2025 microbiology columns - Healthcare Hygiene magazine

Tuberculosis: Principles of Laboratory Detection for Healthcare Professionals

By Priya Dhagat, MS, MLS(ASCP) CM, CIC

This article originally appeared in the Nov-Dec 2025 issue of Healthcare Hygiene magazine.

Tuberculosis remains a leading infectious disease killer despite being an ancient disease dating back thousands of years. According to the WHO, over 10 million people fell ill from TB 1.25 million people died from TB in 2023. The U.S. CDC reported 9,633 cases of TB disease, representing a 15.6% increase in case count compared with 2022. With stark increases in TB disease and deaths, rapid and accurate detection of MTB is critical for prompt clinical diagnosis and treatment as well as for public health and prevention of transmission within highly affected areas. While healthcare professionals are educated routinely about airborne isolation precautions, PPE, use of negative pressure rooms, and other infection prevention measures for MTB, it is equally as important to understand the basics of laboratory detection. In this article, we will take a step into the laboratory and focus on three areas: AFB smear microscopy, MTB culture, and molecular diagnostics.

AFB Smear Microscopy

Mycobacteria are characteristically acid-fast, meaning they resist decolorization by acids routinely used during staining procedures. This is due to their waxy, lipid rich mycolic acid cell wall which retains primary stains. Historically, three specific staining techniques have been used to detect acid-fast bacilli: Ziehl-Neelsen, Kinyoun, and Auramine-rhodamine. Both Zeilh-Neelsen utilize carbol-fushin as a primary stain and a methylene blue counter stain which allows for the visualization of red stained mycobacteria against a blue background when using brightfield microscopy. The difference between these two techniques is the usage of heat: Ziehl-Neelson uses heat to force the stain into the waxy cell wall whereas the Kinyoun stains does not use heat, but rather a higher concentration of carbol-fushin. A newer and more common staining technique uses auramine-rhodamine - a fluorescent stain that contains a mixture of both auramine and rhodamine which increases the contrast between tubercle bacilli and other microorganisms. It uses a potassium permanganate as a counter stain, resulting in yellow or orange mycobacteria against a dark background when using fluorescent microscopy. If AFB are seen in a smear examination after staining, they are counted and classified as 4+, 3+, 2+ or 1+, according to the number of AFB seen under the microscope.

Culture

Mycobacterial culture is the gold standard for detection of MTB. Since mycobacteria are intracellular bacteria that depend heavily on the many host cell nutrients, media must be complex to mimic an environment suitable for growth and detection. There are two categories of media: solid egg and agar-based media and liquid/broth media. These specific media types allow for differentiation of species based on pigment production, colony morphology, and inhibition of contaminating bacteria and fungi. Lowenstein-Jensin media is an egg-based selective solid media that contains proteins and fatty acids required for mycobacterial growth and malachite green to inhibit growth of contaminating bacteria and fungi. Middlebrook media is agar based solid composed of amino acids, vitamins, and other growth factors, along with antibacterial and antifungal properties to enhance the growth of mycobacteria tuberculosis. Like solid media, liquid/broth media contains supplements to enrich growth of mycobacteria but in a shorter time. Average time for growth can be between 12-16 days compared to up to 8 weeks for solid media. Colonies growing on solid media have an off-white or cream color and appear crumbly or dry with many wrinkles. Click here to see images of MTB colonies.

Molecular Diagnostics

Polymerase Chain Reaction (PCR) is a rapid and sensitive method to detect MTB DNA in clinical samples. In general, PCR relies on several key steps to separate, amplify, and detect DNA by either conventional methods (e.g., gel electrophoresis) or in real-time using fluorescent signals. PCR offers high specificity and sensitivity and faster results than smear microscopy and culture but should be used in conjunction with these methods for accurate diagnosis. Antibiotic susceptibly and shortened time to treatment are two major benefits of nucleic acid amplification tests. Read more about molecular diagnostics for MTB detection here.

