Relative resistance of Salmonella serotypes (Typhimurium, Infantis, and Reading) to peroxyacetic acid on chicken wings.

Peroxyacetic acid (PAA) is widely used as an antimicrobial in poultry processing. Recent salmonellosis outbreaks caused by Salmonella Infantis (SI) from chicken products and Salmonella Reading (SR) from turkey products have raised concerns about their enhanced resistance (compared to Salmonella Typhimurium [ST]) to commonly used antimicrobial interventions such as PAA. The objective of this research was to evaluate the efficacy of PAA against Salmonella serotypes (Typhimurium, Infantis and Reading), effect on product color and decomposition of PAA at different pH levels. Fresh chicken wings (0.45 kg) were inoculated with a cocktail (ca. 6 log CFU/mL) of nalidixic acid resistant ST, rifampicin resistant SI and kanamycin resistant SR. Inoculated chicken wings were immersed in PAA solutions (100 or 500 ppm; adjusted to either pH 8.5 or unadjusted natural pH) for either 10 s or 60 min to replicate treatments for chicken parts or whole carcasses, respectively. Treated chicken wings were rinsed in buffered peptone water (100 mL) containing sodium thiosulfate (0.1 %), serially diluted in peptone water supplemented with 200 ppm of nalidixic acid, rifampicin or kanamycin for enumeration of ST, SI, and SR respectively, and plated on APC Petrifilm. Immersion of chicken wings in 500 ppm PAA for 60 min resulted in greater microbial reductions (P ≤ 0.05) of ST, SI, SR of ca. Two log CFU/mL each, compared to 10 s treatment. Regardless of concentration and pH of PAA, increased exposure time (60 min vs. 10 s) resulted in greater reductions (P ≤ 0.05) of ST, SI, SR. ST was slightly more resistant to PAA solutions than S. Infantis and S. Reading (P ≤ 0.05) for all experimental conditions (PAA conc, pH, and exposure times). Faster decomposition of PAA (100 and 500 ppm) was observed at pH 8.5 compared to unadjusted, natural pH (P ≤ 0.05). Product color (lightness, L*) was not affected regardless of the PAA concentration, exposure time or the pH.

Low-dose Perioperative Dexamethasone Improves 24-hour Post-Operative Pain after Anterior Cruciate Ligament Reconstruction.

Introduction: Post-operative pain following anterior cruciate ligament reconstruction remains an important challenge. Steroids are used in various surgical procedures to decrease post-operative nausea, vomiting and pain. However, only a few studies have reported the effect of systemic administration of steroids in controlling postoperative pain after anterior cruciate ligament surgery. Materials and methods: We have conducted a prospective randomised trial with 109 patients divided into two groups to determine if administration of dexamethasone in the perioperative period improves pain in the post-operative period. The patients were divided into two groups: D, treatment (dexamethasone) and P, control placebo (saline). Patients in the D treatment group were given the first dose of 10mg of intravenous dexamethasone intravenously intraoperatively and the second dose on transferring of the patient to the inpatient department. The patients in the placebo P group, were administered normal saline in the perioperative period in a similar manner. Result: Post-operative pain was significantly less in the dexamethasone group at rest and on walking (p<0.001) for the first 24 hours after surgical procedure. Subsequently, the VAS pain scores were almost similar in both groups at 48 and 72 hours. The administration of dexamethasone resulted in less requirement of antiemetic and rescue analgesia medication There was no difference in range of motion and wound complications rate during the follow-up period at six months. No adverse side effect, like osteonecrosis of the hip, was detected. Conclusion: The pain following anterior cruciate ligament reconstruction is severe during the first 24 hours and perioperative administration of dexamethasone can decrease the post-operative pain substantially.

The fate and significance of retained staples after stapled haemorrhoidopexy.

AIM: We studied the fate of staples after stapled haemorrhoidectomy (SH) and reviewed the complications they may cause. METHOD: A survey was carried among 16 surgeons experienced at performing SH to assess their knowledge regarding retained staples. In addition, plain pelvic radiology was used to visualize residual staples at different time intervals after SH in 45 patients. RESULTS: Ten (63%) surgeons informed their patients that the staples would be shed within 3 months of SH. Regardless of the timing of surgery, part of the staple line was radiologically visible in all patients, and in 19 patients (45%) the staple line was completely visible. CONCLUSION: Contrary to the common belief, the majority of staples are retained after SH.

