James R. Carlson

Viral, Nutritional, and Bacterial Safety of Flash-Heated and Pretoria-Pasteurized Breast Milk to Prevent Mother-to-Child Transmission of HIV in Resource-Poor Countries A Pilot Study

Background:Heat-treated breast milk of HIV-positive mothers has potential to reduce vertical transmission. This study compared the impact of flash-heating (FH) and Pretoria pasteurization (PP) on HIV, nutrients, and antimicrobial properties in human milk. Methods:Milk samples were spiked with 1 × 108 copies/mL of clade C HIV-1 and treated with FH and PP. We measured HIV reverse transcriptase (RT) activity before and after heating (n = 5). Heat impact on vitamins A, B6, B12, and C; folate, riboflavin, thiamin, and antimicrobial proteins (lactoferrin and lysozyme) was assessed. Storage safety was evaluated by spiking with Escherichia coli or Staphylococcus aureus. Results:Both methods inactivated ≥3 logs of HIV-1. FH resulted in undetectable RT activity. Neither method caused significant decrease in any vitamin, although reductions in vitamins C and E were noted. Heat decreased immunoreactive lactoferrin (P < 0.05) but not the proportions of lactoferrin and lysozyme surviving digestion. FH seems to retain more antibacterial activity. Both treatments eliminated spiked bacteria. Conclusions:FH may be superior to PP in eliminating all viral activity; both methods retained nutrients and destroyed bacterial contamination. Heat-treated breast milk merits further study as a safe and practical infant feeding option for HIV-positive mothers in developing countries.

Role of bacterial infection in Diet-Induced acute pancreatitis in mice

SummaryThis study was performed to elucidate the role of bacterial infection in acute pancreatitis in young female mice fed a choline-deficient diet supplemented with 0.5% DL-ethionine (CDE diet). Mice were randomly classified into two groups: one had been fed CDE diet alone (nonantibiotic group), the other was fed a CDE diet with oral administration of antibiotics (antibiotic group). Survival rates at 96 and 144 h after introduction of the CDE diet were significantly improved in the antibiotic group, 25.0 and 19.4%, respectively, as compared with 3.6 and 0% in the nonantibiotic group (p, 0.05). Aerobic and anaerobic bacterial cultures of blood, ascites, spleen, and pancreas were taken from living mice 72 h after introduction of the CDE diet. Positive bacterial growth from one or more of the specimens occurred in 29.4% of the nonantibiotic group, and in 10.0% of the antibiotic group. Mice with pancreatic necrosis had a higher positive culture rate, 62.5% in the nonantibiotic group vs 20.0% in the antibiotic group. These results suggest that reduction of intestinal flora in mice inhibits secondary infection caused by bacterial transloca-tion and improves survival in diet-induced hemorrhagic pancreatitis. We believe the development of bacterial infection of gut origin may be a factor influencing mortality in severe pancreatitis.

Therapeutic Regimens in Acute Experimental Pancreatitis in Rats: Effects of a Protease Inhibitor, a β-Agonist, and Antibiotics

The osmolality of contrast injected retrograde into the rat pancreatic duct did not affect the severity of the pancreatitis (Urografin, 1,300 mOsm/kg, and Hexabrix, 580 mOsm/kg). The severity of the pancreatitis induced in rats was assessed by survival rate, histologic grading, wet lung ratio, and serum levels of amylase, lipase, and trypsin-like activity. Rats with pancreatitis induced by retrograde injected Urografin, lipopolysaccharide, taurocholic acid plus enterokinase were treated with either intravenous (i.v.) FUT-175 (Nafamstat Mesilate), FUT-175 administered by retrograde pancreatic injection, i.v. terbutaline, i.v. piperacillin sodium, piperacillin sodium by retrograde pancreatic duct injection, or a combination of FUT-175 plus terbutaline and piperacillin. Survival among the rats was increased and the incidence of pancreatic infection reduced in rats treated with i.v. piperacillin or with a combination of FUT-175 plus i.v. terbutaline, plus i.v. piperacillin compared to controls.

Isolation of acquired immunodeficiency syndrome virus from the placenta.

The etiologic agent of the acquired immunodeficiency syndrome is human immunodeficiency virus. We report here a case of the culturing of this agent from the placenta and suggest the clinical implications.

Coliform Bacteria in Sierra Nevada Wilderness Lakes and Streams: What Is the Impact of Backpackers, Pack Animals, and Cattle?

