Effect of HIV infection on plasma exposure to first-line TB drugs and target attainment in children.

BACKGROUND: Whether HIV infection adversely affects exposure to first-line TB drugs in children is debatable. It is also not known whether HIV infection increases the risk of plasma underexposure or overexposure to TB drugs. This study sought to address these questions.DESIGN/METHODS: Children on TB treatment were enrolled. After 4 weeks on therapy, blood samples were collected at pre-dose, 1, 2, 4, 8, and 12 h post-dose for pharmacokinetic analysis. Plasma drug exposure below and above the lower and upper bounds of the 95% confidence intervals of the reference mean for children were considered underexposure and overexposure, respectively. The effect of HIV infection on drugs exposure and risk of underexposure were examined using multivariate analysis.RESULTS: Of 86 participants (median age: 4.9 years), 45 had HIV coinfection. HIV coinfection was associated with lower pyrazinamide (PZA) and ethambutol exposures in adjusted analysis. Patients with TB-HIV coinfection were three times more likely to have PZA underexposure than those with TB only. Underexposure of rifampin was common irrespective of HIV coinfection status.CONCLUSIONS: HIV coinfection was associated with a higher risk for PZA underexposure in children. This effect should be accounted for in models and simulations to determine optimal PZA dose for children.

Adequacy of WHO weight-band dosing and fixed-dose combinations for the treatment of TB in children.

BACKGROUND: We examined whether the updated WHO weight-band dosing recommendations and fixed-dose combination tablets for the treatment of TB in children achieves recommended calculated dosages and adequate drug plasma exposure.DESIGN/METHODS: Children on first-line TB treatment per WHO guidelines were enrolled. Blood sampling at pre-dose, 1, 2, 4, 8, and 12 h post-dose after at least 4 weeks of treatment was performed. Drugs concentrations were measured using validated liquid chromatography tandem with mass spectrometry and pharmacokinetic parameters calculated using noncompartmental analysis. Plasma drug exposure below the lower limit of the 95% confidence interval of the mean for children was considered low and above the upper limit was high.RESULTS: Of 71 participants, 34 (47.9%) had HIV coinfection. The median calculated dose for isoniazid (INH), rifampin (RIF), pyrazinamide (PZA), and ethambutol (EMB) was 10.0 (range 4.3-13.3), 15.0 (range 8.6-20.0), 30.0 (range 21.0-40.0), and 20.4 (range 14.3-26.7) mg/kg, respectively. Overall, most patients had under-exposure for RIF and PZA and over-exposure for INH and EMB. Drug dose and weight-for-age Z-score were associated with area under the curve from time 0-24 h for all drugs.CONCLUSIONS: Despite adherence to WHO dosing guidelines, low PZA and RIF plasma exposures were frequent in our study population. Higher than currently recommended dosages of RIF and PZA may be needed in children.

Clinical Evaluation of the Safety and Effectiveness of Heptonica: A Ghanaian Hepatorestorative Polyherbal Product.

The incidence of liver diseases is increasing globally, and many patients in developing countries are resorting to the use of herbal products as treatment. This study was aimed at establishing the safety and effectiveness outcomes for patients with deranged liver panel treated with a Ghanaian finished polyherbal product. The product Heptonica is prepared by CPMR from three medicinal plants: Bidens pilosa, Citrus aurantifolia, and Trema orientalis. Fifty (50) participants with clinical and biochemical signs of liver impairment were purposively recruited and treated for a period of 28 days. Participants received Heptonica at a dose of 30 mL 8 hourly after meals for the treatment period. Clinical and biochemical evaluation (liver panel test, renal function test, haematology, and urinalysis) of subjects for the safety and effectiveness of the product was undertaken at days 0 (baseline), 14, and 28. Compared to the baseline values, Heptonica did not have any untoward effect on renal function, haematological parameters, and urine parameters of subjects. Clinical and liver panel results of the participants also improved compared to the baseline: serum aspartate transaminase (AST) (p < 0.0001), alanine transaminase (ALT) (p < 0.0001), gamma-glutamyltransferase (GGT) (p- 0.0013), total bilirubin (p-0.0136), direct bilirubin (p < 0.0001), total proteins (p-0.0409), and alkaline phosphates (p- 0.0284). Level of albumin showed no significant difference within the study period. The outcome of this study indicates Heptonica has hepatorestorative action with no observable toxicity and can be used with confidence as indicated as a liver tonic.

