Drug-resistant infections are outpacing treatments, and WHO says laboratory leaders are vital to protecting antibiotic effectiveness.
For laboratory leaders, the latest WHO report on antimicrobial resistance (AMR) underscores just how critical diagnostic testing, data accuracy, and surveillance capacity have become in the global fight against drug-resistant infections. With one in six bacterial infections now resistant to antibiotics, labs stand on the front lines. They are responsible not only for detecting resistant strains but also for generating the data that informs national and international response strategies.
According to a press release, the “Global antibiotic resistance surveillance report 2025”warns that between 2018 and 2023, “antibiotic resistance rose in over 40% of the pathogen-antibiotic combinations monitored,” with “an average annual increase of 5–15%.”
Data from over 100 countries reported to the WHO Global Antimicrobial Resistance and Use Surveillance System (GLASS) show that growing resistance to essential antibiotics “poses a growing threat to global health.” For the first time, the report presents “resistance prevalence estimates across 22 antibiotics used to treat infections of the urinary and gastrointestinal tracts, the bloodstream and those used to treat gonorrhea.” It examines eight common bacterial pathogens—Acinetobacter spp., Escherichia coli, Klebsiella pneumoniae, Neisseria gonorrhoeae, non-typhoidal Salmonella spp., Shigella spp., Staphylococcus aureus, and Streptococcus pneumoniae—each linked to these major infections.
Resistance Highest in South-East Asia and Eastern Mediterranean
WHO found that the “risk of antibiotic resistance varies across the world.” The highest resistance levels are in the WHO South-East Asian and Eastern Mediterranean Regions, “where 1 in 3 reported infections were resistant.” In the African Region, “1 in 5 infections was resistant.”
According to the report, resistance “is also more common and worsening in places where health systems lack capacity to diagnose or treat bacterial pathogens.”
Tedros Adhanom Ghebreyesus, PhD, WHO director-general noted, “As countries strengthen their AMR surveillance systems, we must use antibiotics responsibly, and make sure everyone has access to the right medicines, quality-assured diagnostics, and vaccines. Our future also depends on strengthening systems to prevent, diagnose and treat infections and on innovating with next-generation antibiotics and rapid point-of-care molecular tests.” (Photo credit: WHO)
Gram-Negative Bacteria Present the Greatest Threat
The WHO report highlights that drug-resistant Gram-negative bacteria are becoming increasingly dangerous worldwide, with the heaviest impact seen in countries least equipped to manage the threat. Among these pathogens, E. coli and K. pneumoniae remain the most common causes of drug-resistant bloodstream infections—serious conditions that can lead to sepsis, organ failure, and death.
Globally, resistance to third-generation cephalosporins—the standard treatment for these infections—has climbed above 40% for E. coli and 55% for K. pneumoniae, and in parts of Africa, it exceeds 70%. Other essential antibiotics, including carbapenems and fluoroquinolones, are also losing effectiveness against E. coli, K. pneumoniae, Salmonella, and Acinetobacter. Once rare, carbapenem resistance is now emerging more frequently, reducing available treatment options and forcing reliance on last-resort antibiotics that are expensive, difficult to obtain, and often unavailable in low- and middle-income countries.
Progress in Surveillance but Major Gaps Remain
Despite these concerning trends, the report noted progress in global surveillance.
“Country participation in GLASS has increased over four-fold, from 25 countries in 2016 to 104 countries in 2023.” However, challenges persist: “48% of countries did not report data to GLASS in 2023,” and “about half of the reporting countries still lacked the systems to generate reliable data.” Many nations facing the highest burden of resistance “lacked the surveillance capacity to assess their antimicrobial resistance (AMR) situation.”
The report links its findings to the “political declaration on AMR adopted at the United Nations General Assembly in 2024,” which set global targets for combating antimicrobial resistance.
The declaration emphasizes strengthening health systems and working with a ‘One Health’ approach coordinating across human health, animal health, and environmental sectors.
WHO is calling on countries to strengthen laboratory systems and build reliable surveillance networks, particularly in underserved regions, to better guide treatment decisions and public health policies. The organization has set a goal for all nations to submit high-quality data on antimicrobial resistance and antibiotic use to the GLASS platform by 2030. Achieving this target will require coordinated efforts to improve data quality, expand geographic coverage, and enhance information sharing. WHO also encourages countries to implement comprehensive strategies to address antimicrobial resistance across all levels of healthcare and to ensure that treatment guidelines and essential medicines lists reflect local resistance trends.
