Clinical Characteristics and Resistance Patterns of Extensively Drug-Resistant Salmonella Typhi in the Pediatric Population of Pakistan: A Prospective Study

Abstract

Objectives:

To describe the demographics, clinical characteristics, antimicrobial resistance profiles, and treatment responses of pediatric XDR Salmonella Typhi in Northern Punjab, Pakistan, and assess their association with disease severity.

Introduction:

XDR Salmonella enterica serovar Typhi poses a major pediatric health challenge in Pakistan due to limited treatment options, relapse risk, and contaminated water exposure.

Methods:

A prospective cohort study was conducted at POF Hospital, Northern Punjab (January 2024-January 2025), enrolling 65 children (1-12 years) with culture-confirmed XDR S. Typhi. Clinical, laboratory, and treatment data were analyzed using Chi-square tests.

Results:

Most patients (60%) were aged 5 to 10 years; vomiting (69.2%) and headache (47.7%) were frequent. All isolates were resistant to ceftriaxone, ciprofloxacin, and first-line agents; meropenem and azithromycin remained 100% and 96.9% susceptible. Severe disease was associated with delayed fever clearance (P < .001) and longer hospitalization (P < .05).

Conclusion:

XDR typhoid demands strengthened stewardship, vaccination, and water safety.

Keywords

antibiotic resistance azithromycin CRP meropenem pediatric typhoid XDR typhoid

Introduction

Extensively drug-resistant (XDR) Salmonella enterica serovar Typhi is defined as resistance to all first-line agents: ampicillin, chloramphenicol, and trimethoprim–sulfamethoxazole alongside fluoroquinolones and third-generation cephalosporins such as ceftriaxone. The phenomenon of multi-drug resistance has restricted our choices of antibiotics. There are still a few antibiotics like azithromycin and the carbapenems, but their usefulness is being threatened more and more.^1,2

Typhoid fever is a systemic bacterial infection caused by Salmonella Typhi and is primarily acquired by ingestion of food or water that has been contaminated. The pathogen breaks through the mucosa of the intestine and spreads by way of the blood, where it is able to invade several multiple organ systems.³ In children, the most common symptoms/presentation of the illness are a sustained high fever, pain in the abdomen, diarrhea or constipation, enlarged spleen and liver (hepatosplenomegaly), and in more extreme cases, certain complications (eg, perforation of the intestine, or the brain becoming infected), or shock due to septicemia. Because the side effects are so similar to other illnesses that induce a fever, achieving rapid diagnosis is notoriously difficult.^4,5

Globally, XDR Salmonella Typhi has emerged as a major pediatric infectious disease concern. In South Asia, where sanitation infrastructure is limited and antibiotic stewardship is poor, resistant strains have spread rapidly.^6,7 Drivers of antimicrobial resistance (AMR) in Salmonella Typhi are multifactorial and well documented in endemic settings. Major contributors include widespread non-prescription access to antibiotics and inappropriate or incomplete treatment courses,⁸ suboptimal typhoid conjugate vaccine (TCV) coverage that allows continued transmission,⁹ and the acquisition and spread of plasmid-mediated extended-spectrum β-lactamase genes (eg, blaCTX-M-15)¹⁰ that facilitate horizontal transfer of resistance across strains.^11,12 Pakistan remains the epicenter of the largest and most persistent XDR typhoid outbreak on record, first detected in Sindh province in 2016.¹³ Since then, resistant strains have been reported across the country, including Northern Punjab, with high pediatric case counts.⁹ Resistance rates now exceed 90% for fluoroquinolones and nearly 50% for ceftriaxone, forcing reliance on azithromycin and carbapenems. Alarmingly, recent surveillance has detected isolates with reduced susceptibility to these last-line drugs.¹⁰

Despite growing genomic data on these resistant strains, clinical research focusing on pediatric populations remains limited. Little is known about the full clinical spectrum, antimicrobial resistance trends, and treatment options that influence disease severity and outcomes in children. This study aims to bridge this gap by describing the demographics, clinical characteristics, antimicrobial resistance patterns, treatment response, outcomes and association with disease severity in Northern Punjab, Pakistan. Our findings aim to support improved case management, guide targeted TCV deployment, and inform pediatric AMR control policies in endemic regions.

Materials and Methods

Study Setting and Duration

This prospective cohort study was conducted from January 1, 2024, to January 1, 2025, between the Department of Pediatrics at a university hospital in Northern Punjab Pakistan, a tertiary care hospital that serves urban and semi-urban settings and referral hospital for the surrounding districts.

Study Population and Sampling

Children between the ages of 2 to 12 years who were admitted to the Department of Pediatrics and were and found to have blood culture confirmed Salmonella enterica serovar Typhi with an extensively drug-resistant (XDR) pattern were the study population. Salmonella Typhi XDR was defined to have resistance to all of the first-line 3 drug classes, that is, aminopenicillins, chloramphenicol, trimethoprim–sulfamethoxazole and fluoroquinolones, and 3rd generation cephalosporins like ceftriaxone.