On-site generated peroxy acetic acid (PAA) technology reduces Salmonella and Campylobacter on chicken wings.

Peroxyacetic acid (PAA) is a widely used antimicrobial during poultry processing that requires to be shipped in a concentrated solution, stored under hazardous conditions and diluted for use. On-site PAA generation using nonhazardous ingredients can help eliminate transportation and storage issues at the processing plant and reduce the risk of occupational hazards. The objective of the proposed research was to 1) evaluate the efficacy of on-site generated PAA in reducing Salmonella and Campylobacter populations compared to the commercially available PAA stock solutions and 2) to perform color measurements to evaluate any deviations between treatments. PAA solutions at 50 and 100 ppm were used for treating the chicken wings. Fresh chicken wings (0.45 kg) were inoculated with a cocktail of nalidixic acid resistant Salmonella Typhimurium (STNR) and gentamicin resistant Campylobacter coli (CCGR) and immersed in PAA solutions (50 and 100 ppm) adjusted to pH 8.5 and 10.0 or 10.5, for either 10 s or 60 min. Treated chicken wings were rinsed for 1 min in chilled BPW (100 mL), serially diluted and plated on APC Petrifilm for Salmonella, spread plated on Campy-cefex agar supplemented with gentamicin (200 ppm) for Campylobacter enumeration. Immersion of chicken wings in 100 ppm PAA for 60 min irrespective of pH levels and PAA solutions resulted in greater microbial reductions (P < 0.05) of Salmonella by 1.68 and 1.42 log CFU/mL for SaniDate, 1.82 and 1.83 log CFU/mL for OxyFusion (on-site generated). For the same treatments, Campylobacter reductions of 1.59 and 1.36 log CFU/mL for SaniDate, 1.63 and 1.71 log CFU/mL for OxyFusion were achieved. The antimicrobial efficiency of PAA was not affected by pH and type of PAA solution. No significant differences (P > 0.05) in color were observed between treatments and controls. On-site generated PAA provides poultry processors an effective, safer, and less hazardous alternative to commercially available PAA solutions, ensuring poultry workers' health and safety.

Proviral DNA of a retrovirus, human T-cell leukemia virus, in two patients with AIDS.

The acquired immune deficiency syndrome (AIDS) is characterized by T-lymphocyte dysfunction and is frequently accompanied by opportunistic infections and Kaposi's sarcoma. Human T-cell leukemia virus (HTLV) is associated with T-cell malignancies and can transform T lymphocytes in vitro. In an attempt to find evidence of HTLV infection in patients with AIDS, DNA from samples of peripheral blood lymphocytes from 33 AIDS patients was analyzed by Southern blot-hybridization with a radiolabeled cloned HTLV DNA probe. Analysis of DNA from both the fresh (uncultured) lymphocytes and from T cells cultured with T-cell growth factor revealed the presence of integrated HTLV proviral sequences in lymphocytes from two of the patients, both of whom had antibody to HTLV. The proviral sequences could not be detected in blood samples obtained from these individuals at a later date, consistent with the possibility that the population of infected cells had become depleted.

Isolation of human T-cell leukemia virus in acquired immune deficiency syndrome (AIDS).

Several isolates of a human type-C retrovirus belonging to one group, known as human T-cell leukemia virus (HTLV), have previously been obtained from patients with adult T-cell leukemia or lymphoma. The T-cell tropism of HTLV and its prevalence in the Caribbean basin prompted a search for it in patients with the epidemic T-cell immune deficiency disorder known as AIDS. Peripheral blood lymphocytes from one patient in the United States and two in France were cultured with T-cell growth factor (TCGF) an shown to express HTLV antigens. Virus from the U.S. patient was isolated and characterized and shown to be related to HTLV subgroup I. The virus was also transmitted into normal human T cells from umbilical cord blood of a newborn. Whether or not HTLV-I or other retroviruses of this family with T-cell tropism cause AIDS, it is possible that patients from whom the virus can be isolated can also transmit it to others. If the target cell of AIDS is the mature T cell as suspected, the methods used in these studies may prove useful for the long-term growth of these cells and for the identification of antigens specific for the etiological agent of AIDS.

