Utilisation of specialist epilepsy services and antiseizure medication adherence rates in a cohort of people with epilepsy (PWE) accessing emergency care.

BACKGROUND: An epilepsy-related attendance at A&E is associated an increased risk of subsequent death within 6 months. Although further work is required to provide a definitive explanation to account for these findings, in the interim it would seem reasonable that services are designed to ensure timely access and provide support at a time of greatest risk. We aim to determine the frequency of patients accessing specialist neurology services following an epilepsy-related admission/unscheduled care episode and consider ASM adherence at the point of attendance. METHODS: Patients were identified retrospectively via the NHS Greater Glasgow and Clyde live integrated epilepsy Dashboard following an unscheduled epilepsy-related admission or A&E attendance between 1st January 2022 and 30th June 2022. We calculated adherence to anti-seizure medication for a period of 6 months prior to admission and defined poor medication adherence as a medication possession ratio of less than 80 %. We evaluated the rate of any outpatient neurology clinic attendance in the subsequent 3, 6 and 12 months following an epilepsy-related unscheduled care episode. Additional clinical information was identified via the electronic patient records. RESULTS: Between 1st Jan 2022 and 30th June 2022, there were 266 emergency care seizure-related attendances. The mean age at attendance was 46 years (range: 16-91). Most of PWE were males (63 %) and 37 % were females. Epilepsy classification-29.3 % had GGE, 41.7% had focal epilepsy, and in 29 % of cases the epilepsy was unclassified. Of the admissions, 107/ 266 (40.2 %) generated follow-up within 6 months of attendance. Poor medication adherence was noted in 54/266 (20.3 %). 28.2 % of cases had input from on-call neurology service during admission/ED attendance, and of those 60 % had ASM adjusted. 18 % of attendances had a background diagnosis of learning disability. One-third of attendances of PWE had a history of mental health disorder 35 % (93/266). 25 % of ED attendances noted an active history of alcohol consumption misuse or/and recreational drug use. 14 (5.5 %) of PWE died during the period of interest (12 months following the last ED visit). In 6/14 (42.3 %) death was associated with poor medication adherence. CONCLUSION: This study demonstrates that a significant proportion of patients who experienced seizure-related admissions/ attendance did not access specialist neurology services in a timely manner. In addition, poor medication adherence remains a problem for a substantial number of people living with epilepsy. Early access to specialist services may go some way to improving care and reducing excessive mortality in PWE by allowing anti-seizure medication to be titrated and poor medication adherence to be addressed in those at greatest risk.

Extravascular Dispersion of Polyvinyl Alcohol Microsphere Particles in Uterine Artery Embolization.

Uterine artery embolization (UAE) is a common minimally invasive treatment of different uterine pathologies, such as fibroids, adenomyosis, and menorrhagia. The procedure involves the injection of embolic agents into the uterine arteries, whereby various particles can be used, such as polyvinyl alcohol (PVA). Complication of UAE is the dispersion of polyvinyl alcohol (PVA) microsphere particles in the uterine body which can lead to a granular vaginal discharge. We report the management of complications of PVA microspheres dispersed from the uterine body causing postprocedural discomfort due to the vaginal passage of microspheres or because of an induced fibroid-size enlargement. The dispersion of the PVA microspheres is one example of a minor UAE complication, which nevertheless causes significant distress to the patient and eventfully requires further surgical interventions.

Rapa Nui (Easter Island) Rano Raraku crater lake basin: Geochemical characterization and implications for the Ahu-Moai Period.