Sunlight exposure, consumption of vitamin D-rich foods and vulvovaginal candidiasis in an African population: a prevalence case-control study.

BACKGROUND/OBJECTIVES: To date, only two studies have investigated the relationship between vitamin D (vitD) deficiency and candidiasis in spite of vitD's antimicrobial and immunomodulatory roles. We examined the relationship between sunlight exposure and consumption of vitD-rich foods, markers of vitD status, and vulvovaginal candidiasis (VVC) in an African population to add to the limited evidence and stimulate additional research. SUBJECTS/METHODS: Three hundred cases (females diagnosed as suffering from VVC) and three hundred controls (females diagnosed as suffering from any condition other than VVC) were selected from three health facilities in Cape Coast, Ghana. Sunlight exposure was assessed in a structured questionnaire with a food frequency questionnaire used to ascertain the frequency of consumption of vitD-rich foods. RESULTS: Self-reported low sunlight exposure was associated with 3.38 (95% CI:1.99, 5.74) increased odds of VVC. Low and moderate sunlight exposure estimated by outdoor visits was also associated with increased odds of VVC. In sensitivity analysis restricted to matched sunlight exposure data, low and moderate exposure was associated with 5.78 (95% CI: 2.57, 12.99) and 3.53 (95% CI: 1.85, 6.75) increased odds of VVC. Odds of VVC increased with decreasing levels of consumption of vitD-rich foods (Likelihood-ratio test trend p = 0.1382). In the joint analysis, low and moderate vitD intake was associated with much higher increased odds of VVC. CONCLUSIONS: Our findings should be confirmed in prospective studies and clinical trials to strengthen the evidence base for preventive action and to also inform clinical decision making.

Surgical treatment of Cesarean scar ectopic pregnancy: efficacy and safety of ultrasound-guided suction curettage.

OBJECTIVES: To assess the efficacy of ultrasound-guided suction curettage for management of pregnancies implanted into the lower uterine segment Cesarean section scar. METHODS: This was a retrospective study including women diagnosed with Cesarean section scar pregnancy at two large tertiary referral early pregnancy units between 1997 and 2014. Surgical evacuation was offered to selected women presenting in the first trimester ≤ 14 weeks' gestation. All procedures were performed transcervically under ultrasound guidance using suction curettage. A modified Shirodkar cervical suture was used in women who required additional measures to secure hemostasis. RESULTS: A total of 232 women with Cesarean section scar pregnancy were seen at the referral units; 191/232 (82.3%) women were treated surgically. The median intraoperative blood loss was 100 mL (range, 10-3000 mL); 9/191 (4.7% (95% CI, 1.7-7.7%)) women required blood transfusion and, in one (0.5% (95% CI, 0-1.5%)), life-saving hysterectomy had to be performed because of uncontrollable intraoperative bleeding. Of the women who attended for follow-up, 7/116 (6.0% (95% CI, 1.7-10.3%)) required a repeat surgical procedure because of retained products of conception. Multivariable analysis showed that the gestational sac diameter (odds ratio (OR), 1.10 (95% CI, 1.03-1.17)) and pregnancy vascularity on Doppler examination (OR, 3.41 (95% CI, 1.39-8.33)) were significant predictors of heavy intraoperative blood loss (> 1000 mL). CONCLUSIONS: Ultrasound-guided suction curettage is an effective method for the treatment of pregnancies implanted into a lower uterine segment Cesarean section scar and is associated with a low risk of blood transfusion and hysterectomy. Copyright © 2016 ISUOG. Published by John Wiley & Sons Ltd.