Abstract Objective.—The presence of coliform bacteria indicates a watershed risk for harboring microbes capable of causing human disease. We hypothesized that water from watersheds that have different human- or animal-use patterns would have differing risks for the presence of coliform bacteria. Methods.—Water was collected in wilderness areas of the Sierra Nevada range in California. A total of 60 sites from lakes or streams were selected to statistically differentiate the risk categories: 1) high use by backpackers, 2) high use by pack animals, 3) cattle- and sheep-grazing tracts, and 4) natural areas rarely visited by humans or domestic animals. Water was collected in sterile test tubes and Millipore coliform samplers during the summer of 2004. Water was analyzed at the university microbiology lab, where bacteria were harvested and then subjected to analysis by standardized techniques. Confirmation was performed with a Phoenix 100 bacteria analyzer. Statistical analysis to compare site categories was performed with Fisher exact test. Results.—Only 1 of 15 backpacker sites yielded coliforms. In contrast, 12 of 15 sites with heavy pack-animal traffic yielded coliforms. All 15 sites below the cattle-grazing areas grew coliforms. Differences between backpacker and cattle or pack-animal areas were significant (P ≤ .05). Only 1 of the 15 wild sites rarely visited by humans grew coliforms. All coliforms were identified as Escherichia coli. All samples grew normal aquatic bacteria of the genera Pseudomonas, Ralstonia, and Serratia and nonpathogenic strains of Yersinia. No correlation could be made with temperature or elevation. Sites below cattle-grazing tracts and pack-animal usage areas tended to have more total bacteria. Conclusions.—Alpine wilderness water below cattle-grazing tracts or areas used by pack animals are at risk for containing coliform organisms. Areas exclusively used by backpackers were nearly free of coliforms.

Risk Factors for Coliform Bacteria in Backcountry Lakes and Streams in the Sierra Nevada Mountains: A 5-Year Study

Abstract Category 1 Continuing Medical Education credit for WMS member physicians is available for this article. Go to http://wms.org/cme/cme.asp?whatarticle=1922 to access the test questions. Objective.—To provide a 5-year longitudinal assessment of risk of acquiring disease from Sierra Nevada Wilderness area lakes and streams. This study examines the relative risk factors for harmful water microorganisms, using coliforms as an indicator. Methods.—Streams and lakes in the backcountry of Yosemite and Kings Canyon National Parks and neighboring wilderness areas were selected and water was analyzed each year over a 5-year period. A total of 364 samples from lakes or streams were chosen to statistically differentiate the risk categories based on land usage, as follows: 1) areas rarely visited by humans (Wild), 2) human day-use-only areas (Day Hike), 3) areas used by backpackers with overnight camping allowed (Backpack), 4) areas primarily impacted by horses or pack animals (Pack Animal), and 5) cattle and sheep grazing tracts (Cattle). Water was collected in sterile test tubes and Millipore coliform samplers. Water was analyzed at the university microbiology lab, where bacteria were harvested and then subjected to analysis using standardized techniques. Statistical analysis to compare site categories was performed utilizing Fisher exact test and analysis of variance. Results.—A total of 364 sampling sites were analyzed. Coliforms were found in 9% (4/47) of Wild site samples, 12% (5/42) of Day Hike site samples, and 18% (20/111) of Backpacker site samples. In contrast, 63% (70/111) of Pack Animal site samples yielded coliforms, and 96% (51/53) of samples from the Cattle areas grew coliforms. Differences between Backpacker vs Cattle or Pack Animal areas were significant at P ≤ .05. All samples grew normal aquatic bacteria. Conclusion.—Surface water from watersheds below cattle areas and those used by pack animals is at high risk for containing coliform organisms. Water from Wild, Day Hike, or Backpack sites poses far less risk for contamination by coliforms.

An analysis of wilderness water in Kings Canyon, Sequoia, and Yosemite national parks for coliform and pathologic bacteria.

OBJECTIVE To determine the prevalence of coliform and potentially pathogenic bacteria in remote backcountry alpine lakes and streams of national parks in the Sierra Nevada mountains. METHODS Water was sampled at 55 predetermined lakes and streams that would stratify the risk, based on sites used by backpackers, sites used by pack animals, and uncontaminated wild areas. Sites were distributed among Kings Canyon (15), Sequoia (17), and Yosemite (23). Water was collected using Millipore bacterial samplers, which provided specific counts of coliform and other bacteria in each water sample and also served as a transport media from the wilderness to the laboratory. On return to the laboratory, bacteria were harvested from the samplers and subjected to specific identification and qualitative analysis using standard microbiology techniques for the analysis of water. RESULTS Coliform bacteria were detected in 22 of the 55 sites. All of these sites were below areas used by backpackers or pack animals. Thirty-three sites were free of coliforms. These sites included both those used lightly by backpackers and those with no human or domestic animal use. All samples contained expected amounts of normal aquatic bacteria including Pseudomonas, Rahnella aquatilis, Serratia spp, and nonpathogenic species of Yersinia. CONCLUSIONS Most sampling sites in these national parks are free of coliform or pathogenic organisms. Low levels of coliform bacteria are found in some bodies of water where the watershed has been affected by human or pack animal travel.

