Developing novel anthelmintics: the stability of cysteine proteinase activity in a supernatant extract of papaya latex.

Plant derived cysteine proteinases (CPs) have long been known to possess anthelmintic properties but have attracted renewed attention recently because of the acute need to discover novel methods for controlling helminth infections as a result of increasing drug resistance. However, surprisingly little is known about the stability of these proteins under typical storage and in vivo exposure conditions. We found that CPs in a supernatant preparation from papaya latex (PLS) were stable during the initial refinement process and when stored under low temperatures, but lost activity during dialysis and within 7 days of storage when kept at ambient temperature (18-20 °C). The enzyme activity in PLS was not affected by repeated freeze-thaw cycles and was also stable under typical in vitro assay conditions at 37 °C used for quantifying effects on helminths. Active enzyme activity was still detectable in the colon 3-4 h after oral administration in rodent models.

Imported Falciparum Malaria: A case series in a tertiary hospital.

INTRODUCTION: Malaysia declared its intent to eliminate malaria by 2020, with a phased goal of achieving zero local transmission. Nonetheless, Malaysia is highl susceptible to malaria importation due to geographical proximity to high-burden countries e.g. Thailand, Myanmar and high influx of foreign workers and students from Asia and Africa. CASE SERIES: We accumulated all malaria cases diagnosed in a tertiary hospital within a period of two years. Three cases were reported, where all of the patients were foreigners with recent travel history to African countries. All of them were infected by P. falciparum, responded to treatment and discharged well. DISCUSSION: This case series highlighted the importance of acquiring recent travel history during history taking and having a high index of suspicions on malaria when dealing with feverish patients originated particularly from African countries.

Dentists' intention to report suspected violence: a cross-sectional study in eight Arab countries.

OBJECTIVES: This study assessed dentists' intention in eight Arab countries to report suspected exposure to violence among patients and factors associated with this intention based on the theory of planned behaviour. METHODS: A cross-sectional study was conducted in 2016 including a convenience sample of dentists practising in public, private and academic sectors in Algeria, Egypt, Jordan, Kuwait, Libya, Palestine, Saudi Arabia and Yemen. Respondents answered a self-administered questionnaire collecting information about personal and professional background and perceived ability to identify victims of violence. The questionnaire assessed (on a scale from 1 to 10 using six negative statements) dentists' perception of healthcare system mandated reporting of suspected violence. Six statements were used to assess professional attitude towards reporting suspected violence. Logistic regression was used to assess the association between intention to report suspected violence and perceived ability, perception and attitude adjusting for confounders. RESULTS: The response rate was 65.2% (n=2936/4506) from general practitioners (70.9%) of mean age=31 years with 56.7% women. Of those, 68.8% intended to report and 52.2% considered themselves able to identify violence victims. The mean (SD) negative perception score=5.3/10 (2.1) and the mean (SD) professional attitude score=7.5/10 (1.9). In multivariate regression, intention to report was associated with professional attitude (OR 1.08, 95% CI 1.03 to 1.14), ability to identify violence victims (OR 1.76, 95% CI 1.45 to 2.12) and negative perception that reporting is not mandated (OR 0.89, 95% CI 0.85 to 0.94). Significant differences existed among countries in intention to report. CONCLUSION: Most dentists intended to report suspected violence and their intention could be explained by the theory of planned behaviour which offers a framework for professional development to support violence victims. Sharing of training resources, policies and guidelines is needed to ensure that practices similar to international guidelines are consistently adopted by dentists across Arab countries.

The anthelmintic efficacy of natural plant cysteine proteinases against the equine tapeworm, Anoplocephala perfoliata in vitro.

Papaya latex has been demonstrated to be an efficacious anthelmintic against murine, porcine, ovine and canine nematode parasites, and even those infecting poultry, and it has some efficacy against rodent cestodes. The active ingredients of papaya latex are known to be cysteine proteinases (CPs). The experiments described in this paper indicate that CPs in papaya latex, and also those in pineapples, are highly efficacious against the equine cestode Anoplocephala perfoliata in vitro, by causing a significant reduction in motility leading to death of the worms. The susceptibility of A. perfoliata to damage by CPs was considerably greater than that of the rodent cestodes Hymenolepis diminuta and H. microstoma. Our results are the first to report anthelmintic efficacy of CPs against an economically important equine helminth. Moreover, they provide further evidence that the spectrum of activity of CPs is not restricted to nematodes and support the idea that these plant-derived enzymes can be developed into useful broad-spectrum anthelmintics.

