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Objective: To determine whether autoclaving suspensions of vaccinia virus, herpes simplex virus (HSV), varicella-zoster virus (VZV), and Bacillus anthracis inactivate infectivity of these agents but allow detection of target DNA by LightCycler polymerase chain reaction (PCR)., Material and Methods: Swabs were inserted into tubes containing serial 10-fold dilutions (10(-1) to 10(-5); 500 microL; 6 samples per dilution) of vaccinia virus, HSV, VZV, or a single suspension of 10(8) colony-forming units of B anthracis (2 samples). One half of the samples were autoclaved, and the remainder were not. An aliquot of each not autoclaved sample served as a control for infectivity., Results: Autoclaving swabs saturated with suspensions of vaccinia virus, HSV, or VZV eliminated the infectivity of these agents; however, DNA was detectable in most autoclaved samples in dilutions of 10(-1) to 10(-4) by LightCycler PCR. All not autoclaved specimens were detected by culture (infectivity) except for VZV and, in most dilutions of 10(-1) to 10(-3), by assay of target DNA by LightCycler PCR. Similarly positive results were obtained for PCR assessment of sporulated B anthracis., Conclusions: Standard autoclaving procedures eliminated the infectivity of viruses (and B anthracis), but target DNA was often retained for detection by LightCycler PCR. Current recommendations indicate that the laboratory diagnosis of smallpox virus infection be performed only within Biosafety Level 4 facilities. We suggest that, in addition to the requirement for immediate coordination with public health officials, the federal government consider expanding the existing guidelines for processing these specimens to encourage immediate collection, autoclaving, and testing by LightCycler PCR to differentiate smallpox virus from other dermal pathogens such as HSV and VZV by specific qualified laboratories.

Rapid-cycle real-time polymerase chain reaction has immediate and important implications for diagnostic testing in the clinical microbiology laboratory. In our experience this novel testing method has outstanding performance characteristics. The sensitivities for detecting microorganisms frequently exceed standard culture-based assays, and the time required to complete the assays is considerably shorter than that required for culture-based assays. We describe the principle of real-time polymerase chain reaction and present clinical applications, including the detection of Bacillus anthracis, the causative agent of anthrax. This latter test is commercially available as the result of a collaborative venture between Mayo Clinic and Roche Applied Science, hence the designation The Mayo-Roche Rapid Anthrax Test.

Several important developments have occurred in recent years in the chemotherapy for and prophylaxis of parasitic infections. Although mefloquine is clearly the most effective agent for prevention of chloroquine-resistant falciparum malaria, its use has been compromised by side effects, both real and imagined. Well-designed studies have shown that side effects occur no more frequently with low-dose mefloquine than with chloroquine. Use of mefloquine in pregnant women has not been associated with birth defects, but the incidence of stillbirths may be increased. Malarone is a new agent that combines atovaquone and proguanil, and it may be as effective as mefloquine; however, it is not yet available in the United States. Several newer agents have appeared in response to the development of multidrug resistant Plasmodium falciparum, especially in Southeast Asia. Halofantrine is available for the treatment of mild to moderate malaria due to P. falciparum and for P. vivax infections. Because of severe toxic effects, use of halofantrine should be restricted to only those unusual and rare situations in which other agents cannot be used. Artemisinin (an extract of the Chinese herbal remedy qinghaosu) and two derivatives, artesunate and artemether, are active against multidrug resistant P. falciparum and are widely used in Asia in oral, parenteral, and rectal forms. The antibacterial azithromycin in combination with atovaquone or quinine has now been reported to treat babesiosis effectively in experimental animals and in a few patients. Azithromycin in combination with paromomycin has also shown promise in the treatment of cryptosporidiosis (and toxoplasmosis when combined with pyrimethamine) in patients with the acquired immunodeficiency syndrome (AIDS). Albendazole is currently the only systemic agent available for treatment of microsporidiosis, an infection primarily of patients with AIDS. In addition, albendazole and ivermectin have emerged as effective broad-spectrum antihelminthics, with albendazole becoming the drug of choice for hydatid disease (echinococcosis), neurocysticercosis, and most intestinal nematode infections (except strongyloidiasis and trichuriasis). Liposomal amphotericin B is the first drug approved by the Food and Drug Administration for the treatment of visceral leishmaniasis.