Quantitative dose-response curves from subcellular lipid multilayer microarrays.

The dose-dependent bioactivity of small molecules on cells is a crucial factor in drug discovery and personalized medicine. Although small-molecule microarrays are a promising platform for miniaturized screening, it has been a challenge to use them to obtain quantitative dose-response curves in vitro, especially for lipophilic compounds. Here we establish a small-molecule microarray assay capable of controlling the dosage of small lipophilic molecules delivered to cells by varying the sub-cellular volumes of surface supported lipid micro- and nanostructure arrays fabricated with nanointaglio. Features with sub-cellular lateral dimensions were found necessary to obtain normal cell adhesion with HeLa cells. The volumes of the lipophilic drug-containing nanostructures were determined using a fluorescence microscope calibrated by atomic-force microscopy. We used the surface supported lipid volume information to obtain EC-50 values for the response of HeLa cells to three FDA-approved lipophilic anticancer drugs, docetaxel, imiquimod and triethylenemelamine, which were found to be significantly different from neat lipid controls. No significant toxicity was observed on the control cells surrounding the drug/lipid patterns, indicating lack of interference or leakage from the arrays. Comparison of the microarray data to dose-response curves for the same drugs delivered liposomally from solution revealed quantitative differences in the efficacy values, which we explain in terms of cell-adhesion playing a more important role in the surface-based assay. The assay should be scalable to a density of at least 10,000 dose response curves on the area of a standard microtiter plate.

The psychological effects and patient acceptability of a test to predict viability in early pregnancy: a prospective randomised study.

OBJECTIVE: To establish if women obtain any measurable short term psychological benefit or perceived benefit from having a test to determine the probability of their pregnancy being on-going when this is uncertain on ultrasound examination. STUDY DESIGN: This was a prospective randomised controlled study conducted January 2012-June 2012 at the EPU of King's College Hospital. The study population was women who conceived spontaneously and had a single intrauterine gestational sac of <20mm mean diameter, with no visible embryo on their first ultrasound scan. Eligible women were randomised to have a test to calculate the probability of viability (cases) or not (controls). Depression and anxiety levels were calculated using the Hospital Anxiety and Depression Score (HADS) and were performed prior to randomisation and seven days later. A repeat scan for pregnancy outcome was performed after one to two weeks as clinically indicated. A sample size of 69 in each group was calculated to have 80% power to detect a probability of 0.362 that an observation in the cases was less than an observation in controls using a Wilcoxon Mann-Whitney rank-sum test with a 0.05 two-sided significance. RESULTS: At recruitment there was no significant difference in anxiety levels between cases and controls. After seven days anxiety levels were significantly lower in cases than controls (p=0.04). Of those who received the probability score, 55/70 (78.6% 95% CI 67.5-86.7%) found it useful and 58/70 (82.9% 95% CI 72.2-90.1) would choose to have the test in a future pregnancy if indicated. CONCLUSIONS: This study has demonstrated that there is evidence of psychological benefit from a simple blood test that gives women the likelihood that their pregnancy will be on-going at the next scan.

Clinical significance of first-trimester chorionic bumps: a matched case-control study.

OBJECTIVE: To determine the clinical significance of a chorionic bump diagnosed by ultrasound in women attending an early pregnancy unit in a teaching hospital. METHODS: This was a retrospective case-control study over an 8-year period (2003-2010). Cases of chorionic bump were identified by searching our early pregnancy database and were matched to controls in a ratio of 1:3. The primary outcome measure was miscarriage vs ongoing pregnancy. Secondary outcomes were gestational age at delivery and the presence or absence of fetal abnormality. RESULTS: A total of 37 798 pregnancies were examined over the study period and 57 pregnancies with a chorionic bump were identified, giving an estimated prevalence of 1.5 per 1000 pregnancies (0.15%; 95% CI, 0.01-0.73%). Of the 52 women with follow-up data, 20 (38.5%; 95% CI, 26.4-52.1%) miscarried vs 31/151 (20.5%; 95% CI, 14.8-27.7%) in the control group (P = 0.01). There were four second-trimester miscarriages in the study group and none in the controls (P < 0.01). Out of 52 pregnancies in the study group there were 32 live births (62%; 95% CI, 47.9-73.6%) vs 118/151 (78%; 95% CI, 70.9-84.0%) in the control group (P = 0.02). There were no differences in preterm delivery rates or fetal anomalies. No significant relationship was found between size of the bump or location in relation to the umbilical cord insertion and risk of miscarriage. CONCLUSIONS: Women presenting to early pregnancy units with a chorionic bump discovered at first-trimester ultrasound examination had approximately double the risk of miscarriage compared with matched controls, the difference being due to a greater number of miscarriages during the second trimester of pregnancy.

