Detection of St. Louis encephalitis virus antigen in mosquitoes by capture enzyme immunoassay.

Surveillance methods that measure St. Louis encephalitis (SLE) virus activity in nature may provide forewarning of its epidemic occurrence in humans. An antigen capture enzyme immunoassay was developed to detect SLE virus in infected mosquitoes. The assay detected purified SLE viral antigen at a concentration of 62 pg/0.1 ml when antigen was incubated overnight; 250 pg/0.1 ml was detected in a single-day assay (antigen incubated for 3 h). The assay detected 67.9 and 70.8% of laboratory-prepared pools of infected mosquitoes after 3 h and overnight incubation, respectively. The sensitivity of the procedure was 90.5% in identifying pools with infectious titers greater than dex 3.0. The specificity of the assay was controlled by retesting positive pools preincubated with SLE virus and normal antibodies, which led to a diminution of signal in the pools containing viral antigen. The procedure was suitably specific in discriminating between SLE and related flaviviruses, detecting only high infectious doses of heterologous antigens.     

Teebi hypertelorism syndrome: report of a family with previously unrecognized findings

We present a family consisting of a mother, a daughter, and a son with Teebi hypertelorism syndrome, including some previously unrecognized manifestations. The clinical findings include a prominent forehead, arched eyebrows, pronounced hypertelorism, long philtrum, mild interdigital webbing, fifth-finger clinodactyly, umbilical anomalies, and hypotonia. The mother and daughter also had ptosis requiring surgical correction. The daughter has bilateral iridochorioretinal colobomas with high hyperopia and a small umbilical hernia. The son has less striking facial features but was born with a small omphalocele, large ASD secundum, PDA, bilateral cryptorchidism right hydronephrosis, and a cystic left kidney. The mother had an umbilical hernia requiring surgical correction as a child and a history of heart murmur. Both children have normal hearing and mild developmental delay. Their high-resolution karyotypes were normal and the FISH for 22q11 microdeletion was negative in the daughter. We conclude that cardiac defects in Teebi hypertelorism syndrome are not rare findings and that eye colobomas and renal anomalies were previously unrecognized.     

Insertion/deletion mutations of type I oculocutaneous albinism in chinese patients from Taiwan

Type I oculocutaneous albinism (OCA1) is an autosomal recessive disorder, which is caused by the reduction or the absence of tyrosinase activity in melanocytes of the skin, hair and eyes. Although tyrosinase mutations of OCA1 have been extensively analyzed in most populations worldwide, there is no systemic study of OCA1 mutation in Chinese patients. By use of single strand conformation polymorphism and direct sequencing, we had detected 21 mutant alleles out of 24 OCA1 chromosomes screened (87.5%). Detected mutant alleles include one splicing site, three insertion/deletion and five missense mutations, of which the splicing site nucleotide alteration (IVS 1-3C>G) and two each of the insertion/deletion (232-233 ins GGG and 861-862 del TT) and missense mutations (Cys 289 Gly and Trp 400 Leu) are novel. The ins/del mutations accounts for about 37.5% in Chinese OCA1 alleles. The 232-233 ins GGG, one of the novel mutations, was found to be most frequent (25%) among the OCA1 alleles in Chinese. Through this study, we found that while some of the OCA mutant alleles were identified in other populations, ethnic difference still exists. Hum Mutat 14:542, 1999.     

Manganese Superoxide Dismutase Gene Polymorphisms in Psoriatic Arthritis

The purpose of the present study is to investigate the role of manganese superoxide dismutase (MnSOD) gene polymorphisms in the susceptibility to psoriatic arthritis. MnSOD gene polymorphisms were determined by polymerase chain reaction/restriction fragment length polymorphisms method in fifty-two patients with psoriatic arthritis and 90 healthy controls. The genotype frequency of MnSOD 1183C/T was significantly higher in patients with psoriatic arthritis than in controls. In contrast, the frequency of MnSOD 1183T/T was significantly decreased in patients with psoriatic arthritis. The phenotype frequency of MnSOD 1183C was significantly increased in patients with psoriatics arthritis in comparison to healthy controls. Therefore, MnSOD 1183C polymorphisms may be a precipitating factor for the development of psoriatic arthritis.     