Objective: To report the first case of acute hepatitis E by a novel isolate acquired in the United States and confirmed by nucleotide sequencing., Material and Methods: We describe the clinical manifestations and the results of associated laboratory studies in a man who was found to have acute hepatitis E infection., Results: A 62-year-old man was hospitalized because of fever, abdominal pain, and jaundice. After an initial evaluation did not provide a cause, his serum was found to be positive for IgG anti-hepatitis E virus (HEV) by three antibody assays. Serum was also positive for HEV RNA by reverse transcriptase polymerase chain reaction (PCR). Sequencing results from the PCR products demonstrated substantial differences at the nucleotide level between this strain and the known Mexican and Burmese strains., Conclusion: On the basis of this initial report, HEV should be considered an etiologic agent in patients with acute non-ABC hepatitis in the United States.

A 33-year-old man with human immunodeficiency virus infection had severe protracted diarrhea. Radiologic assessment disclosed narrowing of the gastric antrum. Biopsy specimens revealed diffuse Cryptosporidium infection of the antral mucosa. Isolated antral narrowing due to Cryptosporidium gastritis should be added to the list of gastrointestinal complications associated with acquired immunodeficiency syndrome (AIDS).

In recent years, introduction of new and more effective agents has improved the overall therapy for parasitic infections. This field, however, is still plagued by numerous problems, including the development of resistance to antimicrobial agents (especially with malaria), unavailability of agents in the United States or lack of approval by the Food and Drug Administration, and major toxicities or lack of experience in pregnant women and children, which limits use in these groups of patients. Widespread resistance of Plasmodium falciparum to chloroquine and other agents has complicated the treatment and prophylaxis of this type of malaria. A combination of quinine and Fansidar is usually effective oral therapy for falciparum malaria; quinidine may be administered if intravenous therapy is needed. Mefloquine, which is currently recommended for prophylaxis against chloroquine-resistant P. falciparum, is also effective for single-dose oral treatment, although this regimen has not yet been approved by the Food and Drug Administration. Metronidazole has been widely used for treatment of gastroenteritis due to Entamoeba histolytica and Giardia lamblia (not approved by the Food and Drug Administration for the latter) and is considered safe and effective. A new macrolide, azithromycin, has been reported to be effective for cryptosporidiosis in experimental animals; currently, no effective therapy is available for human infections. Combinations of sulfonamides with other antifolates, trimethoprim or pyrimethamine, are recommended therapy for Pneumocystis carinii pneumonia or toxoplasmosis, respectively. Therapies for the various types of leishmaniasis and trypanosomiasis are complex, often toxic, and often of limited efficacy. The benzimidazoles are effective for roundworm infections, although thiabendazole has severe toxic effects. The recent introduction of ivermectin has revolutionized the treatment and control of onchocerciasis. Another relatively new agent, praziquantel, is a true broad-spectrum anthelmintic agent that is effective against most trematodes, many adult cestodes, and larval cestodes as well (especially cysticerci of Taenia solium).

Laboratory tests that are usually considered helpful in guiding antimicrobial therapy include antimicrobial agent susceptibility tests, determination of bacterial production of beta-lactamase, and assay of specific antimicrobial levels in serum and other body fluids. Susceptibility tests should be performed primarily on clinically significant isolates from critical specimens (such as blood or other normally sterile body fluids or tissues) with use of standardized methods established by the National Committee for Clinical Laboratory Standards. Reporting of results should be selective so that clinicians are encouraged to use the least expensive but useful agent in a group--for example, first-generation rather than third-generation cephalosporins. Because standardized methods are not available for assays of serum inhibitory and bactericidal activity, the accuracy and clinical utility of these tests are as yet undetermined. Determination of bacterial resistance to antimicrobial agents is the most important goal of susceptibility testing. Special methods are needed to detect methicillin-resistant staphylococci, high-level aminoglycoside- and glycopeptide-resistant enterococci, and antimicrobial-resistant strains of Neisseria gonorrhoeae, Streptococcus pneumoniae, and Haemophilus influenzae. Accurate measurement of serum concentrations of antimicrobial agents is important to ensure that therapeutic levels have been obtained and to avoid excessive levels of potentially toxic agents such as aminoglycosides and glycopeptides, especially when renal function is compromised.

Results of antimicrobial susceptibility testing of anaerobes were reviewed for the last 5 years and compared with two previous surveys from this institution. Allowing for differences in methodology, there appears to be a striking increase in penicillin resistance by the "non-fragilis" Bacteroides. Penicillin concentrations of 50 microgram/ml were required to inhibit 70% of 43 strains tested. The clinical implications of this observation are not known, but isolated reports of therapeutic failure when penicillin was used against the non-fragilis Bacteroides have appeared. Penicillin resistance among Clostridium other than C. perfringens was also noted in the current study. Several strains of clindamycin-resistant Bacteroides fragilis have been observed during the last 2 years, and whether this represents a true increase in resistance will require close scrutiny of clindamycin susceptibility in the future. Future surveillance of antimicrobial susceptibility of anaerobic bacteria will be useful in the detection of any major changes in susceptibility profiles. This knowledge will potentially affect the choice of antimicrobial agent in patients with serious infections caused by anaerobes.

