Wide variation in effectiveness of laboratory disinfectants against bacteriophages.

AIMS: The purpose of this study was to identify an effective disinfectant for the inactivation of the bacteriophages (phages) being used in our laboratory, as published studies on phage inactivation are far from unanimous in their conclusions. METHODS AND RESULTS: The phages studied were three closely related strains of Myoviridae and three strains of Siphoviridae. Three disinfectants which are used commonly in microbiology laboratories were evaluated: Virkon (1%), ethanol (75%) and sodium hypochlorite (2500 ppm available chlorine). The most effective of these was Virkon, which inactivated all six phages rapidly. Ethanol was effective against the Myoviridae but had little effect on the Siphoviridae. Sodium hypochlorite was the least effective of the disinfectants evaluated. CONCLUSIONS: The findings of this study demonstrate a wide diversity in the effectiveness of disinfectants tested for inactivation of phages. SIGNIFICANCE AND IMPACT OF THE STUDY: Of the disinfectants tested Virkon is the most suitable choice for those unable to carry out disinfection validation studies, or where a broad spectrum disinfectant against phages is required. All of the phages in this study showed resilience to inactivation by sodium hypochlorite, and therefore this disinfectant is an unwise choice for use against phage without first assessing its effectiveness.

Survival rate of airborne Mycobacterium bovis.

Despite years of study the principle transmission route of bovine tuberculosis to cattle remains unresolved. The distribution of pathological lesions, which are concentrated in the respiratory system, and the very low dose of Mycobacterium bovis needed to initiate infection from a respiratory tract challenge suggest that the disease is spread by airborne transmission. Critical to the airborne transmission of a pathogenic microorganism is its ability to survive the stresses incurred whilst airborne. This study demonstrates that M. bovis is resistant to the stresses imposed immediately after becoming airborne, 94% surviving the first 10 min after aerosolisation. Once airborne the organism is robust, its viability decreasing with a half-life of approximately 1.5 hours. These findings support the hypothesis that airborne transmission is the principle route of infection for bovine tuberculosis.

Assessment of the fitness impacts on Escherichia coli of acquisition of antibiotic resistance genes encoded by different types of genetic element.

OBJECTIVES: Little is known of the fitness cost that antibiotic resistance exerts on wild-type bacteria, especially in their natural environments. We therefore examined the fitness costs that several antibiotic resistance elements imposed on a wild-type Escherichia coli isolate, both in the laboratory and in a pig gut colonization model. METHODS: Plasmid R46, Tn1 and Tn7 and a K42R RpsL substitution were separately introduced into E. coli 345-2 RifC, a rifampicin-resistant derivative of a recent porcine isolate. The insertion site of Tn1 was determined by DNA sequencing. The fitness cost of each resistance element was assessed in vitro by pairwise growth competition and in vivo by regularly monitoring the recovery of strains from faeces for 21 days following oral inoculation of organic piglets. Each derivative of 345-2 RifC carrying a resistance element was grown in antibiotic-free broth for 200 generations and the experiments to assess fitness were repeated. RESULTS: RpsL K42R was found to impose a small fitness cost on E. coli 345-2 RifC in vitro but did not compromise survival in vivo. R46 imposed a cost both before and after laboratory passage in vitro, but only the pre-passage strain was at a disadvantage in vivo. The post-passage isolate had an advantage in pigs. Acquisition of Tn7 had no impact on the fitness of E. coli 345-2 RifC. Two derivatives containing Tn1 were isolated and, in both cases, the transposon inserted into the same cryptic chromosomal sequence. Acquisition of Tn1 improved fitness of E. coli 345-2 RifC in vitro and in vivo in the case of the first derivative, but in the case of a second, independent derivative, Tn1 had a neutral effect on fitness. CONCLUSIONS: The fitness impact imposed on E. coli 345-2 RifC by carriage of antibiotic resistance elements was generally low or non-existent, suggesting that once established, resistance may be difficult to eliminate through reduction in prescribing alone.

Attaching-effacing lesions associated with Escherichia coli O157:H7 and other bacteria in experimentally infected conventional neonatal goats.

