Cytokine profiles of filarial granulomas in jirds infected with Brugia pahangi

BACKGROUND: A granulomatous inflammatory response develops in jirds infected subcutaneously or intraperitoneally with filarial nematodes namely Brugia pahangi and B. malayi. Previous studies by light and electron microscopy have shown cellular inflammatory responses in and around these granulomas. Furthermore, the cellular inflammatory responses of granulomas found in the lymphatics and peritoneal cavity appear to be similar. The purpose of this study was to determine the cytokine profiles of granulomas in the peritoneal cavity of B. pahangi-infected jirds and to determine whether the granulomas release any proinflammatory cytokines ex vivo. METHODS: A semiautomated quantitative polymerase chain reaction (Q-PCR) was performed on cDNA prepared from the granulomas of infected jirds to study the species-specific mRNA expression of IL-2, IL-4, IFN-γ, IL-5, and IL-10. Genomic DNA was extracted from the granulomas, and parasite DNA was detected by Q-PCR by amplifying the HhaI repeat sequence. The levels of the inflammation-causing cytokines IL-6 and TNFα that were secreted by the granulomas were measured by cell-based assays. RESULTS: Florid granulomas showed higher levels of IFN-γ than other cytokines, linking this Th1 cytokine to the granulomatous inflammation that develops in jirds and humans. IL-4 expression was much lower than that of IFN-γ but higher than that of IL-10. A low level of IL-5 mRNA expression was detectable in all granulomas as was the level of IL-2 expression. The levels of the inflammatory cytokines IL-6 and TNFα, secreted by intact granulomas, spontaneously increased by 48 h after culture. Parasite antigen stimulation and subsequent release of IL-6 and TNFα by the granulomas indicated a moderate increase in the levels of these two cytokines. The amplification of the Brugia HhaI repeat DNA and Wolbachia 16S rDNA indicated worm components and bacterial components in the granulomatous tissue. CONCLUSION: Granuloma development in filarial infections is a complex process involving cellular reactions responding to parasite/bacteria and their components. The interactions between worm-derived granulomas and their hosts are dynamic and multifaceted. The data collected thus far suggest that the expression profiles of many of the measured cytokines in the lymphoid tissues of Brugia-infected jirds are different from those of the cytokines in granulomas. Moreover, granulomas have the ability to secrete the inflammatory cytokines IL-6 and TNFα

Using existing drugs as leads for broad spectrum anthelmintics targeting protein kinases

As one of the largest protein families, protein kinases (PKs) regulate nearly all processes within the cell and are considered important drug targets. Much research has been conducted on inhibitors for PKs, leading to a wealth of compounds that target PKs that have potential to be lead anthelmintic drugs. Identifying compounds that have already been developed to treat neglected tropical diseases is an attractive way to obtain lead compounds inexpensively that can be developed into much needed drugs, especially for use in developing countries. In this study, PKs from nematodes, hosts, and DrugBank were identified and classified into kinase families and subfamilies. Nematode proteins were placed into orthologous groups that span the phylum Nematoda. A minimal kinome for the phylum Nematoda was identified, and properties of the minimal kinome were explored. Orthologous groups from the minimal kinome were prioritized for experimental testing based on RNAi phenotype of the Caenorhabditis elegans ortholog, transcript expression over the life-cycle and anatomic expression patterns. Compounds linked to targets in DrugBank belonging to the same kinase families and subfamilies in the minimal nematode kinome were extracted. Thirty-five compounds were tested in the non-parasitic C. elegans and active compounds progressed to testing against nematode species with different modes of parasitism, the blood-feeding Haemonchus contortus and the filarial Brugia malayi. Eighteen compounds showed efficacy in C. elegans, and six compounds also showed efficacy in at least one of the parasitic species. Hypotheses regarding the pathway the compounds may target and their molecular mechanism for activity are discussed

Effects of Doxycycline on gene expression in Wolbachia and Brugia malayi adult female worms in vivo