Rano Raraku, the crater lake constrained by basaltic tuff that served as the primary quarry used to construct the moai statues on Rapa Nui (Easter Island), has experienced fluctuations in lake level over the past centuries. As one of the only freshwater sources on the island, understanding the present and past geochemical characteristics of the lake water is critical to understand if the lake could have been a viable freshwater source for Rapa Nui. At the time of sampling in September 2017, the maximum lake depth was ~1 m. The lake level has substantially declined in the subsequent years, with the lake drying almost completely in January 2018. The lake is currently characterized by highly anoxic conditions, with a predominance of ammonium ions on nitrates, a high concentration of organic carbon in the water-sediment interface and reducing conditions of the lake, as evidenced by Mn/Fe and Cr/V ratios. Our estimates of past salinity inferred from the chloride mass balance indicates that it was unlikely that Rano Raraku provided a viable freshwater source for early Rapa Nui people. The installation of an outlet pipe around 1950 that was active until the late 1970s, as well as grazing of horses on the lake margins appear to have significantly impacted the geochemical conditions of Rano Raraku sediments and lake water in recent decades. Such impacts are distinct from natural environmental changes and highlight the need to consider the sensitivity of the lake geochemistry to human activities.

Phylogenomic analysis of SARS-CoV-2 from third wave clusters in Malaysia reveals dominant local lineage B.1.524 and persistent spike mutation A701V.

The emergence of a third wave of COVID-19 infection in Malaysia since September 2020 has led to imminent changes in public health prevention and control measures. As high as 96.2% of registered COVID-19 cases and 88.5% of confirmed deaths in Malaysia occurred during this third wave of infection. A phylogenomic study on 258 SARS-CoV-2 full genomes from February 2020-February 2021 has led to the discovery of a novel Malaysian lineage B.1.524. This lineage contains another spike mutation A701V that co-exists with the D614G spike mutation that was predominant in most of the third-wave clusters. The study provides vital genomic insights on the rapid spread of the SARS-CoV-2 variants in Malaysia in conjunction with the presence of a dominant SARS-CoV-2 lineage during the third wave of COVID-19 infection.

Subcutaneous emphysema and pneumomediastinum in patients with COVID-19 disease; case series from a tertiary care hospital in Pakistan.

Since December 2019, the clinical symptoms of coronavirus disease 2019 (COVID-19) and its complications are evolving. As the number of COVID patients requiring positive pressure ventilation is increasing, so is the incidence of subcutaneous emphysema (SE). We report 10 patients of COVID-19, with SE and pneumomediastinum. The mean age of the patients was 59 ± 8 years (range, 23-75). Majority of them were men (80%), and common symptoms were dyspnoea (100%), fever (80%) and cough (80%). None of them had any underlying lung disorder. All patients had acute respiratory distress syndrome on admission, with a median PaO2/FiO2 ratio of 122.5. Eight out of ten patients had spontaneous pneumomediastinum on their initial chest x-ray in the emergency department. The median duration of assisted ventilation before the development of SE was 5.5 days (interquartile range, 5-10 days). The highest positive end-expiratory pressure (PEEP) was 10 cmH2O for patients recieving invasive mechanical ventilation, while 8 cmH2O was the average PEEP in patients who had developed subcutaneous emphysema on non-invasive ventilation. All patients received corticosteroids while six also received tocilizumab, and seven received convalescent plasma therapy, respectively. Seven patients died during their hospital stay. All patients either survivor or non-survivor had prolonged hospital stay with an average of 14 days (range 8-25 days). Our findings suggest that it is lung damage secondary to inflammatory response due to COVID-19 triggered by the use of positive pressure ventilation which resulted in this complication. We conclude that the development of spontaneous pneumomediastinum and SE whenever present, is associated with poor outcome in critically ill COVID-19 ARDS patients.

Phylogenomic analysis of SARS-CoV-2 from third wave clusters in Malaysia reveals dominant local lineage B.1.524 and persistent spike mutation A701V

@#The emergence of a third wave of COVID-19 infection in Malaysia since September 2020 has led to imminent changes in public health prevention and control measures. As high as 96.2% of registered COVID-19 cases and 88.5% of confirmed deaths in Malaysia occurred during this third wave of infection. A phylogenomic study on 258 SARS-CoV-2 full genomes from February 2020-February 2021 has led to the discovery of a novel Malaysian lineage B.1.524. This lineage contains another spike mutation A701V that co-exists with the D614G spike mutation that was predominant in most of the third-wave clusters. The study provides vital genomic insights on the rapid spread of the SARS-CoV-2 variants in Malaysia in conjunction with the presence of a dominant SARS-CoV-2 lineage during the third wave of COVID-19 infection.

