Treatment with bovine hyperimmune colostrum of cryptosporidial diarrhea in AIDS patients.

Cryptosporidium parvum may cause severe, debilitating diarrhea in patients with AIDS. Recent anecdotal reports have suggested that hyperimmune bovine colostrum may be effective. We conducted a double-blind, controlled pilot study of hyperimmune bovine colostrum for diarrhea due to cryptosporidiosis in five AIDS patients. The patients were randomized to receive either hyperimmune or control colostrum by continuous nasogastric infusion for 10 days. All stools were collected, graded, and weighed, and the concentration of oocysts excreted was determined daily. One of the three patients treated with hyperimmune colostrum had a reduction in diarrhea and in the concentration of oocysts excreted. A second treated patient had a modest decrease in the concentration of oocysts excreted. Two patients who received control colostrum also had decreases in the volume of diarrhea but no change in the concentration of oocysts excreted. We conclude that hyperimmune colostrum with high titers of specific anti-Cryptosporidium antibody could be effective in treating patients with cryptosporidiosis. However, more studies of cow colostral immunoglobulin need to be performed so that the efficacy of this treatment can be assessed more thoroughly.

Kinetic properties and inhibition of the dimeric dUTPase-dUDPase from Leishmania major.

Kinetic properties of the dimeric enzyme dUTPase from Leishmania major were studied using a continuous spectrophotometric method. dUTP was the natural substrate and dUMP and PPi the products of the hydrolysis. The trypanosomatid enzyme exhibited a low K(m) value for dUTP (2.11 microM), a k(cat) of 49 s(-1), strict Michaelis-Menten kinetics and is a potent catalyst of dUDP hydrolysis, whereas in other dUTPases described, this compound acts as a competitive inhibitor. Discrimination is achieved for the base and sugar moiety showing specificity constants for different dNTPs similar to those of bacterial, viral, and human enzymes. In the alkaline range, the K(m) for dUTP increases with the dissociation of ionizable groups showing pK(a) values of 8.8, identified as the uracil moiety of dUTP and 10, whereas in the acidic range, K(m) is regulated by an enzyme residue exhibiting a pK(a) of 7.1. Activity is strongly inhibited by the nucleoside triphosphate analog alpha-beta-imido-dUTP, indicating that the enzyme can bind triphosphate analogs. The existence of specific inhibition and the apparent structural and kinetic differences (reflected in different binding strength of dNTPs) with other eukaryotic dUTPases suggest that the present enzyme might be exploited as a target for new drugs against leishmaniasis.

The C-terminus of dUTPase: observation on flexibility using NMR.

The dynamics of the C-terminus of the dUTPases from Escherichia coli and equine infectious anaemia virus (EIAV) were studied by 1H-(15)N nuclear magnetic resonance spectroscopy. The two enzymes differ with regard to flexibility in the backbone of the 15 most C-terminal amino acid residues, some of which are conserved and essential for enzymic activity. In the bacterial enzyme, the residues closest to the C-terminus are highly flexible and display a correlation time in the nanosecond time range. No similar high flexibility could be detected for the C-terminal part of EIAV dUTPase, indicating a different time range of flexibility.

dUTPase from the retrovirus equine infectious anemia virus: specificity, turnover and inhibition.

The kinetic properties of dUTPase from equine infectious anemia virus (EIAV) were investigated. K(M) (1.1 +/- 0.1 microM) and k(cat) (25 s(-1)) were found to be independent of pH in the neutral pH range. Above pH 8.0, K(M) increases slightly. Below pH 6.0, the enzyme is rapidly deactivated. Detergent was found to enhance activity, leaving K(M) and k(cat) unaffected. Compared to the Escherichia coli dUTPase, the EIAV enzyme is equally potent in hydrolyzing dUTP, but less specific. Inhibition of the viral enzyme by the nucleotides dTTP, dUMP and a synthetic analogue, 2'-deoxyuridine 5'-(alpha,beta-imido)triphosphate, is stronger by one order of magnitude.

Transient kinetics of ligand binding and role of the C-terminus in the dUTPase from equine infectious anemia virus.