The sample size of 65 participants was calculated based on a previously reported prevalence of 47% for XDR typhoid,¹⁴ with a 95% confidence level and a 10% margin of error. This calculation ensured sufficient power to detect the prevalence of XDR Salmonella Typhi within the study population while maintaining a reasonable level of precision. Potential selection bias has been minimized by applying consecutive non-probability sampling to ensure that all eligible XDR Salmonella Typhi cases were captured due to presenting during the study period. All children that met the inclusion criteria were enrolled during the admission phase with no cases eligible to be skipped. To identify new culture-positive XDR typhoid cases diagnosed during the study period, a daily trained research officer screened the admission logbook, microbiology alerts, and ward census daily. Before recruiting eligible participants, clinical records and laboratory reports were used to confirm case status. To sustain comprehensive case capture, no patient who met the study criteria during the study period was slacked off the study.

Eligibility Criteria

Inclusion criteria were:

Age between 1 and 12 years

Blood culture positive for Salmonella Typhi with XDR profile

Availability of complete medical records

Exclusion criteria were:

Missing essential clinical or laboratory data

Known immunodeficiency disorders

Documented co-infections such as malaria or dengue

Operational Definitions

Suspected Typhoid Case

A child presenting with fever ≥ 38°C lasting ≥ 3 days and at least 1 symptom such as abdominal pain, vomiting, diarrhea, constipation, or headache, without an alternative diagnosis.⁸

Confirmed Typhoid Case

A patient with blood culture positive for Salmonella Typhi with antimicrobial susceptibility showing an XDR pattern.⁸

Extensively Drug-Resistant (XDR) Salmonella Typhi

The World Health Organization (WHO) defines Extensively Drug-Resistant (XDR) Salmonella Typhi as a strain resistant to first-line antibiotics: ampicillin, chloramphenicol, and trimethoprim–sulfamethoxazole, fluoroquinolones: such as ciprofloxacin and third-generation cephalosporins: such as ceftriaxone.¹³

Treatment Response (Clinical Improvement)

Reduction of fever and symptoms within 72 hours of starting definitive antibiotic therapy.

Treatment Failure

Persistent fever or clinical symptoms beyond 5 days of appropriate treatment, or need to change antibiotic due to non-response.

Clinical Cure

Complete resolution of fever and symptoms without relapse by discharge.

Relapse

Return of fever or symptoms with positive culture within 30 days after completion of therapy.

Disease Severity Classification

Severity was defined according to fever clearance time after initiation of appropriate antimicrobial therapy:

Less severe: Fever resolution within 3 days

Severe: Persistent fever beyond 3 days

Hospital Stay Duration

Short stay: <7 days

Prolonged stay: ≥7 days

Seasonal Definitions (Pakistan)

Summer: April to June characterized by high temperatures, low rainfall, dry weather.¹⁵

Monsoon: July to September characterized by high rainfall, high humidity, seasonal clustering of waterborne diseases including typhoid.¹⁵

Winter: October to March characterized by cooler temperatures, low rainfall, dry to moderate humidity.¹⁵

Sanitation

Adequate Sanitation: Households with flush latrine or safely managed toilet, proper drainage and no reported open defecation.¹⁶

Inadequate sanitation: It includes pit latrines, shared toilets, or absence of structured waste disposal.¹⁶

Water Contamination

A household’s drinking water is considered contaminated if municipal, borehole, or stored water tests positive for bacterial pathogens, including Salmonella Typhi, or if there is documented evidence of unsafe water handling or storage (eg, unprotected storage, no chlorination, or mixing with sewage).¹⁷

Data Collection

Participants were followed prospectively until discharge or complete recovery. Data were extracted from hospital records and entered into structured case record forms. Information collected included:

Demographic data: Age, sex, residence (urban or semi-urban)

Clinical characteristics: Fever duration, gastrointestinal symptoms (diarrhea or constipation), abdominal pain, hepatosplenomegaly, and complications such as intestinal perforation or encephalopathy

Treatment details: Initial empirical therapy, definitive antibiotic regimen based on susceptibility results and use of combination therapy

Outcome measures: Time to fever clearance, total hospital stay, relapse episodes, and survival status at discharge

Laboratory Investigations

Baseline tests involved C-reactive protein (CRP), alanine aminotransferase (ALT), aspartate aminotransferase (AST), serum creatinine, and blood urea nitrogen (BUN).

Microbiological Methods

Blood cultures were processed in the hospital microbiology laboratory using standard aseptic techniques. Antimicrobial susceptibility testing was performed using the Kirby–Bauer disk diffusion method and interpreted according to the Clinical and Laboratory Standards Institute (CLSI) guidelines, 33rd edition, 2023.¹⁸ Tested antibiotics included azithromycin, meropenem, ciprofloxacin, ceftriaxone, ampicillin, chloramphenicol, and trimethoprim–sulfamethoxazole.