The report is accompanied by expanded digital content available in the WHO’s GLASS dashboard, offering global and regional summaries, country profiles based on unadjusted surveillance coverage and AMR data, and detailed information on antimicrobial use.
With resistance trends worsening across regions, laboratory leaders are pivotal to turning the tide on AMR. Expanding diagnostic capabilities, improving data quality, and sharing timely resistance information will be key to shaping effective treatment guidelines and national policies. By advancing surveillance and stewardship from within the lab, clinical professionals can help preserve the power of antibiotics for future generations.
This article was created with the assistance of Generative AI and has undergone editorial review before publishing.
As this therapeutic approach gains regulatory approval, clinical laboratory tests to determine condition of patient’s gut microbiota and monitor therapy will be needed
Some developments in the clinical laboratory industry are less about diagnostic tests and more about novel approaches to therapy. Such is the case with a new carbon bead technology developed by researchers from University College London (UCL) and the Royal Free Hospital intended to remove harmful bacteria toxins from the gut before they leak to the liver. The macroporous beads, which come in small pouches, are delivered orally and could be utilized in the future to treat a number of diseases.
Why is this relevant? Once a new treatment is accepted for clinical use, demand increases for a clinical laboratory test that confirms the therapy will likely work and to monitor its progress.
In collaboration with Yaqrit, a UK-based life sciences company that develops treatments for chronic liver disease, the UCL and Royal Free Hospital scientists engineered the carbon beads—known as CARBALIVE—to help restore gut health. They measured the technology’s impact on liver, kidney, and brain function in both rats and mice.
“The influence of the gut microbiome on health is only just beginning to be fully appreciated,” said Rajiv Jalan, PhD, Professor of Hepatology at UCL in a press release. “When the balance of the microbiome is upset, ‘bad’ bacteria can proliferate and out-compete the ‘good’ bacteria that keeps the gut healthy.
“One of the ways [the ‘bad’ bacteria] do this is by excreting endotoxin, toxic metabolites, and cytokines that transform the gut environment to make it more favorable to them and hostile to good bacteria,” he continued. “These substances, particularly endotoxin, can trigger gut inflammation and increase the leakiness of the gut wall, resulting in damage to other organs such as the liver, kidneys, and brain.”
“I have high hopes that the positive impact of these carbon beads in animal models will be seen in humans, which is exciting not just for the treatment of liver disease but potentially any health condition that is caused or exacerbated by a gut microbiome that doesn’t work as it should,” said Rajiv Jalan, PhD (above), Professor of Hepatology, University College London, in a press release. “This might include conditions such as irritable bowel syndrome (IBS), for example, which is on the rise in many countries.” Though not a clinical laboratory diagnostic test, new therapies like CARBALIVE could be a boon to physicians treating patients with IBS and other gastrointestinal conditions.
Developing the Carbon Beads
The team discovered CARBALIVE is effective in the prevention of liver scarring and injury in animals with cirrhosis when ingested daily for several weeks. They also found a reduced mortality rate in test animals with acute-on-chronic-liver-failure (ACLF).
After achieving success with CARBALIVE in animals, the researchers tested the technology on 28 cirrhosis patients. The carbon beads proved to be safe for humans and had inconsequential side effects.
“In cirrhosis, a condition characterized by scarring of the liver, it is known that inflammation caused by endotoxins can exacerbate liver damage,” Jalan explained. “Part of the standard treatment for cirrhosis is antibiotics aimed at controlling bad bacteria, but this comes with the risk of antibiotic resistance and is only used in late-stage disease.”
The beads, which are smaller than a grain of salt, contain an exclusive physical structure that absorbs large and small molecules in the gut. They are intended to be taken with water at bedtime as harmful bacteria is more likely to circulate through the body at night which could result in damage. The carbon beads do not kill bacteria, which decreases the risk of antibiotic resistance. They eventually pass through the body as waste.
“They work by absorbing the endotoxins and other metabolites produced by ‘bad’ bacteria in the gut, creating a better environment for the good bacteria to flourish and helping to restore microbiome health,” said Michal Kowalski, M.Sc.Eng, Director and VP of Operations at Yaqrit, in the UCL news release.
