were compared with respect to their fluorescein-protein (F:P) ratios and fluorescent-antibody (FA) activities. The whole serum sample treated with 50 mug of FITC per mg of protein was least reactive in FA tests, and its globulin had the lowest F:P. All other conjugates had globulins with F:P ratios that were considered to be adequate for high FA activity. It was found, however, that the prelabeled globulins were considerably less reactive than the postlabeled globulins or the whole serum conjugates. A larger amount of brightly staining reagent per milliliter of original serum could be obtained from labeled whole serum than from postlabeled globulin. Lissamine-rhodamine conjugated to bovine serum albumin (LRBSA) was evaluated as a counterstain to be used in conjunction with FITC-labeled whole antisera. The counterstain was effective in masking nonspecific FITC fluorescence in Formalin-fixed tissues and in culture smears of fungi. Masking was incomplete in culture smears of a bacterium and in blood smears containing a protozoan.     

Comparison of Fluorescent Antibody with Cultural Technique for Isolation of Enteropathogenic Escherichia coli from Swine

In an investigation of hogs as possible reservoirs of human strains of enteropathogenic Escherichia coli (EEC), 92 six-month-old grain- and garbage-fed hogs were examined on the farm and again at the packing plant. Of the 331 specimens obtained by swabbing the rectum, cecum, and edible meat carcass of these hogs, 125 were presumptively positive for EEC when screened by the fluorescent-antibody (FA) technique. These “presumptive positive” specimens then underwent extensive bacteriological examination and complete serological typing. The FA technique proved to be an easier, simpler, and more economical procedure than culture when a large number of specimens were examined for possible EEC serogroups. It was found especially valuable for identification of multiple serogroups of EEC within a single specimen. It also appeared to be more sensitive than cultural examination, since results were not dependent on the presence of large numbers of organisms in the specimen, or even on their viability. However, the FA technique was found to be less specific than culture because of cross-reactivity with antigenically related Enterobacteriaceae when fluorescein-labeled antisera were used. Therefore, any specimen found positive on FA examination should be considered as presumptive positive until confirmed by bacteriological examination and complete serological study.     

Fatty Acid Composition of Muscle,Adipose Tissue and Liver from Muskoxen (Ovibos moschatus) Living in West Greenland

Information about lipid content and fatty acid (FA) composition of muskoxen (Ovibos moschatos) edible tissues is very limited in comparison to other meat sources. Thus, this work aims to present the first in-depth characterization of the FA profile of meat, subcutaneous adipose tissue and liver of muskoxen living in West Greenland. Furthermore, we aim to evaluate the effect of sex in the FA composition of these edible tissues. Samples from muscle (Longissimus dorsi), subcutaneous adipose tissue and liver were collected from female and male muskoxen, which were delivered at the butchery in Kangerlussuaq (West Greenland) during the winter hunting season. The lipid content of muscle, adipose tissue and liver averaged 284, 846 and 173 mg/g of dry tissue, respectively. This large lipid contents confirms that in late winter, when forage availability is scarce, muskoxen from West Greenland still have high fat reserves, demonstrating that they are well adapted to seasonal feed restriction. A detailed characterization of FA and dimethylacetal composition of muskoxen muscle, subcutaneous adipose tissue and liver showed that there are little differences on FA composition between sexes. Nevertheless, the 18:1cis-9 was the most abundant FA in muscle and adipose tissue, reaching 43% of total FA in muscle. The high content of 18:1cis-9 suggests that it can be selectively stored in muskoxen tissues. Regarding the nutritional composition of muskoxen edible tissues, they are not a good source of polyunsaturated FA; however, they may contribute to a higher fat intake. Information about the FA composition of muskoxen meat and liver is scarce, so this work can contribute to the characterization of the nutritional fat properties of muskoxen edible tissues and can be also useful to update food composition databases.     