Minimum inhibitory concentration (MIC) breakpoints applied for interpretation are shown in Table 1. For azithromycin, no official CLSI Enterobacterales breakpoint exists; values used were adopted from published studies on Salmonella Typhi.^19,20

Table 1.

MIC Breakpoints Used for Antimicrobial Susceptibility Interpretation (CLSI 2023).

Antibiotic	Susceptible (S)	Intermediate (I)	Resistant (R)
Ceftriaxone	≤1 µg/mL	2 µg/mL	≥4 µg/mL
Ciprofloxacin	≤0.06 µg/ml	0.12-0.5 µg/mL	≥1 µg/mL
Azithromycin*	≤16 µg/mL	—	≥32 µg/mL
Meropenem	≤1 µg/mL	2 µg/mL	≥4 µg/mL
Ampicillin	≤8 µg/mL	16 µg/mL	≥32 µg/mL
Chloramphenicol	≤8 µg/mL	16 µg/mL	≥32 µg/mL
Trimethoprim–sulfamethoxazole	≤2/38 µg/mL	4/76 µg/mL	≥8/152 µg/mL

For azithromycin, no official CLSI Enterobacterales breakpoint; values adopted from published studies on Salmonella Typhi.

Water Contamination Assessment

Respondents’ contaminated water household status was evaluated based on an administered structured set of questions. Where possible, data from local water quality testing were also analyzed. Primary water source (municipal supply, borehole etc.), water storage practices, and water storage behavior (boiling, chlorination, or filtration) were questions directed to the caregivers of the household. For participants whose households had water testing done by local municipal authorities or privately hired water quality labs within the last 12 months, documented results of tested fecal coliforms and/or Salmonella species were requested and documented.²¹ For households without laboratory reports some water safety practices were assumed based on other existing high risk indicators. For example, lack of evidence of water protective storage, lack of chlorination, reports of mixed sewage and water pipelines, and water which was visually unclean. This method has also been used previously in typhoid outbreak investigations in Pakistan^22,23 and is consistent with the criteria established by WHO/UNICEF Joint Monitoring Program (JMP) for the estimation of unprotected water.²⁴

Data Analysis

IBM SPSS Statistics version 25 was used for data analysis. Continuous variables (eg, CRP, ALT) were expressed as mean ± standard deviation (SD), and categorical variables as percentages and frequencies. Inferential analysis was performed to assess associations between clinical outcomes and disease severity. Fisher’s Exact Test was applied for categorical comparisons due to small cell counts, while Chi-square was used where assumptions were met. A P-value < .05 was considered statistically significant.

Result

Demographic Features

The study included 65 children with blood culture–confirmed XDR Salmonella Typhi as shown in Table 2. Most were aged 5 to 10 years n = 39 (60.0%), with a near-equal male-to-female ratio. Recent travel was reported in n = 31 (47.7%), and n = 40 (61.5%) had contact with a typhoid case. TCV vaccination was documented in n = 28 (43.1%), while n = 20 (30.8%) was unvaccinated. Over half of cases occurred during the monsoon season n = 33 (50.8%). Municipal water was the main source n = 44 (67.7%), and sanitation was inadequate in n = 32 (49.2%) of households. Water contamination was confirmed in n = 25 (38.5%).

Table 2.

Demographic Features of the Study Population (n = 65).

Categories	Characteristic	n	%
Age group (years)	1-5 years	23	35.4
	5-10 years	39	60.0
	11-12 years	3	4.6
Gender	Male	32	49.2
Gender	Female	33	50.8
Vaccination status	Vaccinated (TCV)	28	43.1
	Not vaccinated	20	30.8
	Unknown	17	26.2
Travel in last 30 days	Yes	31	47.7
Travel in last 30 days	No	34	52.3
Contact with typhoid case	Yes	40	61.5
Contact with typhoid case	No	25	38.5
Season of diagnosis	Summer	21	32.3
	Winter	11	16.9
	Monsoon	33	50.8
Primary water source	Municipal	44	67.7
	Borehole	17	26.2
	Other	4	6.2
Sanitation condition	Adequate	33	50.8
Sanitation condition	Inadequate	32	49.2
Water source contamination	Contaminated	25	38.5
Water source contamination	Not contaminated	40	61.5

Clinical Characteristics

Fever was the most frequently reported symptom n = 27 (41.5%), followed by vomiting n = 45 (69.2%), headache n = 31 (47.7%), diarrhea n = 29 (44.6%), abdominal pain n = 22 (33.8%), hepatosplenomegaly n = 21 (32.3%), and constipation n = 12 (18.5%) as shown in Table 3.

Table 3.

Clinical Characteristics Of Pediatric XDR Typhoid Patients (n = 65).

