Search

Several secreted proteins from helminths (parasitic worms) have been shown to have immunomodulatory activities. Asparaginyl-tRNA synthetases are abundantly secreted in the filarial nematode Brugia malayi ( Bm AsnRS) and the parasitic flatworm Schistosoma japonicum ( Sj AsnRS), indicating a possible immune function. The suggestion is supported by Bm AsnRS alleviating disease symptoms in a T-cell transfer mouse model of colitis. This immunomodulatory function is potentially related to an N-terminal extension domain present in eukaryotic AsnRS proteins but few structure/function studies have been done on this domain. Here we have determined the three-dimensional solution structure of the N-terminal extension domain of Sj AsnRS. A protein containing the 114 N-terminal amino acids of Sj AsnRS was recombinantly expressed with isotopic labelling to allow structure determination using 3D NMR spectroscopy, and analysis of dynamics using NMR relaxation experiments. Structural comparisons of the N-terminal extension domain of Sj AsnRS with filarial and human homologues highlight a high degree of variability in the β-hairpin region of these eukaryotic N-AsnRS proteins, but similarities in the disorder of the C-terminal regions. Limitations in PrDOS-based intrinsically disordered region (IDR) model predictions were also evident in this comparison. Empirical structural data such as that presented in our study for N- Sj AsnRS will enhance the prediction of sequence-homology based structure modelling and prediction of IDRs in the future.Communicated by Ramaswamy H. Sarma. more...

Abstract: A patient with unilateral cervical lymphadenopathy suspicious for malignancy underwent a fine needle aspiration. Histology demonstrated mixed inflammatory infiltrates with abundant spirochetes. Sufficient spirochete DNA was extracted from paraffin-embedded tissue sections to obtain the near-complete genome sequence of a macrolide-resistant strain belonging to the SS14 omega strain of Treponema pallidum ., Competing Interests: Conflict of Interest and Sources of Funding: None declared., (Copyright © 2023 American Sexually Transmitted Diseases Association. All rights reserved.) more...

The conference session on Postgraduate Education in Biochemistry and Molecular Biology consisted of wide-ranging presentations and discussions. Approaches, issues, and solutions for postgraduate education and training in countries ranging from the Philippines to Mongolia and the United States were covered., (© 2020 International Union of Biochemistry and Molecular Biology.) more...

We have recently isolated tirandamycin (TAM) B from Streptomyces sp. 17944 as a Brugia malayi AsnRS (BmAsnRS) inhibitor that efficiently kills the adult B. malayi parasites and does not exhibit general cytotoxicity to human hepatic cells. We now report (i) the comparison of metabolite profiles of S. sp. 17944 in six different media, (ii) identification of a medium enabling the production of TAM B as essentially the sole metabolite, and with improved titer, and (iii) isolation and structural elucidation of three new TAM congeners. These findings shed new insights into the structure-activity relationship of TAM B as a BmAsnRS inhibitor, highlighting the δ-hydroxymethyl-α,β-epoxyketone moiety as the critical pharmacophore, and should greatly facilitate the production and isolation of sufficient quantities of TAM B for further mechanistic and preclinical studies to advance the candidacy of TAM B as an antifilarial drug lead. The current study also serves as an excellent reminder that traditional medium and fermentation optimization should continue to be very effective in improving metabolite flux and titer. more...

The therapeutic effects of a controlled parasitic nematode infection on the course of inflammatory bowel disease (IBD) have been demonstrated in both animal and human models. However, the inability of individual well-characterized nematode proteins to recreate these beneficial effects has limited the application of component immunotherapy to human disease. The nematodes that cause chronic human lymphatic filariasis, Brugia malayi and Wuchereria bancrofti, are among the parasites that induce immune suppression. Filarial lymphatic pathology has been shown to involve NF-κB pathway-dependent production of vascular endothelial growth factor (VEGF), and stimulation of VEGF expression has also been reported by interleukin 8 (IL-8) via NF-κB pathways. Previously, we have shown that the filarial asparaginyl-tRNA synthetase (rBmAsnRS) interacts with IL-8 receptors using a combination of extracellular loops that differ from those bound by IL-8. To test the hypothesis that rBmAsnRS might induce an anti-inflammatory effect in vivo, we studied the effects of rBmAsnRS in an established murine colitis model using T-cell transfer mice. T-cell transfer colitis mice treated intraperitoneally with 100 μg of rBmAsnRS four times over 2 weeks showed resolution of cellular infiltration in the colonic mucosa, along with induction of a CD8(+) cellular response. In addition, rBmAsnRS induced a rise in IL-10 production from CD3(+) and lipopolysaccharide (LPS)- and cytosine phosphate guanosine (CPG)-stimulated splenic cells. In summary, this work demonstrates a novel anti-inflammatory nematode protein, supports the hygiene hypothesis, and supports continued refinement of alternative immunotherapies for treatment of IBD. more...