“This prevents these toxins from leaching into other areas of the body and causing damage, as they do in cirrhosis,” he added. “The results in animal models are very positive, with reduction in gut permeability, liver injury, as well as brain and kidney dysfunction.”
Additional Research
The researchers plan to perform further clinical trials in humans to determine if the carbon beads are effective at slowing the progression of liver disease. If the benefits that were observed in lab animals prove to be compelling in humans, the technology may become an invaluable tool for the treatment of liver disease and other diseases associated with poor microbiome health in the future.
According to the American Liver Foundation, 4.5 million adults in the US have been diagnosed with liver disease. However, it is estimated that 80 to 100 million adults have some form of fatty liver disease and are unaware of it. Liver disease was the 12th leading cause of death in the US in 2020 with 51,642 adults perishing from the disease that year.
According to BMC Public Health, globally there were 2.05 million new cases of liver cirrhosis diagnosed in 2019. In that year, 1.47 million people around the world died from the disease.
More research and clinical studies are needed before this novel technology can be used clinically. When and if that happens, the demand for clinical laboratory tests that measure microbiome deficiencies and monitor patient progress during therapy will likely be high.
Shortage could disrupt the ability of clinical laboratories in hospitals and health systems to run certain tests for bloodstream infections
US clinical laboratories may soon experience a “disruption of availability” of BACTEC blood culture media bottles distributed by Becton Dickinson (BD). That’s according to the federal Centers for Disease Control and Prevention (CDC) which issued a Health Alert Network (HAN) Health Advisory to all clinical laboratory professionals, healthcare providers and facility administrators, and other stakeholders warning of the potential shortfall of critical testing supplies.
“This shortage has the potential to disrupt patient care by leading to delays in diagnosis, misdiagnosis, or other challenges in the clinical management of patients with certain infectious diseases,” the CDC stated in the health advisory.
The CDC advises healthcare providers and health departments that use the bottles to “immediately begin to assess their situations and develop plans and options to mitigate the potential impact of the shortage on patient care.”
The advisory notes that the bottles are a key component in continuous-monitoring blood culture systems used to diagnose bloodstream infections and related conditions, such as endocarditis, sepsis, and catheter-related infections. About half of all US laboratories use the BD blood culture system, which is compatible only with the BACTEC bottles, the CDC advisory states.
Infectious disease specialist Krutika Kuppalli, MD (above), Chair of the Infectious Diseases Society of America (IDSA) and a Medical Officer for COVID-19 Health Operations at the World Health Organization, outlined the potential impact of the shortage on healthcare providers and clinical laboratories. “Without the ability to identify pathogens or [their susceptibility to specific antibiotics], patients may remain on broad antibiotics, increasing the risk of antibiotic resistance and Clostridium difficile-associated diarrhea,” she told STAT. “Shortages may also discourage ordering blood cultures, leading to missed infections that need treatment.” (Photo copyright: Loyola University Health System.)
FDA Advises Conservation of Existing BACTEC Supplies
The CDC advisory followed a July 10 notice from the US Food and Drug Administration (FDA) that also warned healthcare providers of “interruptions in the supply” of the bottles. The supply disruption “is expected to impact patient diagnosis, follow up patient management, and antimicrobial stewardship efforts,” the FDA’s letter states. “The FDA recommends laboratories and healthcare providers consider conservation strategies to prioritize the use of blood culture media bottles, preserving the supply for patients at highest risk.”
Hospitals have been warned that the bottle shortage could last until September, STAT reported.
BD issued a press release in which BD Worldwide Diagnostic Solutions President Nikos Pavlidis cast blame for the shortage on an unnamed supplier.
“We understand the critical role that blood culture testing plays in diagnosing and treating infections and are taking all available measures to address this important issue, including providing the supplier our manufacturing expertise, using air shipments, modifying BD manufacturing schedules for rapid production, and collaborating with the US Food and Drug Administration to review all potential options to mitigate delays in supply,” Pavlidis said. “As an additional stopgap measure, our former supplier of glass vials will restart production to help fill the intermittent gap in supply.”
Steps Clinical Laboratories Can Take
The CDC and FDA both suggested steps that clinical laboratories and other providers can take to conserve their supplies of the bottles.