Symptom category	Symptom	n	%
Systemic	Fever	27	41.5
	Headache	31	47.7
Gastrointestinal	Abdominal pain	22	33.8
	Vomiting	45	69.2
	Diarrhea	29	44.6
	Constipation	12	18.5
Hepatosplenic	Hepatosplenomegaly	21	32.3

Laboratory Parameters

Table 4 demonstrates that mean CRP was markedly elevated at 67.17 ± 17.63 mg/L. Liver enzymes were also raised, with mean ALT of 58.05 ± 14.86 U/L and AST of 57.09 ± 14.47 U/L. Mean blood urea was 35.09 ± 9.04 mg/dL, and mean serum creatinine was slightly elevated at 1.10 ± 0.29 mg/dL. These elevations in CRP and liver enzymes may reflect the inflammatory burden and hepatic involvement in pediatric XDR typhoid, suggesting their potential utility as markers of disease severity.

Table 4.

Laboratory Parameters in Pediatric XDR Typhoid Patients (n = 65).

Laboratory test	Mean ± SD	Reference range
C-reactive protein (CRP, mg/L)	67.17 ± 17.63	<10
ALT (U/L)	58.05 ± 14.86	10-40
AST (U/L)	57.09 ± 14.47	10-40
Blood urea (mg/dL)	35.09 ± 9.04	7-20
Serum creatinine (mg/dL)	1.10 ± 0.29	0.4-1.0

Antimicrobial Susceptibility

All isolates were resistant to ceftriaxone, ciprofloxacin, ampicillin, trimethoprim-sulfamethoxazole and chloramphenicol as shown in Table 5. Susceptibility to azithromycin was high n = 63 (96.9%), with resistance observed in 2 isolates n = 2 (3.1%). All isolates remained susceptible to meropenem n = 65 (100%) as shown in Figure 1.

Table 5.

Antibiotic Susceptibility Pattern of XDR Salmonella Typhi Isolates (n = 65).

Antibiotic	Sensitive (n, %)	Intermediate (n, %)	Resistant (n, %)
Meropenem	65 (100%)	—	—
Azithromycin	63 (96.9%)	—	2 (3.1%)
Ceftriaxone	—	—	65 (100%)
Ciprofloxacin	—	—	65 (100%)
Ampicillin	—	—	65 (100%)
Chloramphenicol	—	—	65 (100%)
Trimethoprim–sulfamethoxazole	—	—	65 (100%)

Figure 1.

Antimicrobial resistance profile of XDR Salmonella Typhi (n = 65), demonstrating the proportion of isolates resistant versus sensitive to commonly used antibiotics.

Treatment Response and Outcomes

Table 6 summarizes treatment patterns and clinical outcomes stratified by disease severity. Meropenem monotherapy was used in n = 21 (32.3%), azithromycin in n = 25 (38.5%), and combination therapy in n = 19 (29.2%), with no significant difference in severity between regimens (P = .14). Empirical antibiotics before AST confirmation were administered in n = 34 (52.3%), without significant association with severity (P = .09). Full adherence to therapy was achieved in n = 39 (60.0%) and was significantly higher among less severe cases (P = .04), whereas partial adherence n = 16 (24.6%) and poor adherence n = 10 (15.4%) were more frequent in severe presentations. Rapid fever clearance within ≤3 days occurred in n = 23 (35.4%), all within the less severe group (P < .001). Prolonged hospitalization (≥7 days) was observed in n = 36 (55.4%) and was more common in severe cases (P = .03). Duration of therapy ≥10 days was recorded in n = 36 (55.4%), without significant association with severity (P = .16). Relapse within 30 days occurred in n = 4 (6.2%), exclusively among severe cases (P = .56). When comparing relapse by treatment regimen, n = 2 (50.0%) occurred in azithromycin recipients and n = 2 (50.0%) in those on combination therapy, while no relapse was seen with meropenem monotherapy (P = .36). Adverse effects were predominantly gastrointestinal in n = 44 (67.7%), whereas n = 19 (29.2%) reported none and n = 2 (3.1%) experienced neurological or rash-related symptoms (P = .38). Vaccination status showed no association with severity, with n = 20 (30.8%) unvaccinated, n = 28 (43.1%) vaccinated, and n = 17 (26.2%) unknown (P = .60).

Table 6.

Treatment Response and Outcomes and Their Association with Disease Severity in Pediatric XDR Typhoid (N = 65).