Four conventionally reared goats aged 6 days were inoculated orally with approximately 10(10) colony-forming units (cfu) of a non-verotoxigenic strain of Escherichia coli O157:H7. All remained clinically normal. Tissues were sampled under terminal anaesthesia at 24 (two animals), 48 and 72 h post-inoculation (hpi). E. coli O157:H7 was cultured from the ileum, caecum, colon and rectum of all animals, but the number of bacteria recovered at these sites varied between animals. Attaching-effacing (AE) lesions associated with O157 organisms, as confirmed by immunolabelling, were observed in the ileum of one of the two animals examined at 24 hpi, and in the ileum, caecum and proximal colon of an animal examined at 72 hpi. E. coli O157 organisms were detected at > or =10(5) cfu/g of tissue at these sites. In addition, AE lesions associated with unidentified bacteria were observed at various sites in the large bowel of the same animals. Lesions containing both E. coli O157 and unidentified bacteria (non-O157) were not observed. Non-O157 AE lesions were also observed in the large bowel of one of two uninoculated control animals. This indicated that three (one control and two inoculated) animals were colonized with an unidentified AE organism before the commencement of the experiment. The O157-associated AE lesions were observed only in animals colonized by non-O157 AE organisms and this raises questions about individual host susceptibility to AE lesions and whether non-O157 AE organisms influence colonization by E. coli O157.

Effect of the growth promoter avilamycin on emergence and persistence of antimicrobial resistance in enteric bacteria in the pig.

AIM: To assess the effect of the growth promoter avilamycin on emergence and persistence of resistance in enteric bacteria in the pig. METHODS AND RESULTS: Pigs (treated with avilamycin for 3 months and controls) were challenged with multi-resistant Salmonella Typhimurium DT104 and faecal counts were performed for enterococci, Escherichia coli, S. Typhimurium and Campylobacter (before, during and 5 weeks post-treatment). Representative isolates were tested for antibiotic resistance and for the presence of resistance genes. Avilamycin-resistant Enterococci faecalis (speciated by PCR) were isolated from the treated pigs and continued to be detected for the first week after treatment had ceased. The avilamycin-resistance gene was characterized by PCR as the emtA gene and speciation by PCR. MIC profiling confirmed that more than one strain of Ent. faecalis carried this gene. There was no evidence of increased antimicrobial resistance in the E. coli, Salmonella and Campylobacter populations, although there was a higher incidence of tetB positive E. coli in the treated pigs than the controls. CONCLUSION: Although avilamycin selects for resistance in the native enterococci population of the pig, no resistant isolates were detected beyond 1 week post-treatment. This suggests that resistant isolates were unable to persist once selective pressure was removed and were out-competed by the sensitive microflora. SIGNIFICANCE AND IMPACT OF THE STUDY: Our data suggest the risk of resistant isolates becoming carcass contaminants and infecting humans could be minimized by introducing a withdrawal period after using avilamycin and prior to slaughter.

Naturally acquired attaching and effacing Escherichia coli in sheep.

In a series of experiments involving the inoculation of sheep with Escherichia coli O157:H7, and subsequent detailed histopathological examination of the intestinal mucosa, attaching-effacing (AE) lesions formed by elements of the natural flora were observed in 18% of animals. These incidental AE lesions typically were small and sparse, and were not associated with clinical disease. It was possible to identify further some of the lesional bacteria, revealing that E. coli O115 had formed lesions in one of the seven affected animals, and similarly E. coli O26 had formed some of the lesions in another. As AE strains, source flocks, housing and feed sources were diverse, a common source of lesion-forming bacteria appears to be unlikely. It is postulated that subclinical AE lesions are a mechanism of persistence of AE bacteria in sheep.

An experimental mouse model of progressive atrophic rhinitis of swine.

Pasteurella multocida is responsible for a variety of diseases of veterinary importance, including the pig disease progressive atrophic rhinitis (PAR). The feasibility of using the mouse as an experimental model of PAR was evaluated. We experimentally infected the upper respiratory tract of immature mice with a pig isolate of P. multocida that produces the toxin responsible for causing the nasal lesions characteristic of PAR. We tracked the health status and weight gain of these mice for one month following infection, after which the mice were killed and the integrity of the nasal turbinates was examined. Mice infected with P. multocida appeared healthy throughout the study, although the growth rate of these mice was reduced significantly compared with non-infected control animals. Infected animals also demonstrated marked nasal atrophy analogous to that seen in naturally occurring PAR of swine, with shortening and thinning of the turbinate scrolls and inflammatory cell involvement. The mouse therefore provides a convenient model for the further investigation of PAR of swine.

Rifampicin resistance and its fitness cost in Enterococcus faecium.