Laboratories should strive to prevent contamination of blood cultures, which “can negatively affect patient care and may require the collection of more blood cultures to help determine whether contamination has occurred,” the CDC advised.
In addition, providers should “ensure that the appropriate volume is collected when collecting blood for culture,” the advisory states. “Underfilling bottles decreases the sensitivity to detect bacteremia/fungemia and may require additional blood cultures to be drawn to diagnose an infection.”
Laboratories should also explore alternative options, such as “sending samples out to a laboratory not affected by the shortage.”
The FDA advised providers to collect blood cultures “when medically necessary” in compliance with clinical guidelines, giving priority to patients exhibiting signs of a bloodstream infection.
In an email to STAT, Andrew T. Pavia, MD, Professor of Internal Medicine and Pediatrics at the University of Utah, offered examples of situations where blood culture tests are unnecessary according to clinical guidelines.
“There are conditions like uncomplicated community acquired pneumonia or skin infections where blood cultures are often obtained but add very little,” he told STAT. “It will be critical though that blood cultures are obtained from patients with sepsis, those likely to have bloodstream infections, and very vulnerable patients.”
Hospitals Already Addressing Shortage
STAT reported that some hospitals have already taken measures to reduce the number of tests they run. And some are looking into whether they can safely use bottles past their expiration dates.
Sarah Turbett, MD, Associate Director of Clinical Microbiology Laboratories at Massachusetts General Hospital in Boston, told STAT that her team tested bottles “that were about 100 days past their expiration date to see if they were still able to detect pathogens with the same efficacy as bottles that had not yet expired. They saw no difference in the time to bacterial growth—needed to detect the cause of an infection—in the expired bottles when compared to bottles that had not expired.”
Turbett pointed to a letter in the Journal of Clinical Microbiology and Infection in which European researchers found that bottles from a different brand “were stable for between four and seven months after their expiration dates,” STAT reported.
During a Zoom call hosted by the CDC and the IDSA, hospital representatives asked if the FDA would permit use of expired bottles. However, “a representative of the agency was not able to provide an immediate answer,” STAT reported.
With sepsis being the leading cause of death in hospitals, these specimen bottles for blood culture testing are essential in diagnosing patients with relevant symptoms. This is a new example of how the supply chain for clinical laboratory instruments, tests, and consumables—which was a problem during the SARS-CoV-2 pandemic—continues to be problematic in unexpected ways.
Taking a wider view of supply chain issues that can be disruptive to normal operations of clinical laboratories and anatomic pathology groups, the market concentration of in vitro diagnostics (IVD) manufacturers means fewer vendors offering the same types of products. Consequently, if a lab’s prime vendor has a supply chain issue, there are few options available to swiftly purchase comparable products.
A separate but related issue in the supply chain involves “just in time” (JIT) inventory management—made famous by Taiichi Ohno of Toyota back in the 1980s. This management approach was designed to deliver components and products to the user hourly, daily, and weekly, as appropriate. The goal was to eliminate the cost of carrying large amounts of inventory. This concept evolved into what today is called the “Lean Manufacturing” method.
However, as was demonstrated during the SARS-CoV-2 pandemic, manufacturers and medical laboratories that had adopted JIT found themselves with inadequate numbers of components and finished products.
In the case of the current shortage of BD blood culture media bottles, this is a real-world example of how market concentration limited the number of vendors offering comparable products. At the same time, if this particular manufacturer was operating with the JIT inventory management approach, it found itself with minimal inventories of these media bottles to ship to lab clients while it addressed the manufacturing problems that caused this shortage.
Study results from Switzerland come as clinical laboratory scientists seek new ways to tackle the problem of antimicrobial resistance in hospitals
Microbiologists and clinical laboratory scientists engaged in the fight against antibiotic-resistant (aka, antimicrobial resistant) bacteria will be interested in a recent study conducted at the University of Basel and University Hospital Basel in Switzerland. The epidemiologists involved in the study discovered that some of these so-called “superbugs” can remain in the body for as long as nine years continuing to infect the host and others.
The researchers wanted to see how two species of drug-resistant bacteria—K. pneumoniae and E. coli—changed over time in the body, according to a press release from the university. They analyzed samples of the bacteria collected from patients who were admitted to the hospital over a 10-year period, focusing on older individuals with pre-existing conditions. They found that K. pneumoniae persisted for up to 4.5 years (1,704 days) and E. coli persisted for up to nine years (3,376 days).