An Overview of the Development of New Vaccines for Tuberculosis.

Currently, there is only one licensed vaccine against tuberculosis (TB), the Bacillus Calmette-Guérin (BCG). Despite its protective efficacy against TB in children, BCG has failed to protect adults against pulmonary TB, lacks therapeutic value, and causes complications in immunocompromised individuals. Furthermore, it compromises the use of antigens present in the purified protein derivate of Mycobacterium tuberculosis in the diagnosis of TB. Many approaches, e.g., whole-cell organisms, subunit, and recombinant vaccines are currently being explored for safer and more efficacious TB vaccines than BCG. These approaches have been successful in developing a large number of vaccine candidates included in the TB vaccine pipeline and are at different stages of clinical trials in humans. This paper discusses current vaccination strategies, provides directions for the possible routes towards the development of new TB vaccines and highlights recent findings. The efforts for improved TB vaccines may lead to new licensed vaccines capable of replacing/supplementing BCG and conferring therapeutic value in patients with active/latent TB.

Blood-brain barrier amenable gold nanoparticles biofabrication in aged cell culture medium.

Green fabrication of nanoscale materials is highly desirable because of associated adverse effects with conventional nanomaterial biomedical applications. Moreover, the higher selective nature of the blood-brain barrier (BBB) limits the brain ailments treatment through conventional chemotherapy, thus providing room for nanotechnology-based modalities for BBB traversing. In this contribution, we have biosynthesized gold nanoparticles from the HAuCl4 solution in the aged cells culture medium. This approach is highly facile without any other chemical utilization. The cell culture medium age and cell number can tune the Au nanoparticles (AuNPs) size from 2 to several hundred nm. The 24 h MTT assay and cell uptake studies in vitro and murine models' vital organs (liver, kidney, spleen, lung, and heart) study up to 48 h demonstrated that biosynthesized AuNPs were biocompatible and BBB amenable. Interestingly, the transferrin and cell culture medium isolated proteins were found factors responsible for HAuCl4 solution biomineralization and size control. Moreover, the protein corona on biosynthesized AuNPs could help them traverse BBB both in vitro and in vivo, suggesting their potential applications for brain disease theranostics. In conclusion, the biosynthesis of AuNPs from aged cells medium is highly facile, green, and biocompatible for brain disease theranostics.

Managing People with Diabetes Fasting for Ramadan During the COVID-19 Pandemic: A South Asian Health Foundation Update.

The month of Ramadan forms one of the five pillars of the Muslim faith. Adult Muslims are obligated to keep daily fasts from dawn to sunset, with exceptions. This year Ramadan is due to begin on 23 April 2020 and the longest fast in the UK will be approximately 18 hours in length. In addition, due to the often high-calorie meals eaten to break the fast, Ramadan should be seen as a cycle of fasting and feasting. Ramadan fasting can impact those with diabetes, increasing the risk of hypoglycaemia, hyperglycaemia and dehydration. This year, Ramadan will occur during the global COVID-19 pandemic. Reports show that diabetes appears to be a risk factor for more severe disease with COVID-19. In addition, the UK experience has shown diabetes and COVID-19 is associated with dehydration, starvation ketosis, diabetic ketoacidosis and hyperosmolar hyperglycaemic state. This makes fasting in Ramadan particularly challenging for those Muslims with diabetes. Here, we discuss the implications of fasting in Ramadan during the COVID-19 pandemic and make recommendations for those with diabetes who wish to fast.

Patterns of facial fractures in children.

