By Carol A. Kemper, MD, FACP, Clinical Associate Professor of Medicine, Stanford University, Division of Infectious Diseases; Santa Clara Valley Medical Center, Section Editor, Updates; Section Editor, HIV, is Associate Editor for Infectious Disease Alert.

Tamiflu-Resistant Influenza A Increasing

ProMED-mail post, September 5, 2008; www.promedmail.org

Resistance to oseltamivir (tamifluR) has increased in Influenza A isolates around the world at an alarming rate. Levels of resistance now range from 13% in Chile to 100% (10 of 10 isolates) in Australia. Oseltamivir-resistant strains were initially recognized in Norway, and previously occurred with less than 1% frequency. As reported in Infectious Disease Alert in April 2008, the level of resistance to oseltamivir in 437 isolates of Influenza A/H1N1 identified in 10 European countries from November 2007-January 2008 was 14%. All of the isolates carried the same amino acid substitution (H274Y) of the neuraminidase, which is known to cause high level oseltamivir resistance. At that time, the oseltamivir-resistant strains remained sensitive to zanamivir (Relenza) and amantadine/rimantidine.

Recent reports document the death of a 67-year-old man due to osel-tamivir-resistant Influenza A. He was receiving treatment for chronic lymphocytic leukemia, but remained stable. He presented with bilateral pneumonia and rapidly deteriorated, requiring ventilatory support. Respiratory specimens were positive for Influenza A/H1N1, and he was started on oseltamivir on the sixth hospital day. Despite this, he developed progressive respiratory failure and ultimately succumbed to his illness. Remarkably, by the 13th day of his hospitalization, the laboratory reported sequencing results confirming the H274Y mutation. His isolate was also resistant to amantadine. He had no known contact with any person with influenza.

Studies suggest that the transmission of this strain is not correlated with treatment with oseltamivir; the strain seems to be appearing in countries where use of this agent is not frequent, suggesting that selective pressure is not responsible for circulation of this strain. Even more puzzling are reports that this mutation may diminish the replicative capacity of the organism, making it less virulent, although this is controversial. Obviously, this infection proved fatal for this modestly immunosuppressed man.

Clinicians in the United States should be aware that this strain is now being reported from the United States and Canada, and that oseltamivir may not be effective. Persons with serious respiratory or influenza-like illness should be tested for influenza using newer, rapid screening EIA techniques. Cultures of respiratory secretions are essential for epidemiologic purposes, and your public health department can assist in identifying isolates.

Utility of Newer TB Blood Tests

Pai M, et al. Systematic review: T-cell-based assay for the diagnosis of latent tuberculosis infection: an update. Ann Intern Med. 2008; 149:177-184.

Newer gamma-interferon-based assays for detection of an immune system response to latent infection from M. tuberculosis (LTBI) have many potential benefits over standard tuberculin skin testing (TST). Such assays eliminate the need for two-step skin testing, decreasing personnel time and office visits. In addition, these assays do not cross-react with BCG vaccination, giving them an advantage over TST in BCG vaccinated populations. One of these products, the Quantiferon-TB Gold assay (Cellestis, Victoria, Australia), has been approved by the FDA, and is already being utilized for screening of health care workers. Cellestis also makes a second gamma-interferon-based assay called the QuantiFERON-TB Gold In-Tube test. Another gamma-interferon-based assay, previously available only in Europe, now approved for use in the United States is the T-SPOT.TB test (Oxford Immunotec, Oxford, United Kingdom).

Differences between these assays were discussed in an accompanying editorial.1 The QuantiFERON-TB Gold assay uses whole blood cells, with an undetermined number of white blood cells, and provides quantitation of the total amount of interferon-gamma produced in the supernatant. The T-SPOT.TB test utilizes peripheral blood mononuclear cells and provides quantitation of the level of interferon-gamma produced per PBMC. Both assays can be processed within one day, although the first is somewhat less complicated to perform in the lab. There is no potential for boosting with either assay. Both assays make use of an internal positive control, with a sample well containing a potent nonspecific stimulator of interferon-gamma production. Thus, the failure of the positive control in these assays provides information about the ability of the test subject's T cell function.

Pai et al compared the sensitivity and specificity of the three assays for detection of TB infection based on a meta-analysis of data reported from 38 published studies. To assess the relative specificity of the assays, the study sample had to comprise healthy, low-risk persons without known exposure to TB. To assess the sensitivity of the assays, the data had to be based on individuals with confirmed active tuberculosis. The data was extracted by one of the investigators, and confirmed separately by a second.

