Human-derived probiotic Lactobacillus reuteri demonstrate antimicrobial activities targeting diverse enteric bacterial pathogens.

Lactobacillus reuteri is a commensal-derived anaerobic probiotic that resides in the human gastrointestinal tract. L. reuteri converts glycerol into a potent broad-spectrum antimicrobial compound, reuterin, which inhibits the growth of gram-positive and gram-negative bacteria. In this study, we compared four human-derived L. reuteri isolates (ATCC 55730, ATCC PTA 6475, ATCC PTA 4659 and ATCC PTA 5289) in their ability to produce reuterin and to inhibit the growth of different enteric pathogens in vitro. Reuterin was produced by each of the four L. reuteri strains and assessed for biological activity. The minimum inhibitory concentration (MIC) of reuterin derived from each strain was determined for the following enteric pathogens: enterohemorrhagic Escherichia coli, enterotoxigenic E. coli, Salmonella enterica, Shigella sonnei and Vibrio cholerae. We also analyzed the relative abilities of L. reuteri to inhibit enteric pathogens in a pathogen overlay assay. The magnitude of reuterin production did not directly correlate with the relative ability of L. reuteri to suppress the proliferation of enteric pathogens. Additional antimicrobial factors may be produced by L. reuteri, and multiple factors may act synergistically with reuterin to inhibit enteric pathogens.

Missense mutation of the last nucleotide of exon 1 (G->C) of beta globin gene not only leads to undetectable mutant peptide and transcript but also interferes with the expression of wild allele.

Hemoglobin Monroe (beta globin G->C, codon 30) is a missense mutation. We could not detect either the mutant peptide or transcript in reticulocyte-enriched preparation and in expanded erythroid progenitor cells. By quantitative gene expression assay beta globin mRNA was found to be reduced by more than 70% in all heterozygous subjects with different haplotypes. We conclude that this mutation also interferes with expression of wild type allele.

Familial polycythemia caused by a novel mutation in the beta globin gene: essential role of P50 in evaluation of familial polycythemia.

Two polycythemic subjects from a family with multiple polycythemic subjects were evaluated. Estimation of oxygen affinity of Hb from venous blood gas parameters (P50) revealed low P50 suggesting a high affinity Hb variant. Further work up, which included beta globin gene sequencing, revealed a novel mutation changing a codon to the previously reported high affinity Hb - Hb Johnstown (beta 109 Val->Leu). Polycythemic subjects with high affinity Hb variant are asymptomatic with normal life expectancy. Their differentiation from polycythemia vera (PV) is crucial to avoid therapy which is otherwise reserved for PV patients. We provide an electronic version (in Microsoft excel program) of a previously reported mathematical formula for rapid calculation of P50 from venous blood gases. Estimation of P50 is an essential initial step in the evaluation of a subject with personal and family history of polycythemia.

Infectious enteritis after intestinal transplantation: incidence, timing, and outcome.

AIM: To review the incidence, timing, and outcome of infectious enteritis after intestinal transplantation (IT). METHOD: A retrospective review of all patients undergoing IT at a single institution between 1991 and 2003 was analyze with standard statistical tools. RESULTS: Among 33 IT recipients, 13 (39%) developed 20 culture- or biopsy-proven episodes of infectious enteritis. The recipient demographics were 77% men and median age 2.6 years. Infections were diagnosed at a median of 76 days (32 to 1800) after IT. There were 14 viral (CMV one, rotavirus eight, adenovirus four, EBV one, three bacterial (Clostridium difficile), and three other infections (Giardia lamblia one, cryptosporidium two). Complete resolution was achieved in 17 (94%) infectious after appropriate antimicrobial or conservative therapy. Interestingly, there were six rejection episodes following infectious enteritis. Grafts were lost to rejection after rotaviral enteritis (n = 1) and adenoviral enteritis misdiagnosed as rejection (n = 1). Patient and graft survival were not adversely affected by infections. CONCLUSIONS: Infectious enteritis occurs frequently after IT. Viral agents are the cause in two-thirds of cases. With supportive care and appropriate treatment, resolution is possible in the majority of cases. Differentiating rejection and infection by histopathology can be difficult.

A reversible structural interconversion involving [M(H2pdc)2(H2O)2] . 2H2O (M = Mn, Fe, Co, Ni, Zn, H3pdc = 3,5-pyrazoledicarboxylic acid) and the role of a reactive intermediate [Co(H2pdc)2].

