Development of an immunoassay to measure progesterone using printed biosensors, and its application to the assessment of ovarian function in the numbat (Myrmecobius fasciatus)

A biosensor system was developed to measure progesterone levels in the urine of female numbats (Myrmecobius fasciata) as an index of ovarian function. Screen printed sensors were coated with a monoclonal progesterone antibody, and incubated in a mixture of sample/standard and progesterone-3-CMO-horseradish peroxidase (HRP). The difference in potential between the working and reference electrode was measured, after exposure to an HRP substrate. EIA and biosensor standard curves showed parallelism, and the biosensor gave values similar (r=0.83) to the conventional EIA. Progesterone concentrations at different stages of the oestrus cycle were not significantly different to those obtained by EIA

The potential of phage therapy in cystic fibrosis: Essential human-bacterial-phage interactions and delivery considerations for use in Pseudomonas aeruginosa-infected airways

As antimicrobial-resistant microbes become increasingly common and a significant global issue, novel approaches to treating these infections particularly in those at high risk are required. This is evident in people with cystic fibrosis (CF), who suffer from chronic airway infection caused by antibiotic resistant bacteria, typically Pseudomonas aeruginosa. One option is bacteriophage (phage) therapy, which utilises the natural predation of phage viruses upon their host bacteria. This review summarises the essential and unique aspects of the phage-microbe-human lung interactions in CF that must be addressed to successfully develop and deliver phage to CF airways. The current evidence regarding phage biology, phage-bacterial interactions, potential airway immune responses to phages, previous use of phages in humans and method of phage delivery to the lung are also summarised. © 2017 European Cystic Fibrosis Society

The development of an immunoassay to measure progestogen using printed biosensors, and its application to the assessment of ovarian function in the Numbat (Myrmecobius fasciatus)

A biosensor system using screen printed sensors was developed to measure progesterone as an index of ovarian function, and compared with a standard enzymeimmunoassay (EIA). The sensors were coated with a monoclonal progesterone antibody which cross-reacts with a wide range of progestogens, and incubated in a mixture of sample/standard and progesterone-3-CMO-horseradish peroxidise (Prog/HRP). The endpoint was the change in potential read following the addition of sodium perborate. The assay was optimised in terms of the Prog/HRP concentration, the antibody dilution and incubation times. It was then used to measure progestogen in the urine of five female Numbats (Myrmecobius fasciata). Results were available using the sensors within 20 minutes compared to the standard EIA protocol of 2 hrs. The serial dilution of a urine sample taken at the diestrus stage showed parallelism with the serially diluted standard. There was a significant rise in progesterone (mean ± sem) after mating compared to that seen before for both the EIA (1.31 ± 0.20 to 3.70 ± 0.13 ng/ml) and the sensor (1.83 ± 0.33 to 4.02 ± 0.61 ng/ml), and there were no significant differences between the sensor and EIA results at either stage (all p>0.1). A comparison of the values obtained with the sensors to those obtained with the conventional EIA showed a significant correlation for each of the animals (r=0.82 to 0.99). It is concluded that the biosensor system is a viable alternative to conventional EIA, and provides the advantage of (a) a shorter assay time and (b) greater potential for use in the field

Molecular characterisation and genetic variation of Elephant Endotheliotropic Herpesvirus infection in captive young Asian elephants in Thailand

Elephant Endotheliotropic Herpesvirus (EEHV) is emerging as a new threat for elephant conservation, since being identified as the cause of severe, often fatal, haemorrhagic disease in young Asian elephants. To describe positive cases and the molecular relatedness of virus detected in elephants in Thailand, we re-examined all available of EEHV samples occurring in young elephants in Thailand between 2006 and 2014 (n = 24). Results indicated 75% (18/24) of suspected cases were positive for EEHV by semi-nested PCR. Further gene analysis identified these positive cases as EEHV1A (72%, 13/18 cases), EEHV1B (11%, 2/18) and EEHV4 (17%, 3/18). This study is the first to phylogenetically analyse and provide an overview of most of the known EEHV cases that have occurred in Thailand. Positive individuals ranged in age from one to nine years, with no sex association detected, and occurred across geographical locations throughout the country. All individuals, except one, were captive-born. No history of direct contact among the cases was recorded, and this together with the fact that various subtype clusters of virus were found, implied that none of the positive cases were epidemiologically related. These results concur with the hypothesis that EEHV1 is likely to be an ancient endogenous pathogen in Asian elephants. It is recommended that active surveillance and routine monitoring for EEHV should be undertaken in all elephant range countries, to gain a better understanding of the epidemiology, transmission and prevention of this disease

Successful treatment of a clinical elephant endotheliotropic herpesvirus infection: The dynamics of viral load, genotype analysis, and treatment with acyclovir

This article describes the treatment of clinical elephant endotheliotropic herpesvirus (EEHV) infection in a male Asian elephant (Elephas maximus; approximately 3 yr old), the dynamics of viral load during the active infection, and genetic analysis of the virus. Treatment included injectable acyclovir (12 mg/kg iv, bid), antibiotic, vitamin, and fluids. Quantitative polymerase chain reaction was used to measure the viral levels in blood, which decreased continuously after initiation of intravenous acyclovir. Low levels of virus were detected in the blood for 2 wk, and the virus was undetectable after 1 mo. No complication was observed during the treatment period. This case report suggests that acyclovir, given parenterally, could potentially enhance survival of clinical EEHV-infected individuals