A wide range of secondary biological functions have been documented for eukaryotic aminoacyl-tRNA synthetases including roles in transcriptional regulation, mitochondrial RNA splicing, cell growth, and chemokine-like activities. The asparaginyl-tRNA synthetase (AsnRS) of the filarial nematode, Brugia malayi, is a highly expressed excretory-secretory molecule which activates interleukin 8 (IL-8) receptors via extracellular domains that are different from those used by IL-8. Recent success in determining the complete atomic structure of the B. malayi AsnRS provided the opportunity to map its chemokine-like activity. Chemotaxis assays demonstrated that IL-8-like activity is localized in a novel 80 amino acid amino terminal substructure. Structural homology searches revealed similarities between that domain in B. malayi AsnRS and substructures involved in receptor binding by human IL-8. These observations provide important new insights into how parasite-derived molecules may play a role in the modulation of immune cell function., (Copyright © 2012. Published by Elsevier B.V.) more...

Aminoacyl-tRNA synthetases (AARSs) are a structurally heterogeneous family of enzymes present in prokaryotes, archaea and eukaryotes. They catalyze the attachment of tRNA to its corresponding amino acid via an aminoacyl adenylate intermediate. Errors in protein synthesis will occur if an incorrect amino acid is attached to the tRNA. To prevent such errors, AARSs have evolved editing mechanisms that eliminate incorrect aminoacyl adenylates (pre-transfer editing) or misacylated tRNAs (post-transfer editing). Various AARSs are the targets of natural antibiotics and are considered validated targets for chemotherapy. We have developed a high-throughput screening (HTS) assay measuring the pre-transfer editing activity of pathogen-derived asparaginyl-tRNA synthetase (AsnRS). This was achieved by monitoring the formation of pyrophosphate via cleavage to phosphate, which was quantified by reaction with Malachite Green. L-Aspartate-β-hydroxamate, an asparagine analogue, was most effective in promoting the editing activity of AsnRS from Brugia malayi (BmAsnRS) and Staphylococcus epidermidis (SeAsnRS) with KM values close to 100 mM. The assay sensitivity was enhanced by the thiol agents, DTT and L-Cysteine, which significantly increased the turn-over of aminoacyl adenylate by BmAsnRS, but not SeAsnRS. The HTS assay was used to screen a library of 37,120 natural-product extracts for inhibitors of BmAsnRS. A small number of extracts that inhibited the pre-transfer editing by BmAsnRS was identified for future isolation of the active component(s). The principle of this assay can be applied to all enzymes having a pre- or post-editing activity. more...

Human cytosolic seryl-tRNA synthetase (hsSerRS) is responsible for the covalent attachment of serine to its cognate tRNA(Ser). Significant differences between the amino-acid sequences of eukaryotic, prokaryotic and archaebacterial SerRSs indicate that the domain composition of hsSerRS differs from that of its eubacterial and archaebacterial analogues. As a consequence of an N-terminal insertion and a C-terminal extra-sequence, the binding mode of tRNA(Ser) to hsSerRS is expected to differ from that in prokaryotes. Recombinant hsSerRS protein was purified to homogeneity and crystallized. Diffraction data were collected to 3.13 Å resolution. The structure of hsSerRS has been solved by the molecular-replacement method. more...