A new type of hydrogen bonded networks [M(H2pdc)2(H2O)2] . 2H2O [M = Mn (1), Fe (2), Co (3), Ni (4), Zn (5); H3pdc = 3,5-pyrazoledicarboxylic acid] have been synthesized via hydrothermal reactions and their structures have been characterized. Upon a cooling-heating cycle, these compounds undergo a reversible structural interconversion process via hydration-dehydration: [chemical equation: see text]. The process is associated with distinct color changes. The dehydrated [M(H2pdc)2] (M = Mn, Fe, Co, Ni, and Zn) are amorphous and highly reactive. Further chemical reactions of these reactive intermediates show that they may act as effective precursors towards assembly of new supramolecular compounds that may otherwise be inaccessible by other synthetic routes. An interesting structure containing an "open-box" molecule [Co4(Hpdc)4(py)12] . 4py . 2H2O . 2CH3OH (6) (py=pyridine) has been isolated by using dehydrated [Co(H2pdc)2] as the precursor, and its crystal structure has been analyzed. Crystal data for 1-6: monoclinic, space group P2(1/c) and Z = 2 with a = 10.186(2), b = 12.473(2), c = 6.831(1) A, beta = 108.80(3) degrees (1); a = 9.896(2), b = 12.402(2), c = 6.810(1) A, beta = 108.15(3) degrees (2); a = 9.981(2), b = 12.426(2), c = 6.807(1) A, beta = 108.23(3) degrees (3); a = 9.896(2), b = 12.402(2), c = 6.810(1) A, beta = 108.15(3) degrees (4); a = 10.001(2), b = 12.430(2), c = 6.834(1) A, beta = 108.32(3) degrees (5); a = 9.9617(1), b=18.5080(2), c = 28.4786(3) A, beta = 93.076(1) degrees (6).

3,5-Pyrazoledicarboxylic acid monohydrate

In the title compound, C(5)H(4)N(2)O(4).H(2)O, the 3, 5-pyrazoledicarboxylic acid (H(3)pdc) molecules are joined into one-dimensional chains by O-H.O and N-H.O hydrogen bonds, with distances of 2.671 (2) and 2.776 (2) A, respectively. The one-dimensional chains form a three-dimensional structure via O-H.OW and OW-HW.N hydrogen bonds, with distances of 2.597 (3) and 2.780 (3) A, respectively. In addition to the potential for forming open-channel frameworks, access to the six coordination atoms of H(3)pdc can be directly controlled by varying the pH of the reaction environment, allowing further control over the design and synthesis of novel coordination polymers using various metal centers.

Reactions and reactivity of Co-bpdc coordination polymers (bpdc = 4,4'-biphenyldicarboxylate).

By choosing a suitable metal center, ligand, and solvents, we have revealed several structural transformations involving a polymer precursor. infinity 1[Co(bpdc)(H2O)2].H2O (1) was prepared by reaction of Na2bpdc and Co(NO3)2 in aqueous solution. Immersing 1 in pyridine/water solutions of (2:1) and (8:1) ratios yielded a second one-dimensional structure infinity 1[Co(bpdc)(py)2(H2O)2].2py (2) and a two-dimensional structure infinity 2[Co(bpdc)(py)2].H2O (3), respectively. After heating 1 under N2 to remove all water within the structure, the compound Co(bpdc) (IR) was obtained. When IR was immersed in solutions of pyridine/water (5:4) and in pure pyridine (in air), a third one-dimensional structure of infinity 1[Co(bpdc)(py)2(H2O)2].2py.H2O (4) and 3, respectively, were obtained. Compounds 2-4 easily transformed to 1 when immersed in water. Crystal data for 1: monoclinic, space group C2/c with a = 6.950(1), b = 31.585(6), and c = 6.226(1) A, beta = 95.84(3) degrees, Z = 4. Crystal data for 2: triclinic, space group P1 with a = 9.646(2), b = 10.352(2), and c = 17.031(3) A, alpha = 79.02(3) degrees, beta = 86.88(3) degrees, gamma = 77.16(3) degrees, Z = 2. Crystal data for 3: triclinic, space group P1 with a = 9.137(2), b = 10.480(2), and c = 12.254(2) A, alpha = 102.10(3) degrees, beta = 100.80(3) degrees, gamma = 99.43(3) degrees, Z = 2. Crystal data for 4: orthorhombic, space group Pbcn with a = 13.468(3), b = 16.652 (3), and c = 14.977(3) A, Z = 4.

Interferon alpha-2b in the treatment of chronic hepatitis C: early experience.