Trastuzumab for the treatment of HER2-positive metastatic adenocarcinoma of the stomach or gastro-oesophageal junction.

This paper presents a summary of the evidence review group (ERG) report into trastuzumab for the treatment of human epidermal growth factor receptor 2 (HER2)-positive metastatic adenocarcinoma of the stomach (mGC) or gastro-oesophageal junction. HER2 positivity is defined by immunohistochemistry (IHC)3+ or IHC2+/fluorescence in situ hybridisation (FISH)+. The decision problem addressed was the testing of the whole mGC population with IHC and, for IHC2+ patients, also with FISH, followed by treatment of HER2-positive patients with trastuzumab combined with cisplatin and either capecitabine or 5-fluorouracil (5-FU) [HCX (trastuzumab, cisplatin, capecitabine)/fluorouracil (F)] compared with current standard NHS therapy. The manufacturer's submission contained direct evidence from the ToGA trial, a well-conducted, multinational, phase III randomised controlled trial (RCT) that compared HCX/F with cisplatin and a fluoropyrimidine alone [cisplatin, capecitabine (CX)/F]. HCX/F showed statistically significantly better overall survival in the European Medicines Agency-licensed population subgroup (74%) (hazard ratio 0.65, 95% confidence interval 0.51 to 0.83), corresponding to median survival of 16 months versus 11.8 months. No other evidence exists for the efficacy of any therapy in a known HER2-positive mGC population; other comparisons extrapolate from trials in mixed HER2 status populations. The ERG accepted the manufacturer's view that a meaningful network meta-analysis to establish a comparison for HCX/F compared with current standard NHS therapy [epirubicin, cisplatin, capecitabine (ECX)/epirubicin, oxaliplatin, capecitabine (EOX)/epirubicin, cisplatin, 5-FU (ECF)] was not possible, but was unconvinced by arguments advanced in the alternative narrative synthesis. These involved disregarding evidence from a meta-analysis and interpreting non-significant results of small RCTs comparing epirubicin-containing triplets with cisplatin, 5-FU (CF)/capecitabine (X) doublets as evidence of no difference between triplet and doublet regimens. The high CX/F dose in the ToGA trial was an additional basis for the contention of equivalence. An appropriate de novo economic evaluation, including an economic model that separately compared HCX or trastuzumab, cisplatin, 5-FU (HCF) with the triplet regimens ECX, EOX and ECF, based on a simple, three-state cohort model (progression-free, disease, progression and death), was submitted. Utility weights were applied to estimate quality-adjusted life-years (QALYs). Costs were assessed from an NHS perspective, and incorporated the acquisition and monitoring costs of the alternative regimens, HER2 testing, adverse events and other supportive care costs. An 8-year time horizon was used to represent a lifetime analysis. Results from the ToGA trial were combined with a series of assumptions on relative treatment effects and testing strategies. The manufacturer's results produced an incremental cost-effectiveness ratio (ICER) of £ 53,010 per QALY for HCX versus ECX. Although the manufacturer undertook a detailed set of sensitivity analyses, several alternative model assumptions were not evaluated. The ERG undertook a series of alternative base-case analyses. As a result of these analyses, EOX replaced ECX as the appropriate comparator, and the ICER for the comparison of HCX vs EOX increased to between £ 66,982 and £ 71,636 per QALY. The impact of implementation of alternative testing strategies remained unclear. There is also considerable uncertainty surrounding the true estimate of effectiveness for the comparison between triplet regimens containing epirubicin (ECX/ECF/EOX) and doublet CX/F regimens. Consequently, the view of the ERG was that there is insufficient evidence on the efficacy of HCX/F compared with current NHS standard therapy for an ICER to be determined with any degree of certainty.