Transient kinetics of the equine infectious anemia virus deoxyuridine 5'-triphosphate nucleotide hydrolase were characterized by monitoring the fluorescence of the protein. Rate constants for the association and dissociation of substrate and inhibitors were determined and found to be consistent with a one-step mechanism for substrate binding. A C-terminal part of the enzyme presumed to be flexible was removed by limited trypsinolysis. As a result, the activity of the dUTPase was completely quenched, but the rate constants and fluorescent signal of the truncated enzyme were affected only to a minor degree. We conclude that the flexible C-terminus is not a prerequisite for substrate binding, but indispensable for catalysis.

Amoxicillin-clavulanic acid for treating drug-resistant Mycobacterium tuberculosis.

This report describes two patients with multidrug resistant tuberculosis who were successfully treated with the addition of amoxicillin-clavulanic acid to second-line drugs. Mycobacterium tuberculosis possesses a beta-lactamase contributing to its resistance to beta-lactam antibiotics. The combination of clavulanic acid, a beta-lactamase inhibitor, and amoxicillin has been shown bactericidal for M tuberculosis in vitro. These data suggest that resistant tuberculosis may warrant a trial of treatment including amoxicillin-clavulanic acid.

The emergence of resistant strains of Acinetobacter baumannii: clinical and infection control implications.

A prospective study was undertaken to determine colonization rates, susceptibility profiles, and outcomes in patients with clinical isolates of Acinetobacter baumannii. Fifty percent of patients became colonized with A. baumannii, and 29% of these patients had clinical and colonizing isolates with discordant susceptibility profiles, without apparent relation to antibiotic use. Barrier infection control measures are necessary to prevent nosocomial transmission.

The ESP culture system for drug susceptibilities of Mycobacterium avium complex.

OBJECTIVE: To validate the non-radiometric, broth-based ESP system for determining Mycobacterium avium complex (MAC) susceptibilities. METHODS: MAC isolates from sterile body sites of 20 adult HIV-infected patients who were failing their present MAC regimen were identified. Susceptibilities were determined and comparisons made between the agar proportion method and the ESP system for clarithromycin, ethambutol, sparfloxacin and cycloserine. RESULTS: Ninety-nine percent of the MICS generated by the ESP system user identical to or lower than the MICs determined by the agar proportion METHOD: In vitro resistance was documented by the ESP system for 86% of the drugs that patients were taking at the time of breakthrough, and no resistance was seen to cycloserine, a drug that no patient was taking. CONCLUSIONS: The ESP system, a fast and reliable method for determining MAC susceptibilities, could be used to optimize MAC regimens in a timely fashion, avoid the use of ineffective drugs, minimize emerging resistance and ultimately improve outcome.

Obstacles to penicillin use in treating pneumococcal infection.

OBJECTIVES: To determine the pattern of penicillin use in the treatment of pneumococcal pneumonia, and factors contributing to the use of alternative antibiotics. METHODS: This study included all adult inpatients of St. Vincent's Hospital and Medical Center who had documented pneumococcal pneumonia between December 1998 and October 1999. St. Vincent's is a 600 bed tertiary teaching hospital in New York City. Patients who had Streptococcus pneumoniae isolated from a respiratory tract specimen were identified through microbiology laboratory records. A retrospective chart review of these patients was conducted, and those identified with clinical pneumonia were included in this study. Antibiotic use, patient demographics, resistance data, and clinician awareness of the antibiotic susceptibility results were noted. RESULTS: Sixty adult patients hospitalized with documented pneumococcal pneumonia were identified. Thirteen (21.6%) of the 60 patients received penicillin or ampicillin. Susceptibility results were not noted in the medical record in 21 (35.0%) of the 60 patients, and none received penicillin. High rates of reported penicillin allergy in 8 (13.3%) of the 60 patients, and reluctance to use penicillin when isolates demonstrated intermediate susceptibility in 8 (13.3%) of the 60 patients were observed. CONCLUSIONS: Several remediable obstacles to penicillin use were identified in this study. An increased awareness of susceptibility results by physicians and education of practitioners could have increased the use of penicillin as therapy to two-thirds of these patients.