Metronidazole, a nitroimidazole derivative, is a unique antimicrobial agent that is active against both bacterial and parasitic organisms, although only the anaerobic members of these groups are susceptible. It has been used for the treatment of trichomoniasis for about 20 years and is also effective against amebiasis and giardiasis. More recently, metronidazole has emerged as a principal agent for the treatment of anaerobic bacterial infections. It is highly effective against all species of anaerobes except certain non-spore-forming gram-positive bacilli and cocci and is the only agent rapidly bactericidal against the Bacteroides fragilis group. Clinical studies have proved its efficacy in prophylaxis for elective colorectal surgical procedures and in the treatment of deep abdominal sepsis (usually in combination with another agent, such as an aminoglycoside). Metronidazole is the treatment of choice for nonspecific vaginitis that seems to be a mixed infection due to anaerobes and Gardnerella vaginalis. Adequate concentrations in the blood are produced after both oral and intravenous administration, and the side effects are infrequent and minimal.

Metronidazole, a nitroimidazole derivative, is a unique antimicrobial agent that is active against both bacterial and parasitic organisms, although only the anaerobic members of these groups are susceptible. It has been used for the treatment of trichomoniasis for almost 30 years and is also effective in amebiasis and giardiasis. More recently, metronidazole has emerged as a principal agent for the treatment of anaerobic infections. It is highly effective against all species of anaerobes except certain non-spore-forming gram-positive bacilli and cocci and is the only agent rapidly bactericidal against the Bacteroides fragilis group. The hydroxy metabolite is 65% as effective as metronidazole and may play a major therapeutic role. Clinical studies have substantiated its efficacy for prophylaxis during elective colorectal surgical procedures and the treatment of deep abdominal sepsis (usually in combination with another agent such as an aminoglycoside). Metronidazole is the treatment of choice for bacterial vaginosis and seems to be as effective as vancomycin for treatment of Clostridium difficile-related diarrhea and colitis. Good blood levels are produced after both oral and intravenous administration, and side effects are infrequent and minimal. Metronidazole should not be taken during the first trimester of pregnancy because of concerns about mutagenicity. Tinidazole and ornidazole are recently developed nitroimidazole derivatives that have even greater antimicrobial activity than metronidazole.

Campylobacter-like organisms (provisionally named C. pyloridis) were demonstrated in gastric biopsy specimens by histopathologic analysis and bacterial culture. C. pyloridis organisms were found in 12 of 26 patients (46%) with gastric or duodenal ulcer but in none of 10 healthy volunteers without histologic evidence of gastritis. Iatrogenic antral gastritis, induced by 7 days of treatment with nonsteroidal anti-inflammatory drugs, was not associated with the presence of C. pyloridis. Organisms were found in 6 of 24 patients who had undergone gastric operations, but the prevalence of C. pyloridis was not higher in those with symptoms of alkaline reflux gastritis than in asymptomatic postgastrectomy control patients. We conclude that C. pyloridis is less common in patients with drug-induced and postoperative gastritis than in patients with peptic ulcer.

Laboratory tests that can be helpful in guiding antimicrobial therapy include antimicrobial susceptibility testing, determination of bacterial beta-lactamase activity, assay of serum inhibitory and bactericidal activity, and assay of specific antibiotic levels in serum. When any microorganism is isolated from a normally sterile body fluid in a patient with clinical evidence of infection, susceptibility studies should be performed. The standardized disk test provides results that should be comparable from laboratory to laboratory but has the disadvantage of yielding results expressed only as susceptible, intermediate, or resistant. In contrast, dilution methods allow determination of the minimal inhibitory concentration of an agent, which can be correlated with blood, urine, and other body fluid levels of the antimicrobial agent. Accurate measurement of serum concentrations of antimicrobial agents is important when the margin between therapeutic and toxic levels is narrow, such as for aminoglycosides or vancomycin, and when a patient has renal failure and may have accumulation of high levels of antimicrobial agents that would normally be excreted by the kidneys.

Laboratory tests that can be helpful in guiding antimicrobial therapy include antimicrobial susceptibility testing, determination of bacterial beta-lactamase production, assay of serum inhibitory and bactericidal activity, and assay of specific antibiotic levels in serum. Susceptibility studies should be performed on any microorganism that is isolated from normally sterile body fluid in the presence of clinical evidence of infection. The standardized disk test provides results that should be comparable from laboratory to laboratory. Dilution methods, however, allow determination of the minimal concentration of an agent which inhibits growth, and this value can be correlated with blood, urine, and other body fluid levels of the antimicrobial agent. Determination of serum bactericidal activity is, in effect, an assay of the activity of antimicrobial-containing serum; it indirectly measures the combined effects of susceptibility of the test organism and serum concentration of the antimicrobial agent. Accurate measurement of serum concentrations of antimicrobial agents may be important when treatment includes agents that have a narrow margin between therapeutic and toxic levels such as the aminoglycosides (especially gentamicin) or in patients with renal failure, who may accumulate unusually high levels of antimicrobial agents normally excreted by the kidneys.

