Zika vaccine Phase I trial to begin next month in India

measuring microcephaly. Photo WHO

The Phase I trial will not involve pregnant women.

Bharat Biotech International Ltd in Hyderabad will start a Phase I clinical trial of Zika virus vaccine (MR 766) in two centres in India next month. The inactivated  vaccine being tested in humans will be an African Zika virus strain. “We have already got the approval from DCGI [Drug Controller General of India] in March to carry out the Phase I clinical trial,” says Dr. Krishna Ella, Chairman and Managing Director of Bharat Biotech.

It will be a randomised, placebo-controlled, double-blind trial involving 48 adults, both men and women (who are not pregnant). Each volunteer will receive two doses of the vaccine on days 0 and 30 and followed-up for a year for both safety and immune responses. Three different dose ranges (2.5, 5 and 10 microgram) will be tested.

There will be three arms in the trial and each arm will get one of the three different dose ranges. There will be 16 subjects per arm.

The Phase I human clinical trial is being initiated based on the promising results seen in animals trials. In the animal trials, two doses of the vaccine made using an African Zika virus strain conferred 100% protection against mortality and disease in mice. The protection was the same when the mice were infected with an Asian and an African Zika virus strains.

While the viral load was “undetectable” in the case of vaccinated mice, the amount of virus in unvaccinated mice shot up four days after being infected. All the unvaccinated mice died eight days after infection by the African strain of the virus and 12 days after infection by the Asian strain.

Published in The Hindu on April 18, 2017

India begins caged trials of GM mosquitoes to control dengue, chikungunya


Laboratory studies using GM mosquitoes have been carried out in India since 2012

Outdoor caged trials to demonstrate the efficiency of genetically modified mosquitoes to suppress wild female Aedes aegypti mosquito populations that causes dengue, chinkungunya and Zika were launched on January 23 in Dawalwadi, Badnapur in  Maharashtra’s Jalna district.

Based on the results of the caged trials, which use the Release of Insects carrying Dominant Lethal genes (RIDL) technology, and permission from Indian regulatory authorities, Gangabishan Bhikulal Investment and Trading Limited (GBIT) and Oxitec, plan to conduct open field trials in the country.

The announcement was made on January 23 after the field cage facility was inaugurated by Dr. Soumya Swaminathan, Director General of the Indian Council of Medical Research.

Laboratory-based studies have already been carried out in India since 2012 by GBIT and Oxitec and these studies have demonstrated the compatibility of Aedes aegypti mosquitoes. “The efficiency to kill offspring was over 99% and male mosquitoes imported from the U.K were able to mate with locally available wild female mosquitoes and the longevity of imported mosquitoes was the same as the wild ones,” says Dr. Shaibal Dasgupta, Project Leader, GBIT, Delhi.

Oxitec’s technology uses genetically modified male Aedes aegypti mosquitoes that carry a dominant lethal gene. When male GM mosquitoes mate with wild female mosquitoes the lethal gene is passed on to offspring. The lethal gene in the offspring produces a protein that stops their cells from functioning normally and prevents other genes essential for survival from turning on. This causes the mosquito larvae to die before reaching adulthood.

Since male mosquitoes do not bite humans, the release of genetically modified males will not increase the risk of dengue, chinkungunya and Zika.

mosquito-larvae“The caged trials, which will last 50-55 weeks, will study the efficiency of vector suppression and mating capacity,” Dr. Dasgupta says. “Surveillance [to undertake open field studies] to gather data on predominance of Aedes mosquitoes in the wild has already started. The open field studies, which will be of one-year duration, will be conducted in two villages and two control villages in close vicinity of the caged study area in Jalna.”

“It is a promising technology and India must certainly look at new vector control methods,” says Dr. Swaminathan. “From studies carried out in other countries we know the safety is beyond doubt, but efficiency has to be proved, especially in big cities and towns.”

“There are practical problems of raising large number of mosquitoes needed for vector control — 100-150 [GM] mosquitoes are needed per person for months together. So some innovation in breeding techniques is needed,” says Dr. Swaminathan.