Geldanamycin (GA) is a benzoquinone-containing ansamycin that inhibits heat shock protein 90. GA derivatives are being evaluated as anti-neoplastic agents, but their utility against parasites whose heat shock proteins (Hsps) have homology with human Hsp90 is unknown. The activities of four synthetic GA derivatives were tested in vitro using adult Brugia malayi and Schistosoma japonicum. Two of the derivatives, 17-N-allyl-17-demethoxygeldanamycin (17-AAG) and 17-N-(2-dimethylaminoethylamino)-17-demethoxygeldanamycin (DMAG), are currently in human clinical trials as anticancer drugs. Using concentrations considered safe peak plasma concentrations for these two derivatives, all four derivatives were active against both parasites. The less toxic derivative 17-AAG was as effective as GA in killing S. japonicum, and both DMAG and 5'-bromogeldanoxazinone were more active than 17-AAG against B. malayi. This work supports continued evaluation of ansamycin derivatives as broad spectrum antiparasitic agents. more...

Diagnosis of eukaryotic parasitic infection using antibody-based tests such as ELISAs (enzyme-linked immunosorbent assays) is often problematic because of the need to differentiate between homologous host and pathogen proteins and to ensure that antibodies raised against a peptide will also bind to the peptide in the context of its three-dimensional protein structure. Filariasis caused by the nematode, Brugia malayi, is an important worldwide tropical disease in which parasites disappear from the bloodstream during daylight hours, thus hampering standard microscopic diagnostic methods. To address this problem, a structural approach was used to develop monoclonal antibodies (mAbs) that detect asparaginyl-tRNA synthetase (AsnRS) secreted from B. malayi. B. malayi and human AsnRS amino acid sequences were aligned to identify regions that are relatively unconserved, and a 1.9 A crystallographic structure of B. malayi AsnRS was used to identify peptidyl regions that are surface accessible and available for antibody binding. Sequery and SSA (Superpositional Structural Analysis) software was used to analyze which of these peptides was most likely to maintain its native conformation as a synthetic peptide, and its predicted helical structure was confirmed by NMR. A 22-residue peptide was synthesized to produce murine mAbs. Four IgG(1) mAbs were identified that recognized the synthetic peptide and the full-length parasite AsnRS, but not human AsnRS. The specificity and affinity of mAbs was confirmed by Western blot, immunohistochemistry, surface plasmon resonance, and enzyme inhibition assays. These results support the success of structural modeling to choose peptides for raising selective antibodies that bind to the native protein. more...

Proteins often have multiple functional states, which might not always be accommodated by a single fold. Lymphotactin (Ltn) adopts two distinct structures in equilibrium, one corresponding to the canonical chemokine fold consisting of a monomeric three-stranded beta-sheet and carboxyl-terminal helix. The second Ltn structure solved by NMR reveals a dimeric all-beta-sheet arrangement with no similarity to other known proteins. In physiological solution conditions, both structures are significantly populated and interconvert rapidly. Interconversion replaces long-range interactions that stabilize the chemokine fold with an entirely new set of tertiary and quaternary contacts. The chemokine-like Ltn conformation is a functional XCR1 agonist, but fails to bind heparin. In contrast, the alternative structure binds glycosaminoglycans with high affinity but fails to activate XCR1. Because each structural species displays only one of the two functional properties essential for activity in vivo, the conformational equilibrium is likely to be essential for the biological activity of lymphotactin. These results demonstrate that the functional repertoire and regulation of a single naturally occurring amino acid sequence can be expanded by access to a set of highly dissimilar native-state structures. more...

SLIDE software, which models the flexibility of protein and ligand side chains while docking, was used to screen several large databases to identify inhibitors of Brugia malayi asparaginyl-tRNA synthetase (AsnRS), a target for anti-parasitic drug design. Seven classes of compounds identified by SLIDE were confirmed as micromolar inhibitors of the enzyme. Analogs of one of these classes of inhibitors, the long side-chain variolins, cannot bind to the adenosyl pocket of the closed conformation of AsnRS due to steric clashes, though the short side-chain variolins identified by SLIDE apparently bind isosterically with adenosine. We hypothesized that an open conformation of the motif 2 loop also permits the long side-chain variolins to bind in the adenosine pocket and that their selectivity for Brugia relative to human AsnRS can be explained by differences in the sequence and conformation of this loop. Loop flexibility sampling using Rigidity Optimized Conformational Kinetics (ROCK) confirms this possibility, while scoring of the relative affinities of the different ligands by SLIDE correlates well with the compounds' ranks in inhibition assays. Combining ROCK and SLIDE provides a promising approach for exploiting conformational flexibility in structure-based screening and design of species selective inhibitors. more...