Low dose interferon-alpha therapy for HIV-associated multicentric Castleman's disease.

Successful maintenance therapy with weekly interferon-alpha for HIV-associated multicentric Castleman's disease is described.

Lung cancer in HIV-infected patients: a one-year experience.

The relative incidence and rate of lung cancer in HIV-infected patients compared with the general US population has been a source of controversy. We sought to establish these parameters in a cohort of 2616 HIV-infected patients. Tumour type, stage of disease, patient demographics and immune parameters including viral loads were ascertained. An annual rate of 191 cases/100,000 population was found, which is 3.01 times greater (95% confidence interval [CI]=1.3-7.0) when compared with the general US population and 7.4 times the rate in US males between ages 35 and 54 (95% CI=3.1-17.8). Three patients had CD4 counts greater than 200 cells/mm3 at diagnosis and 2 had undetectable viral loads. The mean age was 44 years and all had advanced disease and short survival. In conclusion, the incidence of primary lung cancer was increased in this cohort of HIV-infected patients and occurred over a wide range of immunosuppression and viral replication.

Effects of components of personal need for structure on occupational strain.

The relationship of two elements of personal need for structure-desire for structure and negative reaction to lack of structure-to vulnerability to occupational strain was investigated. Data obtained from a questionnaire administered to female hospital personnel in Finland and analyzed with structural equation models indicated that a high personal desire for structure had a buffering effect on psychic strain symptoms, whereas a tendency to express negative reactions when confronted with a lack of structure intensified symptoms of strain.

Young-adult children of alcoholic parents: protective effects of positive family functioning.

The occurrence of alcoholism is clustered within families, but the detrimental effect of a positive family history may vary with the degree of family impairment involved. In this study we assessed the effects of family history and family environment on alcohol misuse. From ongoing studies we recruited parents who had a child aged 18-30, 20 with a DSM-III-R alcohol dependence diagnosis, 20 without. The child then completed a multidimensional assessment. The young-adult participants included 20 men and 20 women (mean age = 24.8). Differences by family history were restricted to substance abuse behaviors. While a high level of alcohol problems occurred in both groups, those with an alcohol-dependent parent were more likely to be heavy drinkers and showed more symptoms of alcohol dependence. Overall psychological adjustment did not differ between the groups, however. Alcohol misuse measures did correlate moderately with symptoms of poor emotional health. The most important correlates of alcohol misuse measures in this study were exposure to parental alcoholism, abusive punishment, and psychological symptoms, with some separation of effects in the two subgroups. Psychological symptoms had a stronger relationship with misuse in subjects with social-drinking parents, while abuse was more associated in the group with an alcohol-dependent parent. These results confirm the importance of environmental interactions with familial risk. A biological vulnerability from an alcohol-dependent parent was not sufficient or necessary for the participants in this study to develop alcohol dependence as a young adult, although there was an increased risk. There appear to be strong protective effects of positive family relationships on the potential negative effects of a family history of alcoholism.

Prehatch color stimulation effects on color pecking preferences and color discrimination learning in white leghorn chicks.

Four experiments assessed the effects of stimulating chick embryos with colored light at 2 intensity levels. Both posthatch color pecking preferences (Experiments 1 and 2) and color discrimination learning (Experiments 3 and 4) were unaffected. These results affirm and extend a prior finding of no pre- and posthatch colored light stimulation effect on posthatch color preferences in ducklings. The color pecking preferences found replicated prior findings with chicks. However, they differed from the approach color preferences observed in color discrimination learning.

The role of endoscopy after vertical banded gastroplasty.