The anthelmintic efficacy of natural plant cysteine proteinases against the rat tapeworm Hymenolepis diminuta in vivo.

Hymenolepis diminuta is a natural parasite of the common brown rat Rattus norvegicus, and provides a convenient model system for the assessment of the anthelmintic activity of novel drugs against cestodes. The experiments described in this paper indicate that treatment of rats infected with H. diminuta with a supernatant extract of papaya latex, containing a mixture of four cysteine proteinases, was moderately efficacious, resulting in a significant, but relatively small, reduction in worm burden and biomass. However, faecal egg output was not affected by treatment. In our experiments these effects were only partially dose-dependent, although specific inhibition by E-64 confirmed the role of cysteine proteinases as the active principles in papaya latex affecting worm growth but not statistically reducing worm burden. Data collected for a further 7 days after treatment indicated that the effects of papaya latex supernatant on worm loss and on worm growth were not enhanced. Our findings provide a starting point for further refinement in formulation and delivery, or assessment of alternative natural plant-derived cysteine proteinases in efforts to develop these naturally occurring enzymes into broad-spectrum anthelmintics, with efficacy against cestodes as well as nematodes.

The anthelmintic efficacy of natural plant cysteine proteinases against Hymenolepis microstoma in vivo.

Little is known about the efficacy of cysteine proteinases (CP) as anthelmintics for cestode infections in vivo. Hymenolepis microstoma is a natural parasite of house mice, and provides a convenient model system for the assessment of novel drugs for anthelmintic activity against cestodes. The experiments described in this paper indicate that treatment of H. microstoma infections in mice with the supernatant of papaya latex (PLS), containing active cysteine proteinases, is only minimally efficacious. The statistically significant effects seen on worm burden and biomass showed little evidence of dose dependency, were temporary and the role of cysteine proteinases as the active principles in PLS was not confirmed by specific inhibition with E-64. Worm fecundity was not affected by treatment at the doses used. We conclude also that this in vivo host-parasite system is not sensitive enough to be used reliably for the detection of cestocidal activity of compounds being screened as potential, novel anthelmintics.

The relative anthelmintic efficacy of plant-derived cysteine proteinases on intestinal nematodes.

We examined the in vitro and in vivo efficacy of plant cysteine proteinases (CPs) derived from pineapple (Ananas comosus) and kiwi fruit (Actinidia deliciosa), and compared their efficacy as anthelmintics to the known effects of CPs from the latex of papaya (Carica papaya) against the rodent intestinal nematode, Heligmosomoides bakeri. Both fruit bromelain and stem bromelain had significant in vitro detrimental effects on H. bakeri but in comparison, actinidain from kiwi fruit had very little effect. However, in vivo trials indicated far less efficacy of stem bromelain and fruit bromelain than that expected from the in vitro experiments (24.5% and 22.4% reduction in worm burdens, respectively) against H. bakeri. Scanning electron microscopy revealed signs of cuticular damage on worms incubated in fruit bromelain, stem bromelain and actinidain, but this was far less extensive than on those incubated in papaya latex supernatant. We conclude that, on the basis of presently available data, CPs derived from pineapples and kiwi fruits are not suitable for development as novel anthelmintics for intestinal nematode infections.

The anthelmintic efficacy of natural plant cysteine proteinases against two rodent cestodes Hymenolepis diminuta and Hymenolepis microstoma in vitro.

Little is known about the efficacy of cysteine proteinases (CP) as anthelmintics for cestode infections. We examined the effects of CPs on two rodent cestodes, Hymenolepis diminuta and H. microstoma in vitro. Our data showed that naturally occurring mixtures of CPs, such as those found in papaya latex, and relatively pure preparations of fruit bromelain, papain and stem bromelain, were active in vitro against both juvenile, artificially excysted scoleces, as well as against adult worms of both rodent cestodes. Significant dose-dependent reduction in motility, ultimately leading to death of the worms, was observed with both species, and against both freshly excysted scoleces and 14-day old pre-adult worms. The most effective was fruit bromelain (after 30 min of incubation of juvenile H. diminuta and H. microstoma IC50=63 and 74 µM, respectively, and for pre-adult worms=199 and 260 µM, respectively). The least effective was stem bromelain (after 30 min of incubation of juvenile H. diminuta and H. microstoma IC50=2855 and 2772 µM, respectively, and for pre-adult worms=1374 and 1332 µM, respectively) and the efficacies of papaya latex supernatant and papain were between these extremes. In all cases these values are higher than those reported previously for efficacy of CPs against intestinal nematodes, and in contrast to nematodes, all CPs were effective against cestodes in the absence of exogenous cysteine in incubation media. The CPs appeared to attack the tegument resulting in generalised erosion mainly on the strobila. The scolex was more resistant to CP attack but nevertheless some damage to the tegument on the scolex was detected.