Lymphatic filariasis (elephantiasis) is a global public health problem caused by the parasitic nematodes Wuchereria bancrofti and Brugia malayi. We have previously reported anthraquinones from daylily roots with potent activity against pathogenic trematode Schistosoma mansoni. Here we report the synthesis of novel anthraquinones A-S and their antifilrarial activity. Anthraquinones A-S were synthesized by a single-step Friedel-Crafts acylation reaction between phthalic anhydrides and substituted benzenes. The antifilarial properties of these synthetic anthraquinones were tested against microfilaria as well as adult male and female worms of B. malayi. The most active anthraquinone was K, which showed 100% mortality within 1, 5, and 3 days, respectively, against microfilaria and adult male and female worms at 5 ppm concentration. Albendazole, an oral drug currently used to treat parasitic infections, was used as a positive control. Methylated products of anthraquinones did not affect the microfilaria. Histological examination of treated adult female parasites showed most of the anthraquinones caused marked effects on intrauterine embryos. more...

The etiologies of most autoimmune diseases are not completely understood. Aminoacyl-tRNA synthetases (AARS) are a family of heterogenous enzymes responsible for protein synthesis and whose secondary functions include a role in autoimmune myositis. A subset of patients with idiopathic inflammatory myopathies demonstrate autoantibody against specific cytoplasmic AARS and the human asparaginyl-tRNA synthetase (AsnRS) has been shown to be a potent chemokine that interacts with CCR3 chemokine receptors. One way in which a chemotactic cytoplasmic enzyme might contribute to tissue inflammation is if it were abundant in a specific injured tissue and thereby released to the microenvironment at times of cellular damage. To test this hypothesis, the relative levels of AsnRS mRNA were studied in six human tissues. A 1.6 kbF RNA probe identified highly variable levels of the corresponding mRNA in Northern blot analysis of human lung, brain, heart, skeletal muscle, pancreas and liver. The highest levels of signal were noted in muscle and pancreas. Polyclonal antibody raised against recombinant human AsnRS identified abundant antigenic material in the pancreas, in particular in islet cells. Thus, the local abundance of an endogenous pro-inflammatory autoantigen may provide one explanation for perpetuation or exacerbation of tissue specific immune-mediated pathologies. more...

Lymphatic filariasis caused by infection with Wuchereria bancrofti and Brugia malayi is endemic in 45 of 77 provinces in The Philippines. To prepare the island of Mindoro for mass treatment using diethylcarbamazine and albendazole, complete census data were collected in rural villages. A sample of individuals selected from each of two adjacent villages was examined for microfilaremia. Microfilariae were detected from thin smears in 34 (13%) of 272 patients examined from the village of Bayanan and 10 (3.4%) of 292 in the village of Mangangan (P < 0.01, by chi-square test). In these villages, the majority of those infected were members of the ethnic group known as Mangyans: 33 (97%) of 24 in Bayanan and 7 (70%) of 10 in Mangangan (risk ratio = 89, 95% confidence interval = 33-240, P < 0.001.) In children examined who were less than 10 years of age (n = 165), girls were more commonly infected than boys, even though the proportion of males in the general population was greater. Understanding sociocultural characteristics and related behaviors in future observations among the Mangyan may help to explain local differences in the distribution of filariasis. This information should also be helpful in designing more culturally appropriate strategies for the control of lymphatic filariasis among ethnic minorities in The Philippines. more...