In the case of the RIDL technology, the genetically altered male mosquitoes have to be released in large numbers at regular intervals. Only then can they compete with the wild normal male insects for mating. Since the larvae die before reaching adulthood, the technology is a “self-limiting approach (the genetic modifications are not perpetuated in wild populations)”, notes a piece published in The Lancet (February 1, 2016).

The first open field trial by Oxitec was carried out in 2010 in the Caribbean island of Grahnd Cayman. The trial was successful in reducing the mosquito population by 80%. Similar trials have since been carried out in west Panama, Malaysia and more recently in Brazil with at least 90% reduction in vector population.

Another alternative

“India is looking at another alternative. We are about to sign a memorandum of understanding next month with Monash University for vector control using Wolbachia-infected A. aegypti mosquitoes,” she says. Pilot studies of mosquitoes harbouring the Wolbachia bacterium are being conducted in Colombia, Brazil, Australia, Vietnam and Indonesia to help control the spread of dengue virus.

Vector control using Wolbachia-infected A. aegypti mosquitoes is achieved by using the life-shortening bacteria strain in both male and female mosquitoes. Uninfected wild females embryos fertilised by Wolbachia-infected males fail to develop, while embryos from infected females fertilised by infected or uninfected wild males survive. As Wolbachia is maternally inherited, the bacteria are passed on to offspring immaterial of which male mosquito the female mosquito mated with. Dengue, Zika or chikunguya viruses cannot replicate when mosquitoes have Wolbachia and hence cannot be transmitted to humans when the mosquito bites.

Unlike the RIDL technology, a salient feature of Wolbachia is that it is self-sustainable, making it a very low-cost intervention to mosquito vector control. The downside is that the release of even a single female mosquito infected with Wolbachia bacteria could “potentially lead to the alien Wolbachia spreading in the target population” says a June 2013 report in the journal Pathogens and Global Health.

Published in The Hindu on January 25, 2017

Top health stories of 2016

On the health front, the year 2016 witnessed a few setbacks and but also a few positive developments. Here are the most important health developments of 2016

1. Trial results confirm Ebola vaccine provides high protection

who-ebola-optimizedThe Ebola virus that killed more than 11,300 people in three West African countries — Guinea, Liberia and Sierra Leone has finally met its match.

In December, an Ebola vaccine trial carried out in 2015 in nearly 12,000 people in Guinea was found to be safe and highly protective against the deadly virus.

No Ebola cases were recorded in nearly 6,000 people who received the vaccine, while there were 23 cases in the arm that received the vaccine after a 3-week delay. The trial used an innovative design called a “ring vaccination” approach — the same method that was used to eradicate small pox. In the ring vaccination strategy, whenever a new Ebola case was diagnosed, all the people who may have been in contact with that person were traced and included in the trial.

The vaccine was found to have high efficacy even when the interim results were announced. So everyone included in the trial was offered the vaccine immediately, including children older than 6 years.

A total of 28,616 Ebola cases and 11,310 deaths have been reported in three countries, as on June 2016.

Oraquick2. HIV self-testing gets a shot in the arm

Access to and uptake of HIV diagnosis got a shot in the arm when the WHO in November released new guidelines on HIV self-testing. Lack of an HIV diagnosis has been a major obstacle in the war against HIV.

Today, 40 per cent of all people with HIV (over 14 million) are simply unaware of their status. While more than 18 million people with HIV are currently taking anti-retroviral treatment (ART), and a similar number is still unable to access treatment as many are not aware of their HIV positive status.

Only 30 per cent of men have tested for HIV. As a result, men with HIV are less likely to be diagnosed and more likely to die of HIV-related causes than women.

Testing also remains low among high-risk population such as men who have sex with men, sex workers, transgender people, injection drug users, and people in prisons.  These people comprise nearly 44 per cent of the 1.9 million new adult HIV infections that occur each year.

People can use oral fluid or blood- finger-pricks to discover their status in the comfort of their house with no fear of being subjected to stigma and discrimination. This is bound to encourage more people to get tested.