“These patients not only repeatedly become ill themselves, but they also act as a source of infection for other people—a reservoir for these pathogens,” said Lisandra Aguilar-Bultet, PhD, the study’s lead author, in the press release.
“This is crucial information for choosing a treatment,” explained Sarah Tschudin Sutter, MD, Head of the Division of Infectious Diseases and Hospital Epidemiology, and of the Division of Hospital Epidemiology, who specializes in hospital-acquired infections and drug-resistant pathogens. Sutter led the Basel University study.
“The issue is that when patients have infections with these drug-resistant bacteria, they can still carry that organism in or on their bodies even after treatment,” said epidemiologist Maroya Spalding Walters, MD (above), who leads the Antimicrobial Resistance Team in the Division of Healthcare Quality Promotion at the federal Centers for Disease Control and Prevention (CDC). “They don’t show any signs or symptoms of illness, but they can get infections again, and they can also transmit the bacteria to other people.” Clinical laboratories working with microbiologists on antibiotic resistance will want to follow the research conducted into these deadly pathogens. (Photo copyright: Centers for Disease Control and Prevention.)
COVID-19 Pandemic Increased Antibiotic Resistance
The Basel researchers looked at 76 K. pneumoniae isolates recovered from 19 patients and 284 E. coli isolates taken from 61 patients, all between 2008 and 2018. The study was limited to patients in which the bacterial strains were detected from at least two consecutive screenings on admission to the hospital.
“DNA analysis indicates that the bacteria initially adapt quite quickly to the conditions in the colonized parts of the body, but undergo few genetic changes thereafter,” the Basel University press release states.
The researchers also discovered that some of the samples, including those from different species, had identical mechanisms of drug resistance, suggesting that the bacteria transmitted mobile genetic elements such as plasmids to each other.
One limitation of the study, the authors acknowledged, was that they could not assess the patients’ exposure to antibiotics.
Meanwhile, recent data from the World Health Organization (WHO) suggests that the COVID-19 pandemic might have exacerbated the challenges of antibiotic resistance. Even though COVID-19 is a viral infection, WHO scientists found that high percentages of patients hospitalized with the disease between 2020 and 2023 received antibiotics.
“While only 8% of hospitalized patients with COVID-19 had bacterial co-infections requiring antibiotics, three out of four or some 75% of patients have been treated with antibiotics ‘just in case’ they help,” the WHO stated in a press release.
WHO uses an antibiotic categorization system known as AWaRe (Access, Watch, Reserve) to classify antibiotics based on risk of resistance. The most frequently prescribed antibiotics were in the “Watch” group, indicating that they are “more prone to be a target of antibiotic resistance and thus prioritized as targets of stewardship programs and monitoring.”
“When a patient requires antibiotics, the benefits often outweigh the risks associated with side effects or antibiotic resistance,” said Silvia Bertagnolio, MD, Unit Head in the Antimicrobial resistance (AMR) Division at the WHO in the press release. “However, when they are unnecessary, they offer no benefit while posing risks, and their use contributes to the emergence and spread of antimicrobial resistance.”
Citing research from the National Institutes of Health (NIH), NPR reported that in the US, hospital-acquired antibiotic-resistant infections increased 32% during the pandemic compared with data from just before the outbreak.
“While that number has dropped, it still hasn’t returned to pre-pandemic levels,” NPR noted.
The UPenn researchers have already developed an antimicrobial treatment derived from guava plants that has proved effective in mice, Vox reported. They’ve also trained an AI model to scan the proteomes of extinct organisms.
“The AI identified peptides from the woolly mammoth and the ancient sea cow, among other ancient animals, as promising candidates,” Vox noted. These, too, showed antimicrobial properties in tests on mice.
These findings can be used by clinical laboratories and microbiologists in their work with hospital infection control teams to better identify patients with antibiotic resistant strains of bacteria who, after discharge, may show up at the hospital months or years later.
New technology could enable genetic scientists to identify antibiotic resistant genes and help physicians choose better treatments for genetic diseases
Genomic scientists at the Icahn School of Medicine at Mount Sinai Medical Center in New York City have developed what they call a “smart tweezer” that enables researchers to isolate a single bacterium from a patient’s microbiome in preparation for genetic sequencing. Though primarily intended for research purposes, the new technology could someday be used by clinical laboratories and microbiologists to help physicians diagnose chronic disease and choose appropriate genetic therapies.