Intravascular catheter related infections and antimicrobial susceptibility pattern of isolated bacteria in a tertiary care hospital of Bangladesh.

The aim of this study was to evaluate the rate of bacterial colonisation and catheter related blood stream infections (CRBSI) together with the antibiotic susceptibility patterns in a tertiary care hospital. CRBSI was detected with semi-quantitative and quantitative methods. The antimicrobial susceptible patterns of the isolated organisms were performed by Kirby Bauer disk diffusion method. The rate of catheter colonisation and CRBSI were 42.1% and 14% (16.1/1000 catheter days) respectively. The most common causative pathogens were Pseudomonas sp. (23.7%), Acinetobacter sp. (18.4%), Staphylococcus aureus (13.2%) and Enterobacteriaceae (10.5%). The rate of isolation of methicillin resistance S. aureus, imipenem resistant Pseudomonas sp. and extended spectrum ß lactamase producing Enterobacteriaceae were 60%, 44.0% and 100%. The result of this study would be useful for control and treatment of CRBSI.

The anthelmintic efficacy of papaya latex in a rodent-nematode model is not dependent on fasting before treatment.

In earlier studies of the anthelmintic activity of plant cysteine proteinases (CPs), a period of food deprivation was routinely employed before administration of CPs, but there has been no systematic evaluation as to whether this does actually benefit the anthelmintic efficacy. Therefore, we assessed the effect of fasting on the efficacy of CPs from papaya latex (PL) against Heligmosomoides bakeri in C3H mice. We used a refined, supernatant extract of papaya latex (PLS) with known active enzyme content. The animals were divided into three groups (fasted prior to treatment with PLS, not fasted but treated with PLS and fasted but given only water). The study demonstrated clearly that although food deprivation had been routinely employed in much of the earlier work on CPs in mice infected with nematodes, fasting has no beneficial effect on the efficacy of PLS against H. bakeri infections. Administration of CPs to fed animals will also reduce the stress associated with fasting.

Dissimilar differentiation of mesenchymal stem cells from bone marrow, umbilical cord blood, and adipose tissue.

Mesenchymal stem cells (MSCs) have been investigated as promising candidates for use in new cell-based therapeutic strategies such as mesenchyme-derived tissue repair. MSCs are easily isolated from adult tissues and are not ethically restricted. MSC-related literature, however, is conflicting in relation to MSC differentiation potential and molecular markers. Here we compared MSCs isolated from bone marrow (BM), umbilical cord blood (UCB), and adipose tissue (AT). The isolation efficiency for both BM and AT was 100%, but that from UCB was only 30%. MSCs from these tissues are morphologically and immunophenotypically similar although their differentiation diverges. Differentiation to osteoblasts and chondroblasts was similar among MSCs from all sources, as analyzed by cytochemistry. Adipogenic differentiation showed that UCB-derived MSCs produced few and small lipid vacuoles in contrast to those of BM-derived MSCs and AT-derived stem cells (ADSCs) (arbitrary differentiation values of 245.57 +/- 943 and 243.89 +/- 145.52 mum(2) per nucleus, respectively). The mean area occupied by individual lipid droplets was 7.37 mum(2) for BM-derived MSCs and 2.36 mum(2) for ADSCs, a finding indicating more mature adipocytes in BM-derived MSCs than in treated cultures of ADSCs. We analyzed FAPB4, ALP, and type II collagen gene expression by quantitative polymerase chain reaction to confirm adipogenic, osteogenic, and chondrogenic differentiation, respectively. Results showed that all three sources presented a similar capacity for chondrogenic and osteogenic differentiation and they differed in their adipogenic potential. Therefore, it may be crucial to predetermine the most appropriate MSC source for future clinical applications.