Results are ready within 20 minutes or less. Those with positive results are advised to seek confirmatory tests at health clinics. Twenty three countries currently have national policies that support HIV self-testing. India is yet to approve HIV self-testing.

zika-microcephaly - Photo WHO3. Zika virus and public health emergency of international concern

On November 18, the World Health Organisation declared that Zika virus is no longer a public health emergency of international concern. It was in February that the WHO declared the Zika outbreak and congenital malformations and neurological disorders in newborns believed to be caused by the virus as a global public health emergency. Brazil has been the worst affected by the virus.

A distinct pattern of birth defects caused by Zika infection during pregnancy is now officially known as congenital Zika syndrome.

In November, a study showed that a small group of Zika-infected babies in Brazil who were born with normal-size heads had developed microcephaly five months to a year after birth.

The virus that has caused about 2,300 confirmed cases of microcephaly (a congenital disorder where babies are born with a small head) since May 2015. The link between Zika virus and microcephaly was established in May this year. After the initial spread to 67 countries, including countries in Southeast Asia, the spread has since slowed down.

While Aedes species of mosquitoes is the primary vector that transmits the virus, it can also be transmitted though semen, blood, tears and other blood fluids. Unlike in the case Ebola, scientists are yet to find an efficacious vaccine against Zika virus. And we are yet to fully understand the entire spectrum of neurological problems that may show up in children born with microcephaly. Meanwhile, the WHO has advised that pregnant women refrain from travelling to countries where local transmission of Zika virus is still prevalent.

GeneXpert4. Incidence of TB in India increases sharply

Although global TB deaths dropped by 22 per cent between 2000 and 2015, TB is still one of the top 10 causes of death worldwide. It kills more people than HIV and malaria.

Of the estimated 10.4 million new cases worldwide, the estimated number of new cases in India alone has increased sharply from 2.2 in 2014 to 2.8 million in 2015. The true incidence in India will be known once the national TB prevalence survey scheduled to begin next year is completed. Besides the uptick in incidence, the number of estimated deaths caused by TB has also more than doubled from 220,000 in 2014 to 483,000 in 2015.

The sudden increase in TB incidence in India is due to 34 per cent increase in case notification between 2013 and 2015 by doctors in the private sector. Yet, in 2015, notification by private-sector doctors was only 16 per cent of the total case notification. Of the 2.8 million estimated cases each year in India, only 1.7 million cases both in the public and private sector were notified in 2015. So the remaining 1.1 million cases are simply not known.

anopheles-gambiae-mosquito-james-gathany-cdc5. Sri Lanka becomes malaria-free

In September, Sri Lanka achieved a huge public health success when the WHO declared it as malaria-free. There has been no local transmission nation-wide in Sri Lanka since October 2012. The WHO certifies a country as malaria-free when the chain of local transmission has been interrupted nationwide for at least three years. With this, Sri Lanka joins the ranks of 34 countries that have been certified as malaria-free since 1960s.

With no local transmission currently, Sri Lanka’s focus is on preventing the re-introduction of malaria from outside, particularly countries such as India, which are malaria-endemic. There were 95 re-introduced cased in 2013, 49 cases in 2014 and 36 cases in 2015.

Sri Lanka was able to win the war against malaria by targeting both the vector and the parasite. This was achieved by active detection of cases and targeting parasite carriers by screening people whether they had malaria symptoms or not.

It came to eliminating malaria in 1963 when it reported just 17 cases including six that were due to local transmission.

Zika - antenatal-care. Photo WHO6. Thailand ends vertical transmission of HIV

In June, Thailand became the first country in the Asia-Pacific region to end vertical transmission of HIV from mother to child.

In 2000, an estimated 1,000 children in Thailand were infected with HIV due to vertical transmission. But in 2015, the numbers dropped dramatically to just 85, a decline of more than 90 per cent. The rate of mother-to-child transmission of HIV has been reduced to less than 2 per cent.