Low-dose Perioperative Dexamethasone Improves 24-hour Post-Operative Pain after Anterior Cruciate Ligament Reconstruction

@#Introduction: Post-operative pain following anterior cruciate ligament reconstruction remains an important challenge. Steroids are used in various surgical procedures to decrease post-operative nausea, vomiting and pain. However, only a few studies have reported the effect of systemic administration of steroids in controlling postoperative pain after anterior cruciate ligament surgery. Materials and methods: We have conducted a prospective randomised trial with 109 patients divided into two groups to determine if administration of dexamethasone in the perioperative period improves pain in the post-operative period. The patients were divided into two groups: D, treatment (dexamethasone) and P, control placebo (saline). Patients in the D treatment group were given the first dose of 10mg of intravenous dexamethasone intravenously intraoperatively and the second dose on transferring of the patient to the inpatient department. The patients in the placebo P group, were administered normal saline in the perioperative period in a similar manner. Result: Post-operative pain was significantly less in the dexamethasone group at rest and on walking (p<0.001) for the first 24 hours after surgical procedure. Subsequently, the VAS pain scores were almost similar in both groups at 48 and 72 hours. The administration of dexamethasone resulted in less requirement of antiemetic and rescue analgesia medication There was no difference in range of motion and wound complications rate during the follow-up period at six months. No adverse side effect, like osteonecrosis of the hip, was detected. Conclusion: The pain following anterior cruciate ligament reconstruction is severe during the first 24 hours and perioperative administration of dexamethasone can decrease the post-operative pain substantially.

Effect of particle size and lipid composition of bovine blood high density lipoprotein on its function as a carrier of beta-carotene.

As part of a study on the influence of dietary lipids on vitamin transport and metabolism in lactating cows, we have examined the beta-carotene content and other properties of fractions of the high-density lipoprotein (HDL, density 1.05-1.16 g/ml) of bovine blood. Our purpose was primarily to explain previous results indicating that feeding cows polyunsaturated lipids alters the properties of the HDL and increases the concentration of beta-carotene in the blood but not in the milk. Fractions of HDL of different particle size were prepared by gel-filtration chromatography and the particle diameters measured by electron microscopy. We found that large HDL particles contain more beta-carotene per unit weight than small particles. Furthermore the HDL from cows fed lipid-rich diets with a high proportion of linoleic-acid residues, which had been protected against microbial degradation in the rumen, had a high percentage of HDL particles with large diameters. The blood from these cows had a higher concentration of beta-carotene than before feeding polyunsaturated lipids, but their milk had a lower concentration. We suggest that HDL is the main store of beta-carotene in bovine blood. Moreover the concentration of beta-carotene in blood is increased by feeding polyunsaturated lipids largely because of the increase in the percentage of large HDL particles, which contain more beta-carotene. The effect on the concentration of beta-carotene in milk implies that the transfer mechanism is less efficient as a result of feeding polyunsaturated lipids. This lower efficiency may be due in part to the higher percentage of large HDL particles.

Hydrogen bonding and catalysis: a novel explanation for how a single amino acid substitution can change the pH optimum of a glycosidase.

