Identification and epidemiological typing of Naegleria fowleri with DNA probes.

Naegleria fowleri is a small free-living amoeboflagellate found in warm water habitats worldwide. The organism is pathogenic to humans, causing fatal primary amoebic meningoencephalitis. When monitoring the environment for the presence of N. fowleri, it is important to reliably differentiate the organism from other closely related but nonpathogenic species. To this end, we have developed species-specific DNA probes for use in the rapid identification of N. fowleri from the environment. Samples were taken from the thermal springs in Bath, England, and cultured for amoebae. Of 84 isolates of thermophilic Naegleria spp., 10 were identified as N. fowleri by probe hybridization. The identity of these isolates was subsequently confirmed by their specific whole-cell DNA restriction fragment length polymorphisms (RFLPs). One DNA clone was found to contain a repeated element that detected chromosomal RFLPs that were not directly visible on agarose gels. This enabled the further differentiation of strains within geographically defined whole-cell DNA RFLP groups. N. fowleri DNA probes represent a specific and potentially rapid method for the identification of the organism soon after primary isolation from the environment.     

Survey of "do not resuscitate" orders in a district general hospital.

OBJECTIVE--To evaluate the local use of written "Do not resuscitate" orders to designate inpatients unsuitable for cardiopulmonary resuscitation in the event of cardiac arrest. DESIGN--Point prevalence questionnaire survey of inpatients'' medical and nursing records. SETTING--10 acute medical and six acute surgical wards of a district general hospital. PARTICIPANTS--Questionnaires were filled in anonymously by nurses and doctors working on the wards surveyed. MAIN OUTCOME MEASURES--Responses to questionnaire items concerning details about each patient, written orders not to resuscitate in the medical case notes and nursing records, whether prognosis had been discussed with patients'' relatives, whether a "crash call" was perceived as appropriate for each patient, and whether the "crash team" would be called in the event of arrest. RESULTS--Information was obtained on 297 (93.7%) of 317 eligible patients. Prognosis had been discussed with the relatives of 32 of 88 patients perceived by doctors as unsuitable for resuscitation. Of these 88 patients, 24 had orders not to resuscitate in their medical notes, and only eight of these had similar orders in their nursing notes. CONCLUSIONS--In the absence of guidelines on decisions about resuscitation, orders not to resuscitate are rarely included in the notes of patients for whom cardiopulmonary resuscitation is thought to be inappropriate. Elective decisions not to resuscitate are not effectively communicated to nurses. There should be more discussion of patients'' suitability for resuscitation between doctors, nurses, patients, and patients'' relatives. Suitability for resuscitation should be reviewed on every consultant ward round.     

Avoidance behavior in two sympatric planaria species: effects of conspecific and heterospecific chemical alarm cues

In many aquatic animals, predator avoidance can be stimulated by chemical cues, including those released by injured prey (alarm cues). Alarm cues of both conspecific and heterospecific origin have been identified within several fish taxa, where phylogenetic conservation of the cue-response complex is common. Turbellarian flatworms (planaria) are among the simplest animals known to respond to chemical cues released by injured conspecifics. We examined how two locally sympatric planaria species respond to conspecific and heterospecific chemical cues using macerated tissue suspensions. Brown (Girardia tigrina) and black (Dugesia dorotocephala) planaria both exhibited avoidance behavior when presented conspecific cues. Despite a significant twofold difference in body size (black > brown), stimulus prepared from a single (1×) individual of either species elicited avoidance. Increasing brown planaria cue concentration by macerating two individuals (2×) produced a significant increase in conspecific avoidance. Heterospecific stimuli produced asymmetric results. Black planaria avoided the brown planaria stimulus, but only in the higher concentration (2×) trials. Brown planaria did not consistently exhibit avoidance of the black planaria stimulus and some brown subjects approached and consumed black planarian tissues. Our results expand the demonstrated occurrence of alarm cues among planaria and suggest that avoidance behavior can be mediated by multiple environmental and intrinsic factors in freshwater Turbellaria.     

Phenylpropanoids, phenylalanine ammonia lyase and peroxidases in elicitor-challenged cassava (Manihot esculenta) suspension cells and leaves

BACKGROUND AND AIMS: Control of diseases in the key tropical staple, cassava, is dependent on resistant genotypes, but the innate mechanisms are unknown. The aim was to study phenylpropanoids and associated enzymes as possible defence components. METHODS: Phenylalanine ammonia-lyase (PAL), phenylpropanoids and peroxidases (POD) were investigated in elicited cassava suspension cells and leaves. Yeast elicitor was the most effective of several microbial and endogenous elicitors. Fungitoxicity was determined against the cassava pathogens Fusarium solani, F. oxysporum and the saprotroph Trichoderma harzianum. KEY RESULTS: A single and rapid (> or =2-3 min) oxidative burst, measured as hydrogen peroxide, occurred in elicited cells. PAL activity was induced maximally at 15 h and was preceded by PAL mRNA accumulation, which peaked at 9 h. Symplasmic POD activity increased four-fold in cells, 48 h post-elicitation. POD isoforms (2-7 isoforms, pI 3.1-8.8) were detected in elicited and unelicited cells, extracellular medium and leaves but two extracellular isoforms were enhanced post-elicitation. Also expression of a cassava peroxidase gene MecPOD1 increased in elicited cells. Only anionic forms oxidized scopoletin, with highest activity by isoform pI 3.6, present in all samples. Unidentified phenolics and possibly scopolin increased post-elicitation, but there was no enhancement of scopoletin, rutin or kaempferol-3-O-rutinoside concentration. Fungal germ tube elongation was inhibited more than germination by esculetin, ferulic acid, quercetin and scopoletin. T. harzianum was generally more sensitive than the pathogens and was inhibited by > or =50 microg mL(-1) of ferulic acid and quercetin and > or =10 microg mL(-1) of scopoletin. CONCLUSIONS: Phenolic levels in cells were not enhanced and were, theoretically, too low to be inhibitory. However, in combination and when oxidized they may contribute to defence, because oxidation of esculetin and scopoletin by peroxidase and of esculetin by tyrosinase enhanced their fungitoxicity up to 20-fold.     