Thailand achieved this feat by ensuring that 98 per cent of all pregnant women with HIV had access to antiretroviral therapy. Women with HIV have a 15-45 per cent chance of transmitting the virus to their children during pregnancy, labour, delivery or breastfeeding if they are not on treatment during pregnancy. But the risk drops to just over 1 per cent if antiretroviral medicines are given to both mothers and children throughout the stages when infection can occur.

Treatment to prevent vertical transmission is not 100 per cent certain.  So vertical transmission is considered to be eliminated when a country successfully achieves a reduction in transmission to such a low level that it no longer constitutes a public health problem.

Besides treating pregnant women with HIV with antiretroviral medicines, Thailand witnessed a sharp drop in the number of women in the child-bearing age becoming infected with HIV. Between 2000 and 2014, the annual number of women newly infected with HIV fell from 15,000 to 1,900 — an 87 per cent reduction.

polio7. Polio end game starts across the world

In April, 155 countries including India switch from using a trivalent oral polio vaccine that contains type 1, type 2 and type 3 strains to a bivalent oral polio vaccine that contains only type 1 and type 3 strains. The global vaccine switch took place between April 17 and May 1.

The reason for removing the type 2 strain from the vaccine was to confer better protection against polio. The oral polio vaccine contains live, weakened virus, which on rare occasions can turn virulent and cause vaccine-derived poliovirus. Though wild poliovirus type 2 was eradicated in 1999, all type 2 polio cases have been caused only by vaccine-derived polioviruses.  The type 2 strain in the trivalent OPV had caused over 90 per cent of vaccine-derived poliovirus cases in the world in the last 10 years.

Polio, including vaccine-derived polio, can be eradicated only when oral polio vaccine is eventually withdrawn after wild polio transmission has been stopped. Removing type 2 strain from the vaccine and switching over to a bivalent vaccine is the “first major step” of this withdrawal process.

Published in The Hindu on December 28, 2016

No complacency on Zika

zika-microcephaly - Photo WHO

There have been 2,300 confirmed cases of microcephaly since May 2015. Photo: WHO

The World Health Organisation has declared that the Zika virus no longer constitutes a public health emergency of international concern. This brings to an end the heightened global focus on the virus that has caused about 2,300 confirmed cases of microcephaly (a birth defect manifesting in a smaller head size) since May 2015. The WHO had declared the Zika virus a public health emergency on February 1, considering the high number of neurological disorders reported in Brazil and a similar cluster in French Polynesia in 2014. Among the reasons cited were the unknown causal link between the virus and microcephaly and neurological complications, the possibility of its global spread, lack of vaccines and diagnostic tools, and the lack of immunity to the virus in newly affected countries. The link between Zika and microcephaly was established in May, the hunt for a potent vaccine and reliable diagnostic tool has begun, and scientists have been able to find the routes of transmission. However, the global risk assessment has not changed. The spread of Zika to 67 countries and territories is a grim reminder of the lack of immunity against the virus and the abundance of mosquito vectors. A dozen countries have reported local transmission.

Despite the link between the Zika virus infection and microcephaly being well established, the entire spectrum of challenges posed by the disease is not known. The WHO Emergency Committee has called for sustained research and dedicated resources to address the long-term challenges posed by babies born with microcephaly, but signalling the end of the global emergency may lead to lowering of the global alert. There should be no setback to funding, the global search for effective vaccines and diagnostic tests, and creating awareness about the risk of sexual transmission. For instance, it is not clear why more babies were born with microcephaly in northeast Brazil compared to the rest of the country or why the country had a higher caseload than others. This information is crucial to understanding the link between Zika infection and microcephaly, and thereby to containing incidence where the mosquito vector is predominant. Medical journals should continue to provide free and immediate access to papers on the Zika virus, which played a crucial role in information-sharing. The WHO has said it is “not downgrading the importance of Zika” and that its “response is here to stay”. It now needs to ensure that vigilance remains high despite the decline in incidence.