Fatal fat embolism following amphotericin B lipid complex injection

A case of amphotericin B lipid complex induced fatal fat embolism is described. A 41-year-old Caucasian man with AIDS was undergoing treatment for cryptococcal meningitis with amphotericin B. His course was complicated by renal failure necessitating a change in therapy to amphotericin B lipid complex (Abelcet). At approximately 48 h, the patient developed tachycardia, tachypnea, respiratory failure, decline in hematocrit, thrombocytopenia, and alteration in mental status. Autopsy findings included fat emboli involving heart, lungs, kidney, and brain. To our knowledge, this is the first case report of a fatal fat embolism caused by intravenous liposome drug delivery.     

Enhanced molecular volume of conservatively pegylated Hb: (SP-PEG5K)6-HbA is non-hypertensive

Recent studies have suggested that the "pressor effect" of acellular Hb is a consequence of perturbation of the macro-and microcirculatory system in multiple ways, and that PEGylation is an effective approach for controlling the same. In an attempt to confirm this concept, a new and simple thiolation mediated, maleimide chemistry-based conservative PEGylation protocol has been developed to conjugate multiple copies of PEG-chains to Hb. This approach combines the high reactivity of maleimides towards thiols with the propensity of iminothiolane to derivatize the epsilon-amino groups of proteins into reactive thiol groups, with conservation of their positive charge. One of the PEGylated products, namely (SP-PEG5K)6-HbA, that carries on an average six copies of PEG5000 chains per Hb, is non-hypertensive in hamster top load and in rat 50% exchange transfusion models. This hexa-PEGylated-Hb has (i) a hydrodynamic volume corresponding to that of an oligomerized Hb of 256kDa, (ii) a molecular radius of approximately 6.8 nm, (iii) high oxygen affinity, (iv) lowered Bohr effect, and (v) increased viscosity and colloidal osmotic pressure. These properties of (SP-PEG5K)6-HbA are consistent with the emerging new paradigms for the design of Hb based oxygen carriers and confirm the concept that the "pressor effect" of Hb is a multifactorial event. The thiolation mediated maleimide chemistry-based PEGylation protocol described here for the generation of (SP-PEG5K)6-Hb is simple, highly efficient, and is carried out under oxy conditions. The results demonstrate that a non-hypertensive PEG-Hb can be generated by conjugation of a lower number of PEG chains than previously reported.     

Sequence variation within botulinum neurotoxin serotypes impacts antibody binding and neutralization

The botulinum neurotoxins (BoNTs) are category A biothreat agents which have been the focus of intensive efforts to develop vaccines and antibody-based prophylaxis and treatment. Such approaches must take into account the extensive BoNT sequence variability; the seven BoNT serotypes differ by up to 70% at the amino acid level. Here, we have analyzed 49 complete published sequences of BoNTs and show that all toxins also exhibit variability within serotypes ranging between 2.6 and 31.6%. To determine the impact of such sequence differences on immune recognition, we studied the binding and neutralization capacity of six BoNT serotype A (BoNT/A) monoclonal antibodies (MAbs) to BoNT/A1 and BoNT/A2, which differ by 10% at the amino acid level. While all six MAbs bound BoNT/A1 with high affinity, three of the six MAbs showed a marked reduction in binding affinity of 500- to more than 1,000-fold to BoNT/A2 toxin. Binding results predicted in vivo toxin neutralization; MAbs or MAb combinations that potently neutralized A1 toxin but did not bind A2 toxin had minimal neutralizing capacity for A2 toxin. This was most striking for a combination of three binding domain MAbs which together neutralized >40,000 mouse 50% lethal doses (LD(50)s) of A1 toxin but less than 500 LD(50)s of A2 toxin. Combining three MAbs which bound both A1 and A2 toxins potently neutralized both toxins. We conclude that sequence variability exists within all toxin serotypes, and this impacts monoclonal antibody binding and neutralization. Such subtype sequence variability must be accounted for when generating and evaluating diagnostic and therapeutic antibodies.