Morbidity and mortality among children is usually the result of trauma. Because a child's face is retruded relative to the protecting skull, has a thicker layer of adipose tissue, more elastic bones, flexible sutures lines, the presence of tooth buds within the jaws, and the lack of pneumatisation of the sinuses, the facial bones fracture less commonly than in adults. Our aim was to assess the patterns of such fractures in children who presented to the department of Oral and Maxillofacial Surgery, King Edward Medical University/Mayo Hospital Lahore, Pakistan. All 535 eligible children between the ages of 1-16 years who presented during the two years December 2009 - December 2011 were included in the study. Facial fractures were diagnosed by clinical examination, plain radiographs, and computed tomography, and the pattern of fractures of the facial bones including the frontal bone, orbital bones, maxilla, zygoma, naso-orbito-ethmoidal complex, mandible, and dentoalveolar region was documented. The male:female ratio was 2:1 with 369 male (70%) and 166 female (31%) patients. Fall was the cause in 212 (39%), and in 167 (31%) it was road traffic accidents, while sports were the cause in 135 (25%). The naso-orboto-ethmoid complex was fractured in 37 cases (7%) while 104 children (19%) presented with isolated fractures of the zygomatic bone. The maxilla was fractured in 195 cases (36%), the mandible in 380 (71%), and dentoalveolar trauma was the cause in 256 (50%). The mandible was the bone that was most often fractured (mostly in boys and usually as a result of falls during summer vacations), with the peak occurring in those aged 8-12 years.

The Influence Of Income Level And Remuneration Scheme On Job Satisfaction Among Professional Driving Instructors In Malaysia

@#This study aims to determine the job satisfaction (JS) level among professional driving instructors (PDIs) in the driver training and licensing system in Malaysia. Specifically, it looks at the influence of income level and remuneration scheme on PDI’s satisfaction. The average income of PDI is relatively low with three remuneration schemes: fixed income; fixed income plus commission; and commission only. The study adopted the cross-sectional survey using the 20-item Minnesota Job Satisfaction Questionnaire (MSQ) (short-form) with end-specified 10-point scale. Analysis of the data revealed that PDIs with higher income are more satisfied with their job, t (179) = -3.248, p = 0.001. Similarly, three level One-way ANOVA of JS scores with three remuneration schemes revealed a significant difference of JS across these schemes, F (2,178) = 3.51, p ≤ 0.032. Researchers’ further exploration found a significant interaction between these two variables (income level vs. remuneration scheme), F (5,175) = 4.88, p = 0.001. In general, PDIs with higher income are more satisfied with their job only if they received it as a fixed income (regardless of having commission or not). This insightful finding can be a basis for relevant stakeholders in understanding the influence of income and remuneration scheme towards JS among PDIs in Malaysia.

The efficacy and safety of probiotics for prevention of chemoradiotherapy-induced diarrhea in people with abdominal and pelvic cancer: a systematic review and meta-analysis.

A systematic review and meta-analysis were designed to evaluate the efficacy and safety of probiotics for prevention of chemoradiotherapy-induced diarrhea in people with abdominal and pelvic cancer. We searched the Cochrane Library, PubMed, EMBASE and Web of Science up to November 2015. We also hand searched the citation lists of included studies and previously identified systematic reviews to identify further relevant trials. Odds ratio (OR) was used to compare efficacy, and the pooled OR was estimated using a random effects model; heterogeneity was assessed with Cochran's Q and the Higgins' I2-test. Two reviewers assessed trial quality and extracted data independently. Analysis and bias for each included study were performed using Review Manager 5.2. Nine randomized and placebo-controlled studies (N=1265 participants) were included for assessing efficacy, of which seven were about radiotherapy and two about chemotherapy. Probiotic groups were compared with control groups with respect to the the incidence of diarrhea, OR=0.47 (95% confidence interval 0.28-0.76; P=0.002). Eleven studies, including 1612 people (873 consuming probiotics and 739 not consuming probiotics), were used for the analysis of safety of probiotics. Of the 11 studies, seven studies had no adverse events (AEs) caused by probiotics, whereas four studies reported varying degrees of AEs in their treatment. Probiotics may have a beneficial effect in prevention of chemoradiotherapy-induced diarrhea generally, especially for Grade⩾2 diarrhea. Probiotics may rarely cause AEs.