Hyperimmunoglobulinemia E (HIGE) is associated with various conditions such as atopy, dermatitis, hypersensitivity reactions, and certain parasitic infections. In the course of vaccination initiatives in the province of Napo, eastern Ecuador, blood samples were collected from one of the two remaining rural subgroups of Huaorani Indians who in 1979 were reported to have the world's highest concentrations of IgE. One subgroup of Huaorani, the Dicaron, lives in a protected Amazonian region which has reportedly suffered from extensive pollution after petroleum industry exploration. Plasma was collected from 31 members of the Dicaron (age range 15-75 years), eight non-Dicaron Huaorani, and 16 Quichua Indians from the same province, and tested for IgE, IgG, IgM, IgA, and immunoglobulin allotypes. Subjects were examined for evidence of filariasis, a group of parasitic diseases associated with HIGE. Mean IgE concentration in the Dicaron was measured by CAP ELISA at 11,850 IU/mL (range 5000-33,000) while IgA and IgM concentrations were within normal limits compared to North American controls. IgG levels were slightly elevated and there was no evidence of filariasis. Compared to the Quichua and non-Dicaron Huaorani, two other Amerindian tribes in the Ecuadorian Amazon, the highest concentrations of IgE were recorded from the Dicaron who live within the allegedly polluted section of the Amazon. We conclude that an unexplained HIGE syndrome exists among only one subgroup of Huaorani, the Dicaron. Other eastern Ecuadorian Amerindians, such as the Quichua and resettled Huaorani, have IgE concentrations expected in a population with intestinal helminthiasis. Environmental factors cannot be excluded as the cause of HIGE in the Dicaron. more...

Autoantibodies to five of the aminoacyl-transfer RNA (tRNA) synthetases have been described, and each is associated with a syndrome of inflammatory myopathy with interstitial lung disease (ILD) and arthritis. Serum KS, from a patient with ILD and inflammatory arthritis without evidence of myositis, immunoprecipitated a tRNA that was distinct from that precipitated by any described anti-synthetase or other reported tRNA-related Abs, along with a protein of 65 kDa. KS serum and IgG fraction each showed significant (88%) inhibition of asparaginyl-tRNA synthetase (AsnRS) activity, but not of any of the other 19 aminoacyl-tRNA synthetase activities. Among 884 patients with connective tissue diseases tested, only two other sera were found to immunoprecipitate tRNAs and proteins of identical gel mobility. These two and KS showed identical immunodiffusion lines using HeLa cell extract. The new sera significantly inhibited AsnRS without significant effects on other synthetases tested. Both patients had ILD but neither had evidence of myositis. These data strongly suggest that these three sera have autoantibodies to AsnRS, representing a sixth anti-synthetase. Anti-KS was more closely associated with ILD than with myositis. Further study of this Abs might prove useful in dissecting the stimuli responsible for the genesis of anti-synthetase autoantibodies. more...

Immunoglobulin (Ig) allotypes are polymorphic genetic systems that show distinct racial arrays, thus making them powerful tools for studies of genetic admixture and biological relationships. In the province of Sorsogon, southern Luzon, Philippines, allotyping was completed on 252 persons residing in two neighboring villages. The people demonstrated 14 GM and 3 KM phenotypes. The frequency of homozygous KM3, KM1 and heterozygous KM1,3 was identical in these villages; however, half of the GM phenotypes present in one village were significantly less frequent in the other village. The frequency of KM and GM haplotypes was different from those reported in Filipino aboriginal groups, but similar to a population on Samar Island, the only other Filipino group for which Ig allotype data exist. Variability in the prevalence of parasitic disease such as lymphatic filariasis may in part reflect differences in genetic susceptibility, resulting from allotypic heterogeneity between villages. more...

The republic of Ecuador, which has a population of 10 million, is one of the smallest of the Andean countries in South America. Although it covers only an area of 110,000 square miles, it yields an extraordinary diversity of infectious diseases. Public health problems reflect socioeconomic realities and uniquely diverse climates, cultures, and geography. Equador extends from the Galapagos Islands 600 miles to the west, to inhabited Andean highlands with altitudes over 15,000 feet, and to both the coastal and Amazonian rain forests. Health statistics in Ecuador are widely variable by western standards. Life expectancy is 64-68 years, and infant mortality rates are up to 60 per 1000. The physical and geographic barriers to health care facilities are highly variable in different provinces. This was evidenced dramatically by recent death rates from cholera in 1991-92, which ranged from 0% in Guayaquil to over 50% in isolated highland villages or in the Oriente (eastern provinces). Major cities, provincial borders, and selected topographic features are illustrated in the accompanying map of Ecuador (Fig. 1.) This review reports data on some of the major infectious diseases existent in Ecuador. Emphasis is on the viral, bacterial, protozoal, or helminthic diseases, which are uncommon elsewhere in the world, but which are prevalent, or especially important to public health officials in Ecuador. Table 1 lists reportable disease categories in Ecuador, 1986-1993. Recognition of the diversity of infectious agents endemic to Ecuador may prove useful for diagnosis, treatment, and prevention of infectious diseases in international travelers. more...