The antiviral and immunomodulatory effects of interferon were assessed in the treatment of chronic Hepatitis C in multi-ethnic patients to prevent viral replication and chronic liver damage. Three million units of recombinant interferon alpha-2b were administered three times a week for 48 weeks to a group of 9 active Hepatitis C patients. A clinical response was defined as normalization of serum ALT values. Serum was frozen and stored for Hepatitis C viral assays. Four patients normalized their liver functions. When viral levels were measured only two patients had unmeasurable levels of HCV RNA after treatment. Therapeutic results were observed and much work needs to be done to improve therapy because a serious epidemic is predicted for the future.

Long-term low dose interferon alpha-2b in the treatment of chronic hepatitis B in multi-ethnic patients in Hawaii.

The antiviral and immunomodulatory effects of interferon were assessed in the treatment of chronic hepatitis B in multi-ethnic patients to prevent viral replication and chronic liver damage. Five million units of recombinant interferon alpha-2b were administered three times a week for 48 weeks to a group of 18 chronic active hepatitis B patients. A complete response was defined as seroconversion to anti-HBe and/or loss of HBe antigen. Seroconversion to anti-HBe in 5 of 12 (42%) chronic active hepatitis B patients occurred after 48 weeks of therapy. HBV-DNA decreased to undetectable levels in 8 of 12 (67%) patients. This chronic low-dose interferon administration regimen demonstrated responses comparable to other studies.

The immunologic staging of chronic active hepatitis B patients in Hawaii.

The hepatitis B antigen/antibody levels and natural killer cell activity status of chronic hepatitis B patients identified by the Hawaii State Department of Health were evaluated to select chronically infected hepatitis B patients for interferon therapy and to determine possible immunodeficiencies. The presence of hepatitis Be antigen denotes active replication of the virus. Ninety-five patients were studied: 17/95 (18%) had chronic active hepatitis B, 71/95 (75%) were hepatitis B carriers and 7/95 (7%) had seroconverted. NK activity to the erythroleukemia K562 cell and virus-infected HSV-1 cell of the chronic active and carrier population (P < .05) were lower than that of the control population and those who had spontaneously seroconverted. Of this population 18% were identified with active viral infection and would be candidates for interferon therapy.

Frequency of glucose-6-phosphate dehydrogenase (G6PD) mutations in Chinese, Filipinos, and Laotians from Hawaii.

In a Hawaii Hereditary Anemia Screening Project, 4,984 participants were tested for glucose-6-phosphate dehydrogenase (G6PD) deficiency by a filter paper blood spot fluorescence test. Abnormal samples and suspected heterozygotes were checked by quantitative G6PD assay (normal 4.5 to 14 units/g Hb). G6PD was deficient (< 1.5 units/g Hb) in 188 of 2,155 males; 7 other males had low activity (1.5 to 2.8 units/g Hb). The gene frequency, estimated from males after excluding referred and related cases, was 0.037 for Chinese, 0.134 for Filipinos, and 0.203 for Laotians. Among 2,829 females tested, family data showed 111 females were obliged to be at least heterozygous, regardless of G6PD activity, and 43 others had low G6PD activity. Most heterozygotes probably remained undetected by G6PD screening. In 28 females, activity was under 10%; in another 9 females, activity was < 1.5 units/g Hb. Since only 25 homozygotes would be predicted, this apparent excess of females with deficient activity could be due to unequal X-inactivation in some heterozygotes. DNA analysis by polymerase chain reaction amplification and special analytic procedures revealed 10 different missense mutations in 75 males. The nucleotide 835 A-->T and 1360 C-->T transitions were first detected in this Hawaiian Project; we found that the nucleotide 1360 mutation was the most common cause of G6PD deficiency in Filipinos. This is the first report of G6PD screening and analysis of molecular G6PD mutations in Filipino and Laotian populations.

Two-dimensional phase unwrapping using a minimum spanning tree algorithm.

Phase unwrapping refers to the determination of phase from modulo 2pi data, some of which may not be reliable. In 2D, this is equivalent to confining the support of the phase function to one or more arbitrarily shaped regions. A phase unwrapping algorithm is presented which works for 2D data known only within a set of nonconnected regions with possibly nonconvex boundaries. The algorithm includes the following steps: segmentation to identify connectivity, phase unwrapping within each segment using a Taylor series expansion, phase unwrapping between disconnected segments along an optimum path, and filling of phase information voids. The optimum path for intersegment unwrapping is determined by a minimum spanning tree algorithm. Although the algorithm is applicable to any 2D data, the main application addressed is magnetic resonance imaging (MRI) where phase maps are useful.

A PCR procedure to determine the sequence of large polypeptides by rapid walking through a cDNA library.