Development and characterization of Haemophilus influenzae type b conjugate vaccine prepared using different polysaccharide chain lengths.

Capsular polysaccharide conjugates of Haemophilus influenzae type b (Hib) are important components of several mono- or multi-valent childhood vaccines. However, their access to the most needy people is limited due to their high cost. As a step towards developing a cost effective and more immunogenic Hib conjugate vaccine, we present a method for the preparation of Hib capsular polysaccharide (PRP)-tetanus toxoid (TT) conjugates using optimized PRP chain length and conjugation conditions. Reactive aldehyde groups were introduced into the polysaccharides by controlled periodate oxidation of the native polysaccharide, which were subsequently covalently linked to hydrazide derivatized tetanus toxoid by means of reductive amination. Native polysaccharides were reduced to average 100 or 50kDa polysaccharide and 10kDa oligosaccharides in a controlled manner. Various conjugates were prepared using Hib polysaccharide and oligosaccharide yielding conjugates with polysaccharide to protein ratios in the range of 0.25-0.5 (w/w) and free saccharide levels of less than 10%. Immunization of Sprague Dawley rats with the conjugates elicited specific antibodies to PRP. The low molecular weight PRP-TT conjugates were found to be more immunogenic as compared to their high molecular weight counterparts and the PRP-TT reference vaccine.

Effects of interval exercise training programme on the indices of adiposity and biomarker of inflammation in hypertension: a randomised controlled trial.

AIMS AND OBJECTIVES: Hypertension remains a significant risk factor in cardiovascular morbidity and mortality. The purpose of the present study was to investigate the effects of 8 weeks interval training programme on blood pressure, aerobic capacity (VO max), indices of adiposity and marker of inflammation in black African men with essential hypertension. PATIENTS AND METHODS: Two hundred and forty five (245) male subjects with stage 1 and 2 (systolic blood pressure [SBP] between 140-179 & diastolic blood pressure [DBP] between 90-109 mmHg) essential hypertension were age matched and grouped into experimental and control groups. The experimental (n=140; 58.90 ± 7.35 years) group involved in an 8 weeks interval training (60-79% HRmax) programme of between 45 and 60 minutes, while age-matched control hypertensive (n=105; 58.27±6.24 years) group remain sedentary during this period. All subjects in both groups were on antihypertensive drugs throughout the study period. Cardiovascular parameters (SBP, DBP) & VO max and percent body fat [%BF], waist to hip ratio [WHR] and C-reactive protein [CRP] were assessed. Independent t-test and Pearson correlation test were used in data analysis. RESULTS: Findings of the study revealed significant decreased effects of interval training programme on SBP, DBP, %BF, WHR and CRP and significant increased effect on VO max at p< 0.05. Also, changes in CRP as a result of exercise training significantly and positively correlated with changes in SBP, DBP, %BF, WHR, CRP and negatively correlated with VO max at p< 0.05.

Liquid chromatographic determination of CPZEN-45, a novel anti-tubercular drug, in biological samples.

CPZEN-45 is a new drug candidate being considered for the treatment of tuberculosis (TB). The aim of this study was to develop and validate a reverse-phase high-performance liquid chromatographic (HPLC) method suitable to determine CPZEN-45 concentrations in biological samples. CPZEN-45 was extracted from biological fluids and tissues (plasma, lung and spleen from guinea pig) by sequential extraction with acetonitrile and quantified by a Waters HPLC Alliance System coupled with a ZORBAX Bonus-RP column, guard column and UV detection at 263nm. The mobile phase was 20:80 acetonitrile:ultrapure-water with 0.05% TFA. The CPZEN-45 peak was eluted at 5.1min with no interference from the inherent peaks of plasma, bronchoalveolar lavage (BAL), lung or spleen tissues. Recovery of CPZEN-45 from biological samples was >96% of the spiked amount. The limit of detection was 0.05µg/ml and the limit of quantitation was 0.29µg/ml which was more than 5 and 21 times lower than the reported minimal inhibitory concentration of CPZEN-45 (MIC=1.56µg/ml for Mycobacterium tuberculosis and 6.25µg/ml for MDR-TB, respectively). Thus, HPLC method was deemed reliable, sensitive, reproducible and accurate for the determination of CPZEN-45 concentrations in plasma, BAL, lung and spleen tissues. Therefore, this method was used in subsequent studies in the guinea pig model to determine the disposition of CPZEN-45 after administration of solutions by the IV and SC routes.