Indigenous Indian groups comprise approximately 20% of Ecuador's population, the third largest percentage in all of Central or South America, yet immunogenetic data on these groups are lacking in the literature. In the course of population migration studies, sera collected from 65 Ecuadorians living in the northern province of Esmeraldas were typed for six GM and two KM markers. The study population consisted of 47 Cayapa Indians and 18 blacks of African origin, descendants of slaves imported into the area during the seventeenth century. The Cayapa demonstrated three GM phenotypes, two of which are common to other South American Indian tribes. The frequency of KM1 positive Cayapa Indians (63%) is similar to other South American Indian tribes, but is significantly greater than the Huaorani of eastern Ecuador (2%), the only other Ecuadorian Indian group for whom limited immunoglobulin allotype data are available (chi 2 = 35.8, P < 0.0001). more...

In Ecuador, previous epidemiological studies of onchocerciasis affecting a population consisting of Blacks of African origin and Amerindians of the Chachi tribe, have demonstrated different frequencies of clinical disease manifestations in each racial group. To explore the possibility of differential antibody responses to O. volvulus antigens in this population, a subset of sera from these two groups of Ecuadorians with onchocerciasis was analyzed for antibody to O. volvulus antigens using ELISA and Western blot techniques. Significantly higher levels of immunoglobulin against O. volvulus were demonstrated in Chachi Indians compared to native Blacks of African origin: total Ig (p = 0.049), IgG (P = 0.002) IgG4 (p = 0.019) and IgA (p = 0.035). Western blot analysis of sera from 41 persons demonstrated more intense and more frequent recognition of low molecular weight parasite antigens in the Chachi compared to a similar group of male and female Ecuadorian Blacks. These data suggest that the intensity of antibody responses to O. volvulus antigens may reflect the influence of racial factors in the pathogenesis of onchocerciasis in Ecuador. more...

Strategies for detection and control of onchocerciasis in Africa have included identification of DNA probes and PCR target sequences for sensitive and specific detection of parasites. To evaluate the applicability of published PCR and DNA probe based methods for the study of onchocerciasis in Sudan, we collected adult O. volvulus from geographically distinct regions of Sudan (700 miles apart), Abu Hamed (northern desert) and Raja (southwestern savannah), and we examined the similarities between Sudanese O. volvulus repeats and published versions of the repeat from West African O. volvulus. Amplification of DNA extracted from the Raja O. volvulus strain predictably generated a ladder of products, multiples of the base 150 bp repeat, as has been reported from West Africa. However, amplification of DNA from the Abu Hamed O. volvulus isolate resulted in a series of doublets. The unexpected DNA fragments thus amplified differed in size from the base 150 bp unit by approximately 50 base pairs and was most clearly visualized at 150-200 base pairs. DNA sequence analysis of the amplified repeats in the isolate of O. volvulus from Abu Hamed revealed a variant of the 150 bp tandem repeat which contained an extra 49 bp. The additional 49 bp contained two short repeats of 21 bp and 10 bp, corresponding to bases 99-119 and 128-137 respectively, of the known 150 bp O. volvulus repeat. This work demonstrates a variant of the O. volvulus 150 bp tandem repeat, which easily distinguishes Raja and Abu Hamed isolates of O. volvulus, and which has potential value for differentiating Abu Hamed strains of O. volvulus from other strains in East Africa. more...

The suitability of a polymerase chain reaction assay for Borrelia burgdorferi in epidemiological studies of infected tick populations was evaluated by using 28 strains of Leptospira interrogans and lysates of fixed adult Ixodes tick tissues. Two false positives representing leptospires were differentiated from B. burgdorferi by using an oligonucleotide probe. more...