The pH optima of family 11 xylanases are well correlated with the nature of the residue adjacent to the acid/base catalyst. In xylanases that function optimally under acidic conditions, this residue is aspartic acid, whereas it is asparagine in those that function under more alkaline conditions. Previous studies of wild-type (WT) Bacillus circulans xylanase (BCX), with an asparagine residue at position 35, demonstrated that its pH-dependent activity follows the ionization states of the nucleophile Glu78 (pKa 4.6) and the acid/base catalyst Glu172 (pKa 6.7). As predicted from sequence comparisons, substitution of this asparagine residue with an aspartic acid residue (N35D BCX) shifts its pH optimum from 5.7 to 4.6, with an approximately 20% increase in activity. The bell-shaped pH-activity profile of this mutant enzyme follows apparent pKa values of 3.5 and 5.8. Based on 13C-NMR titrations, the predominant pKa values of its active-site carboxyl groups are 3.7 (Asp35), 5.7 (Glu78) and 8.4 (Glu172). Thus, in contrast to the WT enzyme, the pH-activity profile of N35D BCX appears to be set by Asp35 and Glu78. Mutational, kinetic, and structural studies of N35D BCX, both in its native and covalently modified 2-fluoro-xylobiosyl glycosyl-enzyme intermediate states, reveal that the xylanase still follows a double-displacement mechanism with Glu78 serving as the nucleophile. We therefore propose that Asp35 and Glu172 function together as the general acid/base catalyst, and that N35D BCX exhibits a "reverse protonation" mechanism in which it is catalytically active when Asp35, with the lower pKa, is protonated, while Glu78, with the higher pKa, is deprotonated. This implies that the mutant enzyme must have an inherent catalytic efficiency at least 100-fold higher than that of the parental WT, because only approximately 1% of its population is in the correct ionization state for catalysis at its pH optimum. The increased efficiency of N35D BCX, and by inference all "acidic" family 11 xylanases, is attributed to the formation of a short (2.7 A) hydrogen bond between Asp35 and Glu172, observed in the crystal structure of the glycosyl-enzyme intermediate of this enzyme, that will substantially stabilize the transition state for glycosyl transfer. Such a mechanism may be much more commonly employed than is generally realized, necessitating careful analysis of the pH-dependence of enzymatic catalysis.

Cytoplasmic fragments causing spurious platelet counts in hairy cell leukemia: ultrastructural characterization.

A patient with hairy cell leukemia had a spuriously elevated automated platelet count because of cytoplasmic fragments. The cytoplasmic fragments observed with electron microscopy were found in the same fraction as the platelets (platelet-rich plasma). Ultrastructural examination confirmed the hairy cell cytoplasmic origin of these fragments because of their obvious difference from platelet ultrastructure. Cytoplasmic fragments as a cause of spuriously high automated platelet counts should be considered in all cases of hairy cell leukemia. The blood smear can suggest these factitious events and should be used to confirm the platelet count. It is important to validate the automated platelet count even when reported values are within the normal range, since electronic counting may result in a spurious thrombocytosis or a spuriously normal count.

Rapid renal failure in AIDS-associated focal glomerulosclerosis.

We studied the clinical features, pathologic findings, and course of 18 patients who were found to have glomerular disease at the time of hospitalization with manifestations of acquired immunodeficiency syndrome or acquired immunodeficiency syndrome-related complex at New York University Medical Center, New York, NY, during 1984 through 1987. Focal glomerulosclerosis, characterized by segmental and/or global collapse of capillary walls, was observed in 15 of these patients; mesangial proliferation in 2, and membranous nephropathy in 1. Those with focal glomerulosclerosis typically demonstrated heavy proteinuria without edema or hypertension and progressed rapidly to renal failure in less than 1 year from the time of discovery. This form of focal glomerulosclerosis is characterized by a fulminant course, the collapse type of sclerosis, and the frequent occurrence of uremia without advanced glomerular obliteration. The absence of widespread glomerular sclerosis and the rapid course suggest that unique renal hemodynamic mechanisms may be responsible for the progression.

Cloning, mutagenesis, and structural analysis of human pancreatic alpha-amylase expressed in Pichia pastoris.