A new era in the management of burns trauma in kumasi, ghana.

The aim of the study was to investigate the factors affecting the outcome of treatment of burns patients admitted to the Burns Intensive Care Unit (BICU) of the Komfo Anokye Teaching Hospital, Kumasi, Ghana. Information on patients admitted to the BICU from February 2001 to January 2006 was recorded. Parameters recorded included: admission record and demographics, causes of the injury, burned surface area, laboratory investigations, treatment regime, and record of discharge/death. The data were analysed with SPSS version 12.0 and Spearman's rank correlation. A total of 826 patients were recorded; males (n = 492, 60%) outnumbered females (n= 334, 40%). The mean age was 10.5 ± 5 yr, the majority (n = 441, 53%) in the range 0-10 yr. Flame burns (n = 587, 71%), scalds (n = 209, 25%), and chemicals (n = 19, 2%) were the three significant causes of burn injuries. The mean range of the total body surface area (TBSA) burned was 11-20%; 94% (n = 775) had up to 60% TBSA; 64% (n = 527) had only wound dressings for treatment; 21% (n = 174) had early excision with skin grafting, while 15% (n = 125) had delayed excision with skin grafting. The majority (n = 563, 68%) of the patients stayed for less than 10 days after admission. The mortality rate fell over the years, decreasing drastically between 2001 (20.4%) and 2002 (8.6%) and remaining at single digit level in 2003 (7.6%), 2004 (7.9%), and 2005 (7.4 %). The factors affecting the mortality trends were proper case management, increases in the number of professional medical personnel, and their greater dedication.

Treatment of urinary schistosomiasis: methodological issues and research needs identified through a Cochrane systematic review.

Guidelines recommend praziquantel (PZQ) for the treatment and control of schistosomiasis, with no real alternative. Metrifonate was still widely used against Schistosoma haematobium in the 1990s, and then withdrawn. Experimental studies and clinical trials suggest that artemisinin compounds are active against S. haematobium. In a Cochrane systematic review assessing the efficacy and safety of drugs for treating urinary schistosomiasis, 24 randomized controlled trials (n=6315 individuals) met our inclusion criteria. These trials compared a variety of single agent and combination regimens with PZQ, metrifonate or artemisinin derivatives. The review confirmed that both the standard recommended doses of PZQ (single 40 mg/kg oral dose) and metrifonate (3x7.5-10 mg/kg oral doses administered fortnightly) are efficacious and safe in treating urinary schistosomiasis, but there is no study comparing these two regimens head-to-head. There is currently not enough evidence to evaluate artemisinin compounds. Most of the studies included in the Cochrane systematic review were insufficiently powered, lacked standardization in assessing and reporting outcomes, and had a number of methodological limitations. In this paper we discuss the implications of these findings with respect to public health and research methodology and propose priority research needs.

Determinants of health-seeking behaviour for schistosomiasis-related symptoms in the context of integrating schistosomiasis control within the regular health services in Ghana.