Priya Dhagat, MS, MLS(ASCP) CM, CIC, is an infection preventionist and the associate director of the system-wide Special Pathogens Program within the Department of Emergency Management at New York City Health + Hospitals, overseeing special pathogen preparedness and response efforts across New York City Health + Hospitals frontline healthcare facilities. Additionally, she supports and offers subject matter expertise for infection prevention topics for the National Emerging Special Pathogens Training and Education Center (NETEC).

Neisseria meningitidis Conjunctivitis Among Texas Military Trainees

By Rodney Rohde, PhD, MS, SM(ASCP) CM, SVCM, MBCM, FACSc

This article originally appeared in the Sept-October 2025 issue of Healthcare Hygiene magazine.

Between Feb. 5,2025 and May 9, 2025, there was an outbreak of bacterial conjunctivitis among basic military trainees at Joint Base San Antonio-Lackland in San Antonio, Texas according to a report in the Centers for Disease Control and Prevention (CDC)’s Morbidity and Mortality Weekly Report (MMWR). Among 11,797 trainees who began basic military training (BMT) during that period, 79 cases of mucopurulent (pus-forming) conjunctivitis were identified. Of those 79 cases, 41 (52 percent) were confirmed by culture to be Neisseria meningitidis, an uncommon agent for conjunctivitis. Another 32 (41 percent) were caused by Haemophilus species; the rest were other bacteria or unspecified.

Who Was Affected & Clinical Features

Affected individuals were healthy young adults (military trainees). All had recently received quadrivalent meningococcal vaccine (which covers N. meningitidis serogroups A, C, Y, and W) upon arrival. This vaccine doesn’t target unencapsulated (“non-groupable”) strains. Sex distribution: about 90 percent male, about 10 percent female among the confirmed N. meningitidis cases. Many had symptoms of a preceding upper respiratory infection: about 80 percent of N. meningitidis conjunctivitis cases reported this. The conjunctivitis was typically unilateral (one eye) in about 85 percent of cases.

Microbiology & Strain Information

The N. meningitidis isolates from the first two confirmed cases underwent whole-genome sequencing. They were found to be non-groupable (i.e. lacking the capsule genes usually associated with serogroups A, B, C, Y, W) and were of the same sequence type (ST-32), indicating that the cases were related. These non-groupable strains are less likely to cause invasive meningococcal disease (e.g. meningitis or bloodstream infection) because the capsule is a major virulence factor. The isolates also showed a mutation in the penA gene, consistent with decreased susceptibility to penicillin. No other major antibiotic-resistance genes were noted.

Response, Treatment, and Outcomes

After identification of the first two cases (within about a three-week span in February), the base health surveillance team initiated an investigation and enhanced case finding. They set up active surveillance, encouraged cultures from ocular discharge, and established a registry for cases. Treatment: Most trainees with confirmed N. meningitidis conjunctivitis were treated with topical antibiotics (e.g. moxifloxacin, ciprofloxacin, or erythromycin). Signs of corneal involvement were monitored.

One case had progression to periorbital cellulitis and required hospitalization and intravenous antibiotics—this was after delay in using topical moxifloxacin.
No cases of invasive disease (i.e. meningitis, bacteremia) or corneal ulceration was reported.
Preventive measures: The investigation reviewed hygiene and cleaning practices (dormitories, showers, common areas), training activities, etc. Protocols were followed; no specific environmental source was identified.

Contact tracing and prophylaxis for close contacts were not broadly used, because current guidelines recommend prophylaxis chiefly for invasive disease. Additional vaccination wasn’t deemed necessary given the nature of the strain.