A procedure that uses the PCR to make rapid successive steps through a random-primed cDNA library has been developed to provide a method for sequencing very long genes that are difficult to obtain as a single clone. In each successive step, the portions of partial clones that extend out from the region of known DNA sequence are amplified by two stages of PCR with nested, outward-directed primers designed approximately 50 bases in from the end of the known sequence, together with a general primer based on the sequence of the vector. This procedure has been used to determine the coding sequence of the cDNA for the beta heavy chain of axonemal dynein from embryos of the sea urchin Tripneustes gratilla. By starting from a single parent clone, whose translated amino acid sequence overlapped the microsequence of a tryptic peptide of the beta heavy chain, and making 3 such walk steps downstream and 14 walk steps upstream, we obtained a sequence of 13,799 base pairs that had an open reading frame of 13,398 base pairs. This sequence encodes a polypeptide with 4466 residues of Mr 511,804 that is believed to correspond to the complete beta heavy chain of ciliary outer arm dynein.

Molecular screening for haemoglobin constant spring.

Haemoglobin H/Constant Spring is an important cause of severe haemoglobin H disease, but the Constant Spring protein is difficult to detect by electrophoresis. A technique for allele specific polymerase chain amplification of the 3'-end of the alpha 2 globin gene improved detection of the alpha cs alpha haemoglobin variant in DNA samples by slot-blot hybridisation. The alpha cs alpha mutation was confirmed in subjects that had been previously diagnosed by haemoglobin electrophoresis, and it was also detected in patients who were negative by protein electrophoresis. 10 of 103 unrelated Laotians with HbE were alpha cs alpha heterozygotes. Of these, 3 were negative to the normal probe because they had -alpha 3.7/alpha cs alpha with a single alpha globin deletion. 5 samples did not amplify or hybridise to either probe because they had deletions of both alpha 2 globin regions. The gene frequency for alpha cs alpha is about 0.05 for Laotians. This technique, which is highly specific and sensitive for rapid detection of the alpha cs alpha mutation, is suitable for clinical diagnoses and population studies. The true incidence of alpha cs alpha may prove to be greater than previously suspected from protein electrophoresis.

Plasma amino acid and serum unesterified fatty acid deficits and the effect of nutritional support in chemotherapy treatment.

The deficits in plasma amino acids and serum unesterified fatty acids of cancer patients undergoing chemotherapy and/or radiation therapy were studied to delineate the special requirements of the patients and efficacy of our nutritional therapy. Seven general surgery patients and 13 patients treated by the Head-Neck Service had baseline levels measured as part of their nutritional evaluation prior to surgical treatment of their cancers. Fifteen chemotherapy outpatients maintained on their regular diets had fasting levels analyzed. Twenty-six patients who were admitted for their therapy had their intake of the regular hospital diet supplemented with a low-residue enteral diet formula (Vivonex High Nitrogen Diet); parenteral nutrition was used only if their oral intake was totally inadequate. Baseline and sequential measurements were made of plasma amino acid and serum unesterified fatty acid levels by gas liquid chromatographic techniques. Before operation the patients had normal levels of amino acids except for a significant deficiency of threonine and glycine observed in patients with head-neck tumors. Outpatients with and without hepatic metastases had significantly depressed levels of the essential amino acids valine, leucine, threonine, and methionine and the nonessential amino acids serine, glycine, and proline. The baseline levels of the patients admitted for treatment had similar deficiencies except for more evidence of lysine deficiency. Patients supported with total parenteral nutrition had rapid elevation of the amino acid levels. The patients whose intake was supplemented with the oral diets had improvement in their amino acid levels, but the deficiency in the leucine and threonine fractions persisted up to 4 weeks of therapy. Although the lysine levels were normal when first analyzed, significant differences developed in the patients without hepatic metastases after the start of chemotherapy with return to normal only after chemotherapy was discontinued. Fatty acid levels were not significantly different between the cancer groups except for preoperative elevated oleic acid levels noted in the general surgery tumor group; there were no deficiencies in the essential fatty acids. These studies indicate a need for enteral formulas with adequate branched-chain amino acids and enrichment with threonine and lysine for supplementing the nutrition of the cancer patient who is undergoing chemotherapy.

Relationships of prostaglandin levels and natural killer (NK) cell cytotoxicity of mononuclear cells in cord blood.

Natural killer cell cytotoxicity of mononuclear cells and the corresponding plasma prostaglandins were examined in cord blood. Low NK cell cytotoxicity was demonstrated against three target cells: NK(K562), NK(HSV-1) and NK(Fs). Prostaglandins of the B, E and F series were examined and found to be higher than adult values. A significant correlation (p less than 0.05) between NK cell cytotoxicity and the prostaglandin F series was demonstrated.