Variable	Category	Total n (%)	Disease severity		χ² (do)	P-value
Variable	Category	Total n (%)	Less severe n (%)	More severe n (%)	χ² (do)	P-value
Fever clearance time	≤3 days	23 (35.4)	23 (100.0)	0 (0.0)	—	<.001
Fever clearance time	>3 days	42 (64.6)	0 (0.0)	42 (100.0)	—	<.001
Final treatment regimen	Meropenem only	21 (32.3)	14 (60.9)	7 (16.7)	3.93 (2)	.14
	Azithromycin only	25 (38.5)	6 (26.1)	19 (45.2)
	Combination	19 (29.2)	3 (13.0)	16 (38.1)
Empirical antibiotic before AST	Yes	34 (52.3)	25 (59.5)	9 (39.1)	2.84 (1)	.09
Empirical antibiotic before AST	No	31 (47.7)	17 (40.5)	14 (60.9)	2.84 (1)	.09
Treatment adherence	Full	39 (60.0)	24 (57.1)	15 (65.2)	—	.04
	Partial	16 (24.6)	14 (33.3)	2 (8.7)
	None	10 (15.4)	4 (9.5)	6 (26.1)
Hospital stay duration	<7 days	29 (44.6)	15 (65.2)	14 (33.3)	4.63 (1)	.03
Hospital stay duration	≥7 days	36 (55.4)	8 (34.8)	28 (66.7)	4.63 (1)	.03
Duration of therapy	<10 days	29 (44.6)	17 (73.9)	12 (28.6)	1.95 (1)	.16
Duration of therapy	≥10 days	36 (55.4)	6 (26.1)	30 (71.4)	1.95 (1)	.16
Final outcome	Cured	65 (100.0)	42 (100)	23 (100)	—	—
Relapse (30 days)	Yes	4 (6.2)	0 (0.0)	4 (9.5)	—	.56
Relapse (30 days)	No	61 (93.8)	23 (100.0)	38 (90.5)	—	.56
Relapse rates by treatment regimen	Meropenem only	0 (0.0)	0 (0)	0 (0)	2.05 (2)	.36
	Azithromycin only	2 (3.1)	1 (50)	1 (50)
	Combination	2 (3.1)	0	2 (100)
Reported side effects	None	19 (29.2)	14 (33.3)	5 (21.7)	—	.38
	Gastrointestinal	44 (67.7)	26 (61.9)	18 (78.3)
	Other (Neuro/Rash)	2 (3.1)	2 (4.8)	0 (.0)
Vaccination status	Unvaccinated	20 (30.8)	8 (34.8)	12 (28.6)	1.00 (2)	.60
	Vaccinated	28 (43.1)	8 (34.8)	20 (47.6)
	Unknown	17 (26.1)	7 (30.4)	10 (23.8)

Discussion

This prospective study characterizes the clinical features, laboratory findings, antimicrobial susceptibility profiles, and treatment outcomes of pediatric extensively drug-resistant (XDR) Salmonella enterica serovar Typhi in Northern Punjab, Pakistan. Our results are consistent with molecular surveillance from Lahore, which identified blaCTX-M-15 harboring H58 lineage strains as the predominant cause of XDR typhoid in the region.²⁵ While we did not perform genomic sequencing, the resistance profile observed in our cohort serves as a proxy for H58 XDR strains. School-aged children were most affected, a pattern also noted in other Pakistani studies, likely due to increased environmental exposure, incomplete immunity, and suboptimal typhoid conjugate vaccine (TCV) coverage.^6,26
-28 The seasonal clustering during monsoon months mirrors findings from Sindh, where heavy rains and flooding amplify contamination of municipal water supplies.^15,29

Environmental exposures were significant in our cohort, with contaminated municipal water and inadequate sanitation as major risk factors.³ Similar observations from Hyderabad identified municipal water as the predominant source during the 2016 outbreak.^13,26 These findings emphasize the role of poor water safety and sanitation infrastructure in sustaining XDR typhoid transmission in urban areas.^16,30

Clinically, vomiting, diarrhea, and headache were the most frequent symptoms, aligning with pediatric XDR typhoid presentations reported in Karachi and other regions.²⁷ Notably, only n = 27 (41.5%) presented with fever at admission, which is lower than prior studies This could be related to empirical antibiotic use before hospitalization, which may attenuate febrile responses.³⁰ Elevated CRP and transaminases in our patients are consistent with systemic inflammation and hepatocellular injury patterns previously described in invasive Salmonella Typhi infection.^27,31 Elevated blood urea and creatinine levels, seen in a subset, may indicate dehydration, septic physiology, or possible nephrotoxic drug effects, similar to prior reports from pediatric typhoid cohorts in South Asia.^32,33 These laboratory abnormalities may also serve as markers of disease severity in pediatric XDR typhoid.

The antimicrobial susceptibility profile observed complete resistance to ampicillin, chloramphenicol, ceftriaxone, trimethoprim–sulfamethoxazole and ciprofloxacin, matches national surveillance data confirming the dominance of highly resistant H58 strains. Although genomic confirmation was not performed, the resistance pattern serves as a proxy for H58 XDR lineage.^5,10 Retained meropenem susceptibility n = 65 (100%) and high azithromycin susceptibility n = 63 (96.9%) are reassuring; however, the detection of azithromycin resistance in n = 2 (3.1%) of isolates is concerning.^33,34 Comparable reports from Bangladesh indicate rising azithromycin minimum inhibitory concentrations (MICs), raising fears that loss of this oral option could severely limit pediatric treatment.^10,35,36 Similar concerns have been raised in Karachi, where azithromycin resistance has emerged alongside persistent XDR prevalence.^11,20,29 Beyond clinical implications, the high cost of managing XDR typhoid remains a critical challenge.³⁷ In resource-limited settings, this economic pressure underscores the urgency of preventive measures including vaccination, sanitation, and antimicrobial stewardship to minimize reliance on expensive last-line therapies.^16,33