Published in The Hindu on November 23, 2016

Zika vaccine trials on monkeys show promise

Zika vaccine-Optimized

The Pilot Bioproduction Facility at WRAIR has manufactured a batch of killed Zika virus vaccine to be used in clinical studies. – Photo: WRAIR

Tests involving three different kinds of Zika vaccines on rhesus monkeys have produced good immune responses and caused no adverse effects, raising hopes for the development of an effective vaccine for use in humans. None of the vaccinated monkeys developed infection when later challenged with Zika virus. Also, the transfer of antibodies at higher doses from vaccinated monkeys to Zika-naïve mice and monkeys was found to offer “complete protection” when the animals were exposed to Zika virus.

Three vaccine approaches

The results of the study by a team led by Dan H. Barouch from Harvard Medical School, U.S. were published today (August 5, 2016) in the journal Science.

Compared with DNA vaccine, the killed virus vaccine was found to be “more potent” in rhesus monkeys.

Zika virus strains from both Brazil and Puerto Rico were used for developing the vaccines. The study used three vaccine approaches — a purified killed Zika virus, DNA from the Zika virus woven into a plasmid, and inserting Zika genes into adenoviruses which infect cells and trigger immune responses.

In addition, passive protection was also tested by using antibodies from vaccinated monkeys to protect Zika-naïve mice and monkeys. While all the mice that got the antibodies at a higher dose were completely protected, one of the two monkeys that got the antibodies was completely protected.

measuring microcephaly. Photo WHO

A potent Zika virus vaccine for use in humans will prevent the birth of children with micocephaly. – Photo: WHO

The vaccine study using purified killed virus vaccine was conducted on 16 rhesus monkeys. Eight monkeys got the purified killed virus vaccine and the remaining eight did not get a vaccine. Within two weeks of immunisation, all vaccinated animals developed Zika-specific binding and neutralising antibodies. After a booster vaccine was administered at four weeks, the antibody levels increased substantially. When these animals were exposed to Zika virus, they showed “complete protection”. The virus was not found in blood, urine and other body fluids including cervicovaginal secretions.

In another experiment, 12 rhesus monkeys were immunised with either a DNA vaccine or an adenovirus vector-based vaccine. While the monkeys developed antibodies when both the vaccines were administered, the adenovirus vector-based vaccine “provoked a broader and a more potent antibody response”. Both the vaccines conferred complete protection when the vaccinated monkeys were exposed to Zika virus four weeks after vaccination.

Compared with DNA vaccine, the killed virus vaccine was found to be “more potent” in rhesus monkeys, the authors say. The adenovirus vector-based vaccine offered complete protection after only one dose of the vaccine.

Passive protection through transfer of antibodies from vaccinated animals to Zika-naïve animals offered protection against Zika virus even at relatively low antibody titers.

Phase I human clinical trials using killed Zika virus vaccine and other candidate vaccines are expected to begin later this year.

Published in The Hindu on August 5, 2016

Infectious Zika virus found in saliva and urine

Zika virus - Photo  NIAD

Infectious Zika virus  was found in both urine and saliva samples of two patients in Brazil.

If researchers had earlier reported the presence of Zika virus in urine and saliva samples (besides semen, blood and breast milk), a new study has been able to find infectious Zika virus particles in urine and saliva samples of two patients during the acute phase of infection. While in theory the presence of Zika virus in urine and saliva can lead to yet another route for Zika transmission, at this point in time, it is not clear whether it can actually happen. It is extremely important to know this as it will help in controlling Zika virus transmission.

According to the results of a study published in the journal PLOS Neglected Diseases, the viral concentration was as high as 80 plaque forming units per ml in a saliva sample of a patient in his acute phase of infection.

The urine and saliva samples from nine patients from Rio de Janeiro suspected of Zika virus infection were examined. The samples were collected between January 14, 2016 and February 2, 2016. Six of the seven women were pregnant with gestational ages varying from 18 to 33 weeks.

Of the nine patients studied, Zika virus was detectable in urine and saliva samples in only two patients (patients 1 and patient 6). Generally, whenever urine and saliva samples are collected from the same patients, the viral load is higher in urine than saliva. Yet, the researchers were able to detect more number of plaques in saliva samples. Also, the virus was found in the urine sample of only one patient (patient 1) though the viral load in the urine sample of patient 1 was 100 times lower than patient 6.