OBJECTIVES: The genetic basis of rifampicin resistance and the associated fitness cost in Enterococcus faecium were investigated. METHODS: Twelve spontaneous rifampicin-resistant E. faecium mutants were selected from four parent strains recently isolated from porcine faecal material. The DNA sequence of the complete rpoB gene from the parent strains and of nucleotides -189 to +1785 from the mutants was determined from PCR amplicons. The fitness of the mutants was assessed by determining growth rate, by direct growth competition and by the ability of some of the mutants to survive in the pig intestine. RESULTS: The rpoB genes of the parent strains diverged from each other by 1-10% and each encoded proteins that were 1208 amino acids in length. All mutants had a single amino acid substitution in the region implicated in rifampicin resistance in other organisms. Six mutants carried the substitution H489Y/Q, two mutants carried the substitution R492H, one mutant carried the substitution Q480H, two mutants carried the substitutions S494L and V224I, and one mutant carried the substitutions G485D and V224I. Per generation fitness costs of the mutants ranged from a gain of 2.5% to a cost of 10%. Mutants with the substitution H489Y/Q were the most fit, whereas the double mutants were the least fit. The mutant with the substitution H489Q was able to survive in the pig gut for 12 days. There was some correlation between the rifampicin MIC and fitness cost, with higher MICs being associated with higher fitness costs. CONCLUSIONS: Substitutions in RpoB are associated with rifampicin resistance in E. faecium. The fitness cost of resistance is variable and can sometimes be absent.

The effect of chlortetracycline treatment and its subsequent withdrawal on multi-resistant Salmonella enterica serovar Typhimurium DT104 and commensal Escherichia coli in the pig.

AIMS: To investigate the effect of a therapeutic and sub-therapeutic chlortetracycline treatment on tetracycline-resistant Salmonella enterica serovar Typhimurium DT104 and on the commensal Escherichia coli in pig. METHODS AND RESULTS: Salmonella Typhimurium DT104 was orally administered in all pigs prior to antibiotic treatment, and monitored with the native E. coli. Higher numbers of S. Typhimurium DT104 were shed from treated pigs than untreated pigs. This lasted up to 6 weeks post-treatment in the high-dose group. In this group, there was a 30% increase in E. coli with a chlortetracycline minimal inhibitory concentration (MIC) > 16 mg l-1 and a 10% increase in E. coli with an MIC > 50 mg l-1 during and 2 weeks post-treatment. This effect was less-pronounced in the low-dose group. PCR identified the predominant tetracycline resistance genes in the E. coli as tetA, tetB and tetC. The concentration of chlortetracycline in the pig faeces was measured by HPLC and levels reached 80 microg g-1 faeces during treatment. CONCLUSION: Chlortetracycline treatment increases the proportion of resistant enteric bacteria beyond the current withdrawal time. SIGNIFICANCE AND IMPACT OF THE STUDY: Treated pigs are more likely to enter abattoirs with higher levels of resistant bacteria than untreated pigs promoting the risk of these moving up the food chain and infecting man.

Effects of stressors on immune parameters and on the faecal shedding of enterotoxigenic Escherichia coli in piglets following experimental inoculation.

The study examined the effects of stressors on the responses of 3 and a half-week old piglets that had been given an oral dose of enterotoxigenic Escherichia coli (ETEC) and a novel harmless antigen (ovalbumin). Removal from the sow (WEAN), a short-term cold stressor (12;C for 48 hours) (TEMP) and mixing with non-littermates (MIX) were assessed in terms of the effects on faecal shedding of ETEC, immune responses, weight gain and an ACTH stimulation test. WEAN and TEMP reduced weight gain and all stressors increased faecal shedding of ETEC. All stressors increased the IgG responses to F4(K88)ac antigens and WEAN and TEMP increased the IgA responses to the same antigens, probably as a result of increased intestinal proliferation of ETEC. None of the stressors, however, had significant effects on antibody responses to ovalbumin or on lymphocyte proliferation assays. The results indicate that stressors influence the faecal shedding of ETEC in young piglets by a mechanism that may not involve modulation of immune responses.

Contributory and exacerbating roles of gaseous ammonia and organic dust in the etiology of atrophic rhinitis.