The pooled specificity of both QuantiFERON-TB assays was 99% for non-BCG-vaccinated subjects and 96% for BCG vaccine recipients. The pooled specificity of the T-SPOT.TB test was slightly lower at 93%. The specificity of all three assays compared favorably with TST, which also had a high specificity (97%) in non BCG-vaccinated patients; but provided variable and poorer results in BCG recipients. Hence, the risk of false-positives with any of the newer assays is low, and compares favorably with TST, which still provides excellent specificity in most patients, except those who have received BCG.

The sensitivity of the three assays was more variable: pooled sensitivities for the QuantiFERON-TB Gold and QuantiFERON-TB Gold In-Tube assays were 78% (95% CI, 73% to 82%) and 70% (CI, 63% to 78%), respectively. In contrast, the pooled specificity of the T-SPOT.TB test was 90% (95% CI, 86% to 93%).

Any of these assays, including standard TST, provide variable sensitivity, depending on the population tested, and whether screening for LTBI or active TB infection. All three gamma-interferon-based assays and TST have diminished sensitivity in persons with immune system suppression, especially those with T cell dysfunction. For example, the utility of the QuantiFERON-Gold assay was evaluated in 242 persons with suspected active TB at the San Francisco Public Health Department, 45 of whom were diagnosed with active TB (82% culture-confirmed).2 QuantiFERON-Gold results were positive in 55%, negative in 38%, and indeterminate in 7%, providing a sensitivity of 60% for this group, with a negative predictive value of 86%. Patients with extra-pulmonary disease were more likely to have falsely-negative results compared with those with pulmonary disease (35% vs 4%, p < .05). QuantiFERON-Gold results were positive in only one of three HIV-infected patients with active TB.

Thus, the interferon-based assays are highly specific for detection of LTBI, irrespective of prior BCG vaccination. In immune-competent persons, all three assays are sensitive for the detection of LTBI, and somewhat less sensitive for detection of active TB, although various studies suggest the T-SPOT.TB test may have slightly greater sensitivity. The tests perform differently in patients with cellular immune suppression where, again, the T-SPOT.TB may be somewhat more sensitive. Clinicians should be mindful that a negative interferon-gamma assay does not rule-out a diagnosis of TB, similar to our experience with skin testing.


  1. Richeldi L. An update on the diagnosis of tuberculosis infection. Am J Respir Crit Care Med. 2006:174:736-742.
  2. Dewan PK, et al. Low sensitivity of a whole-blood interferon-gamma release assay for detection of active tuberculosis. Clin Infect Dis. 2007;44:69-73.

Ebola Vaccination of Great Apes

Dolgin E. Baiting Ebola. The Scientist. 2008;22:22.

Scientists and primate biologists have teamed up in Leipzig, Germany, to create a novel vaccine delivery system for delivering a newly developed live-attenuated Ebola vaccine to chimps and gorillas. Ebola is just as deadly for apes and humans, and in 2006, is estimated to have claimed the lives of approximately 5,000 gorillas in a 2,700 square meter area in central Africa.

The most effective Ebola vaccine for apes is a live-attenuated vaccine — if it can be appropriately delivered to the animals. Hypodermic darts are only effective if you can actually find the animals, and using bait risks wasting much of the vaccine on the forest floor. What you really need is some way of getting the animals to eat every last drop of the vaccine, absorbing the virus through cheek epithelial cells where it can quickly stimulate a vigorous immune system response.

But how to deliver a 6 cm. slug of vaccine-impregnated bait to a chimp? Candy! Like children, chimps and other apes love sweets. The scientists set out to perform an (non-randomized) experiment with paraffin-coated bait (to withstand the tropical heat) at the Leipzig zoo. The test baits (sans vaccine) were colored red, yellow, or orange, and flavored with banana, mango, or fig. One gorilla quickly ate all of the baits at once, with no apparent preference. But the chimps went crazy for the mango-flavored baits, which seemed to be a consistent hit.

The next step is to test the sample baits in the wild. There is some concern that placement of the baits is key — lest other forest creatures like rodents sample the baits before the apes get a chance to find them. The stability of the attenuated virus is also a concern, especially if the product does not end up in the right hands and mouths.