An analysis of human pathogens found in horse/mule manure along the John Muir Trail in Kings Canyon and Sequoia and Yosemite National Parks.

OBJECTIVE To determine the prevalence of microorganisms that are potentially pathogenic for humans in horse/mule manure along the John Muir Trail (JMT). METHODS Random samples of horse/mule manure were collected along sections of the JMT in Yosemite, Kings Canyon, and Sequoia national parks (NP), as well as in portions of the Pacific Crest Trail (PCT) and selected JMT/PCT access trails. Convenience samples of wild animal scat found within I mile of trails were also collected. The fresh specimens were individually preserved both in 0.9% saline and polyvinyl alcohol (PVA)-containing tubes and stored at 4 degrees C until time of analysis. Bacteriological analysis was performed using standard microbiology laboratory procedures. PVA samples were stained with trichrome and were then examined by a parasitologist. RESULTS Collection: A total of 186 trail miles were sampled, including 113 on the JMT (Yosemite 37, Kings 53, and Sequoia 23). The PCT samplings included 24 miles, and NP and wilderness area access trails added an additional 49 miles. A total of 102 samples were collected, which included 81 samples from pack animals and 21 identified as having come from wild animals. Pack Animal Bacteria: All plated specimens grew large numbers of commensal gut flora. Potential pathogenic bacteria were found in only 12 samples and included Hafnia alvei (4), Serratia odorifera (1), Citrobacter freundii (1), Escherichia vulneris (1), Clostridium clostridioforme (1), Yersinia enterocolitica (1), Sherwinella putraformus (1), and Enterobacter spp (4). No Escherichia coli O157, Salmonella, or Aeromonas were found. Microscopic examination for protozoal organisms revealed occasional commensal ciliates and I Giardia. Wild Animal Pathogens: One specimen grew Y enterocolitica, and another grew Enterobacter amnigenus. CONCLUSIONS We found a low prevalence of human pathogens in pack animal manure on the JMT.

Antigen detection for human immunodeficiency virus.

The recent development of enzyme immunoassay procedures for the direct determination of human immunodeficiency virus (HIV) antigens has been of significant benefit in both clinical and research applications. The historical development of HIV antigen assays as well as their current and future applications for use in the clinical microbiology laboratory are reviewed. A detailed description of selected commercially available assays is presented, and a comparison is made of various parameters, including sensitivity, specificity, and cost. The use of the HIV antigen assay as an alternative to the reverse transcriptase assay in virus culture applications is also discussed. In addition, the diagnostic and prognostic utility of the HIV antigen assay is considered for various patient groups, including neonatal, high-risk asymptomatic, seronegative, and seropositive patient populations. The use of the HIV antigen assay as an adjunct to anti-HIV antibody testing, as well as its utility in assessing the therapeutic efficacy of antiviral drug therapy, is discussed. The biology of HIV antigen expression and modulation of anti-HIV antibody titers during infection are also discussed in terms of two possible models.

Anti-HTLV-III testing of blood donors: reproducibility and confirmability of commercial test kits.

Since 2% of the cases of Acquired Immune Deficiency Syndrome (AIDS) have been attributed to transfusions of blood and blood products, licensed tests to detect antibody to the human T‐lymphotropic virus type III (anti‐HTLV‐III) have been put into practice to reduce the risk of transfusion associated AIDS. Two commercial ELISA kits (Abbott and ENI) were used to test for anti‐HTLV‐III in 100 coded samples from individuals with AIDS, at high risk for AIDS, or with low risk for AIDS and in 1280 unlinked blood donor serums. From the 100 coded samples, both Abbott and ENI tests identified 51 of 52 coded samples with anti‐ HTLV‐III which were confirmable with Western blot analysis. Initial testing of the donor serums by Abbotts test revealed 20 reactives, of which 5 were repeatably reactive; initial testing by ENIs test revealed 25 reactives, of which 14 were repeatably reactive. However, only 3 donor serums were repeatably reactive by both test kits, out of 17 repeatable reactive by either, and no ELISA positive samples were confirmed by Western blot or IFA. Before a blood donor is notified of “anti‐HTLV‐III reactivity”, tests demonstrating this should be both reproducible and confirmable by at least one additional test.