Our analysis indicates that clinical response, rather than antimicrobial choice, was more closely associated with disease severity in pediatric XDR typhoid. Early fever clearance within ≤3 days was significantly associated with non-severe disease, consistent with reports from Peshawar and Dhaka, where early defervescence correlated with favorable outcomes regardless of regimen type.^35,38 Full adherence to therapy also showed a strong association with reduced severity, in line with Karachi pediatric XDR studies demonstrating that incomplete adherence increased the risk of complications and delayed recovery.^27,38

In contrast, treatment regimen, vaccination status, prior empirical antibiotic exposure, and relapse rates were not significantly associated with severity in our cohort findings comparable to those from genomic and regional investigations in South Asia, which highlight that environmental exposure, healthcare access, and care-seeking delays contribute more to disease severity than antimicrobial choice.^39,40 However, a randomized comparative trial in Pakistani children reported faster fever resolution and lower complication rates with combination therapy, suggesting that therapeutic benefit may vary by patient population and clinical setting.⁴⁰

Additionally, prolonged hospitalization and delayed fever clearance in our study both strong indicators of severe disease, have been similarly identified during XDR typhoid outbreaks outside Pakistan, such as the Beijing waterborne outbreak, where delayed response and severe clinical manifestations were observed.⁴¹ Finally, a recent global synthesis underscores that typhoid outcomes are increasingly determined by host response, vaccination coverage, and public health infrastructure rather than drug class alone, reinforcing the importance of response-based monitoring, adherence promotion, WASH interventions, and robust surveillance frameworks for sustainable XDR typhoid control.⁴²

Potential Avenues for Further Development

Strengths of this study include its prospective design, culture-confirmed diagnoses, and detailed clinical and laboratory characterization. Limitations include its single-center scope, modest sample size, and lack of genomic sequencing, which could have provided additional insights into resistance mechanisms. Additionally, while antimicrobial susceptibility testing was performed using CLSI 33rd edition (2023) disk diffusion guidelines,¹⁸ MIC data particularly for azithromycin were not generated, which may limit global comparability of resistance findings.

Conclusion

Pediatric XDR Salmonella Typhi is a major challenge in Northern Punjab, driven by unsafe water and poor sanitation. School-aged children are most affected, with cases clustering during monsoon season. Meropenem and azithromycin remain effective, but emerging resistance is concerning. Prolonged 14-day regimens improved outcomes and reduced relapse, highlighting the need for adherence. This study highlights the urgent need for an integrated response combining strengthened surveillance, typhoid conjugate vaccination, antimicrobial stewardship, and sustained WASH interventions to contain pediatric XDR typhoid and preserve the effectiveness of remaining treatment options.

Footnotes

Acknowledgements

We gratefully acknowledge the support of NUMS, Wah Medical College, and POF Hospital in facilitating this research.

ORCID iDs

Hamna Khalid

Zoha Rehman

Noof K. Binashikhbubkr

Ethical Considerations

Ethical approval was granted by the Institutional Review Board of Pakistan Ordinance Factories (POF) Hospital. (IRB No. POF/00/9055).

Consent to Participate

As the data used was prospective, approval was obtained from the relevant ethical committee of the hospital. Written informed consent was obtained from the patients, ensuring confidentiality and adherence to ethical guidelines.

Author Contributions

HK, MMK, KR, ZA, NKB: Concept and design. HK, ZR, NR, MDN: Acquisition, analysis, or interpretation of data. HK, NR, ZA, MDN: Drafting of the manuscript. HK, ZR, MMK, KR, MDN, NKB: Critical review of the manuscript for important intellectual content. NR: Supervision. All authors have read and approved the final version of the manuscript and agree to be accountable for all aspects of the work, ensuring its accuracy and integrity.

Funding

The authors received no financial support for the research, authorship, and/or publication of this article.

Declaration of Conflicting Interests

The authors declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

References

Baloch

Abbasi

Shaikh

MMA

Sagar

Memon

Amir

Drug resistant XDR typhoid in children admitted in a tertiary care hospital. Ann Pak Inst Med Sci Shaheed Zulfiqar Ali Bhutto Med Univ. 2024;20(1):69-73. doi:10.48036/apims.v20i1.964

Zaki

Karande

Multidrug-resistant typhoid fever: a review. J Infect Dev Ctries. 2011;5(5):324-337. doi:10.3855/jidc.1405

Khan

Paul

SK.

Sanitation-hygiene knowledge, practices and human health impacts: insights from coastal Bangladesh. Geosfera Indones. 2023;8(2):133-151. doi:10.19184/geosi.v8i2.39584

Kang

Chu

Yoo

, et al. Epidemiology of nontyphoidal salmonella infections in Korean children and genetic factors associated with extra-intestinal invasion: a whole-genome sequencing analysis. Ann Lab Med. 2025;45(3):312-321. doi:10.3343/alm.2024.0378

Kavai

Mutai

Mbae

, et al. Genomic insights into the role of Salmonella Typhi carriers in antimicrobial resistance and typhoid transmission in urban Kenya. PLoS One. 2025;20(5):e0321879. doi:10.1371/journal.pone.0321879

Sijad-ur-Rehman

Ahad

Muhammad

Bibi

Ullah

Antimicrobial sensitivity of Salmonella species in children: a single-centered study. Pak J Med Health Sci. 2023;17(5):162. doi:10.53350/pjmhs2023175162

Jamil

Ur Rehman

Rasheed

DF.