Pattern of facial palsy in a typical Nigerian specialist hospital.

BACKGROUND: Data on incidence of facial palsy is generally lacking in Nigeria. OBJECTIVE: To assess six years' incidence of facial palsy in Murtala Muhammed Specialist Hospital (MMSH), Kano, Nigeria. METHOD: The records of patients diagnosed as facial problems between January 2000 and December 2005 were scrutinized. Data on diagnosis, age, sex, side affected, occupation and causes were obtained. RESULTS: A total number of 698 patients with facial problems were recorded. Five hundred and ninety four (85%) were diagnosed as facial palsy. Out of the diagnosed facial palsy, males (56.2%) had a higher incidence than females; 20-34 years age group (40.3%) had a greater prevalence; the commonest cause of facial palsy was found out to be Idiopathic (39.1%) and was most common among business men (31.6%). Right sided facial palsy (52.2%) was predominant. Incidence of facial palsy was highest in 2003 (25.3%) and decreased from 2004. CONCLUSION: It was concluded that the incidence of facial palsy was high and Bell's palsy remains the most common causes of facial (nerve) paralysis.

Cellular immune responses in mice induced by M. tuberculosis PE35-DNA vaccine construct.

The PE35 (Rv3872) gene of Mycobacterium tuberculosis is present in the region of difference (RD) one that is deleted in all vaccine strains of Mycobacterium bovis bacillus Calmette Guerin. The aim of this study was to clone PE35 DNA into a DNA vaccine plasmid with CMV promoter and interleukin-2 secretory signal and evaluate the recombinant plasmid for induction of antigen-specific cellular responses in mice. DNA corresponding to PE35 was PCR amplified from the genomic DNA of M. tuberculosis H(37) Rv, cloned into pGEMT-Easy vector and sub-cloned into the DNA vaccine vector pUMVC6. BALB/c mice were immunized with recombinant pUMVC6/PE35 and spleen cells were tested for T-helper (Th)1-type (antigen-induced proliferation and secretion of IFN-γ) and Th2-type (IL-5), and anti-inflammatory (IL-10) cytokine responses to pure recombinant PE35 protein and its synthetic peptides. Mice immunized with the recombinant plasmid DNA (pUMVC6/PE35) showed positive Th1-type cellular responses to pure PE35, but not to an irrelevant antigen, i.e. PPE68 (Rv3873). However, the vaccine construct did not induce antigen-specific Th2-type (IL-5) or anti-inflammatory (IL-10) reactivity to PE35. Testing with synthetic peptides showed that Th1-type cells recognizing various epitopes of PE35 were induced in mice immunized with pUMVC6/PE35 DNA. These results suggest that pUMVC6/PE35 may be useful as a safer vaccine candidate against TB.

DNA vaccine constructs expressing Mycobacterium tuberculosis-specific genes induce immune responses.