Morbidity control of schistosomiasis through integration within existing health care delivery systems is considered a potentially sustainable and cost-effective approach. We conducted a questionnaire-based field study in a Ghanaian village endemic for both urinary and intestinal schistosomiasis to determine whether infected individuals self-reported to health centres or clinics and to identify factors that influenced their decision to seek health care. A total of 317 subjects were interviewed about having signs and symptoms suggestive of schistosomiasis: blood in urine, painful urination, blood in stool/bloody diarrhoea, abdominal pain, diarrhoea, swollen abdomen and fatigue within 1 month of the day of the interview. Fever (for malaria) was included as a disease of high debility for comparison. Around 70% with blood in urine or painful urination did not seek health care, whilst diarrhoea, blood in stool, abdominal pain and fever usually led to action (mainly self-medication, with allopathic drugs being used four to five times more often than herbal treatment). On average 20% of schistosomiasis-related signs and symptoms were reported to health facilities either as the first option or second and third alternative by some of those that self-medicated. A few of those who visited a clinic or health centre as first option still self-medicated afterwards. Children under 10 years and adults were more likely to seek health care than teenagers. Also, females were more likely to visit a health facility than males of the same age groups. Socio-economic status and duration of symptoms did not appear to affect health-seeking behaviour. 'Do not have the money' (43%) and 'Not serious enough' (41%) were the commonest reasons for not visiting a clinic, reported more frequently by lower and higher socio-economic classes, respectively, for both urinary or intestinal schistosomiasis. The regular health service shows some potential in passive control of schistosomiasis as some, but far too few, people visit a health facility as first or second option.

Oomycete plant pathogens use electric fields to target roots.

Plant roots generate electrical currents and associated electrical fields as a consequence of electrogenic ion transport at the root surface. Here we demonstrate that the attraction of swimming zoospores of oomycete plant pathogens to plant roots is mediated in part by electrotaxis in natural root-generated electric fields. The zones of accumulation of anode- or cathode-seeking zoospores adjacent to intact and wounded root surfaces correlated with their in vitro electrotactic behavior. Manipulation of the root electrical field was reflected in changes in the pattern of zoospore accumulation and imposed focal electrical fields were capable of overriding endogenous signals at the root surface. The overall pattern of zoospore accumulation around roots was not affected by the presence of amino acids at concentrations expected within the rhizosphere, although higher concentrations induced encystment and reduced root targeting. The data suggest that electrical signals can augment or override chemical ones in mediating short-range tactic responses of oomycete zoospores at root surfaces.

Improved production of adenovirus vectors expressing apoptotic transgenes.

Adenovirus vectors expressing gene products that can induce apoptosis have potential utility in gene therapy applications ranging from the treatment of proliferative diseases to transplantation. However, adenovirus vectors carrying proapoptotic gene products are difficult to produce, as the apoptotic environment is not conducive to adenovirus gene expression and replication. Production of AdFasL/G, an adenovirus vector that expresses high levels of Fas ligand, was severely reduced in the 293 packaging cell line. Increased yields of AdFasL/G were achieved by inclusion of peptide-based caspase inhibitors in the growth medium. However, use of these inhibitors for large-scale production would be difficult and expensive. A screen for gene products that increase the yield of AdFasL/G in 293 cells revealed that the poxvirus serpin CrmA and the adenovirus 14.7K product were able to increase virus yields significantly. Apoptosis induced by AdFasL/G was attenuated in 293CrmA cell lines and virus titers were increased dramatically. However, serial passage of AdFasL/G on 293CrmA cells resulted in the generation of replication-competent adenovirus. To resolve this problem, the CrmA gene was introduced into AE25 cells, an E1-complementing cell line that has limited sequence identity with the vectors. AdFasL/G titers were increased 100-fold on AE25CrmA cells relative to the AE25 cells and RCA contamination was not detectable. In addition, adenovirus vectors that express FADD, caspase 8, and Fas/APO1 were produced efficiently in AE25CrmA and 293CrmA.

Plasmodium falciparum: stage-specific ribosomal RNA as a potential target for monitoring parasite development in Anopheles stephensi.

The transcriptional switch of Plasmodium falciparum ribosomal RNA A gene to the C gene was demonstrated during the developmental transition from the vertebrate blood stage to the invertebrate sporozoite stage. Expression of the sporozoite specific C gene in infected mosquitoes was not detected until Day 10 postinfectious blood meal, the time of mature oocyst formation on the midgut. As a potential target for monitoring malaria parasite development in mosquitoes, oligonucleotide probes based on sequences of small subunit ribosomal RNA were evaluated for specificity and sensitivity by filter blot hybridization against different species and stages of malaria parasites. Probes PfC02 and PfA02 were selected as the most sensitive for sporozoite and blood stage parasites, respectively. Filter blot hybridization using probe PfC02 resulted in sensitivity comparable with microscopic dissection in single mosquitoes, detecting mosquitoes with an average of 1.2 oocysts per gut or as few as 800 salivary gland sporozoites.