Emerging antimicrobial resistance in typhoid fever: challenges in diagnosis and treatment. J Med Health Sci Rev. 2025;2(2):525-532. doi:10.62019/a1v2gq57

World Health Organization. Global antimicrobial resistance and use surveillance system (GLASS) report 2022. Geneva: WHO. 2025. https://www.who.int/publications/i/item/9789240108127

World Health Organization. Antimicrobial resistance: fact sheet. Geneva: WHO. 2023. https://www.who.int/news-room/fact-sheets/detail/antimicrobial-resistance

10.

Hooda

Tanmoy

Nath

, et al. Outbreak of ceftriaxone-resistant Salmonella enterica serovar typhi, Bangladesh, 2024. Emerg Infect Dis. 2025;31(7):1460-1465. doi:10.3201/eid3107.241987

11.

da Silva

Tanmoy

Pragasam

, et al. The international and intercontinental spread and expansion of antimicrobial-resistant Salmonella Typhi: a genomic epidemiology study. Lancet Microbe. 2022;3(8):e567-e577. doi:10.1016/S2666-5247(22)00093-3

12.

Centers for Disease Control and Prevention. Antibiotic use and antimicrobial resistance: facts and statistics. CDC. 2024. https://www.cdc.gov/antibiotic-use/data-research/facts-stats/index.html

13.

World Health Organization. Typhoid fever–Pakistan. Disease Outbreak News. 27 December 2018. https://www.who.int/emergencies/disease-outbreak-news/item/27-december-2018-typhoid-pakistan-en

14.

Ali Shah

Nadeem

Syed

Fatima Abidi

Khan

Bano

Antimicrobial sensitivity pattern of Salmonella Typhi: emergence of resistant strains. Cureus. 2020;12(11):12. doi:10.7759/cureus.11778

15.

Pakistan Meteorological Department. Monthly climate reports 2024. Islamabad: PMD. 2024. http://www.pmd.gov.pk

16.

World Health Organization. WHO global water, sanitation and hygiene (WASH) monitoring and evidence: annual report 2024. WHO; 2025. https://www.who.int/teams/environment-climate-change-and-health/water-sanitation-and-health/monitoring-and-evidence/wash-monitoring

17.

World Health Organization. Drinking water. WHO fact sheet. WHO; 2023. https://www.who.int/news-room/fact-sheets/detail/drinking-water

18.

Clinical and Laboratory Standards Institute. Performance standards for antimicrobial susceptibility testing. 33rd ed. CLSI supplement M100. Clinical and Laboratory Standards Institute. 2023. https://clsi.org/about/news/clsi-publishes-m100-performance-standards-for-antimicrobial-susceptibility-testing-33rd-edition/

19.

Parry

Thieu

Dolecek

, et al. Clinically and microbiologically derived azithromycin susceptibility breakpoints for Salmonella enterica serovars typhi and Paratyphi A. Antimicrob Agents Chemother. 2015;59(5):2756-2764. doi:10.1128/AAC.04729-14

20.

Khan

Kurup

Vinod

Biswas

Pillai

Kumar

Reconsidering azithromycin disc diffusion interpretive criteria for Salmonellae in view of azithromycin MIC creep among typhoidal and nontyphoidal salmonella. J Lab Physicians. 2019;11(1):39-44. doi:10.4103/JLP.JLP_99_18

21.

World Health Organization. Guidelines for Drinking-water Quality: Fourth Edition Incorporating the First and Second Addenda. WHO; 2022. https://iris.who.int/handle/10665/352532

22.

Tajammul

Benson

Ahmed

VanDerslice

Tanner

WD.

Detection of Salmonella Typhi and blaCTX-M genes in drinking water, wastewater, and environmental biofilms in Sindh Province, Pakistan. PLoS Negl Trop Dis. 2025;19(4):e0012963.

23.

Perveen

Amar-Ul-Haque

Drinking water quality monitoring, assessment and management in Pakistan: a review. Heliyon. 2023;9(3):e13872.

24.

World Health Organization. Guidelines for Drinking-Water Quality: First Addendum to the Fourth Edition. WHO; 2017. https://iris.who.int/handle/10665/254636

25.

Jabeen

Saleem

Jahan

, et al. Molecular characterization of extensively drug resistant Salmonella enterica serovar Typhi clinical isolates from Lahore, Pakistan. Infect Drug Resist. 2023;16:2987-3001. doi:10.2147/IDR.S406253

26.