<p>Abstract</p> <p>Background</p> <p>Most filarial nematodes contain <it>Wolbachia </it>symbionts. The purpose of this study was to examine the effects of doxycycline on gene expression in <it>Wolbachia </it>and adult female <it>Brugia malayi</it>.</p> <p>Methods</p> <p><it>Brugia malayi </it>infected gerbils were treated with doxycycline for 6-weeks. This treatment largely cleared <it>Wolbachia </it>and arrested worm reproduction. RNA recovered from treated and control female worms was labeled by random priming and hybridized to the Version 2- filarial microarray to obtain expression profiles.</p> <p>Results and discussion</p> <p>Results showed significant changes in expression for 200 <it>Wolbachia </it>(29% of <it>Wolbachia </it>genes with expression signals in untreated worms) and 546 <it>B. malayi </it>array elements after treatment. These elements correspond to known genes and also to novel genes with unknown biological functions. Most differentially expressed <it>Wolbachia </it>genes were down-regulated after treatment (98.5%). In contrast, doxycycline had a mixed effect on <it>B. malayi </it>gene expression with many more genes being significantly up-regulated after treatment (85% of differentially expressed genes). Genes and processes involved in reproduction (gender-regulated genes, collagen, amino acid metabolism, ribosomal processes, and cytoskeleton) were down-regulated after doxycycline while up-regulated genes and pathways suggest adaptations for survival in response to stress (energy metabolism, electron transport, anti-oxidants, nutrient transport, bacterial signaling pathways, and immune evasion).</p> <p>Conclusions</p> <p>Doxycycline reduced <it>Wolbachia </it>and significantly decreased bacterial gene expression. <it>Wolbachia </it>ribosomes are believed to be the primary biological target for doxycycline in filarial worms. <it>B. malayi </it>genes essential for reproduction, growth and development were also down-regulated; these changes are consistent with doxycycline effects on embryo development and reproduction. On the other hand, many <it>B. malayi </it>genes involved in energy production, electron-transport, metabolism, anti-oxidants, and others with unknown functions had increased expression signals after doxycycline treatment. These results suggest that female worms are able to compensate in part for the loss of <it>Wolbachia </it>so that they can survive, albeit without reproductive capacity. This study of doxycycline induced changes in gene expression has provided new clues regarding the symbiotic relationship between <it>Wolbachia </it>and <it>B. malayi</it>.</p

Effect of improved crop production technology on pigeonpea yield in resource poor rainfed areas.

Twelve on-farm trials were conducted in Nandavaram village of Banaganapalle mandal in Kurnool district, Andhra Pradesh, India, during the 2002/03 rainy season to study the effects of improved production technologies (T1) and farmers' practice (T2) on pigeon pea yield. T1 consisted of a medium-duration high-yielding cultivar (ICPL 87119), sowing rate of 12 kg/ha, seed treatment with thiram (3 g/kg), inoculation with Rhizobium, 20 kg N, 50 kg P2O5/ha, basal application of micronutrient mixture (5 kg borax, 0.5 kg B/ha; 50 kg zinc sulfate, 10 kg Zn/ha; and 200 kg gypsum, 30 kg S/ha), and need-based pest and disease control measures. Inter-cultivation was conducted at 25 and 50 days after sowing to control weeds. One insecticide spray was given at the pod formation stage to control pod borers. T2 included a sowing rate of 10 kg/ha, 12 kg N/ha, and 30 kg P2O5/ha. T1 gave higher yields than T2 and recorded a mean grain yield of 1.61 t/ha, which was 204% higher than that obtained with T2 (0.53 t/ha). T1 also resulted in higher stalk yield (2.93 t/ha) than T2 (1.10 t/ha). The increased grain and stalk yields under were mainly because of increased total dry matter, pod weight, shelling percentage, 100-grain weight and harvest index. T1 recorded a higher mean income (US$290) and a cost-benefit ratio of 2.4

Increased chickpea yield and economic benefits by improved crop production technology in rainfed areas of Kurnool District of Andhra Pradesh, India