Why This Outbreak Matters

Unusual cause: N. meningitidis is classically associated with serious invasive disease but is a rare cause of bacterial conjunctivitis. Conjunctivitis outbreaks more often involve viral or allergic causes.
Congregate living risk: Military trainees live in close quarters with communal dormitories which facilitate close contact and spread. Such settings require heightened surveillance for unusual pathogens.
Implications for vaccine coverage: The quadrivalent vaccine does not cover non-groupable strains. This outbreak underscores that even vaccinated populations can have outbreaks from organisms outside vaccine coverage.
Treatment paradigm: Because the strain was non-groupable and patients were healthy, topical antibiotics (instead of systemic therapy) were effective in nearly all cases. This may inform future guidelines in similar contexts.
Limitations and Questions

Only the first two N. meningitidis isolates had whole-genome sequencing, so it remains possible that other cases involved slightly different strains or virulence factors. No environmental sampling (e.g. from surfaces, facilities) was done. Thus, the precise source or route of transmission remains unidentified. The findings may not generalize to other populations—these were young, immunocompetent trainees, under strict supervision, etc.

Recommendations and What to Do in Similar Situations

In outbreaks of mucopurulent conjunctivitis, especially in congregate settings, clinicians should consider doing cultures of ocular discharge rather than assuming viral or allergic causes. Early microbial diagnosis helps guide treatment. When N. meningitidis is identified, whole-genome sequencing (or at least determinations of capsule gene presence and sequence type) should be used to assess risk of invasive disease and antimicrobial susceptibilities. For healthy people infected with non-groupable N. meningitidis, topical antibiotics may suffice, unless signs of more severe disease emerge. However, disruption or delay in treatment can lead to complications.

Broader Significance and Take-Home Messages

This outbreak is a stark reminder that even less common pathogens can cause outbreaks under the right conditions, especially in high-density living environments like military training bases. Vaccination remains crucial, particularly for preventing invasive disease caused by encapsulated N. meningitidis, but vaccine coverage has limits: non-vaccine strains or non-encapsulated organisms may still cause disease. Laboratory surveillance, including culture and genomic tools, is vital—not only diagnostic but also for guiding response (how aggressive treatment should be; whether prophylaxis needed; vaccination adjustment). Prompt recognition and treatment can prevent complications; in this event, most cases were resolved with topical treatments, with only one more serious case.

Rodney E. Rohde, PhD, MS, SM(ASCP) CM, SVCM,MBCM, FACSc, is the Regents’ Professor, Texas State University System; University Distinguished Chair & Professor, Clinical Laboratory Science (CLS); TEDx Speaker & Global Fellow – Global Citizenship Alliance; Texas State Honorary Professor of International Studies; Associate Director, Translational Health Research Initiative; Past President, Texas Association for CLS.

Source: Outbreak of Neisseria meningitidis Conjunctivitis in Military Trainees — Texas, February–May 2025 MMWR Weekly / September 4, 2025 / 74(33);516–521. https://www.cdc.gov/mmwr/volumes/74/wr/mm7433a1.htm?s_cid=mm7433a1_w

Workforce Reduction and the Impact on Public Health

By Rodney Rohde, PhD, MS, SM(ASCP) CM, SVCM, MBCM, FACSc

This article originally appeared in the March-April 2025 issue of Healthcare Hygiene magazine.

In February 2025, the Centers for Disease Control and Prevention (CDC) faced a significant workforce reduction, with nearly 1,300 probationary employees—approximately 10 percent of its staff—being dismissed as part of a broader federal initiative to downsize government agencies. There are also reports that two essential CDC training programs – the Public Health Associate Program (PHAP) and Laboratory Leadership Service (LLS)—are reportedly being dismantled. Hopefully, the administration’s decision to reverse plans for devastating cuts to the CDC Epidemic Intelligence Service (EIS) is true and going to hold – it’s the right decision.

This move, coupled with substantial funding cuts to medical research institutions like the National Institutes of Health (NIH), has profound implications for the recruitment and retention of future college majors and young professionals in public health, medical laboratory sciences, and research fields.

I have worked in public health, healthcare, and academia for more than 30 years. The first decade of my career was with the Texas Department of State Health Services, Laboratory and Zoonosis Control Division, where I was on the inaugural Oral Rabies Vaccination Program which eliminated canine rabies from Texas saving countless animal and human life. For the past 20 years, I have been at Texas State University where I am a Regents’ Professor and chair of the Medical Laboratory Science Program. It has been my passion in life to introduce students to these amazing college majors and career paths. Now, I’m worried about our future professions.