Pigs reared commercially indoors are exposed to air heavily contaminated with particulate and gaseous pollutants. Epidemiological surveys have shown an association between the levels of these pollutants and the severity of lesions associated with the upper respiratory tract disease of swine atrophic rhinitis. This study investigated the role of aerial pollutants in the etiology of atrophic rhinitis induced by Pasteurella multocida. Forty, 1-week-old Large White piglets were weaned and divided into eight groups designated A to H. The groups were housed in Rochester exposure chambers and continuously exposed to the following pollutants: ovalbumin (groups A and B), ammonia (groups C and D), ovalbumin plus ammonia (groups E and F), and unpolluted air (groups G and H). The concentrations of pollutants used were 20 mg m-3 total mass and 5 mg m-3 respirable mass for ovalbumin dust and 50 ppm for ammonia. One week after exposure commenced, the pigs in groups A, C, E, and G were infected with P. multocida type D by intranasal inoculation. After 4 weeks of exposure to pollutants, the pigs were killed and the extent of turbinate atrophy was assessed with a morphometric index (MI). Control pigs kept in clean air and not inoculated with P. multocida (group H) had normal turbinate morphology with a mean MI of 41.12% (standard deviation [SD], +/- 1. 59%). In contrast, exposure to pollutants in the absence of P. multocida (groups B, D, and F) induced mild turbinate atrophy with mean MIs of 49.65% (SD, +/-1.96%), 51.04% (SD, +/-2.06%), and 49.88% (SD, +/-3.51%), respectively. A similar level of atrophy was also evoked by inoculation with P. multocida in the absence of pollutants (group G), giving a mean MI of 50.77% (SD, +/-2.07%). However, when P. multocida inoculation was combined with pollutant exposure (groups A, C, and E) moderate to severe turbinate atrophy occurred with mean MIs of 64.93% (SD, +/-4.64%), 59.18% (SD, +/-2.79%), and 73.30% (SD, +/-3.19%), respectively. The severity of atrophy was greatest in pigs exposed simultaneously to dust and ammonia. At the end of the exposure period, higher numbers of P. multocida bacteria were isolated from the tonsils than from the nasal membrane, per gram of tissue. The severity of turbinate atrophy in inoculated pigs was proportional to the number of P. multocida bacteria isolated from tonsils (r2 = 0.909, P < 0.05) and nasal membrane (r2 = 0.628, P < 0.05). These findings indicate that aerial pollutants contribute to the severity of lesions associated with atrophic rhinitis by facilitating colonization of the pig's upper respiratory tract by P. multocida and also by directly evoking mild atrophy.

Effect of ovalbumin aerosol exposure on colonization of the porcine upper airway by Pasteurella multocida and effect of colonization on subsequent immune function.

Seventy-three piglets were weaned at 1 week of age, randomly assigned to 10 groups (A to J), accommodated in stainless steel exposure chambers, and exposed continuously to a controlled environment containing aerosolized ovalbumin. The concentrations of ovalbumin dust were as follows (milligrams per cubic meter): A and F, 16.6; B and G, 8.4; C and H, 4.2; D and I, 2.1; E and J, 0. At weekly intervals, the pigs were bled via venipuncture and anesthetized for nasal lavage and tonsilar biopsies performed for subsequent bacteriologic analysis. At 2 weeks of age, the pigs in groups A to E were challenged with toxigenic Pasteurella multocida (10(8) CFU pig(-1)), and at 6 weeks of age, the pigs were euthanatized. At postmortem, the extent of turbinate atrophy was assessed on the snout sections by using a morphometric index. Exposure to aerial ovalbumin resulted in a dose-dependent increase in serum antiovalbumin immunoglobulin G (IgG; P < 0.001) and serum antiovalbumin IgA (P < 0.001). Exposure also caused a significant increase in the numbers of P. multocida organisms isolated from the upper respiratory tract (P < 0.001) and a corresponding increase in turbinate atrophy, as judged by the morphometric index (P < 0.001). Concurrent challenge with P. multocida and ovalbumin resulted in a significant decrease in both the IgG and IgA responses to ovalbumin (P < 0.001). These results show that ovalbumin exposure increases pig susceptibility to P. multocida colonization and that toxigenic P. multocida modifies the serum IgG and IgA responses to ovalbumin in the pig. Both of these effects may enhance the virulence of this respiratory pathogen and so influence the pathogenesis of atrophic rhinitis in pigs.

Effects of ammonia inhalation and acetic acid pretreatment on colonization kinetics of toxigenic Pasteurella multocida within upper respiratory tracts of swine.