The pathogenesis of tissue damage in filarial infections involves the generation of inflammatory mediators by interaction of parasite surfaces with the host immune response. In the presence of a heavy parasite burden, the tremendous potential for development of adverse local or systemic inflammatory responses may be appreciated by knowledge of the cumulative surface area represented by circulating microfilariae. The approximate surface area of a single microfilariae of Onchocerca volvulus was calculated using mathematical formulae for standard geometrical shapes and values derived from serial measurements of electron micrographs. Numerical values for the surface area represented by large numbers of microfilariae are given and compared to surface areas of the host which can harbor many millions of microfilariae. more...

Sparganosis, infection with plerocercoids of the pseudophyllidean tapeworm Spirometra, rarely has been described in Ecuador. We report the details of a human case of sparganosis identified serendipitously in the course of an abdominal hernia repair. The parasite was found moving freely upon the external oblique fascia adjacent to the site of a direct abdominal hernia. The organism was recovered intact, photographed while alive and preserved for subsequent detailed morphological studies. The presumed route of entry into this patient was percutaneous, after application of a poultice of snake flesh to the site of a painful abdominal hernia. The literature on sparganosis in South America is reviewed. This is the second case of human sparganosis reported from Ecuador. more...

Pentamidine, recently released for clinical use, is effective in therapy for the hemolymphatic stage of Gambian trypanosomiasis, antimony-resistant leishmaniasis, and Pneumocystis carinii pneumonia. The mechanism of action is unclear and may differ for different organisms. Trypanosomes actively transport pentamidine intracellularly, and the drug may then interfere with DNA biosynthetics. However, pentamidine appears to kill nonreplicating P. carinii. The mechanism of killing is unexplained. The pharmacokinetics of pentamidine has been incompletely studied in humans. The estimated volume of distribution is 3 liters/kg. Levels in plasma of pentamidine range from 0.3-1.4 microgram/ml after standard 4 mg/kg dosing, with no appreciable increase in drug levels on successive dosing and no correlation between levels and creatinine clearance or adverse reactions. The drug appears to be concentrated in the kidney and excreted in the urine, with levels detectable six to eight weeks after cessation of therapy. Immediate adverse reactions have included hypotension, nausea, and vomiting. Local pain or abscess formation at an injection site, mild azotemia, leukopenia, abnormal findings from liver function tests, and hypoglycemia may also occur. more...

The conditions were examined for in vitro antibody-mediated adherence of granulocytes to microfilariae of Onchocera volvulus and Dirofilaria immitis. Reactivity in human sera from patients in endemic foci in Sudan was specific for O. volvulus and no reactions were observed with heterologous Onchocerca species or with Mansonella perstans. Microfilariae from skin, nodules or adult female worms were satisfactory targets for cell adherence, and the cells involved were almost exclusively eosinophils. The reaction was inhibited by indomethacin but not by nordihydroguaiaretic acid, an inhibitor of leukotriene production. Agents that slowed or stopped microfilarial motility (e.g. nifedipine, lidocaine, chloroquine) inhibited the reaction, probably by reducing target/cell contact. Ivermectin did not enhance the reaction, and in the absence of cells exerted only slight effects on the movement of microfilariae at higher concentrations (greater than 10 micrograms/ml). Antibody activity was labile, and did not persist well through freeze-thaw cycles. Some differences between homologous and heterologous mixtures (microfilariae/cells/serum) were seen but they could not be resolved satisfactorily. There were no apparent geographical differences between microfilariae from different foci in Sudan. In the D. immitis system neutrophils were the dominant cell type adhering to microfilariae, and the activity was stable to storage and freeze-thaw. No enhancement was detectable with diethylcarbamazine. Antibody activity was absorbable with microfilarial antigens and was reduced by agents that inhibited microfilarial motility. In dogs, adherence-mediating antibody was seen only in amicrofilaraemic animals with occult infection, and in only a minority of these sera. In humans the relationship to clinical findings was less clear, but patients with punctate keratitis were the most likely to have positive serum and were the most reactive in the assay. This system may therefore offer some insights into disease mechanisms in vivo, and its molecular mechanisms deserve further characterization. more...

A two-month remission of minimal change disease induced by hepatitis A infection occurred. The remission was substantiated by clinical, biochemical, and pathologic methods. To our knowledge, this is the first such case reported in the literature. more...