A case of giant cell arteritis is reported that was detected by bone marrow biopsy. The histopathologic features of the bone marrow lesion included an enlarged artery, whose wall showed intimal thickening, fibrinoid necrosis, destruction of the internal elastic lamina, and multinucleated giant cells. The demonstration of involvement of bone marrow vessels by giant cell arteritis reaffirms the systemic nature of the disease. Bone marrow biopsy is not the method of choice for establishment of the diagnosis of giant cell arteritis, but it may lead to the diagnosis in rare instances.

Results of antimicrobial susceptibility testing of anaerobic bacteria isolated at the Mayo Clinic were reviewed for 1982 through 1987 and compared with a previous survey during 1977 through 1981 at this institution. Between the earlier and the later period, clindamycin resistance increased in the Bacteroides fragilis group (from 4% of isolates to 8%). We noted continuing penicillin resistance among Bacteroides species other than B. fragilis and rare penicillin resistance among Fusobacterium organisms, with four isolates during the 1982 through 1987 period being beta-lactamase producers. The high levels of resistance to some agents seen in certain Clostridium species in 1977 through 1981 were not as great in the current survey. No major changes were noted in the susceptibilities of C. perfringens, anaerobic non-spore-forming gram-positive bacilli, and anaerobic gram-positive cocci.

Seventy patients with substantiated anaerobic infections were treated parenterally with clindamycin or chloramphenicol in a prospective, double-blind, randomized study. No significant differences in clinical response or toxicity were noted between the two groups of patients.

Laboratory tests that can be helpful in guiding antimicrobial therapy include antimicrobial susceptibility testing, determination of bacterial beta-lactamase production, assay of serum inhibitory and bactericidal activity, and assay of specific antibiotic levels in serum. Susceptibility studies should be performed on any microorganism that is isolated from normally sterile body fluid (blood, cerebrospinal fluid, pleural fluid, synovial fluid) in the presence of clinical evidence of infection. The standardized disk test provides results that should be comparable from laboratory to laboratory. Dilution methods, however, allow determination of the minimum concentration of an agent which inhibits growth (MIC), and this value can be correlated with blood, urine, and other body fluid levels of the antimicrobial agent. Determination of serum bactericidal activity is, in effect, an assay of the activity of antimicrobial-containing serum; it indirectly measures the combined effects of susceptibility of the test organism and serum concentration of the antimicrobial agent. Accurate measurement of serum concentrations of antimicrobials may be important when treatment includes agents that have a narrow margin between their therapeutic and their toxic levels such as the aminoglycosides (especially gentamicin) or in patients with renal failure, who may accumulate unusually high levels of antimicrobials normally excreted by the kidneys.

Moxalactam was administered (20 mg/kg intravenously every 8 hours) as single-drug empiric antimicrobial therapy to 63 patients with bacteremia who were neither neutropenic nor immunosuppressed. Six patients (10%) had microorganisms that were susceptible to moxalactam and resistant to all other antimicrobial agents tested; two patients (3%) had microorganisms that were resistant to moxalactam and other agents tested. Of these 63 patients, 47 (75%) were cured with moxalactam therapy. Nine patients (14%) had breakthrough bacteremia while receiving other antimicrobial therapy and were cured subsequently with moxalactam therapy alone. The two major risk factors for failure of moxalactam therapy were polymicrobial bacteremia and an extrahepatic intra-abdominal source of infection; these two conditions frequently coexisted. Six of nine patients with polymicrobial bacteremia died. Superinfection (one pseudomonal, five enterococcal) was responsible for 6 of the 16 treatment failures. Enterococcal superinfection occurred exclusively among patients who had received relatively prolonged therapy with moxalactam for extrahepatic intra-abdominal infection, especially intraabdominal abscess. These five patients died, and postmortem examination showed that enterococcal superinfection was the major cause of death in all. Mild, reversible adverse reactions associated with use of moxalactam occurred in 14 of the 63 patients (22%). None had clinically overt bleeding. The use of moxalactam alone seems to be safe and effective and a cost-effective alternative empiric antimicrobial therapy for most patients with bacteremia who are not immunosuppressed or neutropenic and who are not at high risk of having Pseudomonas or polymicrobial bacteremia.