RD1 PE35, PPE68, EsxA, EsxB and RD9 EsxV genes are present in Mycobacterium tuberculosis genome but deleted in Mycobacterium bovis BCG. The aim of this study was to clone these genes into DNA vaccine vectors capable of expressing them in eukaryotic cells as fusion proteins, fused with immunostimulatory signal peptides of human interleukin-2 (hIL-2) and tissue plasminogen activator (tPA), and evaluate the recombinant DNA vaccine constructs for induction of antigen-specific cellular immune responses in mice. DNA corresponding to the aforementioned RD1 and RD9 genes was cloned into DNA vaccine plasmid vectors pUMVC6 and pUMVC7 (with hIL-2 and tPA signal peptides, respectively), and a total of 10 recombinant DNA vaccine constructs were obtained. BALB/c mice were immunized with the parent and recombinant plasmids and their spleen cells were tested for antigen-induced proliferation with antigens of M. tuberculosis and pure proteins corresponding to the cloned genes. The results showed that antigen-specific proliferation responses were observed for a given antigen only with spleen cells of mice immunized with the homologous recombinant DNA vaccine construct. The mice immunized with the parent plasmids did not show positive immune responses to any of the antigens of the cloned genes. The ability of the DNA vaccine constructs to elicit cellular immune responses makes them an attractive weapon as a safer vaccine candidate for preventive and therapeutic applications against tuberculosis.

Molecular cloning, expression, purification and immunological characterization of three low-molecular weight proteins encoded by genes in genomic regions of difference of mycobacterium tuberculosis.

The aim of this study was to clone, express and purify three major antigenic proteins, i.e. Rv3874, Rv3875 and Rv3619c, encoded by genes located in regions of difference of Mycobacterium tuberculosis and characterize them for immunogenicity in rabbits. The respective genes were amplified using gene-specific primers and genomic DNA of M. tuberculosis by polymerase chain reaction. The amplified DNA were cloned into pGEM-T Easy and subcloned into pGES-TH-1 vector for high-level expression in Escherichia coli and efficient purification. The results showed that the three fusion proteins, i.e. glutathione-S-transferase (GST)-Rv3874, GST-Rv3875 and GST-Rv3619c, were expressed at high levels and were purified (free of the GST fusion partner) to homogeneity using glutathione-Sepharose and Ni-NTA agarose affinity matrix after cleavage of the column-bound fusion proteins by thrombin protease. The purified recombinant Rv3874, Rv3875 and Rv3619c proteins were immunogenic and induced antigen-specific antibodies in rabbits. Testing of the rabbit sera with overlapping synthetic peptides showed that the antibodies were induced to several epitopes that were scattered throughout the sequence of each protein. These results show immunogenicity of all the proteins for inducing antigen-specific antibodies in rabbits and demonstrate the usefulness of pGES-TH-1 vector for obtaining purified recombinant proteins of M. tuberculosis for immunological characterization.

Species-specific antigenic Mycobacterium tuberculosis proteins tested by delayed-type hypersensitivity response.

OBJECTIVE: To investigate the diagnostic potential of four Mycobacterium tuberculosis antigens encoded by M. tuberculosis-specific region of difference 1 (RD1) region genes (PE35, PPE68, culture filtrate protein 10 [CFP-10], early secreted antigenic target-6 [ESAT-6]) and RD9 region gene Rv3619c, for delayed-type hypersensitivity (DTH) responses in guinea pigs. DESIGN: Recombinant M. tuberculosis proteins were expressed in Escherichia coli and purified to homogeneity by affinity chromatography. Guinea pigs were injected with heat-killed M. tuberculosis and live bacille Calmette-Guérin (BCG), M. avium and M. vaccae. Two to four weeks later, the guinea pigs were challenged intradermally in the flank region with mycobacterial sonicates and purified recombinant proteins. The DTH responses were quantitated by measuring erythema at injection sites after 24 h. RESULTS: All mycobacterial sonicates induced positive DTH responses in guinea pigs injected with M. tuberculosis, M. bovis BCG, M. avium and M. vaccae. Purified proteins PE35, PPE68, CFP10 and ESAT-6 elicited positive DTH responses in the M. tuberculosis-injected group but not in BCG-, M. avium- and M. vaccae-injected guinea pigs, whereas Rv3619c elicited positive DTH responses in the M. tuberculosis- and BCG-injected groups, but not in the M. avium- and M. vaccae-injected guinea pigs. CONCLUSION: The recombinant RD1 antigens induced M. tuberculosis-specific DTH responses. These antigens may therefore be useful in the diagnosis of tuberculosis.