Human pancreatic alpha-amylase (HPA) was expressed in the methylotrophic yeast Pichia pastoris and two mutants (D197A and D197N) of a completely conserved active site carboxylic acid were generated. All recombinant proteins were shown by electrospray ionization mass spectrometry (ESI-MS) to be glycosylated and the site of attachment was shown to be Asn461 by peptide mapping in conjunction with ESI-MS. Treatment of these proteins with endoglycosidase F demonstrated that they contained a single N-linked oligosaccharide and yielded a protein product with a single N-acetyl glucosamine (GlcNAc), which could be crystallized. Solution of the crystal structure to a resolution of 2.0 A confirmed the location of the glycosyl group as Asn461 and showed that the recombinant protein had essentially the same conformation as the native enzyme. The kinetic parameters of the glycosylated and deglycosylated wild-type proteins were the same while the k(cat)/Km values for D197A and D197N were 10(6)-10(7) times lower than the wild-type enzyme. The decreased k(cat)/Km values for the mutants confirm that D197 plays a crucial role in the hydrolytic activity of HPA, presumably as the catalytic nucleophile.

Thymosin beta4 accelerates wound healing.

Angiogenesis is an essential step in the repair process that occurs after injury. In this study, we investigated whether the angiogenic thymic peptide thymosin beta4 (Tbeta4) enhanced wound healing in a rat full thickness wound model. Addition of Tbeta4 topically or intraperitoneally increased reepithelialization by 42% over saline controls at 4 d and by as much as 61% at 7 d post-wounding. Treated wounds also contracted at least 11% more than controls by day 7. Increased collagen deposition and angiogenesis were observed in the treated wounds. We also found that Tbeta4 stimulated keratinocyte migration in the Boyden chamber assay. After 4-5 h, migration was stimulated 2-3-fold over migration with medium alone when as little as 10 pg of Tbeta4 was added to the assay. These results suggest that Tbeta4 is a potent wound healing factor with multiple activities that may be useful in the clinic.

Arnebin-1 accelerates normal and hydrocortisone-induced impaired wound healing.

Wound healing involves inflammation, cell proliferation, matrix deposition, and tissue remodeling. Interaction of different cells, extracellular matrix proteins, and their receptors are mediated by cytokines and growth factors during wound healing. In this study, we have evaluated the effect of arnebin-1, a natural product isolated from Arnebia nobilis, on normal and impaired wound healing in cutaneous punch wound model. Arnebin-1 was applied topically daily on wounds of hydrocortisone-treated or untreated animals. Arnebin-1 significantly accelerated healing of wounds with or without hydrocortisone treatment as revealed by a reduction in the wound width and gap length compared with controls. Arnebin-1 treatment promoted the cell proliferation, migration, and vessel formation to form a thick granulation tissue and re-epithelialization of the wounds. An increase in the synthesis of collagen, fibronectin and transforming growth factor-beta1 was seen in arnebin-1-treated wounds compared with the untreated control. As transforming growth factor-beta1 is known to enhance wound healing, and associated with the wound healing defect in hydrocortisone-treated wounds, the enhanced expression of transforming growth factor-beta1 at both translational and transcriptional level by arnebin-1 may be responsible for the enhancement of wound healing during normal and impaired wound repair. These studies suggest that arnebin-1 could be developed as a potent therapeutic agent for wound healing in steroid-impaired wounds.

Diagnosis of progressive multifocal leukoencephalopathy by brain biopsy with biotin labeled DNA:DNA in situ hybridization.

DNA:DNA in situ hybridization using a cloned JC virus (JCV) DNA probe labeled with biotin confirmed the presence of JCV DNA in formalin-fixed, paraffin-embedded brain biopsies from four cases of progressive multifocal leukoencephalopathy (PML). Only small pieces of tissue were available in each case. Detection of the JC DNA:DNA hybrids was carried out by affinity cytochemistry. JCV DNA was identified predominantly in the nuclei of interfascicular oligodendrocytes in demyelinated areas of the biopsies. JC virus was isolated from one case, and the diagnosis of PML was substantiated in all cases by electron microscopic identification or immunocytochemical labeling of JC viral antigen. In situ hybridization using a biotin labeled JCV DNA probe is a specific, sensitive and convenient method for confirming the diagnosis of PML in suspected cases evaluated by brain biopsy.

Role of vacuolar H(+)-ATPase in interferon-induced inhibition of viral glycoprotein transport.