A total of 31 plasmids, all bearing a gene that encodes a novel, plasmid-mediated Richmond-Sykes class III beta-lactamase designated OHIO-1 and a gene that encodes aminoglycoside 2"-adenyltransferase, have been collected from hospitals in Ohio. By using restriction endonuclease digestion and Southern hybridization, we were able to demonstrate that all these plasmids have a common genetic origin. more...

An Onchocerca volvulus cDNA clone expressing epitopes found in adult and larval parasites, designated lambda RAL-2, was derived from a 1,000-base message present in adult O. volvulus, which encodes a protein with an apparent molecular weight of 17,000. This protein does not appear to be extensively posttranslationally modified. Serum samples from 52 individuals exposed to O. volvulus were examined for antibodies recognizing the lambda RAL-2 recombinant antigen; 77% produced such antibodies. In addition, individuals producing antibodies recognizing the recombinant antigen were significantly less likely to develop some aspects of ocular pathology associated with O. volvulus infection than were individuals who did not do so. These results suggest that recombinant antigens such as that produced by lambda RAL-2 may be useful in attempts to understand the mechanism of O. volvulus-induced ocular pathology. more...

Onchocerca volvulus infection of the residents of two villages in the Upper Atbara River region of Sudan (63.4% prevalence) was found to be medically and socially significant with a remarkable degree of pruritus and dermal pathology present. The severest skin changes, which were often confined to certain areas of the body, were most common in the teenage group (13-19 years). Quantitation of the clinical changes showed that the most severe alterations were present in patients with the lower levels of microfilariae in the skin (0.1-6 mf mg-1) rather than in those with higher levels. Nodules containing adult O. volvulus were found in many individuals, and often in clusters in older patients. Major losses of vision due to O. volvulus were not found, although microfilariae were frequently seen in the cornea and anterior chamber. The commonest corneal change was punctate keratitis, and this was most frequently seen in the teenage group. Trachomatous changes of eyelids and corneas were also seen. The prevalence values of O. volvulus infection in the two villages were significantly increased when, in addition to microfilarial presence in skin biopsies, onchocercal signs such as punctate keratitis, nodule presence and microfilariae in the anterior chamber were taken into consideration. This illustrates the need to consider factors other than skin snip parasitology in obtaining a diagnosis. This study also emphasizes the degree of severity that onchodermatitis can reach and that low levels of dermal microfilariae can be accompanied by very extensive skin changes without any loss of vision. The necessity of considering the severity of skin changes when defining tolerable levels of onchocerciasis in a community is discussed. more...

Epidemiologic studies of plasmid-mediated resistance at the Cleveland Veterans Administration Medical Center revealed that related plasmids had disseminated among members of the family Enterobacteriaceae. We studied the beta-lactamases encoded by these plasmids in Escherichia coli C600 transformants or transconjugants. Substrate and inhibition profiles of the enzymes determined by two of these plasmids suggested an activity resembling TEM-1; however, isoelectric focusing revealed a pI of 7.0. These two plasmids were originally found in a Serratia marcescens (pDS076) and an Enterobacter cloacae (pDS075) strain isolated from the same sink in the medical intensive care unit and later, in an Enterobacter cloacae (pDS142 identical to pDS076) isolate colonizing a patient in the same unit. The plasmids also carried the aminoglycoside resistance determinant, 2"-aminoglycoside nucleotidyl transferase. A 2-kilobase AvaI restriction endonuclease digestion fragment of pSD075 known to carry the beta-lactamase determinant was used as a molecular probe. This probe did not recognize sequences of any plasmid-mediated beta-lactamase tested including the recently described determinants ROB-1, TLE-1, and OXA-4-7. A TEM-1 probe derived from the 0.7-kilobase PstI-EcoRI fragment of pBR322 failed to recognize the new beta-lactamase gene. Four additional Enterobacter cloacae and two Enterobacter aerogenes strains isolated in Columbus, Ohio, have been shown to produce a pI 7.0 beta-lactamase and to carry plasmids recognized by the 2-kilobase probe. These data suggest dissemination of a novel plasmid-mediated beta-lactamase among members of the family Enterobacteriaceae in Ohio and demonstrate the development and utility of a molecular probe for the new determinant. We suggest that the novel beta-lactamase be named OHIO-1. more...