Sixteen on-farm trials in 2002 and 9 trials in 2003 were conducted in Nandavaram and Jillella villages of Banaganapalle mandal in Kurnool district, Andhra Pradesh, India, during the post-rainy season to evaluate the performance of improved production technologies (T1) and farmers' practice (T2) for chickpea production. T1 consisted of improved cultivar (ICCC 37), sowing rate of 60 kg/ha, seed treatment with thiram (3 g/kg), inoculation with Rhizobium, 20 kg N, 50 kg P2O5/ha, basal application of micronutrient mixture (5 kg borax, 0.5 kg B/ha; 50 kg zinc sulfate, 10 kg Zn/ha; and 200 kg gypsum, 30 kg S/ha), and need-based pest and disease control measures. Inter-cultivation was conducted at 25 and 50 days after sowing to control weeds. One insecticide spray was given at the pod formation stage to control pod borers. T2 consisted of a local cultivar, sowing rate of 50 kg/ha, 14 kg N/ha, and 35 kg P2O5/ha. T1 gave higher grain yields and recorded a mean yield of 2.09 t/ha, which was 53% higher than that obtained with T2 (1.37 t/ha). The increased grain yield under T1 was mainly because of greater total dry matter, 100-grain weight and harvest index. T1 also resulted in increased mean income of US$190 and a cost-benefit ratio of 2.9

Persistence of Brugia malayi DNA in vector and non-vector mosquitoes: Implications for xenomonitoring and transmission monitoring of lymphatic filariasis

Abstract. Xenomonitoring (detection of filarial larvae or their DNA in mosquitoes) is a sensitive marker for as-sessing the endemicity of filariasis and a useful tool for evaluating elimination programs. To examine the fate of microfilariae (mf) and filarial DNA in vector competent and non-competent mosquito strains, we compared the detec-tion of Brugia malayi parasites by dissection and by quantitative real-time polymerase chain reaction (PCR) in three different mosquito strains. We conclude that PCR is much more sensitive than dissection for detecting filarial larvae, especially their remnants in mosquitoes. However, parasite DNA can be detected in both vector and non-vector mosquitoes for two weeks or longer after they ingest mf-positive blood. Thus, although xenomonitoring with vector and non-vector mosquito species may be a sensitive method for indirectly detecting filarial parasites in human populations, positive test results for parasite DNA in mosquitoes do not necessarily prove that transmission is ongoing in the study area

Portable infrared imaging for longitudinal limb volume monitoring in patients with lymphatic filariasis

BACKGROUND: The Global Programme to Eliminate Lymphatic Filariasis (LF) emphasizes hygiene, exercise, and other measures to reduce morbidity and disability related to LF. We recently reported that a portable, three-dimensional, infrared imaging system (3DIS) provides accurate limb volume measurements in patients with filarial lymphedema. To assess the practical utility of repeated 3DIS measurements for longitudinal lymphedema management, we examined intraday and day-to-day leg volume changes in adults with filarial lymphedema in southern Sri Lanka. METHODOLOGY AND PRINCIPAL FINDINGS: We assessed 41 participants with lower extremity lymphedema (stages 1-6) in their homes in the mornings (6:00-9:00 AM) and afternoons (2:00-6:00 PM) of three days within one calendar week. Two examiners performed replicate 3DIS volume measurements at each visit. Median coefficient of variation among replicate volume measurements was 1.7% (IQR 1.1% - 2.3%) for left legs and 2.2% (IQR 1.6% - 2.8%) for right legs. Median intraday volume increase was 3.0%. Range among daily volume measurements tended to be lower for afternoon measurements (median 2.25%, IQR 1.4%- 5.4%) than for morning measurements (median 3.0%, IQR 1.4% - 8.4%). CONCLUSIONS AND SIGNIFICANCE: Limb volume measurements by 3DIS are accurate and reproducible, and this technique is feasible for use in patients\u27 homes. We have developed practical suggestions for optimal outcomes with 3DIS. Duplicate measurements should be performed and repeat assessments should be done at approximately the same time of day to minimize bias. Duplicate measures that vary by more than 8% should prompt review of scanning technique with a repeat measurement. With proper training and attention to technique, 3DIS can be a valuable tool for healthcare workers who work with lymphedema patients