Viral detection and recovery from urine and saliva samples might be related to severity of infection as well as the period of specimen collection after the onset of Zika symptoms. The pH of saliva and urine also affects the ability to recover the virus. The pH in urine varies from 4.5 to 8.0 while saliva assumes values near neutral pH. The pH of the urine sample of patient 6, which was mildly acidic at 5.6, might be the reason why no virus was detected.

“The importance of Zika virus in urine for human transmission is unexplored, but the effect of acidic pH on viral viability might represent a serious restriction for viral spreading,” writes Myrna C. Bonaldo, the senior author of the paper, from Instituto Oswaldo Cruz, Fiocruz, Rio de Janeiro, Brazil.

Virus in saliva

At the same time, the very presence of viable Zika virus in saliva “does not always indicate that the virus can be transmitted orally and become epidemiologically relevant”.  This is because saliva contains antiviral molecules.

Despite the protective nature of saliva, ulcer or disruption of oral skin can facilitate the entry of the virus into the body. As the latest study has found infectious Zika virus in saliva samples, there is a possibility of person-to-person oral transmission of Zika virus when the oral cavity mucus membrane is disrupted. More studies are needed to confirm this.

“In conclusion, the detection of infective Zika virus in saliva and urine of patients deserves a more detailed study to establish whether or not these fluids contribute to viral transmission. Surely, these findings will be extremely relevant to prevent and control Zika virus transmission,” the authors write.

A cheap, sensitive diagnostic tool for Zika virus developed

Zika virus - Photo  NIAD

The Zika virus. – Photo: NIAD

Scientists have developed a cheap, highly-reliable, instrument-free, point-of-care Zika virus detection test. The sensitivity of the test is very high — the presence of Zika virus can be detected even if the concentration of the virus is as low as 5 plaque-forming units (PFU). For comparison, the amount of Zika virus present in people showing symptoms ranges from 1,000 PFU per mililitre to 10,00,000 PFU/mL.

And results will be known in less than 40 minutes when the dye turns purple. The higher the concentration of the virus in the sample, the quicker will be the detection. The sensitivity can also be increased by using higher sample volume.

“Our system is particularly suitable for resource-poor settings, where centralized laboratory facilities, funds, and trained personnel are in short supply, and for use in doctors’ offices, clinics, and at home,” says a paper published in the journal Analytical Chemistry.

While the diagnostic test can use any of the body fluids — saliva, blood, urine or semen, the researchers tested the sensitivity using only saliva samples. Zika virus is present in higher concentration in saliva compared with blood.  Also, collecting saliva samples is easy, simple and non-invasive. It is becomes particularly important when the testing has to be done on newborns or just-born babies.

The test is specific to Zika virus and will not turn positive to other flaviviruses such as dengue and chikungunya, the researchers claim. This is because Changchun Liu, the corresponding author of the paper from the University of Pennsylvania, U.S., and his co-researchers identified a highly conserved region (hence will not change even when the virus mutates) of the Zika virus that is identical to 19 strains of the virus circulating in the Americas but very different from other common flaviviruses (like dengue and chikungunya).

The Zika virus used for testing the system was obtained from NIAD’s World Reference Center for Emerging Viruses and Arboviruses.

The experiments were carried out at ambient temperature of 18 degree C. Saliva samples from Zika-infected people were not used for testing the sensitivity of the system. Instead, saliva samples were spiked with Zika virus at various concentrations and then tested. The system relies on the heat producing (exothermic reaction) nature of magnesium-iron alloy reacting with water. Thus electricity is not needed to run the test.

The system uses a custom-made “microfluidic cassette that combines viral nucleic acid capture, concentration, and purification”, and also amplification before detecting the virus.

The microfluidic cassette containing saliva spiked with Zika virus is inserted into a chemically heated cup for 40 minutes. If samples contain as little as 5 plague-forming units, they turn purple, which is easily discernible by naked eyes.

While the current demonstration was restricted to qualitative detection of Zika virus, the method can be expanded to include quantification, they say.