The innate resistance of Trypanosoma copemani to human serum

Trypanosoma copemani is known to be infective to a variety of Australian marsupials. Characterisation of this parasite revealed the presence of stercorarian-like life-cycle stages in culture, which are similar to T. rangeli and T. cruzi. The blood incubation infectivity test (BIIT) was adapted and used to determine if T. copemani, like T. cruzi and T. rangeli, has the potential to grow in the presence of human serum. To eliminate any effects of anticoagulants on the complement system and on human high density lipoprotein (HDL), only fresh whole human blood was used. Trypanosoma copemani was observed by microscopy in all human blood cultures from day 5 to day 19 post inoculation (PI). The mechanism for normal human serum (NHS) resistance in T. copemani is not known. The results of this study show that at least one native Australian trypanosome species may have the potential to be infective for humans

Lung Deposition of 99mTc-Radiolabeled Albuterol Delivered through a Pressurized Metered Dose Inhaler and Spacer with Facemask or Mouthpiece in Children with Asthma

Background: Research on the use of a pressurized metered dose inhaler (pMDI) with spacer (pMDI/spacer) in children has indicated oral inhalation via the spacer mouthpiece is more efficient than the combination of oral and nasal inhalation that occurs when a pMDI/spacer is used with a facemask. Changes in pMDI formulations and developments in spacer and facemask designs have highlighted the need for new comparative studies of spacer use, particularly focusing on the age at which children can be taught to transition from use of a pMDI/spacer with facemask to use of the spacer mouthpiece. Methods: Twelve children aged 3–5 years (7 males) with stable asthma were recruited. Of these, 10 children (6 males) completed both arms of the study. A transmission scan of each compliant subject was taken using a 37 MBq 99mTc flood source. Actuations (2–3) of a 99mTc-radiolabeled albuterol pMDI were administered through an antistatic spacer (OptiChamber Diamond) via either a facemask (medium LiteTouch facemask), or the spacer mouthpiece. The subject's inhalation pattern was simultaneously recorded using a pMDI Datalogger, and narrative data relating to tolerance and compliance were documented. Anterior and posterior planar scintigraphic scans were taken immediately after aerosol administration.Results: Mean (SD) lung deposition (% total dose) was 18.1 (9.1)% with the facemask and 22.5 (7.9)% with the spacer mouthpiece (p>0.05). Peripheral lung deposition (expressed as peripheral:central (P:C) ratio) was higher in 7 out of 10 children with the facemask compared with the spacer mouthpiece: 1.3 (0.26) vs. 1.2 (0.35); (p=0.11). Head and neck deposition was higher with use of the facemask compared with the spacer mouthpiece: 19.7 (10.6)% vs. 10.8 (5.3)% (p=0.011). Conclusions: Lung deposition achieved using the spacer with facemask was higher than previously reported, with a difference of only 4.4% of total dose measured compared to the deposition with mouthpiece. This may be due to a combination of factors including pMDI formulation, and use of an antistatic spacer with a flexible, well-fitting facemask

Subclinical infection of captive Asian elephants (Elephas maximus) in Thailand with elephant endotheliotropic herpesvirus

Elephant endotheliotropic herpesvirus (EEHV) infection is a conservation threat to the endangered Asian elephant (Elephas maximus), causing fatal hemorrhagic disease in juvenile elephants throughout the world, including Thailand. This study revealed a subclinical EEHV1 infection rate of 5.5% in healthy captive Asian elephants in Thailand (n = 362). The virus was detected in all age classes above one year old, in both sexes, and across the country – even in facilities with no history of hemorrhagic disease (EEHV HD). Subclinical EEHV infection in Thailand urgently requires proper health management

Recombinant Jembrana disease virus gag proteins identify several different antigenic domains but do not facilitate serological differentiation of JDV and nonpathogenic bovine lentiviruses

In Indonesia, it is suspected that there are two bovine lentiviruses circulating in the cattle population: a pathogenic Jembrana disease virus (JDV), and a nonpathogenic bovine immunodeficiency-like virus (BIV). Both viruses cross-react antigenically and cannot be differentiated by current serological tests using JDV antigens. To identify possible type-specific epitopes, a series of recombinant protein constructs including the matrix, capsid and nucleocapsid proteins were produced from JDV gag and the expressed proteins were tested by Western blot using JDV and BIV hyperimmune sera. JDV matrix and truncated capsid proteins were recognised by both JDV and BIV hyperimmune sera indicating that there were multiple cross-reactive epitopes present in JDV gag. At least three epitopic regions were identified in these constructs, including the major homology region, by monoclonal antibody binding studies. JDV nucleocapsid recombinant protein was not recognised by either JDV or BIV hyperimmune sera and none of the recombinant gag proteins were able to differentiate between JDV positive sera from Jembrana disease endemic and Jembrana disease-free areas. Additionally, a 40 amino acid recombinant subunit protein encompassing the region recently found to contain an epitope unique to BIV [Zheng, L., Zhang, S., Wood, C., Kapil, S., Wilcox, G.E., Loughin, T.A., Minocha, H.C., 2001. Differentiation of two bovine lentiviruses by a monoclonal antibody on the basis of epitope specificity. Clin. Diagn. Lab. Immunol. 8, 283–287] was tested but was not recognised by either JDV positive sera from Jembrana disease-endemic or Jembrana disease-free areas