Impact on Recruitment of Future College Majors

The allure of a stable career in public health and medical research has traditionally attracted many students to these fields. However, the recent job cuts and funding reductions send a discouraging message to prospective students. The perception of job instability and diminished federal support may deter individuals from pursuing degrees in these areas, exacerbating existing workforce shortages.

Prior to these developments, the medical laboratory profession was already experiencing a critical shortage. The Bureau of Labor Statistics projected a 13 percent increase in demand for medical laboratory scientists and technicians up to 2026, nearly double the average growth rate for other occupations. Despite this demand, there was a significant gap between job openings and the number of graduates, with approximately 4,900 students graduating annually to fill more than 9,000 positions, resulting in a 46 percent vacancy rate.

The recent federal actions are likely to intensify this gap. Educational institutions may struggle to attract students to programs perceived as leading to unstable careers. Moreover, reduced funding for research could lead to the downsizing or closure of academic programs, further limiting the pipeline of qualified professionals entering the workforce.

Challenges in Retention of Young Professionals

For those already in the field, the current climate presents significant retention challenges. Job insecurity, increased workloads due to understaffing, and diminished morale can lead to burnout and attrition. The recent CDC layoffs, for instance, have created an atmosphere of uncertainty among remaining employees, potentially prompting them to seek opportunities in more stable sectors.

Studies have shown that job satisfaction in medical laboratory professions is closely tied to workload, recognition, and compensation. In environments where staffing is inadequate, the remaining employees often face increased workloads, leading to exhaustion and a higher likelihood of errors. This scenario not only affects employee well-being but also compromises the quality of public health services.

Furthermore, the professional socialization of early-career medical laboratory scientists plays a crucial role in retention. A study highlighted that new professionals often face a theory-practice gap, where their educational experiences do not align with workplace realities. This disparity can lead to dissatisfaction and a sense of unpreparedness, increasing the likelihood of leaving the profession.

Broader Implications for Public Health and Research

The downsizing of federal health agencies and cuts to research funding have ripple effects beyond immediate employment concerns. They undermine the nation's capacity to respond to public health crises and stifle innovation in medical research. CDC, for example, plays a pivotal role in managing disease outbreaks and providing critical health information. A reduced workforce hampers its ability to effectively carry out these functions, potentially compromising public health.

Similarly, funding cuts to the NIH threaten ongoing and future research projects. Scientists have expressed concerns that reduced support endangers patients, threatens jobs, and undermines America's leadership in science and innovation. Legal challenges have temporarily halted some of these cuts, but the uncertainty continues to loom over the research community.

Importantly, those of us in academic programs who prepare these future professionals know that we can’t simply “create a new professional overnight.” It can take many years to matriculate through an academic program that includes rigorous coursework, laboratory sessions, field work and/or clinical and medical rotations. Then, these professionals need time to “mature and become seasoned” veterans to mix with the newer professionals. Without this type of educational and training model, we lose our pipeline which is already struggling to fill new employment spots.

Strategies to Mitigate Negative Outcomes

Addressing these challenges requires a multifaceted approach:

Advocacy for Stable Funding: Professional organizations, academic institutions, and industry stakeholders must advocate for sustained or increased funding for public health and research. Highlighting the long-term benefits of investment in these areas can help influence policy decisions.
Enhancing Job Security and Satisfaction: Employers should focus on creating supportive work environments that offer job security, competitive compensation, and opportunities for professional development. Recognizing and addressing factors that contribute to burnout can improve retention rates.
Educational Outreach and Support: To attract future professionals, educational institutions should provide clear pathways to stable careers in public health and research. This includes offering scholarships, mentorship programs, and real-world experiences that align with job market demands.
Public Awareness Campaigns: Raising public awareness about the critical role of public health and research professionals can generate broader support for these fields. Showcasing success stories and the tangible impacts of their work can inspire the next generation to pursue these careers.
The 2025 CDC job losses and similar reductions in medical laboratory and research positions pose significant challenges to the recruitment and retention of professionals in these vital fields. Proactive measures are essential to ensure a robust workforce capable of safeguarding public health and advancing medical knowledge. In summary, cutting jobs in the medical laboratory sector of healthcare, CDC’s Epidemic Intelligence Program, and other similar public health programs would likely lead to weaker disease surveillance, slower outbreak responses, greater economic costs, and increased national security risks. The long-term impact could be more severe pandemics and a less prepared public health workforce.