The nature, causes, and visual outcome of ocular trauma requiring posterior segment surgery at a county hospital.

Fifty consecutive, severe ocular trauma cases with posterior segment involvement underwent vitreoretinal surgery over a 14-month period at a county hospital. Forty-five (90%) of the patients were boys and men, and the mean age was 27 years. The injuries were the result of assaults in 37 cases (74%), industrial accidents in six (12%), accidents involving children at play in six (12%), and child abuse in one (2%). Twenty-five (50%) of the injuries were inflicted by firearms, by knives or sharp objects in ten (20%), by baseball bats in two (4%), by broken glass in two (4%), and unknown in 11 (22%). Twenty-six (52%) were penetrating injuries, of which eight were double perforations. There were ten (20%) intraorbital foreign bodies and three (6%) intraocular foreign bodies. Forty-three (86%) of the cases had initial visual acuities of 20/400 or worse. After a minimum of six months' follow-up, 48.8% improved two or more lines visually, and 69.4% were anatomic successes. Unlike other ocular-trauma studies, where industrial accidents and sports-related injuries were the most common causes of the trauma, assaults with firearms were the most common cause of ocular trauma at this county hospital. The visual and anatomic surgical results, however, were similar to those of other ocular-trauma studies.

Plasmodium berghei: quantitation of in vitro effects of antimalarial drugs on exoerythrocytic development by a ribosomal RNA probe.

A stage-specific ribosomal RNA probe has been used to quantitate exoerythrocytic development of Plasmodium berghei in primary cultures of mouse hepatocytes. Parasite rRNA could be detected as soon as 6 hr after sporozoite invasion and was increased during schizogony to a maximum at 48 hr, when mature schizonts were identified by microscopy. As few as 10 exoerythrocytic schizonts could be detected by filter blot hybridization, followed by autoradiography and liquid scintillation counting. By hybridizing the culture rRNA samples with either parasite-specific or universal rRNA probes, the in vitro tissue schizonticidal activity and hepatotoxicity of primaquine, two of its analogues, and pyrimethamine, could be assessed. After a 48-hr exposure of the culture to serial dilutions of each drug, a quantitative relationship was demonstrated between the decrease of the parasite rRNA and the increase of the drug concentrations. No significant parasite-specific rRNA could be detected at the concentration achieving complete inhibition of schizont formation but causing no cytotoxic effects on host hepatocytes. In contrast to microscopic-based assays, this molecular approach provides an objective and quantitative in vitro method for rapid screening and evaluation of tissue schizonticidal antimalarials.

Delayed diagnosis of diabetic retinopathy in black and Hispanic patients with diabetes mellitus.

We examined 51 black and 35 Hispanic consecutive patients with noninsulin-dependent diabetes mellitus from similar socioeconomic backgrounds who were evaluated initially for diabetic retinopathy. The mean ages were 58 years for the Hispanics and 53 years for the blacks (P greater than .05). Sixty-five percent of the Hispanics were women, as were 68.6% of the blacks (P greater than .05). The mean duration between the diagnosis of the diabetes mellitus and the initial diagnosis of the retinopathy was 11.5 years for the Hispanics and 12.2 years for the blacks (P = .671). The percentage of patients with severe diabetic retinopathy (preproliferative or proliferative) at the time of initial diagnosis was 42.9% for the Hispanics and 37.3% for the blacks (P greater than .10). Although no significant differences were found between the two groups, the high percentage of patients in both groups (42.9% and 37.3%) with severe diabetic retinopathy at the time of initial presentation suggests that earlier patient referral from primary-care physicians and better patient education about the need for earlier eye examinations should be encouraged in these two groups.