Yousafzai

Karim

Qureshi

, et al. Effectiveness of typhoid conjugate vaccine against culture-confirmed Salmonella enterica serotype Typhi in an extensively drug-resistant outbreak setting of Hyderabad, Pakistan: a cohort study. Lancet Glob Health. 2021;9(8):e1154-e1162. doi:10.1016/s2214-109x(21)00255-2

27.

Shahid

Mahesar

Ghouri

Noreen

A review of clinical profile, complications and antibiotic susceptibility pattern of extensively drug-resistant (XDR) Salmonella Typhi isolates in children in Karachi. BMC Infect Dis. 2021;21:900.

28.

Bilal

Khan

Rehman

Hameed

Yang

Antibiotic resistance in Pakistan: a systematic review of past decade. BMC Infect Dis. 2021;21:244. doi:10.1186/s12879-021-05906-1

29.

Irfan

Afridi

Fatima

Zafar

Muhammad Pervaiz

Farooq

Prevalence of extensively drug resistant Salmonella Typhi and its susceptibility against meropenem, tigecycline, fosfomycin and azithromycin among clinical isolates from a tertiary care hospital laboratory: susceptibility in extensively drug resistant

Salmonella Typhi. Pak J Health Sci. 2024;5(8):49-54. doi:10.54393/pjhs.v5i08.1787

30.

Hanif

Bai

Rehman

Memon

Ashfaq

Emerging trends of resistance of typhoid fever in paediatric population: a hospital based study. J Liaquat Univ Med Health Sci. 2021;20(01):21-25. doi:10.22442/jlumhs.2021.00764

31.

Tashfeen

Asif

Farooq

Haematological profile derangements in patients due to non-resistant and resistant typhoid fever. Pak Armed Forces Med J. 2021;71(5):1615-1618. doi:10.51253/pafmj.v71i5.4359

32.

Liaqat

Jalil

Liaqat

Zafar

Shah

Frequency of multi-drug resistant (MDR) and extended drug resistant (XDR) Salmonella Typhi in patients presenting with culture proven enteric fever at tertiary care hospital. J Med Health Sci Rev. 2025;2(2):e240604. doi:10.62019/2w3dca69

33.

Carey

Keddy

Dilley

Tipple

Parkhill

von Gottberg

, et al. Spontaneous emergence of azithromycin-resistant Salmonella Typhi in South Asia. Clin Infect Dis. 2021;72(5):e120. doi:10.1093/cid/ciaa1773

34.

Qureshi

Naveed

Yousafzai

, et al. Response of extensively drug resistant Salmonella Typhi to treatment with meropenem and azithromycin in Pakistan. PLoS Negl Trop Dis. 2020;14(10):e0008682. doi:10.1371/journal.pntd.0008682

35.

Tanmoy

Hooda

Sajib

MSI

, et al. Trends in antimicrobial resistance amongst Salmonella Typhi in Bangladesh: a 24-year retrospective observational study (1999-2022). PLoS Negl Trop Dis. 2024;18(10):e0012558. doi:10.1371/journal.pntd.0012558

36.

Herekar

Sarfaraz

Imran

Ghouri

Shahid

Mahesar

Clinical spectrum and outcomes of patients with different resistance patterns of Salmonella enterica. Pak J -med Sci. 2022;38(2):356-361. doi:10.12669/pjms.38.ICON-2022.5789

37.

Asghar

Khan

Séraphin

, et al. Exploring the antimicrobial resistance profile of Salmonella Typhi and its clinical burden. Antibiotics. 2024;13(8):765. doi:10.3390/antibiotics13080765

38.

Mumtaz

Irfan

Siraj

, et al. Whole-genome sequencing of extensively drug-resistant Salmonella enterica serovar typhi clinical isolates from the Peshawar region of Pakistan. J Infect Public Health. 2024;17:271-282. doi:10.1016/j.jiph.2023.12.002

39.

Tagg

Pinto

Hatcher

, et al. Azithromycin-resistant Salmonella enterica serovar typhi carrying mph(A) — first US cases identified via PulseNet. Lancet Microbe / J Clin Microbiol. 2024;39:69-72. doi:10.1016/j.jgar.2024.08.005

40.

Siddiqi

Rais

Zafar

, et al. Treatment of 21st-century typhoid fever in children with mono vs combination drug therapy: an open-label randomized comparative trial. Int J Endorsing Health Sci Res. 2021;9(3):384-394. doi:10.29052/IJEHSR.v9.i3.2021.384-394

41.

Wang

Jin

, et al. Extensively drug-resistant (XDR) Salmonella Typhi outbreak by waterborne infection - Beijing Municipality, China, January-February 2022. China CDC Weekly. 2022;4:254-258. doi:10.46234/ccdcw2022.062

42.

Buzilă

Dorneanu

Trofin

Sima

Iancu

LS.

Assessing Salmonella Typhi pathogenicity and prevention: the crucial role of vaccination in combating typhoid fever. Int J Mol Sci. 2025;26(9):3981. doi:10.3390/ijms26093981