Pigs reared in intensive production systems are continuously exposed to ammonia released by the microbial degradation of their excrement. Exposure to this gas has been shown to increase the severity of the disease progressive atrophic rhinitis by facilitating colonization of the pig's upper respiratory tract by Pasteurella multocida. The etiological mechanism responsible for this synergy was investigated by studying the colonization kinetics of P. multocida enhanced by ammonia and comparing them with those evoked by an established disease model. Three-week-old Large White piglets were weaned and allocated to five experimental groups (groups A to E). Pigs in groups A and B were exposed continuously to ammonia at 20 ppm for the first 2 weeks of the study. Pigs in group C were pretreated with 0.5 ml of 1% acetic acid per nostril on days -2 and -1 of the study. On day 0 all the pigs in groups A, C, and D were inoculated with 1.4 x 10(8) toxigenic P. multocida organisms given by the intranasal route. The kinetics of P. multocida colonization were established by testing samples obtained at weekly intervals throughout the study. The study was terminated on day 37, and the extent of turbinate atrophy was determined by using a morphometric index. The results of the study showed that exposure to aerial ammonia for a limited period had a marked effect on the colonization of toxigenic P. multocida in the nasal cavities of pigs, which resulted in the almost total exclusion of commensal flora. In contrast, ammonia had only a limited effect on P. multocida colonization at the tonsil. The exacerbation of P. multocida colonization by ammonia was restricted to the period of ammonia exposure, and the number of P. multocida organisms colonizing the upper respiratory tract declined rapidly upon the cessation of exposure to ammonia. During the exposure period, the ammonia levels in mucus recovered from the nasal cavity and tonsil were found to be 7- and 3.5-fold higher, respectively, than the levels in samples taken from unexposed controls. Acetic acid pretreatment also induced marked colonization of the nasal cavity which, in contrast to that induced by ammonia, persisted throughout the time course of the study. Furthermore, acetic acid pretreatment induced marked but transient colonization of the tonsil. These findings suggest that the synergistic effect of ammonia acts through an etiological mechanism different from that evoked by acetic acid pretreatment. A strong correlation was found between the numbers of P. multocida organisms isolated from the nasal cavity and the severity of clinical lesions, as determined by using a morphometric index. The data presented in the paper highlight the potential importance of ammonia as an exacerbating factor in respiratory disease of intensively reared livestock.

Immunisation regimes used to elevate serum IgE in sheep.

Six different immunisation regimes were used in an attempt to elevate the concentration of ovine IgE. The response was measured by a passive cutaneous anaphylaxis (PCA) test. These regimes included infection with Ascaris suum and the combination of infection and parenteral administration of Ascaris antigens and ovalbumin in adjuvants. The regime producing the highest titre of IgE was the combination of Ascaris infection with parenteral administration of Ascaris extract in Bordetella pertussis. This regime consistently produced a transient high titre (1:1280-1:2560) of serum IgE 9-12 days after immunisation. The response to ovalbumin was transient and the maximum PCA titre obtained was 1:10.

Synergistic role of gaseous ammonia in etiology of Pasteurella multocida-induced atrophic rhinitis in swine.

One-week-old Large White piglets were weaned and allocated to 14 experimental groups, each composed of five animals. Each group was housed in a separate Rochester exposure chamber and exposed continuously to gaseous ammonia at either 0, 5, 10, 15, 25, 35, or 50 ppm (two groups per exposure level). One week after ammonia exposure commenced, the pigs from one group at each exposure level were inoculated intranasally with 9 x 10(7) CFU of Pasteurella multocida type D. After a further 4 weeks of exposure, all the pigs were euthanized and the extent of turbinate degeneration was assessed by using a morphometric index (J.T. Done, D. H. Upcott, D. C. Frewin, and C. N. Hebert, Vet. Rec. 114:33-35, 1984) and a subjective scoring system (Ministry of Agriculture, Fisheries and Food, Atrophic Rhinitis: a System of Snout Grading, 1978). Exposure to ammonia at a concentration of 5 ppm or greater resulted in a significant increase in the severity of turbinate atrophy induced by P. multocida compared with that occurring in pigs kept in 0 ppm of ammonia. This effect was maximal at 10 ppm but decreased progressively at concentrations above 25 ppm. Regression analysis revealed a significant relationship between the severity of turbinate degeneration and the number of P. multocida organisms isolated from the nasal epithelium at the end of the experiment (R2 = 0.86). These findings suggest that exposure to ammonia facilitates the growth and/or survival of P. multocida within the upper respiratory tract of the pig, thereby contributing to the severity of the clinical disease atrophic rhinitis. Furthermore, exposure of pigs to ammonia at 10 ppm or greater, in the absence of either P. multocida or Bordetella bronchiseptica, induced a mild but statistically significant degree of turbinate atrophy. The findings of this study demonstrate that exposure to ammonia, at concentrations within the range encountered commonly in commercial piggeries, contributes to the severity of clinical lesions associated with atrophic rhinitis.