Human Metapneumovirus: A Common Virus Sparking Concerns

By Priya Dhagat, MS, MLS(ASCP) CM, CIC

This article originally appeared in the January/February 2025 issue of Healthcare Hygiene magazine.

Human Metapneumovirus (HMPV) is surging in China, making headlines as a “mystery illness” that has sparked fears of a new outbreak on the rise comparable to the COVID-19 pandemic. Recent reports from China indicate that rates of numerous flu-like illnesses are increasing and cases of HMPV infection have risen among people who are younger than 14 years old and living in northern Chinese provinces. While any major increase in respiratory disease is concerning, it is critical to note that HMPV infections typically coincide with colder weather and increased indoor activity, making it consistent with seasonal trends of other viruses circulating this time of the year such as the flu, COVID-19, and RSV.

HMPV was first discovered by Dutch scientists in 2001. The virus was isolated from nasopharyngeal aspirates from children and infants suffering from respiratory infection that could not be identified from routine diagnostic assays for known respiratory viruses. Full genome sequencing revealed that HPMV is genetically related to avian pneumoviruses, indicating probable zoonotic spillover from an avian reservoir. HMPV is an enveloped, single-stranded RNA virus belonging to the Pneumoviridae family, which also includes respiratory syncytial virus (RSV). Since it was first detected, HMPV has been isolated on every continent and from individuals of all ages.

In the United States, HMPV infections begin circulating in winter and last through spring. Similar to other respiratory viruses, HMPV is spread through secretions from coughing and sneezing, close personal contact with someone who is sick (e.g., touching or shaking hands), or touching objects or surfaces and then touching the mouth, nose, or eyes. HMPV contains several proteins that are responsible for infecting airway epithelial cells and spreading into the respiratory tract which induces an immune response that leads to pulmonary inflammation. HMPV generally causes mild illness that can be treated by supportive care. Symptoms begin as a common cold after an incubation period of three to six days, but may intensify and cause upper and lower respiratory tract infections characterized by high fever, wheezing, severe cough, difficulty breathing, tachypnea, bronchiolitis and pneumonia, and hypoxia. Infants, the elderly, and those who have weakened immune systems are at risk for serious illness or hospitalization. Healthcare personnel should follow contact and standard precautions, which includes wearing a mask, when caring for patients hospitalized with HPMV.

Symptoms are often indistinguishable from other common respiratory viruses, so testing for it may not be considered. However, CDC recommends that clinicians consider testing patients with severe respiratory illness using upper airway or lower respiratory tract specimens. HMPV is commonly included in multi-pathogen PCR respiratory panels.

The National Respiratory and Enteric Virus Surveillance System (NREVSS) monitors and tracks viral activity in the United States from participating U.S. laboratories in effort identify patterns and potential outbreaks. As of Dec. 28, 2024, HMPV accounted for 1.94 percent of positive tests reported to NREVSS, compared to 1.54 percent the previous week.

The surge of cases in China is a reminder of the seasonality of certain respiratory viruses and emphasizes the importance of routine infection prevention measures such as:

Washing hands often with soap and water for at least 20 seconds
Avoid touching eyes, nose, or mouth with unwashed hands
Avoid close contact with people who are sick
Cover mouth and nose when coughing and sneezing
Avoid sharing cups and eating utensils with others
Stay at home if sick
Clean frequently touched surfaces (e.g., doorknobs, shared toys)
For more information on HPMV:

Human Metapneumovirus - StatPearls - NCBI Bookshelf

About Human Metapneumovirus | Human Metapneumovirus | CDC

Zoonotic Origins of Human Metapneumovirus: A Journey from Birds to Humans - PMC