Since 1984, a total of 99 patients underwent vertical banded gastroplasty (VBG) through protocol (pouch 8 ml in size, band 4.3 cm in circumference) to treat morbid obesity. Follow-up was obtained in 95 patients. Thirty upper gastrointestinal endoscopies were performed post-operatively in 17 patients. Indications were nausea/vomiting in 11, epigastric pain in 4, acute obstructive symptoms in 4, and miscellaneous in three. Findings included food impaction in 10, distal esophagitis in 8, gastritis in 4, and a normal examination in 2. Only 4 of 10 food impactions were associated with an excessively narrowed gastroplasty outlet. Eight patients had an excessively narrowed gastric stoma: two became asymptomatic with dietary modification only and six underwent dilation therapy (dilator range from 8 to 18 mm in diameter) with immediate resolution of symptoms in four of six. One of the two patients unresponsive to dilation was lost to follow-up, and the other required surgical revision after multiple dilation sessions.

The protein p30, encoded at the gag-pro junction of mouse mammary tumor virus, is a dUTPase fused with a nucleocapsid protein.

A ribosomal frameshift at the gag-pro junction of mouse mammary tumor virus (MMTV) gives rise to the protein p30. The protein consists of two domains, the zinc-finger-containing nucleocapsid (NC) protein portion with 95 residues and a C-terminal extension comprising 154 residues. The C-terminal domain shows similarity in sequence with the enzyme dUTPase from other sources. In this paper, we demonstrate that p30 is a functional dUTPase. Overproduction of the NC protein in Escherichia coli, using the native frameshift sequence at the gag stop codon, caused a detectable expression of dUTPase ascribed to a low frequency of readthrough. By a 1-base insertion, eliminating the gag stop codon and fusing the gag and pro reading frames, a plasmid, pET-3d-NCDU, directing overexpression of p30, was constructed. The overproduced protein, purified by phosphocellulose chromatography, shows both zinc-binding and dUTPase activity. Analytical gel filtration and sequence homology to other dUTPases suggest a trimeric assembly of p30 subunits. MMTV thus possesses two different forms of the nucleocapsid protein, the ordinary NC protein and the p30, having the NC protein connected to a domain of dUTPase.

dUTPase from the retrovirus equine infectious anemia virus: high-level expression in Escherichia coli and purification.

Deoxyuridine 5'-triphosphate nucleotidohydrolase (dUTPase, EC 3.6.1.23) catalyzes the hydrolysis of dUTP to dUMP and pyrophosphate, and plays important roles in nucleotide metabolism and DNA replication. The dUTPase gene of the retrovirus equine infectious anemia virus (EIAV) was cloned and overexpressed in Escherichia coli using the T7 RNA polymerase expression system. The recombinant vector (pET-3a/EDU), constructed by mutagenic PCR, was transformed into E. coli BL21 (DE3) pLysS cells, resulting in expression of EIAV dUTPase at about 40% of the extracted protein. This level of overproduction is very high compared to previous reports on heterologous expression of dUTPases in E. coli. A one-step purification procedure using phosphocellulose chromatography results in a homogeneous preparation of the enzyme in a yield of 45 mg liter-1 of bacterial culture. The purified EIAV dUTPase, run on a sodium dodecyl sulfate-polyacrylamide gel electrophoresis, shows an apparent molecular mass of 15.1 kDa in accordance with the gene structure. The isoelectric point (pI) was determined to 5.6. Gel filtration under nondenaturating conditions gives a retention volume corresponding to a molecular mass of 40.6 kDa, suggesting a trimeric organization of the enzyme. The amino acid composition and amino-terminal sequence of the recombinant dUTPase are in agreement with predictions from the DNA sequence.

Vitamin A deficiency in non-vitamin-supplemented patients with AIDS: a cross-sectional study.

The prevalence of vitamin A deficiency and its association with dietary retinol intake in patients with AIDS was assessed in a cross-sectional study. Sixty eligible patients with AIDS provided serum samples that were analyzed for retinol content. Exclusion criteria included current use of vitamin supplements (57% of the 140 willing to participate) and pregnancy (none). Past dietary intake was determined using a standardized food intake frequency questionnaire. The prevalence of hyporetinemia was 22%. This was a 241-fold greater prevalence than that of a representative sample of the U.S. population, after adjusting for age and sex. There was a positive association between serum retinol status and dietary intake, but 27% of those with adequate intake had serum retinol levels below the normal range. These findings suggest that regardless of intake, patients with AIDS may represent a population at considerable risk of vitamin A deficiency.