The role of immunoperoxidase staining in diagnostic cytology

A survey was made of the immune staining characteristics of 60 malignant neoplasms. Cytologically positive smears from each case were tested against a panel of six antibodies (alpha-antichymotrypsin, carcinoembryonic antigen, cytokeratin, desmin, vimentin and S-100 protein). The smears were decolorized and stained with polyclonal sera using the standard avidin-biotin immunoperoxidase procedure. In selected cases, the application of Diatex compound for partition of smears was necessary to obtain optimal results. Most staining reactions reflected the histogenesis of the neoplasms. However, more than one of five reactions was nonconclusive due to background staining, scanty cellularity or poor cytoplasmic preservation; furthermore, the 238 reactions scored as positive or negative included 33 unexpected positives and 15 unexpected negatives. In a series of 20 additional cases, selective immunoperoxidase staining was used in an attempt to solve specific diagnostic problems; the results in 13 of 15 cases with conclusive staining agreed with the cytologic impression. It is concluded that standard immunoperoxidase techniques can contribute to the solution of certain diagnostic problems in cytology; however, the results should be interpreted with caution and with full knowledge of the limitations of the technique.     

Facial attractiveness signals different aspects of “quality” in women and men

We explored the relationships between facial attractiveness and several variables thought to be related to genotypic and phenotypic quality in humans (namely fluctuating asymmetry (FA), body mass index (BMI), health, age). To help resolve some controversy around previous studies, we used consistent measurement and statistical methods and relatively large samples of both female (n=94) and male (n=95) subjects (to be evaluated and measured), and female (n=226) and male (n=153) viewers (to rate attractiveness). We measured the asymmetry of 22 traits from three trait families (eight facial, nine body and five fingerprint traits) and constructed composite asymmetry indices of traits showing significant repeatability. Facial attractiveness was negatively related to an overall asymmetry index in both females and males, with almost identical slopes. Female facial attractiveness was best predicted by BMI and past health problems, whereas male facial attractiveness was best predicted by the socioeconomic status (SES) of their rearing environment. Composite FA indices accounted for a small (<4%) but usually significant percentage of the variation in facial attractiveness in both sexes, when factors related to asymmetry were controlled statistically. We conclude that, although facial attractiveness is negatively related to developmental instability (as measured by asymmetry), attractiveness also signals different aspects of “quality” in the two sexes, independent of FA.     

Altered Fecal Microbiota Composition Associated with Food Allergy in Infants

Increasing evidence suggests that perturbations in the intestinal microbiota composition of infants are implicated in the pathogenesis of food allergy (FA), while the actual structure and composition of the intestinal microbiota in human beings with FA remain unclear. Microbial diversity and composition were analyzed with parallel barcoded 454 pyrosequencing targeting the 16S rRNA gene hypervariable V1-V3 regions in the feces of 34 infants with FA (17 IgE mediated and 17 non-IgE mediated) and 45 healthy controls. Here, we showed that several key FA-associated bacterial phylotypes, but not the overall microbiota diversity, significantly changed in infancy fecal microbiota with FA and were associated with the development of FA. The proportion of abundant Bacteroidetes, Proteobacteria, and Actinobacteria phyla were significantly reduced, while the Firmicutes phylum was highly enriched in the FA group (P < 0.05). Abundant Clostridiaceae 1 organisms were prevalent in infants with FA at the family level (P = 0.016). FA-enriched phylotypes negatively correlated with interleukin-10, for example, the genera Enterococcus and Staphylococcus. Despite profound interindividual variability, levels of 20 predominant genera were significantly different between the FA and healthy control groups (P < 0.05). Infants with IgE-mediated FA had increased levels of Clostridium sensu stricto and Anaerobacter and decreased levels of Bacteroides and Clostridium XVIII (P < 0.05). A positive correlation was observed between Clostridium sensu stricto and serum-specific IgE (R = 0.655, P < 0.001). The specific microbiota signature could distinguish infants with IgE-mediated FA from non-IgE-mediated ones. Detailed microbiota analysis of a well-characterized cohort of infants with FA showed that dysbiosis of fecal microbiota with several FA-associated key phylotypes may play a pathogenic role in FA.