Cassava: an appraisal of its phytochemistry and its biotechnological prospects

The present state of knowledge of the phytochemistry of small molecules isolated from the roots and leaves of cassava, Manihot esculenta Crantz (Euphorbiaceae), is reviewed. Cassava roots are an important source of dietary and industrial carbohydrates, mainly eaten as a source of starch, forming the staple food to over 500 million; additionally, the roots have value as a raw material for industrial starch production and for animal feed giving the crop high economic value, but it suffers markedly from post-harvest physiological deterioration (PPD). The hydroxycoumarins scopoletin and its glucoside scopolin as well as trace quantities of esculetin and its glucoside esculin are identified from cassava roots during PPD. The biotechnological prospects for cassava are also reviewed including a critical appraisal of transgenic approaches for crop improvement, together with its use for bioethanol production, due to cassava's efficient ability to fix carbon dioxide into carbohydrate.     

Characterisation and carriage ratio of Clostridium difficile strains isolated from a community-dwelling elderly population in the United Kingdom

Background

Community-associated Clostridium difficile infection (CDI) appears to be an increasing problem. Reported carriage rates by C.difficile are debatable with suggestions that primary asymptomatic carriage is associated with decreased risk of subsequent diarrhoea. However, knowledge of potential reservoirs and intestinal carriage rates in the community, particularly in the elderly, the most susceptible group, is limited. We have determined the presence of C.difficile in the faeces of a healthy elderly cohort living outside of long-term care facilities (LCFs) in the United Kingdom.

Methods

Faecal samples from 149 community-based healthy elderly volunteers (median age 81 years) were screened for C.difficile using direct (Brazier''s CCEY) and enrichment (Cooked Meat broth) culture methods and a glutamate dehydrogenase (GDH) immunoassay. Isolates were PCR-ribotyped and analysed for toxin production and the presence of toxin genes.

Results

Of 149 faecal samples submitted, six (4%) were found to contain C.difficile. One particular sample was positive by both the GDH immunoassay and direct culture, and concurrently produced two distinct strain types: one toxigenic and the other non-toxigenic. The other five samples were only positive by enrichment culture method. Overall, four C.difficile isolates were non-toxigenic (PCR-ribotypes 009, 026 (n = 2) and 039), while three were toxigenic (PCR-ribotypes 003, 005 and 106). All individuals who had a positive culture were symptom-free and none of them had a history of CDI and/or antibiotics use in the 3 month period preceding recruitment.

Conclusions

To our knowledge, this is the first study of the presence of C.difficile in healthy elderly community-dwelling individuals residing outside of LCFs. The observed carriage rate is lower than that reported for individuals in LCFs and interestingly no individual carried the common epidemic strain PCR-ribotype 027 (NAP1/BI). Further follow-up of asymptomatic carriers in the community, is required to evaluate host susceptibility to CDI and identify dynamic changes in the host and microbial environment that are associated with pathogenicity.     

Colour pattern and inhibition of aggression in the cichlid fish Astronotus ocellatus

Oscars Astronotus ocellatus defeated in combat often undergo a colour pattern change in which the normal olive-green to brown body coloration darkens to near black, interrupted with irregular white barring. Dummy oscars were used in two complementary tests which showed that the dark-and-banded colour pattern inhibits aggression in conspecifics under two different experimental designs and with two different subject groups.     

Histology of melanic flank and opercular color pattern elements in the firemouth cichlid,Thorichthys meeki

Dark melanic color pattern elements, such as bars, stripes, and spots, are common in the skin of fishes, and result from the differential distribution and activity of melanin‐containing chromatophores (melanophores). We determined the histological basis of two melanic color pattern elements in the integument of the Firemouth Cichlid, Thorichthys meeki. Vertical bars on the flanks were formed by three layers of dermal melanophores, whereas opercular spots were formed by four layers (two lateral and two medial) in the integument surrounding the opercular bones. Pretreatment of opercular tissue with potassium and sodium salts effectively concentrated or dispersed intracellular melanosomes. Regional differences in epidermal structure, scale distribution, and connective tissues were also identified. J. Morphol. 2013. © 2013 Wiley Periodicals, Inc.     

Male mate preference and size-assortative pairing in the convict cichlid

Male convict cichlids Cichlasoma nigrofasciatum presented three females simultaneously were found to prefer larger females, even if a female's size exceeded their own. This indicates that some mechanism other than male choice, such as female choice or intrasexual competition, must contribute to male-larger assortative pairing in convict cichlids. Despite a preference for larger females, males continued to consort with smaller females when available, and a females attractiveness was a function of her size relative to other females.