We have shown previously that interferon-beta (IFN-beta) induces the alkalinization of trans-Golgi network (TGN) and inhibits the transport of G protein of vesicular stomatitis virus (VSV) in L(B) cells and gD protein of herpes simplex virus (HSV-1) in LMtk- cells transfected with gD cDNA. The vacuolar H(+)-ATPase (V-ATPase) is responsible for maintaining pH in TGN, and V-ATPase-mediated acidification is required for normal transport of proteins. To examine whether alkalinization caused by IFN is mediated through V-ATPase, the activity of V-ATPase was determined in IFN-treated cells by coupling ATP hydrolysis to NADH oxidation. Bafilomycin (Baf) was used as positive control, as it specifically inhibits V-ATPase. The activity of V-ATPase was reduced in IFN-treated or Baf-treated cells compared with untreated cells. Doses of IFN-beta or Baf that neither alter pHi nor inhibit the transport of viral glycoproteins concomitantly inhibited the transport of G and gD proteins in TGN, as demonstrated by indirect immunofluorescence studies, and raised the pH of TGN as demonstrated by a decrease in the uptake of DAMP. Further, the effect of Baf on IFN-induced antiviral activity against VSV was examined to correlate the biologic significance of these findings. Data showed that Baf significantly enhances (5-50-fold) the IFN-induced antiviral activity as demonstrated by viral titers from supernatants. These findings suggest that the inhibition of transport of G and gD proteins by IFN-beta, may be related to the inhibition of V-ATPase-mediated acidification of TGN.

Mechanism of enhancement of the antiviral action of interferon against herpes simplex virus-1 by chloroquine.

Using double immunofluorescence, we have shown previously that interferon (IFN) treatment inhibits the transport of herpes simplex virus-1 (HSV-1) gD from the Golgi complex to the plasma membrane in the virus infected and gD cDNA transfected LMtk-cells. In the present study, we quantitated the gD protein on the cell surface and localized the gD protein in the trans-Golgi network (TGN). The results showed 10-fold less fluorescence for the gD protein on the cell surface in IFN-treated LMtk-cells. Subcellular fractionation studies demonstrated that gD was associated with TGN-enriched membranes. Gold labeling for DAMP distribution using electron microscopy showed that IFN raised the pH of TGN. IFNs induced alkalinization of TGN may be related to the block in the transport of HSV-1 gD. Earlier we reported that a subeffective dose of chloroquine (CHL) or IFN does not change the pHi. However, both CHL and IFN together raise the pHi significantly. To study the biologic significance of the finding, the effect of these subeffective doses of IFN and CHL on the antiviral activity and the transport of the gD protein was studied. Results suggested that CHL enhance the antiviral activity of IFN against HSV-1 and concomitantly increase the inhibition of HSV-1 gD transport. This IFN-induced increase in pHi of the TGN may also explain the inhibitory effect of IFN reported on the terminal steps of some of the enveloped viruses.

Metastatic meningioma. Hemangiopericytoma or angioblastic meningioma?

The question of whether meningeal hemangiopericytoma is a variant of meningioma ("angioblastic meningioma") or a nosologically distinct entity remains controversial. We present the case histories of an intracranial meningioma and of a meningeal hemangiopericytoma, both of which developed extracranial metastases. The metastatic lesions in both cases were studied by electron microscopy, which demonstrated pericytomatous differentiation in one instance and meningothelial differentiation in the other. This report supports the opinion that meningeal hemangiopericytomas and meningiomas of the CNS are distinct pathological entities.

Long-term effects of etretinate on the liver in psoriasis.

Etretinate is an aromatic retinoid and derivative of vitamin A soon to be approved for general use in the U.S. as therapy for severe psoriasis. We report on liver morphology and function in 18 subjects who received the drug for at least 5 years as part of a clinical trial. The majority (14) suffered no or mild and reversible structural liver changes; mild transient elevations in serum triglyceride and liver enzymes were noted occasionally. Of the remaining four patients, mild periportal fibrosis was documented in two, another had changes similar to chronic active hepatitis, and a fourth had cirrhosis that was unrelated to alcohol use. Liver function data, cumulative drug dose, and treatment duration were generally not reflective of these changes. The results of this study suggest a need for periodic liver biopsy to monitor patients on long-term etretinate therapy.