Cross-reactivity of ovine IgE with anti-human IgE.

Cross-reactivity between ovine and human IgE was investigated using monoclonal antibodies to human IgE heavy chain. Western immunoblotting, reverse cutaneous anaphylaxis (RCA), passive cutaneous anaphylaxis inhibition (PCAI) and reverse passive cutaneous anaphylaxis (RPCA) tests were all used to assess cross-reactivity between the monoclonals and ovine IgE. No cross-reactivity was demonstrated using Western immunoblotting, RCA and PCAI tests. However, evidence for homology in the region of the molecule concerned with mast cell binding was demonstrated by RPCA performed in sheep skin primed with human myeloma IgE.

Isolation of porcine IgE, and preparation of polyclonal antisera.

High Titre, Ascaris specific, porcine reaginic antibody was induced by combining parasitic infestation and antigen challenge. The reagin was characterised as being heat labile at 56 degrees C, possessing long term skin sensitising activity, and as having physicochemical properties similar to IgE in other species. Porcine IgE was purified by conventional immunochemical techniques, and an epsilon chain isolate prepared using SDS-PAGE gel excision and electro elution. Antisera raised against the intact IgE molecule and its epsilon chain were rendered monospecific by immunoabsorption and characterised by reverse cutaneous anaphylaxis (RCA) testing, passive cutaneous anaphylaxis inhibition, immunohistology and Western blotting. These antisera are currently being used to study the regulation of IgE responses in the pig.

Isolation and preparation of antisera to ovine IgE.

Elevated levels of ovine reaginic antibody were induced by immunization with Ascaris suum antigens. The PCA activity of this antibody persisted in the skin of sheep for 20 days and was abolished by heating to 56 degrees C, suggesting that it was immunoglobulin E. Attempts to isolate IgE from this hyperimmune serum by gel filtration, ion exchange and affinity chromatography resulted in the preparation of a PCA-positive fraction containing proteins with molecular weights of 70, 56 and 22 kD on SDS-PAGE. This preparation was used to raise an antiserum in a rabbit to IgE which was rendered specific by absorption. An antiserum to the heavy chain of ovine IgE was also raised in a rabbit by immunization with a fraction prepared from the 68- to 80-kD region of SDS-PAGE by excision and electroelution. Both antisera were positive in reverse cutaneous anaphylaxis tests and recognized a single protein with a molecular weight of 70-72 kD on SDS-PAGE immunoblotting. The presence of this protein in reaginic sheep serum, the molecular weight of its heavy chain, its heat lability and long-term skin-sensitizing ability are characteristic of IgE.

Durable power of attorney for health care. A survey of senior center participants.

Fifty-nine healthy senior center participants were interviewed to determine how, when, and why the durable power of attorney for health care is being used. The 21 users of the durable power of attorney for health care executed a form for the expected reasons; however, a majority had not given a copy of the form to their physician and few had discussed details of their preferences with the proxy. Of the 38 nonusers, the most frequent reasons for not executing a durable power of attorney for health care were: lack of awareness of the form, procrastination, and difficulty choosing a proxy. The new Patient Self-Determination Act requiring hospitals to inform patients of advance directives will help to overcome some of the obstacles in use of the durable power of attorney for health care; however, community education must still be encouraged.

Ingestion of grapefruit lowers elevated hematocrits in human subjects.

This study was based on in vitro observations that naringin isolated from grapefruit induced red cell aggregation and evidence that clumped red cells are removed from the circulation by phagocytosis. The effect on hematocrits of adding grapefruit to the daily diet was determined using 36 human subjects (12 F, 24 M) over a 42-day study. The hematocrits ranged from 36.5 to 55.8% at the start and 38.8% to 49.2% at the end of the study. There was a differential effect on the hematocrit. The largest decreases occurred at the highest hematocrits and the effect decreased on the intermediate hematocrits; however, the low hematocrits increased. There was no significant difference between ingesting 1/2 or 1 grapefruit per day but a decrease in hematocrit due to ingestion of grapefruit was statistically significant at the p less than 0.01 level.