The researchers designed their new technology—called mEnrich-seq—to improve the effectiveness of research into the complex communities of microorganisms that reside in the microbiomes within the human body. The discovery “ushers in a new era of precision in microbiome research,” according to a Mount Sinai Hospital press release.
“Imagine you’re a scientist who needs to study one particular type of bacteria in a complex environment. It’s like trying to find a needle in a large haystack,” said the study’s senior author Gang Fang, PhD (above), Professor of Genetics and Genomic Sciences at Icahn School of Medicine at Mount Sinai Medical Center, in a press release. “mEnrich-seq essentially gives researchers a ‘smart tweezer’ to pick up the needle they’re interested in,” he added. Might smart tweezers one day be used to help physicians and clinical laboratories diagnose and treat genetic diseases? (Photo copyright: Icahn School of Medicine.)
Addressing a Technology Gap in Genetic Research
Any imbalance or decrease in the variety of the body’s microorganisms can lead to an increased risk of illness and disease.
In researching the microbiome, many scientists “focus on studying specific types of bacteria within a sample, rather than looking at each type of bacteria present,” the press release states. The limitation of this method is that a specific bacterium is just one part of a complicated environment that includes other bacteria, viruses, fungi and host cells, each with their own unique DNA.
“mEnrich-seq effectively distinguishes bacteria of interest from the vast background by exploiting the ‘secret codes’ written on bacterial DNA that bacteria use naturally to differentiate among each other as part of their native immune systems,” the press release notes. “This new strategy addresses a critical technology gap, as previously researchers would need to isolate specific bacterial strains from a given sample using culture media that selectively grow the specific bacterium—a time-consuming process that works for some bacteria, but not others. mEnrich-seq, in contrast, can directly recover the genome(s) of bacteria of interest from the microbiome sample without culturing.”
Isolating Hard to Culture Bacteria
To conduct their study, the Icahn researchers used mEnrich-seq to analyze urine samples taken from three patients with urinary tract infections (UTIs) to reconstruct Escherichia coli (E. Coli) genomes. They discovered their “smart tweezer” covered more than 99.97% of the genomes across all samples. This facilitated a comprehensive examination of antibiotic-resistant genes in each genome. They found mEnrich-seq had better sensitivity than standard study methods of the urine microbiome.
They also used mEnrich-seq to selectively examine the genomes of Akkermansia muciniphila (A. muciniphila), a bacterium that colonizes the intestinal tract and has been shown to have benefits for obesity and Type 2 diabetes as well as a response to cancer immunotherapies.
“Akkermansia is very hard to culture,” Fang told GenomeWeb. “It would take weeks for you to culture it, and you need special equipment, special expertise. It’s very tedious.”
mEnrich-seq was able to quickly segregate it from more than 99.7% of A. muciniphila genomes in the samples.
Combatting Antibiotic Resistance Worldwide
According to the press release, mEnrich-seq could potentially be beneficial to future microbiome research due to:
Cost-Effectiveness: It offers a more economical approach to microbiome research, particularly beneficial in large-scale studies where resources may be limited.
Broad Applicability: The method can focus on a wide range of bacteria, making it a versatile tool for both research and clinical applications.
Medical Breakthroughs: By enabling more targeted research, mEnrich-seq could accelerate the development of new diagnostic tools and treatments.
“One of the most exciting aspects of mEnrich-seq is its potential to uncover previously missed details, like antibiotic resistance genes that traditional sequencing methods couldn’t detect due to a lack of sensitivity,” Fang said in the news release. “This could be a significant step forward in combating the global issue of antibiotic resistance.”
More research and clinical trials are needed before mEnrich-seq can be used in the medical field. The Icahn researchers plan to refine their novel genetic tool to improve its efficiency and broaden its range of applications. They also intend to collaborate with physicians and other healthcare professionals to validate how it could be used in clinical environments.
Should all this come to pass, hospital infection control teams, clinical laboratories, and microbiology labs would welcome a technology that would improve their ability to detect details—such as antibiotic resistant genes—that enable a faster and more accurate diagnosis of a patient’s infection. In turn, that could contribute to better patient outcomes.
